4 5951913224361215540

Mariagrazia Stracquadanio · Lilliana Ciotta
Metabolic
Aspects of PCOS
Treatment with
Insulin Sensitizers
123
Metabolic Aspects of PCOS
Mariagrazia Stracquadanio • Lilliana Ciotta
Metabolic Aspects
of PCOS
Treatment with Insulin Sensitizers
Mariagrazia Stracquadanio Lilliana Ciotta
Obstetrics and Gynecological Pathology Obstetric and Gynecological Pathology
P.O. “S. Bambino”, University of Catania P.O. “S. Bambino”, University of Catania
Catania Catania
Italy Italy
ISBN 978-3-319-16759-6 ISBN 978-3-319-16760-2 (eBook)

DOI 10.1007/978-3-319-16760-2
Library of Congress Control Number: 2015938769
Springer Cham Heidelberg New York Dordrecht London

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Acknowledgments
A special thanks to “Oxford University” and “John Radcliffe Hospital – Cairns

Library” for allowing the collection of the extended bibliography used.
v
Contents
1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.1 PCOS Origins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.2 Definition and Epidemiology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2 Etiopathogenesis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
2.1 Genetics of PCOS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
2.2 PCOS Physiopathology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
2.3 Role of Insulin in the Pathogenesis of PCOS . . . . . . . . . . . . . . . . . . 12
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
3 Clinical Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
3.1 Endocrine Aspects of PCOS. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
3.1.1 Endocrine Pattern . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
3.1.2 Clinical Endocrine Features . . . . . . . . . . . . . . . . . . . . . . . . . 23
3.2 Metabolic Aspects of PCOS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
3.2.1 The Role of the Adipocyte in Linking PCOS
to Metabolic Syndrome . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27
3.2.2 The Role of Vitamin D in the Development
of Metabolic Syndrome in PCOS Women . . . . . . . . . . . . . . 30
3.2.3 Metabolic Syndrome and Associated Disorders . . . . . . . . . 31
3.2.4 Role of Insulin Resistance in Infertility
and Pregnancy Outcome. . . . . . . . . . . . . . . . . . . . . . . . . . . . 39
3.3 PCOS Phenotype in Different Ages . . . . . . . . . . . . . . . . . . . . . . . . . 43
3.3.1 Adolescence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
3.3.2 Fertile Period . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45
3.3.3 Premenopausal and Postmenopausal Period . . . . . . . . . . . . 45
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46
4 Psychological Implications of PCOS . . . . . . . . . . . . . . . . . . . . . . . . . . . 63
4.1 PCOS Symptoms and Psychological Correlation. . . . . . . . . . . . . . . 63
4.1.1 Obesity and Body Image . . . . . . . . . . . . . . . . . . . . . . . . . . . 63
4.1.2 Hirsutism . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64
4.1.3 Infertility and Sexual Life . . . . . . . . . . . . . . . . . . . . . . . . . . 64
vii
viii Contents
4.2 PCOS and Mental Disorders. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65

4.2.1 Mood Disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65
4.2.2 Anxiety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66
4.2.3 Eating Disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
5 Diagnosis and Assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
5.1 Differential Diagnosis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
5.2 Risk Factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
5.3 Clinical–Endocrine Features. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
5.3.1 Oligomenorrhea and Anovulation . . . . . . . . . . . . . . . . . . . . 72
5.3.2 Hirsutism . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
5.3.3 Acne . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
5.4 Endocrine Blood Tests . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74
5.5 Ultrasound Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76
5.6 Clinical–Metabolic Features. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78
5.7 Metabolic Blood Tests . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
5.7.1 Glucose Metabolism Assessment and Calculation
of Insulin Resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
5.7.2 Lipid and Hepatic Profile . . . . . . . . . . . . . . . . . . . . . . . . . . . 83
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84
6 PCOS Therapy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
6.1 Diet and Exercise . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
6.1.1 PCOS Dietary Recommendations . . . . . . . . . . . . . . . . . . . . 90
6.1.2 Glycemic Index (GI) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 92
6.1.3 Glycemic Load (GL) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 96
6.1.4 Insulin Index. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97
6.1.5 Exercise . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97
6.2 Insulin-Sensitizing Agents and Statins . . . . . . . . . . . . . . . . . . . . . . . 98
6.2.1 Thiazolidinediones . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
6.2.2 Metformin. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
6.2.3 Statins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112
6.3 Inositol and Other Supplements . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113
6.3.1 Inositol and Its Isomers . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113
6.3.2 Antioxidants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
6.3.3 Vitamin D . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
6.3.4 Glucomannan . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
Erratum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E1
Introduction
1
1.1 PCOS Origins
Polycystic ovary syndrome (PCOS) is not a recent disorder, but it seems to be very
old. Going back to the early history, Hippocrates and Soranus of Ephesus reported
that “many women with masculine and robust aspect don’t menstruate and they
don’t become pregnant” [1, 2].
Some authors suggest that the origin of PCOS began in Paleolithic communities,
in which environmental stressful factors favored the survival of the “thrifty geno-
type”: it was represented by males and females with the greatest capacity for energy
storage necessary to face fasting periods [3, 4].
Moreover, subfertility among nomadic hunters gave some benefits: women could
care only for one child, and a lower parity may have reduced the death rates of these
women and the risk of progeny abandonment, as delivery-related complications
were a major cause of mortality in reproductive-age women.
On the other hand, during the Neolithic revolution, when communities started to
be sedentary, PCOS genotype may have survived because of its robustness, with
some gene variants over the years, as well shown by the heterogeneity of PCOS
phenotypes and genotypes.
Moreover, since the eighteenth century it was noticed that signs of hyperandrogen-
ism were associated with metabolic abnormalities, such as increased visceral fat [5].
Jean Vague, physician and professor at University of Marseille, introduced the
term “android obesity” to define the abdominal fat accumulation, which is the
typical male pattern of body fat distribution, associated with increased diabetes
and cardiovascular risk [6]. Later, it was realized that lots of hyperandrogenic
women were obese with increased visceral fat; they had increased insulin response
during OGTT (oral glucose tolerance test), and they presented with acanthosis
nigricans [7, 8]: all of these are signs of insulin resistance, and these observations
were the starting point for the study of the association between insulin resistance
and PCOS.
© Springer International Publishing Switzerland 2015 1

M. Stracquadanio, L. Ciotta, Metabolic Aspects of PCOS: Treatment with Insulin
Sensitizers, DOI 10.1007/978-3-319-16760-2_1
2 1 Introduction
1.2 Definition and Epidemiology
Polycystic ovary syndrome (PCOS) is a heterogeneous endocrine and metabolic

disorder, characterized by chronic anovulation/oligomenorrhea, hyperandrogenism,
and insulin resistance.
In accordance with the most used guidelines drawn in Rotterdam in 2003 by
ESHRE/ASRM (European Society for Human Reproduction and Embryology/
American Society for Reproductive Medicine) [9], PCOS diagnosis can be raised
only after the exclusion of other known causes of hyperandrogenism and amenor-
rhea (hyperprolactinemia, non-classic congenital adrenal 21-hydroxylase defi-
ciency, thyroid dysfunction, androgen-secreting neoplasm, Cushing’s syndrome)
and when there are at least two of the three following parameters:
1. Oligomenorrhea or anovulatory cycles with menstrual irregularities

2. Elevated levels of circulating androgens or clinical manifestations of
hyperandrogenism
3. Ultrasound evidence of micropolycystic ovaries
The previous criteria processed by NIH (National Institute of Health) in 1992

included both:
1. Clinical and/or biochemical hyperandrogenism

2. Menstrual dysfunction
Τhe most recent (2006) AES (Androgen Excess Society) criteria [10] includes all
of the following conditions:
1. Clinical and/or biochemical hyperandrogenism

2. Ovarian dysfunction and/or micropolycystic ovaries
Approximately 85–90 % of women with oligomenorrhea have PCOS, while

30–40 % of women with amenorrhea suffer from PCOS [11].
More than 80 % of women showing symptoms of androgen excess have PCOS
[12]. Roughly 90–95 % of anovulatory women presenting to infertility clinics have
PCOS. The syndrome is present in approximately 5–10 % of reproductive-age
women, and it is considered the most frequent endocrine abnormality in females.
As there are significant variations in the clinical appearance of PCOS, its preva-
lence may be different among populations.
It is 4.8 and 8 % in white and black women in southeastern United States [13],
6.8 % in white women in Greece [14], 6.5 % in white women in Spain [15], 6.3 %
in South Asian in Sri Lanka [16], and 5 % in Thai women [17].
Some groups have showed that the frequency of PCOS varies depending on the
diagnostic criteria used: for example, the prevalence estimations using the
Rotterdam criteria are two to three times greater than those achieved using NIH
criteria [18–21].
References 3
For example, in China, Chen et al. reported that the prevalence in South Chinese
population was 2.2 % based on NIH criteria [22], while in a cross-sectional epide-
miologic investigation conducted in ten provinces of China, the prevalence of PCOS
using the Rotterdam criteria was 5.61 % [23]. The difference may depend on the
size of the sample and ethnicity too. The Rotterdam-PCOS group appeared to be
more than 1.5–2 times larger than the group classified as NIH-PCOS [24].
References
1. Hanson AE (1975) Hippocrates: disease of women 1. Signs (Chic) 1:567–584
2. Temkin O (1991) Soranus’ gynecology. The Johns Hopkins University Press, Baltimore
3. Chakravarthy MV, Booth FW (2004) Eating, exercise, and “thrifty” genotypes: connecting the
dots toward an evolutionary understanding of modern chronic diseases. J Appl Physiol
96:3–10
4. Neel JV (1962) Diabetes mellitus: a “thrifty” genotype rendered detrimental by “progress”?
Am J Hum Genet 14:353–362
5. Morgagni J (1765) De Sedibus et Causis Morborum per Anatomen Indagata (The seats and
causes of diseases investigated by anatomy), 2nd edn, Tomus primus. Sumptibus
Remondinianis, Patavii
6. Vague J (1947) La differentiation sexuelle Facteur determinant des formes de l’obesite’. Presse
Med 55:339–340
7. Dunaif A, Hoffman AR, Scully RE et al (1985) Clinical, biochemical, and ovarian morpho-
logic features in women with acanthosis nigricans and masculinization. Obstet Gynecol
66:545–552
8. Flier JS, Eastman RC, Minaker KL et al (1985) Acanthosis nigricans in obese women with
hyperandrogenism. Characterization of an insulin-resistant state distinct from the Type A and
B syndromes. Diabetes 34:101–107
9. Rotterdam ESHRE/ASRM-Sponsored PCOS Consensus Workshop Group (2004) Revised
(2003) consensus on diagnostic criteria and long- term health risks related to polycystic ovary
syndrome. Fertil Steril 81:19–25
10. Azziz R, Carmina E, DeWailly D et al (2006) Position statement: criteria for defining polycys-
tic ovary syndrome as a predominantly hyperandrogenic syndrome: an androgen excess soci-
ety guideline. J Clin Endocrinol Metab 91:4237–4245
11. Hart R (2007) Definitions, prevalence and symptoms of polycystic ovaries and the polycystic
ovary syndrome. In: Allahbadia GN, Agrawal R (eds) Polycystic ovary syndrome. Anshan,
Ltd, Kent, pp 15–26
12. Azziz R, Sanchez L, Knochenhauer ES et al (2004) Androgen excess in women: experience
with over 1,000 consecutive patients. J Clin Endocrinol Metab 89(2):453–462
13. Azziz R, Woods KS, Reyna R et al (2004) The prevalence and features of the polycystic ovary
syndrome in an unselected population. J Clin Endocrinol Metab 89(6):2745–2749
14. Diamanti-Kandarakis E, Kouli CR, Bergiele AT et al (1999) A survey of the polycystic ovary
syndrome in the Greek island of Lesbos: hormonal and metabolic profile. J Clin Endocrinol
Metab 84(11):4006–4011
15. Asuncion M, Calvo RM, San MJ (2000) A prospective study of the prevalence of the polycys-
tic ovary syndrome in unselected Caucasian women from Spain. J Clin Endocrinol Metab
85(7):2434–2438
16. Kumarapeli V, Seneviratne RA, Wijeyaratne CN et al (2008) A simple screening approach for
assessing community prevalence and phenotype of polycystic ovary syndrome in a semi-urban
population in Sri Lanka. Am J Epidemiol 168(3):321–328
17. Vutyavanich T, Khaniyao V, Wongtra-Ngan S (2007) Clinical, endocrine and ultrasonographic
features of polycystic ovary syndrome in Thai women. J Obstet Gynaecol Res 33(5):677–680
4 1 Introduction
18. March WA, Moore VM, Willson KJ et al (2010) The prevalence of polycystic ovary syndrome
in a community sample assessed under contrasting diagnostic criteria. Hum Reprod
25(2):544–551
19. Mehrabian F, Khani B, Kelishadi R, Ghanbari E (2011) The prevalence of polycystic ovary
syndrome in Iranian women based on different diagnostic criteria. Endokrynol Pol
62(3):238–242
20. Tehrani FR, Simbar M, Tohidi M et al (2011) The prevalence of polycystic ovary syndrome in
a community sample of Iranian population: Iranian PCOS prevalence study. Reprod Biol
Endocrinol 9:39
21. Yildiz BO, Bozdag G, Yapici Z et al (2012) Prevalence, phenotype and cardiometabolic risk of
polycystic ovary syndrome under different diagnostic criteria. Hum Reprod 27(10):
3067–3073
22. Chen X, Yang D, Mo Y et al (2008) Prevalence of polycystic ovary syndrome in unselected
women from southern China. Eur J Obstet Gynecol Reprod Biol 139(1):59–64
23. Zhao Y, Qiao J (2013) Ethnic differences in the phenotypic expression of polycystic ovary
syndrome. Steroids 78:755–760
24. Broekmans FJ, Knauff EA, Valkenburg O et al (2006) PCOS according to the Rotterdam con-
sensus criteria: change in prevalence among WHO-II anovulation and association with meta-
bolic factors. BJOG 113(10):1210–1217
Etiopathogenesis
2
2.1 Genetics of PCOS
PCOS is a multifactorial polygenic disease (interaction between several genetic and

environmental factors), with a heritability of ∼70 %. It is intrinsically difficult to
study by a genetic point of view, and most of the current literature (>70 studies
based on the candidate gene approach) is inconclusive, with many studies resulting
inconsistent, controversial, and without a clear consensus [1].
In the first studies on the genetic basis of PCOS, both maternal and paternal pat-
terns of inheritance are suggested: the incidence of oligomenorrhea and polycystic
ovaries was found to be increased in first-degree relatives of PCOS patients com-
pared with controls, and males in those families had increased hairiness according
to the questionnaire, suggesting an autosomal dominant pattern of inheritance [2].
Recently, the inheritance was confirmed by some authors who found that PCOS
was present in 35 % of the mothers and 40 % of the sisters of PCOS patients [3].
Moreover, increased incidence of insulin resistance in the fathers and brothers of
PCOS women [4] has been considered as the “male phenotype” in PCOS families.
The genes involved in the pathogenesis of hyperandrogenism are expressed in a
variable way depending on the factors predominating in every different ethnic popu-
lations; this explains the phenotypic variability of hyperandrogenic disorders.
Another theory is that the features of PCOS families result from nongenetic inheri-
tance, and they are related to environmental factors that are present only in the
affected families.
Ibanez hypothesized that some insults during pregnancy may induce to intrauter-
ine growth retardation, which probably induces a “thrifty phenotype” in small for
gestational age babies. These have a high risk of suffering from insulin resistance,
which may result in hypertension, glucose intolerance, adrenal axis hyperactivity
with relative cortisol excess, functional hyperandrogenism, and PCOS later in life,
especially if they are exposed to environmental factors such as a sedentary lifestyle
and a diet rich in saturated fat [5].

Sensitizers, DOI 10.1007/978-3-319-16760-2_2
6 2 Etiopathogenesis
These environmental factors may cluster in certain families because exercising

and dieting are greatly influenced by parental lifestyle. The metabolic abnormalities
of the “thrifty phenotype” can induce additional insult to the pregnancies of these
SGA (small for gestational age) and PCOS women, and these defects might be trans-
mitted to another generation without the participation of any genetic abnormality.
On the other hand, if small for gestational age babies have healthy habits, insulin
resistance and its consequences might be improved, and theoretically, their fetuses
will not be exposed to a hostile metabolic environment during pregnancy, prevent-
ing nongenetic inheritance of these conditions. However, intrauterine growth restric-
tion might be influenced by genetic variants as well, and the most likely scenario is
represented by an interaction between predisposing genetic abnormalities and unfa-
vorable environmental conditions [6].
Thus, even if several studies conducted in families of women with PCOS have
demonstrated the genetic basis of the syndrome, nowadays a genetic pattern cer-
tainly involved in PCOS predisposition has not been identified.
Most studies have included different kinds of genes: those related to androgen
biosynthesis and action and their regulation, genes involved in insulin resistance
and associated disorders, and also genes involved in chronic inflammation and
atherosclerosis.
Among the genes involved in androgen biosynthesis, there are:
• CYP17: This gene encodes the P450c17α enzyme, which catalyzes the conver-
sion of pregnenolone and progesterone into, respectively, 17-hydroxypregnenolone
and 17-hydroxyprogesterone and of these steroids into dehydroepiandrosterone
and androstenedione. In the past, the hyperactivity of this enzyme was correlated
to hyperandrogenism [7].
CYP17 is located in chromosome 10q24.3, and its promoter encloses a T/C SNP
at 34 bp from the transcription start that might regulate enzyme activity. Some
studies hypothesized that this polymorphism was associated with polycystic ova-
ries morphology on ultrasound [8, 9], and it was found that PCOS patients homo-
zygous for C alleles of this polymorphism showed increased serum testosterone
levels [10, 11].
On the contrary, other studies suggested that this is a polymorphism without
functional consequences for the development of polycystic ovaries and hyperan-
drogenism [12–14]. Besides, no significant evidence for linkage or association
was found in a family-based genome study [15].
• CYP11A: This gene is located at 15q24 and encodes the cholesterol side chain
cleavage enzyme, important for the conversion of cholesterol into progesterone,
which is the first step in adrenal and ovarian steroidogenesis. A VNTR polymor-
phism, consisting in repeats of a (tttta)n pentanucleotide at −528 bp from the
ATG start of translation site in the CYP11A promoter, might play a role in the
pathogenesis of PCOS [16].
Some studies confirmed its association with polycystic ovaries and hirsute
women [16, 17], while other studies did not demonstrate linkage with the
CYP11A locus in PCOS patients or association of CYP11A VNTR alleles with
2.1 Genetics of PCOS 7
hyperandrogenism [18]. Moreover, recent experiments involving a large number

of subjects concluded that the existence of associations between CYP11A pro-
moter variation and androgen-related phenotypes had been considerably overes-
timated in previous studies [19].
• CYP19: This gene encodes aromatase, which converts androgens in estrogens.
This enzyme maybe has a decreased activity in granulosa cells and follicles of
PCOS women, and the consequent androgen excess might contribute to abnor-
mal follicle development [20, 21]. On the contrary, no evidence for linkage of
CYP19 with PCOS was found in other English and American studies [15, 16].
• LH Gene: LH hypersecretion is present in almost 50 % of PCOS women, and
two mutations, Trp8Arg and Ile15Thr, could be the cause of an abnormal LH β
molecule [22]. The first PCOS GWAS (genome-wide association studies) identi-
fied LH/choriogonadotropin receptor (LHCGR) as a susceptibility gene for
PCOS: the interaction of LHCGR and its ligand, LH, plays a fundamental role in
the folliculogenesis of mammals. A study suggested that LHCGR might partici-
pate in the physiopathology of PCOS by deviations in the methylation statuses of
its promoter CpG sites, a hypomethylation in particular [23].
• SHBG Genes: Sex hormone-binding globulin (SHBG) controls the admission of
testosterone and estradiol to target tissues.
Decreased SHBG is an important feature of hyperandrogenic women, causing
increased tissue androgen availability [24].
Recently, an association between a (TAAAA)n polymorphism in the promoter of
the SHBG gene and PCOS has been reported. Longer alleles (more than eight
repeats) were frequent in Greek PCOS patients, while non-hyperandrogenic
women presented with a higher frequency of shorter alleles [25].
The second group of genes includes those involved in insulin resistance and met-
abolic disorders, which are:
• INSR (Insulin Receptor Gene): Insulin resistance represents the major metabolic
aspect of PCOS. INSR contains several polymorphisms, but most of them are
silent or are located in intronic regions and are present with similar frequencies
in patients with polycystic ovaries and hyperandrogenism and in controls [26].
Polymorphism in exon 17 of the tyrosine kinase domain is the only one found,
but it was not associated to insulin resistance [27]. On the other hand, it was
found that a C/T SNP at the tyrosine kinase domain of INSR is associated with
PCOS, but further studies are needed to confirm it [6].
• INS: Pancreatic β-cell dysfunction in PCOS women seems to have a genetic ori-
gin as well. It was found that women with menstrual irregularities and/or hirsut-
ism and polycystic ovaries, who were homozygous for class III alleles, were
more frequently anovulatory and had increased BMI and fasting insulin com-
pared with women homozygous for class I alleles. Paternal transmission of class
III alleles from heterozygous fathers to anovulatory PCOS patients is more fre-
quent than maternal transmission of the allele [28–30], and in addition, class III
alleles predisposed these patients to both PCOS and type 2 diabetes mellitus.
However, other studies were not able to prove this [31, 32], and unluckily the
INS locus was not associated with PCOS in an American linkage study on PCOS
patients [15].
• Insulin Growth Factor System Genes: IGFs, their receptors, binding proteins, and
proteases are important for the normal development of the ovary [33].
They are peptide hormones secreted having important functions such as
mediation of growth hormone action, stimulation of growth of cultured cells,
stimulation of the action of insulin, and involvement in development and
growth. IGFs stimulate ovarian cellular mitosis and steroidogenesis, inhibit
apoptosis, and might be related to the development of functional hyperan-
drogenism and PCOS [34].
In particular, IGF-2 stimulates adrenal and ovarian androgen secretion: the
increased frequency of homozygosis for these alleles could contribute to hyper-
androgenism in PCOS patients [35].
• Peroxisome Proliferator-Activated Receptor-γ (PPAR-γ): They are members of
the nuclear receptor superfamily of ligand-activated transcription factors [36].
These genes are involved in adipocyte differentiation, lipid and glucose metabo-
lism, and atherosclerosis [37]. The human PPAR-γ gene is composed of nine
exons; recent studies have indicated that the modified Ala12 allele is involved in
increased insulin sensitivity by enhanced suppression of lipid oxidation, enabling
more efficient glucose disposal [38].
• Calpain-10: It is an enzyme that has an important role in insulin secretion and
action [39]. The 112/121 haplotype combination of the University of Chicago
single nucleotide polymorphisms (UCSNP)-43, UCSNP-19, and UCSNP-63 in
the gene encoding calpain-10, located at 2q37.3, has been reported to increase
the risk for diabetes [40]. Some authors found no association between this hap-
lotype and PCOS patients [41, 42], while recently a Spanish study reported an
association between PCOS and USCNP-44 [43, 44].
More recently, genes encoding inflammatory cytokines have been identified as

target genes for PCOS, as pro-inflammatory genotypes and phenotypes are also
associated with obesity, insulin resistance, type 2 diabetes, PCOS, and increased
cardiovascular risk.
• Paraoxonase (PON1): The PON1 gene is mainly expressed in the liver and
encodes for serum PON1, which is an antioxidant high-density lipoprotein-
associated enzyme. Liver PON1 mRNA expression is influenced by genetic and
environmental factors, and both androgens and pro-inflammatory mediators
decrease liver PON1 expression [45].
Homozygosis for T alleles of the −108C/T polymorphism in PON1 was more
frequent in PCOS patients compared with non-hyperandrogenic women. Patients
homozygous for −108T alleles of PON1 had increased hirsutism scores, total
testosterone, and free testosterone and androstenedione levels related to those
carriers of −108C alleles [35]. Nowadays, it is well known that oxidative stress
2.1 Genetics of PCOS 9
may damage insulin action. Indeed, reduced serum PON1 activity might
contribute to the insulin resistance of PCOS patients [46].
• TNF-α: In vitro, this growth factor stimulates proliferation and steroidogenesis
in theca cells and helps insulin and IGF-1 to exert their effects on the ovary [47].
Nine polymorphisms in the TNF-α gene were studied (−1196C/T, −1125G/C,
−1031T/C, −863C/A, −857C/T, −316G/A, −308G/A, −238G/A, and −163G/A),
but no differences between patients and controls were found: only lean hyperan-
drogenic patients showed increased serum TNF-α levels [48]. This finding might
imply that TNF-α gene does not have a major role in PCOS etiology but could be
a modifying factor for phenotypic features [6].
• TNFR2 Gene (TNFRSF1B): TNFR2 mediates most of the metabolic effects of TNF-α
[49]. The 196Arg allele of the Met196Arg (676T/G) polymorphism in exon 6 of this
gene was more frequent in patients with PCOS compared with healthy controls, and it
was hypothesized that it was responsible for modulating TNF-α in target tissues [50].
• IL-6: This cytokine seems to be implicated in insulin resistance mechanism, and
increased levels were found in peritoneal fluid of anovulatory PCOS patients,
suggesting a role in the pathogenesis of hyperandrogenic disorders [51]. Common
polymorphisms in both subunits of the IL-6 receptor were studied, and the
Arg148 allele of the Gly148Arg polymorphism in the gp130 gene was more fre-
quent in controls compared with hyperandrogenic patients: control women had
lower 11-deoxycortisol and 17-hydroxyprogesterone concentrations and a sig-
nificant decrease in free testosterone levels, suggesting that this polymorphism
might have a protective effect against androgen excess [52].
Moreover, there are also other genetic structural variations that regulate gene and
phenotype expression, such as telomeres: they are at the ends of eukaryotic chromo-
somes and are specialized chromatin structures composed of highly conserved tan-
dem hexameric nucleotide repeats—TTAGGG—that extend for several kilobases
[53]. Telomeres shorten progressively with each cell division, and their length is
largely inherited and modulated by a variety of genetic and environmental factors
[54]. Short telomeres can cause chromosomal instability, and this could be the rea-
son of genetic mutations and chromosome abnormalities.
There is a correlation between oxidative stress and PCOS and between oxidative
stress and telomere length. For this reason, it has been hypothesized that telomere
length plays an important role in the pathophysiology of PCOS.
In a Chinese study, the mean telomere length was measured in a large cohort of
PCOS patients and controls, and the association between telomere length and this
endocrine–metabolic disease was analyzed. A significant reduction of telomere
length was observed in PCOS patients compared with healthy controls. Individuals
with the shorter telomere length had significantly higher disease risk than those with
the longest telomere length, after adjustment for age. One possible mechanism for
the shortened telomeres in PCOS patients is that some etiological factors of PCOS,
such as androgen excess, abdominal adiposity, insulin resistance, and obesity, could
contribute to raised oxidative stress that leads to telomere shortening. This could
represent a negative feedback cycle in which shortened telomeres, in turn, affect

endocrine-, metabolic-, or reproductive-related gene expression and worsen the
abnormal metabolic phenotypes of the disease [55].
2.2 PCOS Physiopathology
It has been shown that polycystic ovary presents a greater number of small antral
follicles (2–9 mm in diameter) than the normal ovary. This morphological scenario
could be the consequence of a potential dysregulation of the recruitment mechanism
of primordial follicles that, on the contrary, are present in physiological number.
On the other hand, the final pathway of follicular growth, which is gonadotropin
dependent, is blocked in the majority of PCOS patients, and it is the basis of anovu-
lation and oligo-/amenorrhea.
In a normal cycle, only the dominant follicle responds to LH action when it
reaches 10 mm in diameter. In PCOS patients, the response to LH occurs inappro-
priately in smaller follicles; a large number of antral follicles reach a terminal dif-
ferentiation before the appropriate time, producing a larger amount of steroids and
inhibin B that have a negative feedback on the production of FSH: the result is the
arrest of follicular growth.
As underlined before, the etiology of this syndrome is still partly unknown, but
it is likely to be multifactorial. The most significant theories are explained below:
• Exaggerated Adrenarche: It is possible that PCOS might be established and

maintained in response to an abnormal adrenal hypersecretion of androgens due
to congenital adrenal enzyme deficiency [56].
Yen suggested an etiopathogenetic model, which provides, in response to a stress
condition, a transient adrenal androgen hypersecretion, triggering an abnormal
pattern of the pituitary gonadotropins’ pulsatility. As puberty progresses, the
adrenal cortex is replaced by ovaries in maintaining the hypersecretion of
androgens.
Finally, the increase in ovarian androgen level changes adrenal specific enzyme
activities involved in the process of steroidogenesis [57].
• Abnormal Secretion of Gonadotropins: The high levels of LH in women with
PCOS are due to greater amplitude of the peaks of this hormone and its
increased frequency of pulsatility; on the contrary, the average concentration
of FSH is mostly decreased. The high levels of LH are not caused by an
inability of the hypothalamic-pituitary axis to respond to the negative feedback
exerted by estrogen, but it might be caused by the high pituitary sensitivity to
LH-RH. The chronically elevated and acyclic levels of estrogens in PCOS
patients may, in turn, increase both the basal levels of LH and LH response to
GnRH.
Moreover, an elevated endogenous opioid tone might cause an exceeding GnRH
release with a following abnormal LH pulsatility, causing increased level of LH-
dependent ovarian androgens [58].
2.2 PCOS Physiopathology 11
• Rosenfield’s Hypothesis: Rosenfield suggested that PCOS results from a hyper-

activity of cytochrome P450c17α in the ovarian theca cells. This enzymatic
complex binds progesterone and converts it sequentially in 17-hydroxyproges-
terone (via a 17α-hydroxylation) and androstenedione (via a C-17,20-lytic
activity). The steroidogenetic route particularly involved in the ovary is the Δ-4
pathway.
Moreover, at adrenal level, cytochrome P450c17α forms 17-ketosteroids, espe-
cially using the Δ-5 steroidogenetic pathway, and it creates more dehydroepian-
drosterone than androstenedione. An abnormal regulation of this enzyme activity,
therefore, both at ovarian and adrenal levels, could explain the androgenic hyper-
function of both glands, as occurs in PCOS.
Rosenfield proposed three hypotheses to explain the hyperactivity of this enzy-
matic complex:
1. The hyperactivity is the result of an increased LH release, characteristic of
PCOS.
2. The action of LH on theca cells is increased and amplified, even in the pres-
ence of normal levels of LH.
3. In PCOS women, ovarian theca cells might work in a way more similar to the
testicular Leydig cells rather than those of the normal ovarian theca
cells, because in the ovarian stroma, some “aberrant” cells (called “lipid cell
rest”) in which an abnormal steroidogenic secretory pattern is established,
could exist.
However, according to Rosenfield, the hyperactivity of cytochrome P45017α
cannot be the unique cause of PCOS, but it is part of a more complex etiopatho-
genetic model [59].
• Hyperestronemia: Increased levels of estrone (E1), characteristic of polycystic
ovary, are able to modify the normal patterns of gonadotropins’ pulsatility. This
high E1 level in PCOS women is generally the result of an increased ovarian
production of androstenedione (A) and its conversion into E1 by a specific FSH-
dependent enzyme called aromatase.
This enzyme is present in adipose tissue; thus, overweight or obese women have
a greater amount of enzyme and, consequently, more estrone compared to
normal-weight subjects. Alternatively, estrone levels might be increased in lean
women with high production of androstenedione. The part of testosterone con-
verted to estrone is very poor, and probably for this reason, the hypertestostero-
nemia per se is not able to affect significantly the gonadotropins’ pulsatility [60].
Many women with PCOS are overweight or obese: these conditions are usually
associated with low levels of SHBG. This globulin binds both testosterone and
estradiol: thus, in conditions in which SHBG is reduced, consequently, estradiol
free fraction (the most biologically active) is increased.
This condition causes a negative impact on the release of FSH with consequent
alteration of folliculogenesis process and increased release of LH, which is fol-
lowed by an increased ovarian androgen synthesis.
In addition, the increase of androgen plasma levels contributes to the reduction
of hepatic biosynthesis of SHBG.
• Hyperinsulinemia: High level of insulin accelerates the development of granu-

losa cell LH responsiveness by amplifying the induction of LH receptors, and
thus, it induces a block of follicular growth with multiple small follicle
formation.
The role of insulin is properly discussed in Sect. 2.3.
2.3 Role of Insulin in the Pathogenesis of PCOS
Insulin controls glucose homeostasis stimulating glucose uptake by tissues that are
responsive to insulin (adipocytes, skeletal and cardiac muscle) and by suppressing
hepatic glucose production [61, 62]. In addition, insulin decreases free fatty acid
levels by suppressing lipolysis [63], and it promotes cell growth and differentiation
[64].
“Insulin resistance” is defined as “a decreased ability of insulin to mediate its
metabolic actions on glucose uptake, glucose production and lipolysis, requiring
increased amounts of insulin to achieve its proper metabolic action.”
In fact, increased circulating insulin levels characterize insulin resistance if pan-
creatic β-cells are functionally intact [65].
Insulin exerts its function by binding to its cell surface receptor; ligand binding
induces auto-phosphorylation of the insulin receptor on specific tyrosine residues,
and this actives its intrinsic kinase activity, while serine phosphorylation inhibits it
[66, 67].
The tyrosine-phosphorylated insulin receptor phosphorylates, in turn, intracel-
lular substrates, such as IRS 1–4, Shc, and APS to start signal transduction
[68–70].
Insulin stimulates glucose uptake by translocating GLUT-4 (the insulin-
responsive glucose transporter) from intracellular vesicles to the cell surface [68,
70].
This pathway is mediated by activation of PI3K and Akt/PKB, which also leads
to serine phosphorylation of GSK3 (glycogen synthase kinase 3), resulting in inhi-
bition of its kinase activity: this inhibition causes dephosphorylation of glycogen
synthase, increasing glycogen synthesis, and also dephosphorylation of eIF2B
which increase protein synthesis [64, 70].
Insulin has also an important mitogenic action: it stimulates cell growth and dif-
ferentiation through the MAPK-ERK pathway [64].
This route is activated by insulin receptor-mediated phosphorylation of Shc or
IRS, which stimulates a cascade of serine/threonine kinase resulting in stimulation
of MAP kinase and MAPK-ERK 1/2. ERK 1/2 translocates to the nucleus and phos-
phorylates transcription factors to start cell growth and differentiation.
This mitogenic pathway can be altered without affecting the metabolic actions of
insulin and vice versa [64].
Insulin signaling can be terminated by dephosphorylation of the receptor by
tyrosine phosphatases; in addition, serine phosphorylation (mediated by serine
2.3 Role of Insulin in the Pathogenesis of PCOS 13
kinases) of the insulin receptor and its substrates can decrease insulin signaling as
well [64, 70].
There is a post-binding defect in insulin signaling in PCOS women, resulting in
marked insulin sensitivity decrease. The defect is due to serine phosphorylation of
the insulin receptor and IRS-1 secondary to intracellular serine kinases. This causes
a decreased activation of PI3K mediated by insulin and resistance to the metabolic
actions of insulin too [71].
Moreover, supporting this theory, it was shown that serine kinase inhibitors cor-
rected the phosphorylation defect, underlining the role of a serine kinase extrinsic to
the insulin receptor as the cause of decreased receptor auto-phosphorylation. This
defect in the first phases of the insulin signaling pathway is present in adipocytes
[72, 73] and skeletal muscle [71, 74], which are the most important target tissues for
glucose uptake stimulated by insulin.
Even if obesity is the major contributing factor for insulin resistance in PCOS
women, dysfunction in post-receptor mechanism action could be a good explana-
tion for insulin-resistant lean/normal-weight PCOS women.
Moreover, ovarian granulosa lutein cells could be considered a selective target
tissue too, in which insulin resistance is selective, affecting only the metabolic but
not the mitogenic action of insulin.
In addition, it has been taken into consideration the crucial role of serum fetuin-α
in the inhibition of insulin receptor tyrosine kinase activity [75].
It is a carrier protein like albumin, and a recent study has shown that fetuin-α
serum levels are higher in PCOS women, having probably a role in triggering the
processes that lead to insulin resistance and androgen excess in PCOS [76].
Furthermore, it was supposed that hyperinsulinemia might be the result of a
decreased insulin clearance or of an increased insulin secretion [77, 78].
Insulin clearance is receptor mediated; thus, insulin-resistant patients are sup-
posed to have a decreased clearance because of intrinsic or acquired decrease in
receptor number and/or function [78, 79].
Some authors have shown that fasting hyperinsulinemia in PCOS women is the
result of a combination of increased basal insulin secretion and decreased hepatic
insulin clearance [80, 81].
Lots of evidence demonstrate a direct insulin action on ovarian steroidogenesis
and the importance of the insulin signaling pathway in the control of ovulation.
Obviously, insulin receptors are present both in normal and polycystic ovary syn-
drome women. IGF-1 (insulin growth factor 1) is synthetized by the ovary, and its
receptor is a tyrosine kinase with few structural and functional homologies with the
insulin receptor [82, 83].
Insulin can bind to the IGF-1 receptor activating it, and IGF-1 can bind to and
activate the insulin receptor [84, 85].
The affinity of the IGF-1 receptor for insulin is less than it is for IGF-1 and vice
versa; despite this, the two receptors can assemble together to form a hybrid tetramer,
which is able to bind insulin and IGF-1 in the same way. Therefore, some insulin action
on the ovary may be mediated by IGF-1 or hybrid insulin–IGF-1 receptor [86, 87].
Some studies have shown that insulin action on steroidogenesis in granulosa and
theca cells is mediated via insulin receptor, both in normal and PCOS women [88, 89].
Moreover, in PCOS granulosa cells, increased insulin levels might cause premature
LH receptor expression in small follicles, leading to premature granulosa terminal
differentiation and the arrest of follicular growth, which is the basis for anovulation.
In normal theca cells, insulin and LH activate 17α-hydroxylase activity of
P450c17α, a crucial enzyme in the regulation of androgen biosynthesis encoded by
CYP17, via PI3K signaling; inhibition of MAPK-ERK1/2 signaling has no effect on
17α-hydroxylase activity [89].
It seems that PCOS theca cells are more responsive to the androgen-stimulating
insulin actions rather than normal controls [90].
Physiologically, insulin acts as a “co-gonadotropin” to increase androgen syn-
thesis induced by LH in theca cells [91–93] and to boost FSH-mediated estrogen
production and LH-induced luteinization in granulosa cells [94].
Furthermore, human studies have demonstrated that insulin can increase circu-
lating androgen levels in PCOS women: insulin infusion during euglycemic clamp
studies increased androgen level without altering gonadotropin secretion, suggest-
ing a direct effect on steroidogenesis [95, 96].
Suppressing insulin levels leads to decreased testosterone levels in women with
PCOS, while there is an increase in SHBG levels [97–99]. Thus, low insulinemia is
the basis for normal to low androgen production in the ovary and for increasing
SHBG levels which leads to low circulating active androgen levels too.
The correlation between PCOS, insulin, hyperandrogenism, and ovarian dys-
function is well exemplified in Fig. 2.1.
Moreover, insulin action on adrenal androgen production and gonadotropin
secretion is not yet well known. Lowering insulin levels with ISD (insulin-sensitizing
drugs) resulted in DHEAS decrease in PCOS women [100, 101]; other studies also
suggested that insulin resistance and consequent hyperinsulinemia cause a reduced
pituitary sensitivity to GnRH, contributing to anovulatory syndrome [102, 103].
According to all these findings, insulin could be defined as a “reproductive hor-
mone” as well.
The central paradox in the pathophysiologic association between hyperinsu-
linemia and hyperandrogenemia in PCOS is that the ovary remains sensitive to insu-
lin activity and consequent androgen production, despite a systemic insulin
resistance: it is the so-called selective insulin resistance theory [104].
On the other hand, androgens can produce insulin resistance by direct effects on
the skeletal muscle and adipose tissue insulin action, by altering adipokine secre-
tion, and by increasing visceral adiposity, even if these effects on insulin actions are
modest [105].
Additionally, adipose tissue in PCOS women is characterized by hypertrophic
adipocytes and impaired lipolysis and insulin action. TNF-α, as well as other adipo-
kines involved in insulin resistance, is altered in these kinds of patients [106].
Adiponectin applies insulin-sensitizing properties by stimulating fatty acid oxi-
dation and reducing hepatic gluconeogenesis: some studies hypothesized that its
dysregulation could be implicated in the pathogenesis of insulin resistance [107].
References 15
PCOS
Insulin receptor binding Post-binding abnormality
Insulin-resistance
Hyperinsulinemia
Liver SHBG synthesis Free androgens
Abnormal LH pulsatility
LH
Ovary
Folliculogenesis disruption
Theca cells (prematur follicular atresia and
Androgen production antral follicle arrest)
(CYP450-17α activity)
Fig. 2.1 Correlation between PCOS, hyperinsulinemia, hyperandrogenism, and ovarian

dysfunction
TNF-α is secreted by adipose tissue macrophages and has pro-inflammatory prop-

erties: it causes serine phosphorylation of the insulin receptor substrate (IRS-1).
IGFBP3 is secreted by hepatic Kupffer cells and inhibits insulin-stimulated glu-
cose uptake by dephosphorylating insulin receptor.
Both TNF-α and IGFBP3 might inhibit transcription of adiponectin, contributing
to insulin resistance [108]: in fact low levels of adiponectin were found in patients
with PCOS [109].
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Clinical Features
3
3.1 Endocrine Aspects of PCOS
Polycystic ovary syndrome (PCOS) is a chronic and self-perpetuating endocrine

disorder, whose clinical, endocrine, and metabolic manifestations affect the whole
life course of a patient. In PCOS, in fact, we can distinguish two sides of the same
coin: endocrine and metabolic aspects.
A polycystic ovary appears enlarged with a thickened albuginea that has a porce-
lain appearance. In the subcapsular layer, there are many follicles measuring
2–10 mm in diameter, reduced number of granulosa cells, and a characteristic theca
cell hyperplasia.
Thus, the fundamental abnormality is the presence of a raised number of follicles
recruited with primary maturation block and increased atretic follicles.
3.1.1 Endocrine Pattern
3.1.1.1 Gonadotropins
PCOS is considered a normo-gonadotropic normo-estrogenic anovulatory disorder,
but it is characterized by elevated LH serum concentrations with an inverted FSH/
LH ratio [1].
PCOS follicles are present in large numbers, but they are arrested at an early to
mid-developmental state and fail to mature even when they are exposed to normal
FSH levels [2–4]. On the other hand, FSH levels do not increase during the early
follicular phase to stimulate follicular maturation [5].
The resulting low estrogen and progesterone levels do not produce a negative
feedback on LH secretion, and this is the major cause for the high serum LH con-
centrations in women with PCOS [6].
Despite these findings, gonadotropin levels have never been included in any of
the diagnostic criteria for PCOS, especially because of the pulsatile nature of LH
release [7–9].

Sensitizers, DOI 10.1007/978-3-319-16760-2_3
22 3 Clinical Features
3.1.1.2 Sex Hormones

Hyperandrogenemia is the biochemical feature of PCOS. Elevated circulating
androgen levels are observed in 80–90 % of women with oligomenorrhea [10].
In particular, a decreased SHBG (sex hormone-binding globulin) production with
a consequent increase in free testosterone levels is reported. Furthermore, some
authors suggest that, vice versa, SHBG levels are decreased in PCOS due to the
effects of testosterone and insulin of decreasing hepatic production of SHBG [11, 12].
Ovaries are the main sources of increased androgens in PCOS, but even adrenal
androgen excess is a common feature of the syndrome (approximately 20 % of
PCOS women): an increased secretion of adrenocortical precursor steroids basally
and in response to ACTH, such as pregnenolone, 17-hydroxyprogesterone (17-
OHP), dehydroepiandrosterone (DHEA), and androstenedione (A), was demon-
strated [13, 14].
It has been suggested that androgens enhance apoptosis in the granulosa cells of
preantral and early antral follicles [15]. Moreover, a study found that the exposure
to excessive androstenedione stimulates a premature luteinization of granulosa
cells, most likely due to the loss of communication between the oocyte and the
granulosa cell [16].
Due to the pulsatility of LH, only one blood parameter is not enough for the
PCOS diagnosis, and there is no unanimous consensus on which androgen blood
levels should be considered for a precise diagnosis (total or free testosterone, testo-
sterone/SHBG ratio, or androstenedione). Usually, elevated levels of only DHEA or
17-OHP may exclude the diagnosis of PCOS [17].
3.1.1.3 Estrogens and Progesterone

Estradiol levels are constant, without the normal mid-cycle increase, while the
levels of estrone are increased because of extraglandular aromatization of increased
circulating androstenedione levels [18–20].
As a consequent of anovulation, progesterone levels are low in PCOS women;
moreover, some authors reported that endometrial responsiveness to progesterone is
reduced in PCOS women [21, 22] and that total endometrium PR (progesterone
receptor) expression is higher in women with PCOS who have anovulation com-
pared to women with PCOS who still ovulate [23].
Furthermore, the increased PR expression in epithelial cells is greater than that in
stromal cells in women with PCOS, suggesting that lower binding of progesterone
in stromal cells could lead to the promotion of estradiol-induced epithelial cell pro-
liferation in PCOS women.
It has been hypothesized that lack of progesterone-induced and PR-mediated
stromal cell proliferation could be a cause of progesterone resistance in PCOS
patients [24].
3.1.1.4 AMH
Anti-Mullerian hormone (AMH) belongs to the transforming growth factor-β (TGF-
β) superfamily. In women, AMH is produced by the granulosa cells of follicles from
the stage of the primary follicle to the initial formation of the antrum. In female
3.1 Endocrine Aspects of PCOS 23
newborn, AMH is undetectable, but it increases gradually until puberty, remaining

stable in the reproductive period [25].
Reduction of AMH levels in serum is the first indication of a decline in the fol-
licular reserve of the ovaries. Moreover, AMH concentration remains stable during
the cycle [26].
Since AMH level reflect the number of developing follicles, its measurement
may be used as a marker of ovarian follicle damage in PCOS.
AMH levels are also probably related to the follicular arrest, during the selection
process of the dominant follicle: AMH inhibits the recruitment of primordial follicles
into the pool of growing follicles and decreases their receptiveness to FSH [27–29].
The first studies regarding AMH levels in PCOS women showed that AMH
levels are higher than in healthy controls [30, 31]. Subsequent data indicated that
these levels are related to increased number of small antral follicles of 2–5 mm
diameter [32]: this correlation was found to be the strongest one [33].
The cause of the increased AMH production in PCOS is unknown: it is mainly
ascribed to the increased production of AMH by each follicle, and it is not just a
consequence of an increased follicle number, suggesting intrinsic granulosa cell
dysregulation in PCOS [34, 35].
AMH levels are increased in proportion to PCOS clinical severity, as reflected by
the antral follicle count [36, 37].
Furthermore, blood AMH appears to be associated with androgen levels, and so
it has been proposed as a diagnostic marker for ovarian hyperandrogenism [38].
Some studies demonstrated, in fact, that AMH is positively correlated with total
testosterone levels in normal-weight PCOS women [39].
AMH levels, as written before, decrease with age in women with normal ovula-
tory cycles; in PCOS women, this decline has a slower reduction rate, and it could
be because of a decelerated ovarian aging, probably due to the negative effect of
AMH on the recruitment of primordial follicles.
3.1.2 Clinical Endocrine Features
The clinical scenario of PCOS is very heterogeneous, and the symptoms are related
to the ovarian dysfunction and hyperandrogenism.
This section describes the clinical characteristics of a PCOS woman, while the
diagnostic pathway can be found in Chap. 6.
3.1.2.1 Menstrual Disorder

Since menarche, or after a short period, menstrual cycles show an irregular rhythm.
In many cases they gradually distance themselves from each other, up to result in
oligomenorrhea or in permanent amenorrhea. Menstrual dysfunction in women
affected by PCOS may manifest in different ways, but the most common way is
anovulation with erratic bleedings.
Although the presence of oligomenorrhea indicates ovulatory dysfunction,
apparent eumenorrhea does not completely rule out anovulation [40].
Therefore, ovarian dysfunction usually manifests as oligomenorrhea/amenorrhea

resulting from chronic oligo-ovulation/anovulation. The majority of women com-
plaining oligomenorrhea (up to 80–90 %) are affected by PCOS [41].
A significant relationship between the degree of menstrual dysfunction and the
degree of insulin resistance present was observed. After adjusting for BMI, age, and
race, all PCOS subjects with menstrual cycles longer than 35 days had significantly
higher mean HOMA-IR levels than controls, with those with cycle length longer
than 3 months having the highest one [42]. Confirming these findings, it was
reported also that among PCOS women insulin resistance was significantly worse in
amenorrheic patients [43].
As consequent, prolonged anovulation can be the cause of dysfunctional uterine
bleeding, which may mimic regular menstrual cycles.
In addition, the chronic anovulation implies prolonged estrogen excess (par-
ticularly in obese phenotype women) and lack of progesterone, resulting in
atypical endometrial hyperplasia, which is the precursor of endometrial carci-
noma [44, 45].
It is generally recommended that greater than four cycles per year may protect
the endometrium [46].
3.1.2.2 Infertility
PCOS is the most common cause of anovulatory infertility: 90 % of women atten-
ding infertility clinic for anovulation disorder are affected by PCOS.
Despite these data, 60 % of women with PCOS are fertile, while time to conceive
is often increased [41].
Moreover, infertile PCOS women are overweight in 90 % of cases.
Fifty percent of PCOS women experience recurrent pregnancy loss [47]: it is not
clear whether these defects are caused by uterine dysfunction itself, by possible
interrupted interaction between uterine cells and the developing embryo, or by
insulin-related disorder.
The new guidelines suggest that PCOS is a risk factor for infertility only in the
presence of oligo-ovulation or anovulation. However, there are no clear data about
the fertility of PCOS patients who have normal ovulatory function [48].
3.1.2.3 Hirsutism
Hirsutism is defined as the presence of excessive terminal hairs in areas of the body
that are androgen dependent and usually hairless or with limited hair growth, such
as the face, chest, areolas, and abdomen [49].
Terminal hair is different from “vellus” hair, because the latter is the prolonged
version of “lanugo” (the hair that covers fetuses and is shed gradually after birth)
which covers all body surface except lips, palms, and soles; specifically, terminal
hair is the pigmented, longer, coarser hair that covers the pubic and axillary areas,
scalp, eyelashes, eyebrows, male body, and facial hair [50].
Hirsutism should be differentiated from hypertrichosis, which is the overgrowth
of vellus in a nonsexual pattern distribution, usually related to persistence of the
highly mitotic anagen phase of the hair cycle [51, 52].
3.1 Endocrine Aspects of PCOS 25
Terminal hair growth requires androgen stimulation, specially testosterone and

dihydrotestosterone (DHT) that can bind to the androgen receptor and promote hair
follicle changes [50, 53].
Androgens, in fact, are the most significant hormones associated with hair growth
modulation. They are necessary for terminal hair and sebaceous gland development and
cause differentiation of pilosebaceous units into either a terminal hair follicle or a seba-
ceous gland. They are involved in keratinization, increased hair follicle size, hair fiber
diameter, and the proportion of time that terminal hair spends in the anagen phase [54].
Thus, hyperandrogenemia is the cause of hirsutism, but the percentage of hair
growth is not proportional to the degree of hyperandrogenism, supporting the
important role for androgen receptor localization (keratinocytes, sebaceous glands,
hair dermal papilla cells) and sensitivity in the development of hair patterns [55].
3.1.2.4 Acne and Seborrhea

Sebaceous glands are also androgen-dependent structures: sebocytes are highly sen-
sitive to androgen signaling, which is worsened in PCOS, leading to the develop-
ment of acne and seborrhea [56].
Androgens stimulate sebocyte proliferation (particularly in the mid-back, chin,
and forehead) and secretion of sebum, which is a mixture of lipids including
glycerides, squalene, free fatty acids (FFA), and cholesterol [57].
Local bacteria complicate the process by secreting lipolytic enzymes: they break
down those triglycerides produced in the sebocyte; these FFAs are released into
sebaceous ducts by apocrine glands, and they are responsible for the typical unpleas-
ant odor [58].
3.1.2.5 Androgenic Alopecia

An opposite clinical feature is androgenic alopecia, which is a disorder characte-
rized by miniaturized hair, due to an increased telogen/anagen ratio, and associated
to genetic susceptibility related to increased 5α-reductase activity in the hair folli-
cle. This increased enzymatic activity promotes the local conversion of testosterone
into DHT, which has an increased androgen action.
Seventy percent of women with alopecia areata have PCOS with elevated levels
of androstenedione and testosterone [59]. The balding pattern is mainly in the fron-
tal and parietal scalp zones, while the occipital area has a great hair density [60].
3.1.2.6 Other Clinical Features

In rare cases, virilization patterns can be observed: they include increased size of
clitoris, muscle mass hypertrophy, deep voice, temporal balding, and masculine
aspect. In these cases, however, a lower ovarian or adrenal androgen-secreting neo-
plasm must be excluded.
Moreover, in PCOS women, nails could be affected by alterations, in the form of
onycholysis [61] (separation of the nail plate from the nail bed caused by disruption
of the onychocorneal band) and onychorrhexis [62] (splitting of nails in lengthway
bridges). Nowadays the association of these nail diseases with hyperandrogenemia
is not completely understood.
In literature, an unusual case of atypical oral hirsutism secondary to PCOS was

described: a 19-year-old girl complained of the appearance of hairs on the sulcular
epithelium of the retroincisor palatal papilla, relapsing after surgical excision.
PCOS diagnosis was confirmed by clinical data (oligomenorrhea, face hirsutism,
and acne), by serum studies, and by the symptom improvement after combined
hormonal therapy [63].
3.1.2.7 PCOS and Thyroid Dysfunction

The most prevalent autoimmune disease in women is autoimmune thyroiditis (AIT),
with a prevalence ranging from 4 to 21 %: it depends on age [64], diagnostic crite-
ria, genetic differences, geographical origin, and iodine intake [65, 66]. In the past,
a German study underlined the association between PCOS and AIT, but the patho-
genesis of this relationship is not clear.
Few explanations were suggested, but none of these appears to be conclusive:
• Probable common genetic predisposition

• Imbalance between estrogens and progesterone, and the consequences of the
stimulatory effect of estrogens on the immune system [67]
• Low-grade inflammatory state characteristic of PCOS
Recently, it has been shown that there is a higher prevalence of subclinical hypo-
thyroidism in young women with PCOS compared with that reported for the general
young women population [68].
Moreover, Mueller et al. [69] observed that PCOS patients with subclinical
hypothyroidism had a higher prevalence of IR and a higher BMI.
3.2 Metabolic Aspects of PCOS
“Metabolic flexibility” is the capacity of the body to rapidly switch from predomi-
nant lipid oxidation (with high rates of fatty acid uptake in low-insulin conditions)
to predominant glucose oxidation and storage with suppression of lipid oxidation in
high-insulin conditions [70].
Few studies showed that obese and/or diabetic and/or insulin-resistant individu-
als, as compared with healthy lean individuals, have an impaired metabolic flexibi-
lity [71, 72].
Insulin resistance and the associated metabolic abnormalities are frequent
findings in women with polycystic ovary syndrome [73].
Many women with PCOS meet the criteria for the metabolic syndrome (MS),
as they report a higher incidence of hypertension, dyslipidemia, and visceral
obesity [74].
Up to 43 % of nondiabetic PCOS women meet MS criteria before the end of their
fourth decade, and most of them before the end of their third decade of life [75, 76].
This prevalence is four times higher than that observed in women aged
20–30 years and twice that of women between the ages of 30 and 40 years [77].
3.2 Metabolic Aspects of PCOS 27
The prevalence of metabolic syndrome is similar across racial backgrounds [78].

Moreover, it was found that the prevalence of metabolic syndrome is higher in
adolescent girls with PCOS: 37 % against 5 % of control non-PCOS girls [79].
The most common phenotypes in parents of adolescents with PCOS were found
to be excessive weight and metabolic syndrome, particularly in fathers in whom the
prevalence of MS and central obesity was 1.5–2-fold greater than expected in the
general population [80].
The essential components of MS include insulin resistance or central obesity
with at least two of hypertension, elevated triglycerides, decreased HDL-C levels,
or elevated fasting glucose [81].
Insulin resistance, and consequent compensatory hyperinsulinemia, appears to
be the central pathophysiologic mechanism that links PCOS to its metabolic disor-
ders; in fact, few studies reported that PCOS women are more insulin resistant than
controls who are matched for age and BMI [78].
Disturbance in the insulin’s ability to bind to its receptor or in the transport
mechanism across the cell membrane may lead to a state of a reduced sensitivity to
insulin, or insulin resistance. Furthermore, pancreatic β-cell secretory dysfunction
has also been reported [82, 83], and a reduction in hepatic insulin extraction contri-
butes to the high insulin levels as well [84, 85].
Compensatory hyperinsulinemia is important in the development of meta-
bolic abnormalities and also contributes to the high androgen levels, peculiar of
PCOS women. As largely described in Chap. 2, it is important to remember that
insulin binds to its receptor on the ovarian theca cell and it acts enhancing
LH-stimulated androgen production [86]. Moreover, insulin can also act indi-
rectly to raise free testosterone serum concentration by inhibiting the hepatic
production of SHBG [12].
Although obesity is a major factor for the development of insulin resistance in
PCOS, it is now well known that a component of insulin resistance is independent
of body weight [87].
To underline the link between PCOS and metabolic syndrome, it is important to
report that coronary heart disease, as well as cerebrovascular disease, is more com-
mon in postmenopausal PCOS patients. Persisting high androgen levels through the
menopause, obesity, and maturity-onset diabetes mellitus are proposed as the main
mechanisms accounting for the increased risk [88].
3.2.1 The Role of the Adipocyte in Linking PCOS to Metabolic

Syndrome
Adipose tissue is nowadays considered not only a storage tissue but also a proper
endocrine organ, metabolically active [89–92].
Adipose tissue responds to chronic changes in energy balance and nutrient con-
tent by altering the proliferation of pre-adipocytes, their differentiation into mature
adipocytes, the growth and hypertrophy of adipocytes, and, finally, their apoptosis
and necrosis [93]. In addition, rates of angiogenesis, extracellular matrix
remodeling, and the relative distribution of the resident immune cell population in
adipose tissue are modified in response to changes in nutritional status [94]. Thus,
it is evident that adipose tissue acts as an enormous endocrine organ, secreting a
variety of signaling molecules that regulate feeding behavior, energy spending,
metabolism, reproduction, and endocrine and immune function [95].
Adipocytes secrete adiponectin, leptin, visfatin, tumor necrosis factor alpha
(TNF-α), interleukin 6 (IL-6), plasminogen activator inhibitor-1 (PAI-1), resistin,
and angiotensinogen: thus, adipose tissue results to be metabolically active [96, 97].
Adiponectin is a 244-amino acid protein that is expressed in white adipose tissue [98].
This adipokine expression within adipocytes is downregulated in obesity [99],
and the result is that serum levels of adiponectin are inversely correlated with body
weight [100].
Adiponectin has insulin-sensitizing, anti-atherogenic, and anti-inflammatory
properties [101, 102]. It is well known that adiponectin has an important role in
mediating the effects of increased fat mass on insulin sensitivity [103]: in fact, its
low serum levels seem to be involved in conditions associated with insulin resi-
stance, such as type II diabetes and obesity [104–106]; moreover, lower levels of
serum adiponectin are present in PCOS women [107].
It also has been reported that adiponectin inhibits theca cell androgen produc-
tion: suppressed levels of adiponectin may allow enhanced ovarian androgen pro-
duction in PCOS women [108].
Leptin controls the fat disposition modulating its accumulation in the heart, liver,
and kidneys; besides, it is involved in the control of vascular tone by producing a
pressure action and opposing the NO-mediated relaxing function [109]: this could
be associated with cardio-metabolic syndrome.
In humans, there is a strong association between the percentage of body fat and
serum leptin levels [110]; some authors found that hyper-leptinemia has a positive
relationship with insulin-resistant PCOS women [111], even if more studies are
needed to confirm it.
Hyper-leptinemia seems to lower the sensitivity of dominant ovarian follicles to
insulin-like growth factor 1 (IGF-1), which is implicated in the mechanism of
anovulation [112].
Visfatin is a multifunctional protein that plays a number of roles including the
regulation of metabolism and inflammation, and it is also involved in the insulin
resistance mechanism [113, 114].
Few recent studies have demonstrated that visfatin levels are significantly higher
in PCOS women comparing to the healthy controls [115], even when considering
only the overweight and obese subgroups [116].
It has been demonstrated that in women with PCOS, adipocyte diameter is 25 %
greater than the diameter of adipocytes taken from obese control women with com-
parable BMI: adipocyte seems to be hypertrophic [117].
Adipocyte hypertrophy in PCOS may be a consequence of variations in storage
and/or adipocyte lipolytic capacity. Thus, obesity in women with PCOS is mainly
characterized by an increase in fat cell size (hypertrophic obesity) rather than an
increase in fat cell number (hyperplastic obesity) [118].
In PCOS subcutaneous adipocytes, there is a reduced catecholamine-mediated

lipolysis [119], and maybe there is an implication of testosterone as a possible con-
tributory factor in this process [120]. Greater lipolysis within visceral adipocytes
results in hepatic insulin resistance through increased hepatic influx of portal free
fatty acids; reduced lipolysis within subcutaneous adipocytes is likely to be one
explanation for adipocyte hypertrophy and consequent insulin resistance [108].
Studies on the molecular insulin signaling pathways within PCOS adipocytes
have demonstrated that the number of insulin receptors and the affinity of these
receptors for insulin are normal [121–123]. Moreover, there are evidences that, in
PCOS adipocytes, basal auto-phosphorylation of the insulin receptor β-subunit is
normal, but insulin-dependent auto-phosphorylation is significantly reduced [123].
The existing literature suggests a large number of possible defective post-insulin
receptor molecular mechanisms that may explain adipocyte’s insulin resistance in
PCOS, although the actual mechanism involved and the determinants of adipocyte
size in PCOS are not fully understood [108].
In the past, Danforth hypothesized that the inability to differentiate sufficient
new subcutaneous adipocytes in response to chronic excessive energy intake may
explain the metabolic dysfunction observed in some obese women: thus, a defi-
ciency in either the proliferation or differentiation capacity of adipocytes leads to
the redistribution of fat from subcutaneous to visceral depots and also to other tis-
sues such as the liver and skeletal muscle, where the ectopic fat causes insulin resi-
stance [124].
Furthermore, steroidogenic activity within the adipocyte plays a crucial role
in the development of PCOS, particularly the hyperandrogenemia associated
with PCOS: this could be the link between obesity and hyperandrogenemic fea-
tures [108].
Although the predominant source of raised androgens in PCOS women is
ovarian, adrenal androgen secretion is also important [125]; moreover, peripheral
conversion of androstenedione and DHEAS accounts for up to 50 % of circula-
ting testosterone in PCOS women, and the major peripheral site is the adipose
tissue [126].
The 5α-reductase enzyme is present in the adipocyte cell and it converts testo-
sterone into the more potent 5α-dihydrotestosterone (DHT), and it is also involved
in the catabolism of cortisol [125].
It is supposed that PCOS women have enhanced peripheral 5α-reductase activity
compared with age and BMI-matched control women [127–130]. This causes an
increased production of DHT, increased catabolism of cortisol, and consequent
reduced feedback of cortisol on the pituitary corticotroph cells [125].
Furthermore, increasing adipose tissue mass is directly associated with increa-
sing levels of angiotensin II from the increased secretion of angiotensinogen by
adipose tissue; this increase could contribute to hypertension and worsen insulin
resistance [131].
On the other hand, few molecules secreted by adipocytes are involved in macro-
phage function, including monocyte chemoattractant protein-1 (MCP-1), macro-
phage migration inhibitory factor, and macrophage inflammatory protein (MIP)-1α
which are upregulated in obesity [132]. Adipose tissue resident macrophages from
obese women are activated and also express few proteins such as MIP-1α, MCP-1,
and related inflammatory cytokines, which may play a role in the development of
obesity-induced insulin resistance [118]. Most adipose tissue TNF-α, inducible
nitric oxide synthase, and IL-6 seem to be expressed by adipose tissue macrophages,
rather than adipocyte [94]. IL-6 inhibits lipoprotein lipase activity, stimulates aro-
matase activity, and increases the hepatic production of triglycerides [133]. IL-6 is
stimulated by TNF-α: the latter stimulates C-reactive protein (CRP), which is highly
associated with obesity, insulin resistance, and endothelial dysfunction; PCOS
women seem to have higher levels of CRP [134, 135] and, specifically, of hs-CRP
(high-sensitive CRP), which is the most specific marker [136].
Elevation of these inflammatory markers is in accord with the hypothesis that
atheroma formation is primarily an inflammatory condition [137].
3.2.2 The Role of Vitamin D in the Development of Metabolic

Syndrome in PCOS Women
Few evidences suggest that vitamin D deficiency could be a causal factor in the
pathogenesis of metabolic syndrome in PCOS women [138].
It is well known that the vitamin D receptor gene regulates 3 % of the human
genome, including genes essential for glucose and lipid metabolism and blood pres-
sure regulation [139–141].
In fact, clinical studies had reported insulin resistance and obesity association
with hypovitaminosis D [138, 142, 143].
The mechanism underlying the association between low vitamin D levels and
insulin resistance is not completely assumed. The suggested hypotheses are the
following:
• Vitamin D may have a positive effect on insulin action by stimulating the expression
of insulin receptor and improving insulin receptiveness for glucose transport [140].
• Vitamin D regulates extracellular and intracellular calcium, which is important
for insulin-mediated intracellular processes in insulin-responsive tissue (skeletal
muscle and adipose tissue) [140].
• Vitamin D has a modulating effect on the immune system, so hypovitaminosis D
might have a pro-inflammatory action, which is associated with insulin resi-
stance [144, 145].
Moreover, it is not fully understood whether vitamin D insufficiency results from

obesity and/or whether obesity is a consequence of hypovitaminosis D.
Despite there is not a clear consensus regarding its optimal value, a level of
30 ng/ml indicates a sufficient vitamin D status [139]; concentrations of 20–30 ng/
ml are considered as vitamin D insufficiency, while a level less than 20 ng/ml
represents a vitamin D deficiency [139].
In a recent study, 72.8 % of PCOS women showed values below the abovemen-
tioned normal cutoff; a significant association of hypovitaminosis with increased
levels of both fasting and stimulated glucose and insulin, elevated HOMA-IR, and
incidence of MS was demonstrated [146].
Furthermore, vitamin D levels were significantly lower in hirsute woman, as
shown in previous study [138, 147]. For example, distress caused by the excessive
hair might lead to hypovitaminosis D because of the decreased sun exposure of
hirsute women.
Vitamin D receptor is present in keratinocytes of the outer root sheath as well as
in cells of the bulge, indicating an important role of vitamin D in hair follicle
cycling [144].
3.2.3 Metabolic Syndrome and Associated Disorders
3.2.3.1 Visceral Obesity

The prevalence of obesity in PCOS varies from approximately 10 % up to 50 %
[148, 149].
A likely explanation for the mechanism underlying the development of obesity
in women with PCOS is the combined effect of a genetic predisposition to obesity
in the context of an “obesogenic” environment (poor diet and reduced exercise).
The development of obesity in PCOS patients, in turn, amplifies and unmasks the
biochemical and clinical abnormalities characteristic of this condition [125].
Obese PCOS women have lower levels of SHBG, DHEAS, DHEA, IGF-1, and
HDL and higher LDL compared with the nonobese PCOS controls [150].
It is important to underline that all overweight/obese people are not insulin resi-
stant, and those who are insulin resistant are not all obese.
Previous researches have studied the importance of fat distribution patterns as risk
factor for cardiovascular and metabolic disease such as diabetes mellitus [151, 152].
In fact, gluteo-femoral obesity is less associated with insulin resistance than is
central or android obesity [153].
The gynoid type of fat distribution, where fat accumulates around the hips,
thighs, and buttocks, is developed during female puberty and is maintained during
the fertile phase [152, 154].
Approximately 50–60 % of PCOS women are characterized by a so-called
“android” distribution of body fat, whereby a disproportionate quantity of adipose
tissue is distributed in the visceral depot [155, 156]: they have a higher trunk/periphery
fat ratio [157].
This upper body fat distribution has been explained mainly by androgen excess
[158], and it is an independent factor of BMI [159]: the pathogenic mechanisms
involved have not yet been defined.
In women affected by PCOS, android body fat distribution per se contributes to
hyperandrogenemia, through its adverse effects on insulin sensitivity and conse-
quent ovarian co-gonadotropic effects of hyperinsulinemia.
Hyperinsulinemia itself contributes to obesity by the anabolic effect on fat
metabolism through the adipogenesis process: the result is an increased uptake of
glucose into adipocytes, the production of triglycerides, and the inhibition of
hormone-sensitive lipase [160].
Therefore, there is a vicious cycle in which android fat produces android fat and
exacerbates the predisposition toward weight gain [125].
Visceral fat, or abdominal fat, is metabolically distinct from subcutaneous fat; it
is resistant to the anti-lipolytic effects of insulin and releases excessive amounts of
free fatty acids, which leads to IR in the liver and muscle. In response to it, in the
liver, there is an increased gluconeogenesis, and in the muscle there is an inhibition
of insulin-mediated glucose uptake [161–163].
Excess fat itself contributes to IR at the level of the adipocyte: when fat cells
become too large, they are unable to store additional lipids, and then, fat is stored in
the muscle, liver, and beta cells of the pancreas [164]. Visceral fat also produces
excess of 11-beta-hydroxysteroid-dehydrogenase-1 [163], an enzyme that converts
inactive cortisone to the biochemically active cortisol: the latter is able to promote
central adiposity and IR [165].
3.2.3.2 Dyslipidemia
The probability of a metabolic disorder in families of PCOS patients is 2.7-fold
higher compared with normal families, and the relative risk for developing dysli-
pidemia is 1.8 [166].
Dyslipidemia is reported in up to 70 % of patients who have PCOS, according to
the National Cholesterol Education Program (NCEP) guidelines [167].
Dyslipidemia in PCOS women seems to be well understood, but which are the
determinant factors of this pattern? Insulin, estrogens, and androgens are each well
known to alter lipoprotein lipid metabolism [168]. All of them influence hepatic
lipase activity, which is important in reductive metabolism of intermediate-density
lipoproteins to small dense LDL particles; greater activity of this enzyme was found
in PCOS women [169].
Insulin stimulates lipogenesis in arterial and adipose tissues via an increased
production of acetyl CoA and the entry of glucose and triglycerides [170].
PCOS women have higher Apo-CIII levels compared to non-PCOS controls
[171]: understanding its metabolism is helpful to deeply comprehend the patho-
physiology of dyslipidemia in PCOS.
In states of IR, it has been shown an increased synthesis of ApoC-III [172]: in PCOS, with
central obesity more free fatty acids flow into the portal vein and more glucose is available,
causing altered apolipoprotein lipid metabolism. The ratio of ApoC-II/CII is increased and
triglycerides carried in VLDL are broken into more atherogenic small LDL particles, which
circulate and enter the arterial wall to initiate inflammation. With elevated triglycerides,
VLDL lipolysis is slowed, causing greater residence time for ApoB, remnant particles, LD
particles and small LDL-particles. ApoC-I, recently shown to be elevated in normal-weight
PCOS women, blocks lipoprotein lipase, cholesterol ester transferase, lecithin cholesterol
acyltransferase, VLDL receptors and LDL receptors in the liver. All of these events lead to
more exposure of the blood vessel wall to entry of atherogenic particles with the potential
for setting inflammation and atherogenesis. [168]
Moreover, insulin increases the levels of HMG-CoA reductase, the rate-limiting

enzyme in the synthesis of cholesterol: this effect may contribute to the raised cho-
lesterol level, which is also a feature of hyperinsulinemia [173].
Hyperandrogenism and lipid metabolism are closely related: in fact, it has been
observed that testosterone has a deleterious effect on lipid profile [174, 175].
Testosterone has been involved in lowering HDL-C levels, an effect attributed to the
upregulation of two genes implicated in the catabolism of HDL: scavenger receptor
B1 (SR-B1) and hepatic lipase [176].
The most common lipid profile found in PCOS individuals is characterized by
[176–182]:
• Increased levels of LDL cholesterol (especially raised amounts of types III and
IV small LDL particles [176])
• Increased VLDL cholesterol
• Increased triglycerides
• Reduced levels of HDL cholesterol (particularly decreased HDL2, the most anti-
atherogenic HDL subtype)
Particularly, a study has demonstrated that the incidence of high triglycerides

increased progressively from the lean to the obese PCOS women, and the incidence
of low HDL was three times higher in the overweight than in the lean PCOS sub-
group [183].
HDL-C has several functions: inhibition of LDL-C oxidation, transport of cho-
lesterol from peripheral cells to the liver, anti-apoptotic effects, and antithrombotic
and antioxidant effects. For this reason, low HDL-C is considered an independent
cardiovascular risk factor: thus, women with PCOS may have a higher cardiovascu-
lar risk than normal women at the same BMI level [184].
Furthermore, these lipid disorders are exacerbated among those women who
develop glucose intolerance in association with PCOS; in fact, 88 % of women with
PCOS and IGT or type II diabetes have an abnormal lipid profile, compared with
58 % of women with PCOS and normal glucose tolerance [177].
Even PCOS adolescents have less favorable blood lipid profiles, with higher
LDL-C and lower levels of HDL-C, and they appear to be more insulin resistant
than their peer control with higher fasting C peptide levels [185].
3.2.3.3 NAFLD (Nonalcoholic Fatty Liver Disease)

NAFLD represents a disease spectrum ranging from steatosis hepatitis (SH or
NAFL) to nonalcoholic steato-hepatitis (NASH), characterized by hepatocyte
injury, inflammation, and fibrosis, which can progress to cirrhosis in 25 % of cases,
with its long-term complications, such as portal hypertension, liver failure, and
hepatocellular carcinoma [186].
The hepatic steatosis is histopathologically characterized by the accumulation of
triglycerides, both in the form of macro- and microvesicles, in more than 5 % of
hepatocytes. These “fatty hepatocytes” are usually peri-venular located, and they
are mainly present at the level of the “portal areas.” The pool of fatty acids available
for the synthesis of triglycerides is related to the balance between their formation
and utilization. The deposition of triglycerides into the hepatocytes depends on both
of these reactions: thus, the development of a fatty liver is the consequence of a
dysfunction in different metabolic pathways.
There are important relationships between peripheral insulin resistance and

hepatic fat deposition. High intake of calories in sedentary individuals who are
genetically susceptible induces a state of IR, which involves an increased lipolysis,
as a result of free fatty acid and TNF-α circulating levels and of lower levels of
adiponectin: this increases both insulin resistance and circulating levels of free fatty
acids. The raised fat deposition in the liver induces insulin resistance by itself, acti-
vating abnormal insulin intracellular signals. Both these processes lead to an
increased hepatic insulin resistance and deposition of fatty acids.
These events, in turn, cause a dysregulation of some sterol regulatory proteins
(SREBP-1C) and, probably, of ghrelin. This pathogenic mechanism is responsible
for inducing a de novo lipogenesis in the liver.
Thus, it is well evident that NAFLD is a complex and multifactorial disease, and
it is currently the most common cause of liver disease and high enzyme levels in
clinical practice. Probably a state of IR plays critical part in both the development
and progression of the liver disease. It seems that an important role is assumed by
oxidative stress and some adipokines, such as TNF-α and adiponectin. Moreover,
the nature of the epidemic NAFLD, as well as its clear association with obesity and
metabolic syndrome, makes the altered lifestyle and a sedentary lifestyle conditio-
ning factors in the development of this disease even in teenagers [187].
Even if liver biopsy is the “gold standard” for distinguishing between simple
steatosis and NASH, and for disease severity assessment, the diagnosis of fatty liver
is clinically performed by ultrasound:
1. Absent: the echogenicity of the liver parenchyma is greater than or equal to that of
the cortex of the kidney; there is a clear view of the intrahepatic venous system.
2. Mild fatty liver: slight increase of fine echoes in the liver parenchyma with nor-
mal visualization of the intrahepatic venous circulation.
3. Moderate hepatic steatosis: moderate and widespread increase of fine echoes in
the liver parenchyma with impaired visualization of intrahepatic venous system.
4. Severe fatty liver: marked increased echogenicity of the liver parenchyma with
deficiency or absence of visualization of intrahepatic venous circulation [187].
The extent of steatosis is related to the degree of insulin resistance [188].

Elevated liver enzymes have been used as a noninvasive surrogate marker of
NAFLD, provided that other potential causes of liver disease (chronic viral hepati-
tis, alcohol-induced liver disease, etc.) have been excluded.
The typical pattern of abnormal liver biochemical profile includes increased
serum aminotransferases, with a predominant increase of alanine aminotransferase
(ALT), relative to aspartate aminotransferase (AST), accompanied by elevated
γ-glutamyl transpeptidase levels (γ-GT) [189, 190].
Elevated ALT, above the level of 35 U/l, has been detected in 30 % of over-
weight/obese PCOS women [191, 192].
Both adult and pediatric patients with NASH are commonly asymptomatic.
Rarely, patients may present with persistent right upper quadrant pain or chronic
pain in the umbilical region. On physical examination, more than 90 % of patients
with NASH are found to be obese, and acanthosis nigricans has been reported in
36–49 % of patients [193].
A number of studies have demonstrated a high risk of hepatic steatosis in women

with PCOS [187, 192, 194–196].
A recent study has shown that women with hyperandrogenic PCOS have evi-
dence of increased liver fat, compared with PCOS women with normal androgens
or with healthy controls [197].
In one of our studies, an elevated percentage of NAFLD both in lean and obese
women, with a low rate of hepatomegaly (8.3 %) and 15 % of elevated liver enzymes,
was found. In fact, liver enzyme impairment is not always associated to the presence
of NAFLD, and vice versa [187].
Moreover, in contrast to what happens in other population subgroup where only
a minority of patients who suffer from NAFLD progress to NASH, the prevalence
of advanced liver disease (NASH with fibrosis) in women with PCOS is higher
[192, 198] even in the adolescent population [199].
For these reasons, from a clinical point of view, it seems advisable to closely fol-
low women with PCOS and insulin resistance, particularly in the presence of body
weight alterations.
3.2.3.4 Hypertension
Hyperinsulinemia may contribute to the hypertension (which is part of the meta-
bolic syndrome) by several mechanisms:
• Stimulating the renin–angiotensin–aldosterone system and consequently increa-

sing renal sodium reabsorption [200, 201]
• Causing an increased intracellular sodium and calcium [202]
• Inducing vasoconstriction by stimulation of the sympathetic nervous system
[203–205]
• Stimulating the release of IGF-1 that may contribute to the development of
hypertension by causing vascular smooth muscle hypertrophy [137]
The prevalence of hypertension in PCOS women increases with BMI as indepen-

dent factor [206].
Women with PCOS were found to have an increased left atrial size and left ven-
tricular mass index, with a reduced left ventricular ejection fraction [207], which is
directly related to the degree of insulin resistance; this finding may represent early
remodeling as a prelude to overt cardiac dysfunction [208].
The results of many studies are controversial: in a few of them, both systolic and
diastolic blood pressures are normal [124, 209–212], while in other studies, mean
arterial pressures and ambulatory systolic pressures are elevated in women with
PCOS compared with non-PCOS controls [212].
PCOS patients appear to be at increased risk for developing hypertension, at least
later in life if it doesn’t occur during the reproductive age.
For menopausal or climacteric women with a previous history of PCOS, this
prevalence varies from 28.1 to 39 % [209, 213], while for patients who are in
their third or fourth decades of life, the prevalence varies from 3.8 to 22 %
[213–215].
This difference of prevalence according to the age range is probably a conse-
quence of aging itself.
3.2.3.5 Diabetes
PCOS women are at high risk of progression to impaired glucose metabolism and
type II diabetes; a family history of type II diabetes is present in a large percentage
of women affected by PCOS, suggesting the important role of the genetic pattern in
the development of the syndrome.
Generally, glucose levels remain normal in PCOS despite insulin resistance,
because of compensatory pancreatic β-cell insulin production resulting in hyperin-
sulinemia. However, some patients have a genetic susceptibility to pancreatic β-cell
failure and, over time, develop elevations in glucose when pancreatic β-cell insulin
production can no longer overcome the insulin resistance [216].
A study reported that 35 % of patients with PCOS had impaired glucose tole-
rance (IGT) and 10 % had type II diabetes (T2DM) by the age of 40 [217].
A very recent study showed that the conversion rate from IGT to T2DM in
women with PCOS was higher than that in the general population of women with
IGT: 2–10.75 % vs 1–7 % per year [218].
Moreover, it was clearly shown that the risk for IGT or T2DM in women with
PCOS was amplified (fourfold increase) in the presence of obesity, highlighting the
role of patient weight in the development of glucose metabolism disorders [218].
A Korean study found that nonobese patients with PCOS presented a higher
prevalence of elevated glycated hemoglobin than nonobese controls [219].
Both IGT and T2DM are very significant cardiovascular risk factors in women.
Once the diagnosis of diabetes is made, the relative risk of cardiovascular disease in
women increases fourfold to sevenfold, with a greater risk of cardiovascular disease
and heart failure compared to men with diabetes [220].
With regard to mortality rates, diabetes may be a more prominent contributing
cause of death in women with PCOS compared with the general population [221].
3.2.3.6 Obstructive Sleep Apnea

Obstructive sleep apnea (OSA) is a known cardiovascular risk factor and is one of
the major causes of chronic sleep disruption. It is characterized by episodic partial
or complete upper airway obstruction during sleep, leading to intermittent hypoxia,
sleep fragmentation, and a reduction in the quantity of deep non-rapid eye move-
ment (NREM) sleep or “slow wave sleep” (SWS). This alteration has been associated
with cortisol levels rising [222].
OSA has been independently related to glucose intolerance and insulin resistance
even after adjustments for obesity and age [223–227], and PCOS women are 5–30
times more likely to have this disorder compared to controls [167, 176, 228–231].
The mechanism by which PCOS increases the risk of OSA remains unclear.
The high prevalence of this disorder cannot be fully attributed to excess adipo-
sity, as reported in some studies: in two studies the severity of OSA did not correlate
with BMI [229–231], and in another one, even after controlling for BMI, women
with PCOS were 30 times more likely to have breathing disorders during sleep and
9 times more likely to have daytime sleepiness than the control women [230].
On the other hand, according to a recent study, nonobese women with PCOS do
not seem to be at increased risk of OSA: this raised risk is only present among the
obese women [232]. Other studies have also indicated that obese women with PCOS
who have OSA are more insulin resistant compared to obese women with PCOS
who do not have OSA [231, 233, 234].
Insulin resistance, in fact, seems to be a stronger predictor of OSA, more than age,
BMI, or circulating testosterone concentration [229]: the role of androgen elevation
in the pathogenesis of OSA in women with PCOS remains controversial [229, 230].
Another potential pathogenic mechanism is the “low progesterone theory”: it has
been estimated that the upper airway resistance is lower during the luteal phase,
when usually progesterone is higher, compared with follicular phase when proge-
sterone is low [235]. Progesterone promotes its effects through direct stimulation of
respiratory drive [236] and enhancement of the upper airway dilator muscle activity
[237] by which it reduces airway resistance. Because women with PCOS have
usually anovulatory cycles, and so circulating progesterone concentrations reflect
the constantly lower levels of the follicular phase, this may contribute to the high
prevalence of OSA in PCOS [238].
3.2.3.7 Plasma Viscosity and Pro-thrombotic State

In healthy individuals, there is equilibrium between the hemostatic coagulation and
fibrinolytic systems: thrombosis results from an imbalance between these complex
systems [239]. The hemostatic system plays an important role in cardiovascular
disease: for example, acute events often precipitate by thrombosis developing on a
ruptured arterial plaque.
Plasma viscosity is an important hemorheologic variable, and it is mainly deter-
mined by several macromolecules, such as fibrinogen, immunoglobulins, and lipo-
proteins [240].
Plasma viscosity is an indicator of blood flow in the network of small blood ves-
sels that constitute microcirculation. An elevated plasma viscosity indicates
increased resistance to blood flow in most tissues of the body [241].
Chronic hyperviscosity is able to impair microcirculation and promote target
organ damage [242], and it is considered an independent predictor of cardiac events
and mortality [243–245].
In a recent study, plasma viscosity is not connected to serum androgen levels, but it
is correlated with serum fasting insulin and cholesterol levels, which appear to be higher
in hirsute women compared to the matched for age and BMI healthy controls [246, 247].
Mild or chronic hyperviscosity is very frequent in older patients with metabolic
syndrome (MS) and insulin resistance (IR) [248–250]. A deterioration of plasma
viscosity was found to be present even in young, slightly overweight, PCOS women
with IR who might be exposed to the same risk factors for cardiovascular diseases
as older obese patients with MS. For this reason, plasma viscosity might be useful
in the assessment of cardiovascular risk in young women with PCOS, in addition to
plasma cholesterol and atherogenic index (triglycerides/HDL-C) [247].
Hyperinsulinemia contributes to the pro-thrombotic state by reducing fibrinoly-
sis and raising the level of PAI-1 (plasminogen activator inhibitor-1). The increase
of the latter in PCOS women seems to be independent of BMI: elevated levels, in
fact, were observed in lean PCOS women too [137].
The procoagulant state is, in part, due to platelet hyperactivity, which was observed
in lean women with PCOS [251] and in type II diabetic patients [252]. Why?
Platelets are involved in acute thrombosis, initiation of atheroma, and modula-
tion of inflammatory responses, and they contribute to endothelial dysfunction
[253]. Platelets are able to adhere to intact activated endothelium in the absence of
exposed extracellular matrix proteins [254]. These adherent platelets could have a
critical role in atherogenesis phenomenon, by secreting chemokines CCL5, CXCL4,
and IL-1 [255].
Normally, platelet activation is counterbalanced by inhibitory signaling cascades
that are activated by endothelial-derived NO and prostacyclin (PGI2), which modu-
late excessive activation [256].
Platelet hyperactivity seems to be related to acute hypertriglyceridemia: in fact,
high levels of triglycerides might decrease the production of endothelial NO and
PGI2, acting as a stimulator of platelet activation [257].
3.2.3.8 Chronic Inflammation, Endothelial Function,

and Atherosclerosis
The presence of cardiovascular risk factors such as obesity, insulin resistance, and
dyslipidemia may predispose PCOS women to coronary heart disease, but the topic
is still controversial [137].
One of the early signs of cardiovascular lesions is the endothelial injury [258].
Several authors have reported precocious anatomical and functional arterial changes
in PCOS women [259–261].
A positive correlation was demonstrated between abnormal endothelial function
and testosterone levels in hyperandrogenic insulin-resistant women [262], while
others have reported no differences for increased cardiovascular risk [263].
Mechanisms involved in the development of endothelial dysfunction could be
the following:
• Reduced synthesis and release of nitric oxide (NO) [264].

• Enhanced inactivation of NO after its release from endothelial cells [265].
• Enhanced synthesis of vasoconstricting agents [266].
• Insulin itself acts directly on the vascular endothelium and the smooth muscle
cells by a hypertrophic effect.
Insulin stimulates both endothelin-1 and NO activity in the skeletal muscle cir-
culation: an imbalance between the release of these factors may be involved in the
pathophysiology of endothelial dysfunction.
In normal women, aging per se is associated with progressive attenuation of
nitric oxide signaling; in PCOS women, these changes are present in early adult life,
predisposing polycystic ovarian syndrome patients to premature atherosclerosis; in
fact, high levels of plasma ADMA were found: endogenous NO synthase inhibitor
NG-NG-dimethyl-L-arginine (ADMA) is a biochemical marker/mediator of endo-
thelial dysfunction [267].
Furthermore, the important role of obesity in the mechanism of endothelial dys-

function in PCOS women was shown: in humans, adiponectin enhances endothelium-
dependent and endothelium-independent vasodilatation, reduces levels of TNF-α,
and diminishes its effects on endothelial cells [268, 269]. This, in turn, reduces
neointimal thickening and proliferation of smooth muscle cells, inhibits endothelial
cell proliferation and migration, inhibits endothelial effects of oxidized LDL, and
attenuates growth factor effects on smooth muscle cells [270–273].
Nowadays, it is clear that PCOS is a pro-inflammatory state, and emerging data
suggest that chronic low-grade inflammation supports the development of meta-
bolic aberration and ovarian dysfunction [274, 275].
CRP is the most reliable circulating marker of chronic low-grade inflamma-
tion in PCOS [276]. Recently, CRP was found to be a direct promoter of the
atherosclerotic processes and endothelial cell inflammation leading to athero-
thrombosis [137].
CRP has a direct role in the vascular inflammatory process stimulating the release
of inflammatory cytokines and increasing endothelial expression of cellular adhe-
sion molecules, which mediate leukocyte migration [277].
Findings of a study suggest that increased cardiovascular risk may be seen in
83.3 % of the PCO women with CRP >2.42 mg/l [278].
CRP values <1 mg/l are considered low risk, 1–3 mg/l are considered intermedi-
ate risk, and 3–10 mg/l are considered high risk for cardiovascular disease [279].
Anatomic evidence of early coronary and other vascular diseases in PCOS
women has been reported: it seems that PCOS patients have increased carotid artery
intima-media thickness (IMT) compared with age-matched control women [280].
Increased IMT has been linked to cardiovascular risk factors including dyslipi-
demia and obesity, and it is considered an independent predictor of stroke and
myocardial infarction [78].
The role of hs-CRP in predicting increased carotid intima-media thickness is not
independent of BMI in PCOS [280].
Coronary artery calcification, another marker of atherosclerosis, is more com-
mon in women with PCOS than in controls, even after adjustment for the effects of
age and BMI [281–283].
3.2.4 Role of Insulin Resistance in Infertility and Pregnancy

Outcome
As pointed out previously, PCOS is the most common cause of anovulatory inferti-
lity: 90 % of women attending infertility clinic for anovulation disorder are affected
by PCOS, and the rising incidence of PCOS women who have been subjected to
IVF had permitted several studies on their oocyte quality.
On the other hand, without considering IVF pregnancies, increased incidence of
pregnancy complications such as miscarriage, gestational diabetes mellitus (GDM),
preeclampsia, preterm delivery, and perinatal mortality has been reported in
polycystic ovary syndrome (PCOS) pregnancies: an increase of two to four times

was noticed [284].
3.2.4.1 Oocyte Quality

Obesity has been associated with lower levels of anti-Mullerian hormone (AMH),
which can indicate a decrease in ovarian reserve or available secondary follicles in
obese women [285]. Obese women also have lower levels of LH than normal-weight
women, and an independent positive association between LH and AMH levels has
been demonstrated [36]. Moreover, women with BMI >25 have lower excretion of
gonadotropins and luteal phase progesterone metabolites, implying that obesity has
a negative effect on corpus luteum function [286].
Studies directly examining oocyte quality have suggested that an altered mater-
nal metabolic environment results in an abnormal follicular fluid microenvironment,
with a subsequent poor oocyte and embryo quality.
Women with higher BMI had increased levels of insulin, lactate, triglycerides,
and CRP in the follicular fluid and decreased levels of SHBG [287], indicating that
the maternal metabolic environment has a direct effect on the ovarian follicular
microenvironment [288].
The increased CRP in the follicular fluid indicates inflammation and increased
oxidative stress, with consequent decreased developmental potential in the oocyte
[287, 289]. A recent analysis has showed that obese PCOS women have smaller
oocyte size compared with the control group [290], but nowadays the effect of
oocyte size on developmental competence and pregnancy outcome is unknown.
3.2.4.2 Recurrent Pregnancy Loss

Recurrent pregnancy loss (RPL) is defined by two or more failed pregnancies and
it is found in 1–5 % of couples during pregnancy, and 50 % of these cases remain
unexplained [291]. The incidence rate between PCOS and recurrent miscarriage is
not clear because of its large variation in different studies [292–295]. Some authors
have reported that PCOS women have a 33 % chance of spontaneous abortion
[293, 296].
The following are the two most reasonable mechanisms:
1. LH hypersecretion: inappropriate LH secretion during the follicular stage might

cause premature oocyte maturation through inhibition of oocyte maturation
inhibitor [297]. The hyperandrogenemia secondary to increased LH levels
impacts on ovarian folliculogenesis, resulting in abnormal granulosa cell func-
tion and follicular atresia.
Moreover, the abnormal endocrine environment might exert an influence on
the endometrium, and the ultimate cause of miscarriage could be secondary to
endometrial non-receptivity [298].
It has also been suggested that PCOS is associated with an endometrial
inflammatory reaction affecting implantation on the basis of raised levels of CRP
[280, 299, 300]. In fact, the low-grade chronic inflammation status could be the
expression of an abnormal immune regulation during pregnancy, with an increase
in the frequency and the extent of immune-mediated placental pathologies that
probably reduce the maternal immunological permissiveness to trophoblastic

invasion and placentation in PCOS women [301, 302].
A recent study demonstrated an increased incidence of placental lesions such
as chronic villitis and intervillositis [302].
2. Hyperinsulinemia has been proposed as the route for the effect of obesity on
some reproductive abnormalities, probably through its effect on androgen pro-
duction. Hyperinsulinemia seems to adversely affect the preimplantation envi-
ronment by decreasing the expression of glycodelin and IGF-binding protein-1
[303], which may play a role in inhibiting the endometrial immune response of
the embryo and facilitating adhesion processes at the feto-maternal interface
[291].
In a few studies, a positive relationship between HOMA2-IR and spontaneous
abortion suggests IR as a significant predictor of pregnancy loss [304, 305].
Recent studies consider the occurrence of hypofibrinolysis associated with
high plasminogen activator inhibitor-1 (PAI-1) the reason for RPL [306, 307].
The effects of elevated PAI-1 may also be worsen by elevated homocysteine
[308]; in fact, some studies propose a possible association between insulin resi-
stance (IR) and hyperhomocysteinemia (HHcy) [309] due to a documented
increased incidence of the latter in PCOS women [310]. Apart from the throm-
bogenic effect of elevated Hcy on pregnant PCOS women (resulting in micro-
thrombus formation causing placental dysfunction), recent findings have
implicated the adverse effect of HHcy on the defect in folliculogenesis [311],
embryo quality [312], oocyte number, and maturation [313].
A recent study clearly showed that PCOS and pregnancy affect the hemostatic
indices independently; the significant interaction between PCOS and pregnancy
only affects the activity levels of factor (F)VIII and factor (F)X. As already high-
lighted, when nonpregnant women with PCOS become pregnant, they are likely
to be in a more pro-thrombotic state than healthy women who get pregnant, as
the activities of FVIII and FX and the levels of Von Willebrand factor (WF) and
PAI-1 (which are involved in the coagulation cascade) are significantly higher in
PCOS pregnant women [284].
Moreover, as well explained previously, there is a strict connection between
hyperinsulinemia and adipose tissue function in PCOS women. The already cited
adiponectin and adiponectin receptors are involved in the female reproductive
tract; the mechanism by which adiponectin system regulates implantation and
early pregnancy remains unknown. It has been reported that a failure on adipo-
nectin system leads to a suboptimal uterine decidualization and pregnancy loss
in obesity and PCOS [314].
On the contrary, other studies did not confirm the association between early preg-
nancy loss and PCOS [315].
3.2.4.3 Gestational Diabetes

In normal pregnancy, maternal carbohydrate metabolism adapts to offer the fetus an
adequate and continuous glucose supply despite intermittent maternal intake. The
physiologic changes include pancreatic β-cell hyperplasia and an initial increase in
insulin sensitivity followed by a progressive insulin resistance.
β-cell hyperplasia seems to be induced by prolactin and human placental lacto-

gen; on the contrary, production of “diabetogenic hormones,” such as GH and CRH,
contributes to insulin resistance. This maternal insulin resistance, in turn, shunts
nutrients to the fetus.
From the third trimester, fasting glucose concentrations are 10–20 % lower, post-
prandial glucose concentrations are significantly elevated and prolonged, and
fasting insulin level is double that of nonpregnant women [216].
Thus, normal pregnancy induces a state of insulin resistance, and because women
with PCOS have a high incidence of IR, they have an increased risk of developing
gestational diabetic complications [316].
GDM is defined as carbohydrate intolerance that either begins in or is first recognized
in pregnancy [317]. Its pathophysiology includes both insulin resistance and abnormali-
ties of β-cell glucose sensitivity, which leads to inadequate insulin response [318].
According to a large study, PCOS women have a 2.4-fold increased odds of
gestational diabetes, independent of age, race, and multiple gestation [319]. Further-
more, this increased risk occurs independent of obesity [320].
In fact, during the pregnancy, hyperinsulinemic women with PCOS develop
more easily impaired glucose tolerance or gestational diabetes: the compensatory
mechanism (reduced glucose clearance and/or defects of insulin action at receptor
and post-receptor sites) that leads to prepregnancy hyperinsulinemia may more
easily fail during pregnancy [321].
Investigators have shown that women with GDM with higher glucose values at
OGTT, higher mean blood glucose, and worse glycemic control are at higher risk of
preterm delivery [322].
Moreover, it was found that women with PCOS and GDM had a 3.5-fold higher
risk for impaired glucose metabolism after delivery [323].
3.2.4.4 Pregnancy-Induced Hypertension

A meta-analysis showed that PCOS women have a higher risk of developing
pregnancy-induced hypertension: this risk was also present after excluding all
studies in which a higher BMI, multiple pregnancy rates, and a lower parity among
women with PCOS were reported. Women with PCOS also demonstrated an
increased incidence of preeclampsia of an order similar to that associated with mul-
tiple pregnancies. Moreover, older (age >30 years) women with PCOS are more
susceptible to PIH than the younger women [324].
As widely explained in the previous chapter, hyperinsulinemia can cause endo-
thelial dysfunction, and this association suggests a placental insufficiency in PCOS
women [320], due to a vascular maladaptation: in fact, a study showed that arterial
elasticity is impaired during the first trimester, while it decreases during the second
and third trimester. It was also reported that systolic, diastolic, and mean arterial
pressures were elevated throughout the pregnancy and that 27 % of the women with
PCOS developed pregnancy-induced hypertension [325].
Furthermore, hyperandrogenemia in early second trimester and throughout preg-
nancy is associated with subsequent preeclampsia [326–330], and preeclampsia in a
previous pregnancy is associated with elevated androgen levels later in life [330]. In
fact, maternal androgen levels are higher in complicated compared to uncomplicated
3.3 PCOS Phenotype in Different Ages 43
pregnancies in PCOS women [301]. This hypothesis was based on in vitro studies of
preeclamptic placentas that were found to have decreased ability to aromatize andro-
gens to estriol, compared to placentas from normal pregnancies [331].
3.2.4.5 Neonatal Outcome

Despite the risk of macrosomia due to the increased risk of gestational diabetes, the
prevalence of SGA seems to be increased in PCOS women: 12.8 % vs 2.8 % of
healthy controls, according to a Dutch study [332].
The cause can be found in the placental dysfunction or could be influenced by the
mild raised number of preterm labor (1.75-fold higher risk).
Neonates of PCOS women have a 2.31 times higher risk of admission to inten-
sive care unit and three times higher perinatal mortality than newborns of healthy
women. Perinatal morbidity could be explained by prematurity and intrauterine
growth retardation due to the placental dysfunction [333].
3.3 PCOS Phenotype in Different Ages
PCOS clinical and biochemical presentations and its metabolic consequences vary
with age (Table 3.1) [334, 335].
3.3.1 Adolescence
The clinical presentation of chronic anovulation varies by age, with amenorrhea and
oligomenorrhea being common among adolescents [336].
Menstrual irregularities and insulin resistance are common and usual features of
normal puberty period, and they can make the diagnosis of PCOS in this period of
life difficult [337].
Menstrual irregularity is common in the early years after menarche, and oligo-
anovulation may be absolutely normal [338]: this is due to the immaturity of the
hypothalamic–pituitary–ovarian (HPO) axis. An old study showed that 80 % of the
cycles were anovulatory in the first year after menarche, 50 % in the third, and 10 %
in the sixth: it is generally accepted that it may take up to 5 years after menarche for
the HPO axis to reach maturation [339, 340].
The serum concentrations of sex hormones increase with age, from premenar-
chal to post-menarchal [339].
Table 3.1 PCOS features in different ages

Adolescence Fertile period Perimenopausal period
Chronic anovulation Periods became Increased IGT, type II diabetes,
more regular hypertension, obesity, metabolic
Oligomenorrhea/amenorrhea Increasing levels syndrome
of insulin
resistance
Furthermore, concomitant eating disorders are frequent during these ages, and
secondary amenorrhea can be associated with anorectic behavior in adolescents
[341]. As ultrasound images have shown, uterine growth continues several years
after menarche, and the average ovarian volume increases from early childhood
until the age of 16 [342].
Regarding the ovarian morphology, the difference between a multifollicular
appearance and polycystic ovarian morphology in adolescents is difficult to define
[342]. About 80 % of girls have this USS finding and the presence of polycystic
ovarian morphology in a non-hyperandrogenic adolescent should be considered
normal [343].
Mild hair growth can be also considered a normal component of the late stages
of puberty and early adolescence, because it can persist for several years; therefore,
the diagnosis is often not made until later in life, when endocrine and metabolic
dysfunctions have been firmly established [344].
In fact, the most important finding for clinical hyperandrogenism in female ado-
lescents is progressive hirsutism [344]: acne and alopecia were not suggested as
clinical markers for the diagnosis of PCOS in adolescents [345].
Premature pubarche, or the development of pubic and axillary hair before age
8 years, may be an early sign of PCOS [346]. Premature pubarche may occur as a
result of some adrenal androgen disorders, but it could also be due to an idiopathic
early activation of adrenal androgen secretion. However, not all girls with PCOS
experience premature adrenarche; persistent hyperandrogenism remains a distinct
feature of girls with premature pubarche who go on to develop PCOS, and the
hyperandrogenism is exacerbated if a child develops obesity [347].
On the other hand, puberty period is normally associated with a mild insulin
resistance: this is called “physiological peri-pubertal hyperinsulinemia,” which
together with increased GH levels is responsible for the “pubertal growth spurt”; the
result is an accelerated bone, muscle, and adipose tissue growth.
Moreover, adolescent hyperandrogenemia is associated with a reduction in
peripheral tissue insulin sensitivity and compensatory hyperinsulinemia, which
implies an increase in the risk of type II diabetes [348]. The increased prevalence of
obesity in the younger population leads to long-term consequences for cardiovascu-
lar disease at relatively young ages.
There is a strong inverse relationship between reported age and weight at
menarche, suggesting that girls who were overweight had an earlier menarche, while
those who were thin, compared with their peers, experienced a later menarche [349].
Earlier menarche in girls with PCOS might be expected based on findings that
overweight girls experience earlier pubarche, thelarche, and menarche than those
with a normal BMI [350, 351].
According to all these findings, a definitive diagnosis of PCOS in adolescents
should require all three Rotterdam elements (not just 2 out of 3) [345].
3.3 PCOS Phenotype in Different Ages 45
3.3.2 Fertile Period
PCOS remains stable only during early adult age (18–30 years), but after that time,
it changes in ovarian and adrenal function and in metabolic regulation modifying
the presentation of the syndrome [352].
The menstrual cycles may become regular with age in women with PCOS [353,
354]: the development of a new balance in the polycystic ovary, caused merely by
follicle loss through ovarian aging, can explain the occurrence of regular cycles in
older patients with PCOS [354]. In a study of aging women with PCOS comparing
those who became regular with those still menstruating irregularly, a lower follicle
count for women with PCOS was predictive of the achievement of regular men-
strual cycles with age [355], confirming that a decrease in the size of the follicle
cohort from ovarian aging is largely responsible for the regular menstrual cycles in
aging PCOS women [355]. The decrease in both ovarian volume and follicle num-
ber, caused by the aging, results in loss of PCO morphology [356].
The production of androgens in women may decrease because of ovarian aging
or decreased production by the adrenal glands over time [357].
Normally, there is a marked decrease in adrenal androgen secretion, including
androstenedione and DHEAS, between the ages of 40 and 45 years [358]; andro-
gens levels also decline 20–30 % in women with PCOS.
A recent study, consisting in a 20-year follow-up of PCOS women, showed the
inability to diagnose the disorder in about 10 % of women who had PCOS dia-
gnosed 20 years earlier [359].
3.3.3 Premenopausal and Postmenopausal Period
Hyperandrogenism partially resolves before menopause in women with PCOS

[360], but a recent study showed that adrenal androgen secretion also remains pro-
nounced up to menopause in women with PCOS, indicating that exposure to hyper-
androgenism persists for a long time in these women [361]: they have an elevated
androgen to estrogen ratio.
It seems probable that long-lasting hyperandrogenism may magnify the unfavora-
ble hormonal and metabolic changes related to menopause and expose these women
to increased health risks [362].
Ovarian volume and follicle number decrease with age in women with and with-
out PCOS [363].
AMH levels decreased with an increase in age in both the PCOS cases and
normo-ovulatory controls [364]. AMH measurement could be useful in the predic-
tion of the menopausal transition [365, 366]. Using AMH as a predictive marker, the
reproductive lifespan of PCOS women is an average of 2 years longer than that of

normo-ovulatory women [364].
Furthermore, aging may also be associated with a defect in insulin action [367].
In fact, age is an important risk factor for developing metabolic disorders and insu-
lin resistance. Aging may also be associated with a defect in insulin action that is
manifested by decreased whole-body tissue sensitivity to insulin without a change
in tissue responsiveness [367]. The glucose intolerance may reflect part of the aging
process. In elderly subjects, the severity of carbohydrate intolerance is directly cor-
related with the degree of peripheral insulin resistance [368].
A recent study has demonstrated that impaired glucose metabolism, enhanced
ovarian androgen secretion, and chronic inflammation observed in premenopausal
PCOS women persist after menopause [362].
As result, it is clearly noted that the most common symptoms in senior age are
those related to metabolic syndrome.
Despite the longer exposure to cardiovascular risk factors, it is still difficult to
demonstrate an increased risk of morbidity and mortality in women with PCOS: only
two studies tried to study PCOS long-term outcomes, but no increased cardiovascu-
lar morbidity or raised risk of death, up to age 70 years, was pointed out [221, 369].
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Psychological Implications of PCOS
4
Agata Ando’ and Antonio Maria D’Alessandro
Nowadays, quality of life (QoL) is widely considered an important parameter for

evaluating the quality and outcome of health care, particularly for patients suffering
from chronic disorders: polycystic ovary syndrome is one of these.
Clinical symptoms of PCOS could compromise women’s quality of life and have
a strong negative effect on mood, psychological well-being, and sexual satisfaction.
Physically visible PCOS symptoms are more likely to provoke distress in
younger women than older women [1].
The “American College of Obstetricians and Gynecologists” suggests that, in
view of the high prevalence rate of depression and persistence of new cases in PCOS
population, an initial evaluation of all PCOS women should also include assessment
of mental health disorders.
The PRIME-MD PMQ (Primary Care Evaluation of Mental Disorders Patient
Health Questionnaire) [2] is suitable to evaluate eating disorders [3]; furthermore,
its interpretation and scoring are very simple.
4.1 PCOS Symptoms and Psychological Correlation
4.1.1 Obesity and Body Image
Dissatisfaction with body image is one of the major causes for psychological disor-
ders even in a healthy population; most women affected by PCOS are overweight,
and having a high BMI exposes them to several appearance-related challenges.
Some studies showed that PCOS women have lower quality of life and over-
weight was the largest contributor to poor QoL [4]. In fact, health-related quality of
life questionnaires in women with PCOS have shown that excess weight and diffi-
culties with losing weight are the foremost concerns [5].
Moreover, by using PCOSQ (Health-Related Quality of Life Questionnaire for
Women with Polycystic Ovary Syndrome), it was demonstrated that higher levels

Sensitizers, DOI 10.1007/978-3-319-16760-2_4
64 4 Psychological Implications of PCOS
of BMI related with lower scores (reported by respondent), which is indicative of

several weight-related concerns [6].
Personal negative judgments regarding own body appear to be associated with
the difficulty to begin close and romantic relationships.
Women with PCOS report that they are not happy with the way they look or the
way that clothes fit them and consequently do not feel their body is sexually
appealing [8]: these feelings are negatively associated with self-esteem, body sat-
isfaction, and fear of negative appearance evaluation [9].
In fact, a poor body representation in PCOS women may be conditioned by cul-
tural influences as it has been shown that android fat pattern, commonly associated
with PCOS, is considered unattractive in many cultures [10, 11].
4.1.2 Hirsutism
Women with PCOS recognize excessive hair growth (especially on face) as the
second most severe symptom negatively affecting on their life satisfaction [12].
Some women, in fact, describe themselves using masculine terms such as “beard”
or “mustache,” and they are frustrated because they look at their bodies as a failure
of their femininity [13, 14].
The presence of facial hair is one of the most essential and visible differences
between men and women: hair on a female face reflects a symbolic transgression
between the two genders [15].
As shown in a qualitative study, hirsute women feel “slaves of their own body”
and describe this condition as a “prison” [16]. Moreover, looking in the mirror very
often could represent an obsessive-compulsive behavior [17].
4.1.3 Infertility and Sexual Life
Characteristic symptoms of PCOS occur during a life period in which relationships,

marriage, and having a child play an important role: for this reason, changes in
femininity are likely to mean an increased risk of psychological distress [7].
As any cause of infertility, even PCOS could lead to exaggerated emotional
states depending on lots of variables such as period of time spent in trying to con-
ceive and number of attempted therapies.
Several factors predicting the impact of PCOS-associated infertility upon
HRQoL (health-related quality of life) have been identified: PCOS women who
had been pregnant but had miscarriage experience reported the lowest scores
on the infertility field, exceeding those who had been unsuccessful in having
pregnancy [6].
Some patients are infertile and are subjected to social pressure due to the impor-
tance given to having children by the society.
Having a partner who supports the hope of having a child was found to be a
protective factor and improves the emotional well-being of PCOS patients [18].
4.2 PCOS and Mental Disorders 65
Moreover, according to a study, even adolescent girls with PCOS are 3.4 times
more likely than healthy girls to be “worried about their ability to become pregnant
in the future” compared to the controls; however, this fear was not associated with
odds of having sexual intercourse [19, 20].
Menstrual irregularities are associated to low feminine identity too [13]. Oligo-/
amenorrhea can have important social consequences, especially in many Muslim
backgrounds. For example, the tenets of Islam decree that menstruating women are
not allowed to pray [20]. If a woman prays every day, without the expected monthly
stop of 4–5 days, her social entourage will be aware that she is experiencing men-
strual irregularities [21].
PCOS has also a negative effect on sexual functioning, even when data are
adjusted for BMI; the main reason is the low self-esteem and constant concerns
about their appearance. Based on the study of Elsenbruch et al. “women with PCOS
did not differ from others in the frequency of their sexual activity and sexual
thoughts; they were less satisfied with their sexual life and found themselves less
attractive thinking that their partners find them less attractive and remain sexually
unsatisfied while being with them” [7].
Moreover, in another study a substantial portion of women with PCOS reported
that they most often took the initiative to have sexual intercourse in the relationship
[22, 23]. Could this be related to the increased testosterone levels in PCOS women?
No associations were found. An alternative psychological explanation is that some
women with PCOS felt that their partners were not attracted by them [7].
4.2 PCOS and Mental Disorders
4.2.1 Mood Disorders
Mood disorders include major depressive disorder (MDD), dysthymic disorder, and
depression not otherwise specified based on DSM-IV [24].
In healthy people, depression can cause or exacerbate clinical symptoms such as
fatigue, poor sleep, and changes in appetite and weight. In those with chronic ill-
ness, depression can have more insidious consequences, influencing the expression
and course of disease [25].
Several studies have been investigating the association between PCOS and
depression. The result is that PCOS women reported more depressive symptoms
compared with the control group [7, 26] and scored above average on question-
naires assessing depression [27, 28].
The prevalence of depression in women with PCOS is high, ranging from 28 to
64 % [29–31]. Studies found that 14 % of women suffering from PCOS reported
suicidal ideation. This percentage is high as what has been reported from other
chronic medical conditions and much higher than in the general population [32].
Despite this, there are discordant opinions about the real cause: neither androgeni-
zation nor excessive hair growth showed significant correlation with depression [27]. In
fact, it was not observed any significant differences in total or free testosterone levels or
in the adrenal androgen DHEAS between depressed women with PCOS and non-
depressed women with PCOS [33].
Two-thirds of women with PCOS show weight problems, but it is not properly
correlated only to PCOS: in fact, high BMI might increase depression in the normal
population as well [34–36].
Some studies found depressed women with PCOS to have a higher evidence of
insulin resistance and impaired fasting glucose than PCOS women without depres-
sion [27, 33].
There are plausible physiological connections between depression and insulin
resistance; in fact, depression has been associated with increased cortisol, amplified
sympathetic activity, decreased central nervous system serotonin, and increased
inflammatory markers: these features are also associated with insulin resistance [37].
Depression is also associated with behaviors that worsen insulin resistance,
including unhealthy eating and physical inactivity. These findings may explain why
depression predisposes to diabetes [38].
In view of all these data and because the peak incidence of depression is during
the reproductive years, gynecologists have to be able to identify and treat women
with PCOS who have depression.
4.2.2 Anxiety
According to the DSM-IV, diagnostic criteria for GAD (generalized anxiety disor-
der) include excessive anxiety and apprehension about events or activities, occur-
ring more days than not, for at least 6 months; abnormal anxiety becomes a problem
when it occurs without any recognizable motivation or when the stimulus does not
warrant that kind of reaction [39].
Anxiety symptoms could be identified in one-third of PCOS patients, especially
social phobia [32, 35, 40]. It has been associated mainly with hirsutism [17], acne
[41], obesity [42], and infertility [43].
The prevalence of anxiety in women with PCOS ranges from 34 to 57 % [31, 44].
Fears reported by hirsute women are mainly categorized as “social phobia” or
anxiety-evoking situations, such as meeting strangers, attending parties, shopping,
and mixing at work [6].
PCOS women with higher anxiety scores showed significantly elevated insulin
resistance and FAI (free androgen index) values than PCOS with lower anxiety
score, independently out of BMI [45].
Some authors have suggested that adolescents with PCOS are at higher risk for
anxiety symptoms related to the clinical signs of hyperandrogenism. In a study of
hirsute 13–18-year-old girls, anxiety was diagnosed in 26 % compared with 10 % in
the control girls [46]. Furthermore, successful treatment of hirsutism leads to a
reduction of time spent on hair removal with a consequent improvement in anxiety
score [47].
The risk of developing coexistent depression and anxiety in women with PCOS
is unknown [39]. An interesting study found that 15 % of PCOS patients had
References 67
coexistent anxiety and depression. Coexisting anxiety in depressed patients may

worsen the outcome increasing the risk of suicide, worsening overall symptoms,
conferring a poorer response to treatment, increasing the number of medically unex-
plained symptoms, and increasing functional disability [48].
Most women with PCOS reported sleep disorders: a partial explanation for this
finding might be that sleep apnea is common in obese women with PCOS [49];
androgen excess and subnormal estrogen levels and visceral adiposity may be
involved in sleep disturbances [50].
4.2.3 Eating Disorders
Association between PCOS and eating disorder has been suggested, mainly corre-
lated to the body image dissatisfaction. 6 % of women with PCOS fall into the
bulimic range [51], and moreover, PCOS was more frequently found among bulimic
women [52]. Compared to the general population, eating disorders seem to be more
prevalent in PCOS population: 12.6 % bulimia and 1.6 % anorexia.
Moreover, an epidemiological cohort study of eating disorders among hirsute
women showed a high prevalence of untreated eating disorders, especially EDNOS
(eating disorders not otherwise specified) and bulimia nervosa; hirsute women with
an eating disorder had high levels of comorbid depression and anxiety: they suffered
from lower self-esteem and considered themselves as more hirsute than they really
were [53].
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Diagnosis and Assessment
5
Currently the ESHRE/ASRM or Rotterdam criteria are the agreed international

diagnostic criteria for PCOS [1].
PCOS diagnosis can be raised only after the exclusion of other known causes of
hyperandrogenism and amenorrhea and when there are at least two of the three fol-
lowing parameters:
1 . Oligomenorrhea or anovulatory cycles with menstrual irregularities

2. Elevated levels of circulating androgens or clinical manifestation of

hyperandrogenism
3. Ultrasound evidence of micropolycystic ovaries
5.1 Differential Diagnosis
First of all, to establish a differential diagnosis is a primary goal when a patient

complains of menstrual disorders, infertility, hyperandrogenism, and overweight/
obesity, in order to identify all possible clinical scenarios that are characterized by
symptoms and signs similar to PCOS features.
These are the following:
• Hyperprolactinemia: history of galactorrhea, spontaneous or induced.

• Thyroid dysfunctions: frequent symptoms are hot or cold intolerance, tremors,
diffuse scalp hair loss, weight change, and textural skin changes.
• Ovarian/adrenal androgen-secreting tumors: symptoms of deep virilization such
as increased libido, deepened voice, and clitoromegaly.
• Non-classic congenital adrenal 21-hydroxylase deficiency: this disorder is
caused by a partial adrenal enzyme defect that leads to impaired cortisol produc-
tion, compensatory elevation in adrenocorticotropic hormone, and subsequent
excess androgen production. Premature pubarche could be a clue symptom.

Sensitizers, DOI 10.1007/978-3-319-16760-2_5
72 5 Diagnosis and Assessment
• Cushing’s syndrome: late-onset hirsutism, mood or sleep disturbance, hyperpig-

mented striae, easy bruising, thin/fragile skin, facial plethora, supraclavicular
fullness, excessive thirst, and increased susceptibility to infections.
• Virilizing drugs: anabolic steroids, glucocorticoids, valproic acid, etc.
• Simple obesity.
• Premature ovarian failure or stress amenorrhea.
It is important to assess the onset and evolution of hyperandrogenism signs

because a rapid onset (2–6 months) is suspicious of androgen-secreting neoplasms,
while a slow onset and evolution (especially during the adolescence) is more pecu-
liar of PCOS. Moreover, it is relevant to investigate the possible intake of virilizing
drugs.
5.2 Risk Factors
Anamnesis is important to assess the presence of various risk factors, such as:
• Family medical history positive for:

–– Type II diabetes
–– Hyperandrogenism
–– Impaired glucose tolerance
–– Hyperinsulinemia
–– Obesity
–– Metabolic syndrome
–– Preeclampsia
–– Gestational diabetes
• Personal medical history positive for:
–– Early pubarche
–– Overweight/obesity
–– Macrosomia
–– Sedentary lifestyle
–– Poor dietary habits
5.3 Clinical–Endocrine Features
5.3.1 Oligomenorrhea and Anovulation
Oligomenorrhea is defined as menstrual periods occurring at intervals of greater

than 35 days, with only four to nine periods in a year.
During the early post-menarche years, the menstrual cycles can last between 21
and 45 days [2]. The characteristic menstrual regularity of the adult female is usually
reached several years following menarche; according to some studies, the persistent
5.3 Clinical–Endocrine Features 73
presence of cycles longer than 45 days, 3–5 years following the menarche, suggests
the presence of ovulatory dysfunction in adolescent girls [3].
Progesterone levels <5 ng/mL in days 20–24 (luteal phase) of the menstrual
cycle is a good cutoff to diagnose an anovulatory cycle. In contrast, a patient can be
diagnosed as anovulatory after ascertaining anovulation in at least two subsequent
cycles, in the presence of hypoprogesteronemia.
5.3.2 Hirsutism
Hirsutism can be assessed through the Ferriman–Gallwey score [4] that evaluates
the presence of the terminal hair in the upper lip, chin, chest, upper and lower back,
upper and lower abdomen, thighs, and arms.
A score of 0–4 is assigned to each area examined, based on the visual density of
terminal hairs, such that a score of 0 represents the absence of terminal hairs, a score
of 1 minimally evident terminal hair growth, and a score of 4 extensive terminal hair
growth. Terminal hairs can be distinguished clinically from vellus hairs primarily
by their length (i.e., >0.5 cm), coarseness, and pigmentation. On the contrary, vellus
hairs generally measure <0.5 cm in length and are soft and nonpigmented.
Integrated scores from all body areas beyond 15 points are related to a hirsutism
diagnosis, although current recommendations suggest the use of 95th percentile of
the score in specific populations, adapting to ethnic groups, hair pattern, and age-
related features, in order to properly diagnose hirsutism [4].
This score system has limitations because of the subjective nature of the assess-
ments and the difficulty of evaluating women who have cosmetically removed their
hairs [5].
Moreover, the F–G score was developed in Caucasian adult women and may not
be applicable to younger women from different ethnic backgrounds (e.g., for Indian
women) [6].
5.3.3 Acne
The two commonly used measures to assess the severity of acne are grading and
lesion counting, but no grading system has been universally accepted [7].
In 1956, Pillsbury, Shelley, and Kligman published the earliest known grading
system [8], which includes the following:
• Grade 1: comedones and occasional small cysts confined to the face

• Grade 2: comedones with occasional pustules and small cysts confined to the
face
• Grade 3: many comedones and small and large inflammatory papules and
pustules, more extensive but confined to the face
• Grade 4: many comedones and deep lesions tending to coalescence and canalize
and involving the face and the upper aspects of the trunk
A more recent and complete system is the one created, in 1997, by Doshi,
Zaheer, and Stiller [9], called “Global Acne Grading System (GAGS)”. This
system divides the face, chest, and back into six areas (forehead, each cheek,
nose, chin, chest, and back) and assigns a factor to each area on the basis of
size.
Each type of lesion is given a value depending on severity:
• No lesions = 0
• Comedones = 1
• Papules = 2
• Pustules = 3
• Nodules = 4
The score for each area (local score) is calculated using the formula:
Local Score = Factor ´ Grade ( 0 - 4 )

The global score is the sum of local scores, and acne severity was graded using the
global score. A score of 1–18 is considered mild; 19–30, moderate; 31–38, severe;
and >39, very severe.
5.4 Endocrine Blood Tests
Blood tests should be done within 10 days from the beginning of a menstrual cycle,
during the early follicular phase. Many studies suggest that hyperandrogenemia
may be the most useful diagnostic feature in adolescents because menstrual irregu-
larities, ovarian morphology, and clinical hyperandrogenism do not correlate
strongly with PCOS in this population [10, 11], even if there is a physiological
increase in androgen levels during puberty [12, 13].
The following are the blood substrates and their values characteristic of PCOS.
As explained previously, in the meanwhile, it is crucial to assess other blood
values (TSH, fT3, fT4, anti-TPO, anti-Tg, prolactin, DHEAS, 24 h urinary cortisol
and creatinine) in order to exclude other pathologies:
• LH ≥10 mUI/mL
• LH/FSH ratio ≥2.5
• Estradiol ≥30 pg/mL
• 17-OHP ≤2 ng/mL
If the value is >2 ng/mL (6 nmol/l), it is suspicious of non-classic congenital
adrenal 21-hydroxylase deficiency (NCAH), and ACTH test is required: it is an
acute adrenal stimulation test that measures 17-OHP before and 60 min after the
intravenous administration of an adrenocorticotropic hormone analog. If the
5.4 Endocrine Blood Tests 75
stimulated 17-OHP exceeds 30 nmol/l, and preferably 45 nmol/l, the diagnosis

of NCAH is confirmed [14].
• Androstenedione ≥2.5 ng/mL
• SHBG ≤15 nmol/l
• Testosterone ≥1 ng/mL
A serum testosterone level >200 ng/dL is highly suggestive of an adrenal or
ovarian tumor. If serum testosterone is elevated despite a normal DHEAS level,
an ovarian source is more likely. If a DHEAS level >700 mcg/dL is present
despite a normal serum testosterone level, an adrenal source should be sus-
pected as the cause of hirsutism [4].
Mildly elevated serum testosterone and DHEAS are often present in functional
ovarian hyperandrogenism (FOH) and late-onset congenital adrenal hyperplasia
(CAH).
A very recent study has revealed that PCOS patients with co-elevation of andro-
stenedione and testosterone have impaired indices of insulin sensitivity com-
pared with those with normal androgens or milder hyperandrogenemia [15].
• FAI: free androgen index or FAI is a ratio used to determine abnormal androgen
status in humans. The ratio is the total testosterone level divided by the sex
hormone-binding globulin (SHBG) level and then multiplying by a constant,
usually 100. The concentrations of testosterone and SHBG are normally mea-
sured in nanomoles per liter, while FAI has no units [16, 17].
FAI = 100 ´ ( Total Testosterone / SHBG )

The majority of testosterone in the blood does not exist as free molecule, while
half is tightly bound to sex hormone-binding globulin, and the other half is
weakly bound to albumin. Only a small percentage is unbound (<3 % in
females and <0.7 % in males). Since only free testosterone is able to bind to
tissue receptors to exercise its effects, it is believed that free testosterone is the
best marker of a person’s androgen status. However, free testosterone is diffi-
cult and expensive to measure, and many laboratories do not offer this
service.
The free androgen index is intended to give a guide to the free testosterone level,
but it is not very accurate. Consequently, there are no universally agreed “normal
ranges,” and levels slightly above or below quoted laboratory reference ranges
may not be clinically significant.
Typical values for the FAI in women are 7–10 [18].
• A serum AMH ≥35 pmol/l (or ≥5 ng/mL) appears to be more sensitive and spe-
cific than a USS follicle count >19 [19].
There is not yet an international consortium that validates the threshold for AMH.
In another recent study, Lin et al. have divided all patients into three groups: high
AMH (>11 ng/mL), moderate AMH (4–11 ng/mL), and low AMH (<4 ng/mL).
As the AMH level increased, the prevalence of PCOS increased significantly
from 21 % in the low-AMH group to 37 % in the moderate-AMH group and
80 % in the high-AMH group [20].
5.5 Ultrasound Features
Polycystic ovarian morphology (PCOM) is an important element for the diagnosis

of polycystic ovarian syndrome in adult women. The Rotterdam consensus defined
PCOM as the presence of 12 or more follicles of 2–9 mm in diameter and/or an
ovarian volume greater than 10 mL in at least 1 ovary (Fig. 5.1).
Some studies have shown that a combination of these characteristics is better
than one, to give greater sensitivity and specificity [21–23].
The subjective aspect of the ovaries, their follicular distribution, or the appearance
of the stroma is not considered as important:
• Ovarian volume: there are many formulas available for the calculation of ovarian
volume, but investigators stated that it should be calculated on the basis of the
simplified formula for an ellipsoid: 0.5 × length × width × thickness of the ovary
[21, 23–25].
• Number of follicles: the adoption of the above-cited criterion for defining a poly-
cystic ovary is different from the methodology used in prior works, which
attempted to define PCOM on the basis of the presence of at least ten follicles
arranged peripherally around an echodense stroma [26] in a single US imaging
plane. The key technical requirement for the assessment of the number of folli-
cles is that the number of antral follicles present throughout the entire volume of
the ovary must be counted [27].
• Stromal echogenicity and volume: one of the features of polycystic ovary is the
increased stromal echogenicity [26]. However, the intrinsic echogenicity of the
ovarian stroma is no different in PCOS than in the normal ovary; the subjective
impression of increased stromal echogenicity is due to the increased stromal
Fig. 5.1 Polycystic ovarian morphology

5.5 Ultrasound Features 77
volume, which positively correlates with serum androgen levels [28]. At the
moment, no standardized method is available for this determination. Because
overall ovarian volume correlates well with stromal volume in polycystic ovaries
[28] and is more easily to evaluate, the determination of overall ovarian volume
is a reliable surrogate for ovarian stromal assessment [27].
Moreover, the Rotterdam consensus states that, wherever possible, ultrasounds

should be carried out endo-vaginally particularly in obese patients [1] because of its
better resolution in comparison to the transabdominal route.
In fact, data suggest that, compared with transvaginal US, the appearance of
polycystic ovaries may not be detected at transabdominal US in up to 30 % of
women with PCOS [29].
All ultrasounds have to be carried out in the early follicular phase (3rd–6th day)
in women with regular menstrual cycles or in patients with oligomenorrhea (often
after MAP test); in case of amenorrhea, the study can be carried out on any given day.
A history of oral contraceptive use should be obtained, since oral contraceptive
use causes a decrease in ovarian size, decreasing the sensitivity of US evaluation.
The above-reported criteria are of limited value in the adolescent population
[30]. In primis, transvaginal USS is inappropriate in virginal patients, diminishing
the quality of imaging obtained [31]. Moreover, ovarian morphology changes dur-
ing the lifespan, with a maximum size and antral follicle count around menarche
[32, 33]: in fact, the mean ovarian volume is larger in young women. The consensus
group also cites the difficulty in distinguishing a polycystic ovary from what has
traditionally been referred to as a multifollicular ovary, defined as an ovary in which
there are six or more follicles (usually 4–10 mm in diameter) with normal stromal
echogenicity and described as the common appearance of ovaries in adolescents
[26]. Given these physiological features, many adolescents might meet the adult
criteria for PCOM [34]. For the explained reasons, there is a lack of high quality
evidence on which to base a recommendation on appropriate criteria for ultrasound
diagnosis of PCOM in adolescents [30]: thus, USS results should be interpreted
with caution [34].
Moreover, it is known that ovarian volume and follicle number decrease linearly
with age in women with PCOS and controls, but some studies showed that the fol-
licle number was higher at all ages in PCOS compared with control women [35, 36]:
thus, regarding the follicle number count, it is necessary to create an age-based
criteria to define PCOM [33].
On the other hand, some studies showed that these ultrasonographic findings are
frequent in young women and decrease with age [33, 37, 38]. In adults, these USS
findings may be present in 10–20 % of healthy women with regular menstrual cycles
and without clinical hyperandrogenism [39, 40].
Other data suggest that 23 % of women of reproductive age will have findings
of PCOM [39] and only 5–10 % of these patients will have classic symptoms of
PCOS [41].
The use of polycystic ovaries as an inclusion/exclusion criterion for a diagnostic
test study is controversial; much of the debates, in fact, have arisen from reports of
unusually high rates of polycystic ovaries in healthy women of reproductive age

using the ultrasound-based criteria supported by the Rotterdam consensus [38].
According to a very recent study, an average value of 26 or more follicles per
ovary is a reliable threshold for detecting PCOM in women with frank symptoms of
PCOS; a lower follicle threshold may be required to detect milder variants of the
syndrome [42].
Thus, given the significant number of women of reproductive age who have
PCOM as defined on the basis of US criteria, actually it would be ideal and prudent
to write this observation in the report: “Findings meet the Rotterdam Consensus
definition for polycystic ovaries. In the absence of ovulatory dysfunction or either
clinically or biochemically diagnosed hyperandrogenism, findings are not specific
and do not indicate the presence of polycystic ovarian syndrome” [27].
Summarizing, Lee and Rausch in a recent study propose the “pertinent USS
reporting parameters in PCOS” [27]:
• Separate reporting for each ovary

• Number and size range of follicles
• Size of the largest follicle should be measured in three axes and the average
diameter calculated
• Documentation of any follicle >10 mm or corpus luteum (presence of either sug-
gest the necessity of repeating USS during the next menstrual cycle)
• Ovarian volume, calculated with the simplified formula for an ellipsoid: (0.5 ×
length × width × thickness)
5.6 Clinical–Metabolic Features
As largely explained in the previous chapters, a metabolic evaluation is necessary

for every PCOS patient. From a clinical–metabolic point of view, physician should:
1. Observe the presence of obesity and evaluate the distribution of body fat (gynoid
or android) by assessing
• BMI = weight / height2 kg/m2 (Table 5.1)
• Waist/hip ratio (WHR) >0.80
According to the World Health Organization [43], the waist circumference
(WC) should be measured at the midpoint between the lower margin of the
last palpable rib and the top of the iliac crest, using a stretch-resistant tape that
provides a constant 100 g tension. WC should be <88 cm.
Hip circumference should be measured around the widest portion of the but-
tocks, with the tape parallel to the floor [44].
WHR and BMI are the easiest anthropometric indices to use in clinical prac-
tice to check the effects of adipose tissue on the metabolic profile. However,
BMI and WHR are indirect methods of assessing body composition and, in
some cases, inaccurate, but they permit a first diagnostic level.
5.6 Clinical–Metabolic Features 79
Table 5.1 WHO—weight Weight classification BMI (kg/m2) cutoff

and BMI classification
Underweight <18.5
Mild thinness <16
Moderate thinness 16–16.9
Severe thinness 17–18.4
Normal range 18.5–24.9
Overweight ≥25
Pre-obese 25–29.9
Obesity ≥30
Obese—class I 30–34.9
Obese—class II 35–39.9
Obese—class III ≥40
Particularly, waist circumference should be examined in every PCOS patient

with normal BMI, to identify individuals at higher risk for developing meta-
bolic disorders [45].
• Recently, a Brazilian study group tried to develop equations that utilize
anthropometric measures to estimate intra-abdominal fat (IAF) and total
abdominal fat (TAF) in obese women with PCOS [46]. The anthropometric
variables used are AC (abdominal circumference), WC (waist circumfer-
ence), CC (chest circumference), and NC (neck circumference):
–– Abdominal circumference (AC): it was measured at the point correspon
ding to the navel.
–– Waist circumference (WC): the patient was asked to stand upright, and a
measure of the smallest curvature between the ribs and the iliac crest was
taken without compressing the tissues [47].
–– Trunk circumference (TC): it is measured on the posterior part of the trunk,
3 cm under the armpit. The patient must place her arms parallel to the body [48].
–– Neck circumference (NC): it must be measured at the upper margin of the
thyroid cartilage [49].
The model proposed for TAF was: 4.63725 + 0.01483 × AC – 0.00117 × NC –
0.00177 × CC (R2 = 0.78); the model proposed for IAF was: 1.88541 +
0.01878 × WC + 0.05687 × NC – 0.01529 × CC (R2 = 0.51).
Authors showed that these equations had good correlation with the real value
measured by CT scan and so can be used in clinical practice [46].
• A Korean study group proposed the visceral adiposity index (VAI) to reflect
visceral adiposity and insulin resistance. It is a mathematical model that uses
simple anthropometric (BMI and WC) and functional (triglycerides and HDL)
parameters [50].
The VAI is calculated using the following formula:
é WC / ( 36.58 + (1.89 ´ BMI ) ) ù ´ ( TG / 0.81) ´ (1.52 / HDL ) , where the tri-
ë û
glycerides and HDL concentrations are expressed in mmol/l [50].
This study observed that the VAI was an independent determinant of visceral
adiposity and can replace visceral CT scanning [51].
More than these indirect methods, there are also different direct imaging tech-
niques, more sophisticated and accurate, such as DEXA, MRI, CT scan,
impedentiometry, and ultrasound scan.
• DEXA (dual-energy X-ray absorptiometry): it is based on the ability of the
tissues to attenuate a dual-energy-level photon beam (70 and 140 kV), accor
ding to the mass and density of the tissue passed through. Currently DEXA is
considered a reliable test for the assessment of body composition (BF%)
because of its high sensitivity and reproducibility in the quantification of lean
and fat mass, both in the whole body and region by region; thus, DEXA is
extremely useful for the assessment of body fat distribution and allows an
accurate distinction between abdominal fat (android) and gluteal–femoral fat
(gynoid).
Moreover, it is a method without any radiological risk, as the photon beam is
collimated and the exposure dose emitted is minimal (<1 mrem). Accordingly,
it is an investigation that can be frequently repeated, in order to do a very
close control of the body.
On the other hand, a recent study reported that BMI and WC were more accu-
rate than BF% for classifying the studied metabolic disorders. The BF% cut-
offs most frequently cited by international scientific literature are BF% ≥30
and BF% ≥35 for women: they produce a high rate of false positives [52].
• CT scan: CT scan, compared to DEXA, offers the advantage of a discrimina-
tion between subcutaneous and visceral fat, and it is able to quantify the pro-
portion of fatty tissue that best correlates with cardiovascular and metabolic
risk, which is the visceral fat.
The adipose tissue areas are calculated by computing the fat area surfaces
with an attenuation range of −190 to −30 Hounsfield units. The abdominal
visceral fat area (VFA) is measured by drawing a line in the muscle wall sur-
rounding the abdominal cavity, at the level of L4–L5 vertebral bodies [53].
The cutoff value of visceral fat area associated with an increase risk of obesity-
related disorder, according to the literature, is 103.8 cm [2] (sensitivity
74.5 %, specificity 64.7 %) [54].
Thus, women with VFA >100 cm2 are at high risk of metabolic syndrome
[51].
However, the high cost, long scan times (possible artifacts due to involuntary
movements by the patient), and exposure to a non-negligible radiological risk
constitute a strong limitation, and so a repeated use for the measurement of
body composition is not recommended.
• Impedentiometry: this method estimates the body composition based on the
physical principle of the different electrical conduction of the tissues, in rela-
tion to their content of water and electrolytes. The conduction of an electric
current is in fact much higher in lean than fat tissue. By applying a weak
electrical current to the body and detecting the impedance presented by the
body itself to the passage of this current, it is possible to calculate the quantity
of total body water (TBW). The advantages of this technique are easy way to
use, the absolute noninvasiveness, and measurement speed. The limitations of
5.7 Metabolic Blood Tests 81
the method relate to the situations with changes in hydration body and/or in
the electrolyte concentration.
2. Finally, from a clinical–metabolic point of view, the physician should verify the
presence of hypertension or clinical signs of hyperinsulinemia such as acantho-
sis nigricans (AN).
AN is a skin lesion appearing thickened and velvety brown streaking to a
leathery, verrucous, papillomatous change. It usually occurs on the neck or in
skinfolds. Microscopically, AN is characterized by an increased number of mela-
nocytes, with papillary hypertrophy and hyperkeratosis [45]. Benign acanthosis
nigricans usually correlates to insulin resistance or obesity [55, 56].
A very recent Chinese study showed that the presence of AN correlates to
insulin resistance and reduced HDL-C level in PCOS patients with normal
BMI. Because of its easy identification, AN could be a reliable marker of IR, but
it lacks the sensitivity to become a noninvasive diagnostic tool for IR in women
with normal BMI [45].
5.7 Metabolic Blood Tests
As reported previously, ~50–70 % of women with PCOS are characterized by

hyperinsulinemic insulin resistance, which plays a causative role in the develop-
ment of the metabolic syndrome.
The American College of Obstetricians and Gynecologists currently recom-
mends screening women with PCOS for glucose intolerance and lipid
abnormalities.
Metabolic Blood Tests Recommended in PCOS Women

• OGTT (biannual rescreening) or HOMA-IR
• Total cholesterol + LDL-C + HDL-C
• Triglycerides
• Transaminases
5.7.1 G
lucose Metabolism Assessment and Calculation
of Insulin Resistance
First of all, a correct evaluation of carbohydrate metabolism is necessary, and so it

is important to remember which are the normal glycemic and insulinemic values.
• Fasting glycemia:
–– Normal, 70–100 mg/dL
–– “Grey zone,” 100–126 mg/dL
–– Diabetes, >126 mg/dL
• Fasting insulinemia: <10 mUI/mL
The World Health Organization (WHO) criteria for impaired fasting glucose
(IFG) differ from the American Diabetes Association (ADA) criteria because the
normal range of glucose is differently defined.
The WHO has defined the upper limit of normal at less than 110 mg/dL.
However, fasting glucose levels of 100 mg/dL and higher have been shown to
significantly increase complication rates, and so the ADA has accordingly lowered
the upper normal limit to a fasting glucose under 100 mg/dL.
• WHO IFG criteria: fasting plasma glucose level from 6.1 mmol/l (110 mg/dL) to
7 mmol/l (126 mg/dL).
• ADA IFG criteria: fasting plasma glucose level from 5.6 mmol/l (100 mg/dL) to
7 mmol/l (126 mg/dL) [57].
• OGTT (oral glucose tolerance test): the most accurate method to diagnose insu-
lin resistance is the OGTT after 75 g glucose challenge, even in adolescent
women [58]. Normal values are the following:
–– Glycemia:
• Fasting, 70–100 mg/dL
• 60 min after glucose administration, <180 mg/dL
• 120 min after glucose administration, <140 mg/dL
Impaired glucose tolerance (IGT) is defined when glucose level is >140 mg/dL 2
h after glucose load, but <200 mg/dL.
Diabetes is defined when glycemia is >200 mg/dL 2 h after glucose load.
–– Insulinemia:
• Fasting, <10 mUI/mL
• 60 min after glucose administration, <60 mUI/mL
• 120 min after glucose administration, ≈10 mUI/mL
Of course, the majority of PCOS patients are not diabetic yet, but only insulin
resistant; insulin resistance is defined when insulin value 1 h after OGTT is
>60 mUI/mL and/or its level is not very close to the fasting insulin value after 2 h
post-glucose administration.
It has been suggested that an OGTT should be performed every 2 years for those
with normal glucose tolerance and annually if IFG or IGT is present [59].
Glucose screening recommendations for PCOS women are summarized in
Table 5.2
• HOMA index: OGTT is not a very confortable method, and it is also expensive
and time-consuming; for this reason, the need for a simple way of measuring
insulin resistance has led to the creation of a large number of insulin sensitivity
indices [60, 61]. The most used model is the HOMA index.
The homeostatic model for assessment of insulin resistance (HOMA-IR) is a
simple and noninvasive method of estimating insulin sensitivity from the steady
glucose and insulin concentrations measured under fasting conditions.
It was calculated using the following formula [62]:
Fasting glycemia ( mmol / l ) ´ Fasting insulinemia ( mUI / mL ) / 22.5

5.7 Metabolic Blood Tests 83
Table 5.2 Glucose assessment in PCOS patients—screening recommendations

AACE (American Association of OGTT for all PCOS patients aged >30 year. Periodically
Clinical Endocrinologists) reassess
ACE (American College of
Endocrinology)
RCOG (Royal College of Screen all by fasting glucose regularly. If fasting glucose
Obstetricians and Gynaecologists) >100 mg/dL or BMI >30 kg/m2 or positive family
history, screen by OGTT
ACOG (American College of OGTT for all women with PCOS. Repeat every 2 years
Obstetricians and Gynecologists)
AE-PCOS (Androgen Excess and OGTT for PCOS women with BMI >30 kg/m2 or in lean
Polycystic Ovary Syndrome women with advanced age (>40 years), personal history
Society) of gestational diabetes, or family history of diabetes
mellitus type II
HOMA index values for percentiles 50–75 ranged from 2.07 to 2.83 [63].
• Glucose/insulin ratio: this simple measure of insulin resistance in PCOS women has
been correlated well with more complicated dynamic tests of insulin action [64].
It has been reported that a fasting G:I ratio of 4.5 or less is a measure of IR in obese
PCOS women, and this cutoff value has a sensitivity of 95 %, specificity of 84 %,
positive predictive value of 87 %, and negative predictive value of 94 % [65].
• Diabetes screening
Recently, a study group from Holland proposed a stepwise screening for glucose
metabolism abnormalities by fasting glucose for all women with PCOS and sub-
sequent OGTT screening for diabetes in the small proportion of PCOS women
with fasting glucose concentration between 110 and 126 mg/dL only, without
compromising early diagnosis of diabetes [66].
However, validation of this new screening algorithm is waited.
Previously, it has been shown that fasting glucose rather than OGTT underesti-
mates the prevalence of diabetes mellitus type II in PCOS women by >50 % [67].
Hemoglobin A1c is a commonly used marker of chronic glycemia, and it reflects
the average blood glucose levels over a 2–3-month period [68].
ADA suggests HbA1c levels as a screening tool for diabetes and prediabetes in
the general population with cutoff levels of 6.5 and 5.6 %, respectively [69], even
if other studies stated that it is insensitive for prediabetes [68].
5.7.2 Lipid and Hepatic Profile
Dyslipidemia is common in PCOS and is present in up to 70 % of subjects [59, 70].

The AE-PCOS Society consensus statement [59] recommends a complete lipid
and hepatic profile in all patients with PCOS. Pathological values are:
• Total cholesterol >200 mg/dL

• LDL cholesterol >130 mg/dL
• HDL cholesterol <50 mg/dL

• Triglycerides >150 mg/dL
• AST >30 U/l
• ALT >35 U/l
• γ-GT >38 U/l
The fatty liver index (FLI) is an algorithm based on BMI, waist circumference,
triglycerides, and γ-GT and might serve as a simple and accurate predictor of
hepatic steatosis in general population. FLI <30 rules out fatty liver disease, while
FLI >60 indicates fatty liver disease [71]. FLI is calculated by the following
formula:
 e0.953×loge( triglycerides ) + 0.139×BMI + 0.718×loge(g − GT ) + 0.053× waist circumference −15.745 


1+ e  ×100
0.953× loge ( triglycerides ) + 0.139× BMI + 0.718× loge (g − GT ) + 0.053× waist circumference −15.745 
 
A recent study revealed that high FLI levels are a common finding in obese
PCOS women and are closely linked to metabolic syndrome. Thus, FLI might
be a useful index to identify PCOS women at high metabolic and hepatic risk in
whom a very careful surveillance is needed and who might benefit from lifestyle
counseling [72].
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PCOS Therapy
6
Gynecologists usually treat PCOS only as an endocrine disorder, without recognition

of the very important part that insulin resistance plays in the syndrome.
In this section, the way to treat PCOS from a metabolic point of view, without
dwelling on the use of oral contraceptives and antiandrogen drugs, will be
discussed.
Lifelong strategies that improve the care of women with PCOS are essential,
because of the chronic nature of the syndrome and the young age at which all the
symptoms begin to manifest [1].
A valid therapeutic protocol for PCOS includes diet, physical exercise, and
insulin-sensitizing agents such as metformin and inositol.
For example, in fact, a normal BMI is associated with a positive fertility out-
come, and fertility specialists recommend achieving this BMI before IVF (in vitro
fertilization): in fact, these techniques are invasive and expensive and have low suc-
cess rates, so it seems logical to improve BMI and to support hormonal balance
through diet, exercise, and nutrition supplements [2].
6.1 Diet and Exercise
As explained previously, a few evolutionary biologists suppose that many genetic

hormonal tendencies contributing to PCOS have their origin in the switch from
the pre-agrarian age diet to the current diet. The rapidly increasing rates of diabe-
tes, heart disease, and PCOS coincide with the rapid changes in the modern human
diet [2].
All women suffering from PCOS benefit from dietary therapy and exercise; in
fact, dietary and lifestyle interventions are considered among the first-line treat-
ments for PCOS.
There is no PCOS diet that will reverse the syndrome, but there are several
dietary principles that a patient should follow to improve the symptoms.

Sensitizers, DOI 10.1007/978-3-319-16760-2_6
90 6 PCOS Therapy
Weight reduction leads to improvements of insulin sensitivity [3] and lipid pro-
file [4]; it ameliorates hyperandrogenism (SHBG increase, FAI and testosterone
decrease) and menstrual cycle rhythm [4–6], with reductions in adiposity from the
truncal–abdominal area [5]. Moreover, there is evidence that these changes exert
important beneficial effects also in the longer term on disorders such as type II dia-
betes mellitus, cardiovascular disease, and certain cancers (endometrial, breast, and
colon cancer), compared with weight loss alone [7–9].
In most of the dietary studies in women with PCOS, improvements in metabolic
and reproductive outcomes have been closely related to improvements in insulin
sensitivity, suggesting that dietary modification (qualitative and quantitative)
designed to improve insulin resistance may produce greater benefits than those
achieved by energy restriction alone [7].
Clinicians prescribing lifestyle modifications must consider the patient’s capa-
city to sustain diet and exercise adherence and weight maintenance over time for the
clinical benefits on PCOS to continue.
Considering how difficult it is for many patients to change their lifestyle, phar-
maceutical modification of weight control could be an additional necessary thera-
peutic tool, such as the lipase inhibitor orlistat [10].
In some studies on overweight and obese women with PCOS, the use of orlistat
has demonstrated an improvement in both metabolic and hormonal parameters
[11, 12].
Orlistat is an antiobesity drug with minimal systemic absorption, and therefore,
any effect of this drug is a result of weight loss and not the direct effect on ovaries.
The proposal therapeutic dose is 120 mg three times daily, before each meal, for
3 months, during which the patient must be able to lose at least 5 % of its total
weight.
6.1.1 PCOS Dietary Recommendations
1. Reduce total calories consumed to standard levels for sex, age, and activity:
calories requests are higher for women with higher BMI and increase in relation
to activity. It is often useful to initially focus on the eating pattern and the
macronutrient content of the diet rather than to try to promote both healthy eat-
ing and weight loss too quickly [8]. Energy consumption can be reached by
limiting nutrient intake or by increasing calories expenditure.
A daily calories deficit of 200 kcal/day will prevent weight gain; a deficit of
500 kcal/day is needed for the average person to lose 0.5 kg/week, while a
1,000 kcal deficit is needed for 1 kg weight loss/week [8].
2. Reduce refined carbohydrates in favor of complex carbohydrates. “Refined” car-
bohydrates refer to a carbohydrate-based food that has been processed to strip it
of some of its original fiber and unpackaged to produce a more “refined” pro-
duct. For example, sugar cane and corn on the cob are whole foods, but the table
sugar that is processed out of sugar cane and the cornstarch or high fructose corn
syrup processed out of the corn are some refined carbohydrates [2].
6.1 Diet and Exercise 91
A period of relatively strict carbohydrate restriction helps at the beginning of

the diet; a recent study demonstrated that a reduced-carbohydrate diet results in
lower measures of β-cell responsiveness and circulating insulin (27 % reduction
in fasting insulin) when compared with a standard higher-carbohydrate diet [13].
Other studies have reported improvements in LDL cholesterol particle size,
LDL concentration, and postprandial blood lipid profile [14–16].
On the other hand, low-carbohydrate diets have been associated with deleteri-
ous effects on lipid profile when used long term [17], and so severe carbohydrate
restriction should be use as a short-term measure to achieve weight loss [8].
3. Eat low-glycemic index (GI) foods: a few studies have shown that a low-GI diet
can improve insulin resistance as well as many of its metabolic consequences
including increasing HDL and plasminogen activator inhibitor-1 levels [18, 19].
Moreover, several epidemiological studies have also associated a low-GI diet
with reduced risk of CVD and type II diabetes [20, 21].
A high-GI diet, on the other hand, has been shown to worsen postprandial
insulin resistance [22]: in fact, a recent study showed that a low-GI diet improves
insulin sensitivity and menstrual regularity more than a conventional healthy,
moderate- to high-GI diet despite similar weight loss [23].
4. Increase fiber to improve glucose regulation: fiber helps to slow the digestion of
carbohydrates and improves insulin resistance [24, 25].
5. Increase high-protein foods: it was demonstrated that proteins consumed at
breakfast (compared with lunch or dinner) lead to a greater initial and sustained
feeling of fullness, increased satiety, and reduced concentrations of the appetite-
regulating hormone ghrelin [26–28].
Adequate protein intake is important to protect lean body mass and to increase
muscle in response to exercise [8]. Actually, there is little evidence to suggest
benefits of high-protein diets on insulin resistance, and a number of studies in
women with PCOS have failed to show significant long-term benefits of a high-
protein diet on weight loss or insulin sensitivity [16, 29]; there are also concerns
about the safety of high-protein, low-carbohydrate diets including the effects of
kidney function and bone mineral density [7].
6. Increase food rich in omega-3 essential fatty acids (PUFAs): they have an impor-
tant role in human cell metabolism; an US study investigated the positive effects
of a polyunsaturated fatty acid (PUFA)-rich diet in PCOS patients [28], but fur-
ther research is required to determine real beneficial and harmful effects of vari-
ous PUFAs in insulin-resistant populations.
7. Meal timing: the frequency and regularity of eating patterns are important, even
if there are small data in the literature.
One of the largest studies [29] conducted revealed that those who ate frequently
during the day had higher intakes of carbohydrates, fibers, and a range of micro-
nutrients, while those who ate less frequently had higher intakes of fat, chole-
sterol, protein, and sodium.
Other studies showed that a regular meal frequency leads to higher post-
prandial energy expenditure, lower energy intake, and improved impaired
insulin sensitivity compared with irregular eating in 2-week interventions
92 6 PCOS Therapy
[30]. In a further study, breakfast consumption was associated with a lower

energy intake and improved insulin sensitivity compared with breakfast
omission [31].
Data in literature show that a diet with 50 % of total calories from carbohydrates
(with a low glycemic index), 30 % from fat (mostly mono- and polyunsaturated fat,
less than 10 % from saturated fat), 20 % from proteins, and high in fiber is the most
appropriate for patients with PCOS [32].
The optimal frequency of food intake has yet to be determined: however, a regu-
lar pattern with low intake from snacks is advisable [8], and high-calorie intake at
breakfast with reduced intake at dinner is suggested, because it leads to reduced
overall insulin levels [33–35].
6.1.2 Glycemic Index (GI)
It has been shown that eating foods with a low GI improves glucose control in
women with PCOS and diabetes.
The glycemic index indicates the rate in which glycemia increases after taking a
quantity of “X” food containing 50 g of carbohydrates.
Foods with carbohydrates that break down quickly during digestion and release
glucose rapidly into the bloodstream tend to have a high GI; foods with carbohy-
drates that break down more slowly, emitting glucose more gradually into the
bloodstream, tend to have a low GI [2].
The concept was developed by Dr. David J. Jenkins and colleagues [36] in
1980–1981 at the University of Toronto in their research to find out which foods
were best for people with diabetes. A lower glycemic index suggests slower rates
of digestion and absorption of the foods’ carbohydrates and may also indicate
greater extraction from the liver and periphery of the products of carbohydrate
digestion.
A lower glycemic response usually relates to a lower insulin demand but not
always and may improve long-term blood glucose control and blood lipids [37].
The glycemic index of a food is defined as the incremental area under the 2-h
blood glucose response curve (AUC) following a 12-h fast and ingestion of a food
with a certain quantity of available carbohydrate (usually 50 g). The AUC of the
test food is divided by the AUC of the standard (either glucose or white bread,
giving two different definitions) and multiplied by 100. The average GI value is
calculated from data collected in ten human subjects. Both the standard and test
food must contain an equal amount of available carbohydrate. The result gives a
relative ranking for each tested food [38]. The GI Symbol Program is an indepen-
dent worldwide GI certification program that helps consumers identify low-GI
foods and drinks. The symbol is only on foods or beverages that have had their GI
values tested according to the standard and meet the GI Foundation’s certification
criteria as a healthy choice within their food group. GI cutoffs are listed in
Table 6.1.
Table 6.1 Glycemic index Glycemic index cutoffs

cutoffs
High ≥70
Moderate 50–70
Low <50
Of course, the glycemic index has also its limitations: the index calculations are
not accurate because the behavior of foods in different individuals can change, and
judging the diet by GI alone does not give the whole portrait of the diet [2].
Moreover, GI values depend on how foods are cooked: cooked carrots have a
higher GI than raw carrots because cooking breaks down the fiber and the glucose
can be absorbed much more quickly. Cooking with a bit of salt or vinegar may lower
the GI of many vegetables because this causes many molecules, not just the sugars,
to be broken down, which results in trapping some of the starches in complex struc-
tures that are digested more slowly [2].
Furthermore, for some people, a food consumed in the morning on an empty
stomach will spike the blood sugar more than the same food eaten later in the day
after having breakfast: patients with good blood sugar control in general will show
less of a spike in blood sugar than someone with poor blood sugar control [2].
PCOS women should follow some useful GI advices for their daily diet:
• Eat five to ten different whole fresh fruits, vegetables, and legumes each day.
• Avoid a diet that consists predominantly of the food highest on the glycemic
index.
• Substitute foods high on the GI with foods lower on the GI: for example, eat
boiled green beans (GI of 15) instead of boiled potatoes (GI of 100) with dinner
(Table 6.2).
• Increase fiber intake: fiber helps to slow the digestion of carbohydrates and
improves insulin resistance. If a food high on the GI is loved, patient should take
care not to consume it often and aim to eat only a small portion of it combined
with high-fiber foods that reduce the glycemic index [24, 25].
• Eat legumes to lower the high-GI foods in the meals: legumes are low on the GI
and contain an impressive amount of fiber and good-quality protein, which can
serve to blunt the glycemic load. Moreover, legumes contain pinitol, a relative of
D-chiro-inositol, noted for improving insulin resistance [2].
• Avoid overeating foods high on the glycemic index. The GI of a food can be tem-
pered by the quantity consumed. For example, a piece of candy might have a very
high glycemic index, but eating just one little piece will not result in a high glyce-
mic load on the body; if the patient eats two pieces of white toast, jam, brown
potatoes, and a sugar- or corn syrup-sweetened fruit drink for breakfast, she is
putting a high glycemic load on her body, and the blood sugar will remain high for
several hours as her body works to process the large amount of high-GI foods [2].
• Evaluate the whole meat, rather than individual food items, to make sure the
patient is preparing meals that will not spike her blood sugars.
94 6 PCOS Therapy
Table 6.2 Foods’ glycemic Foods’ glycemic index

index list
Sweeteners
Corn syrup 100
Table sugar (sucrose) 100
Rice syrup 65
Honey 54
Fructose 10
Stevia 0
Grains
White rice 90
Rice cakes 84
Wild rice 81
Corn chips 72
Cornmeal 70
Couscous 65
Brown rice 55
Pop corn 55
Whole wheat 48
Whole amaranth 35
Bread
Rice bread 100
Polenta 98
Baguettes 95
Doughnuts 76
Croissant 70
White bread 70
Pancakes 67
Kamut bread 54
Rye bread 50
Pasta, whole grain 44
Wheat germ 15
Cereals
Instant oats 92
Puffed rice 85
Grape-Nuts 67
Oat bran 15
Nuts and seeds
Chestnuts 60
Peanut butter 40
Sesame seeds 35
Almonds 15
Hazelnuts 15
Pistachios 14
Table 6.2 (continued) Foods’ glycemic index

Walnuts 14
Peanuts 14
Legumes and beans
Fava beans 50
Black beans 35
Hummus 35
White beans 35
Lentils 29
Soybeans 18
Green beans 15
Tofu 14
Vegetables
Potatoes, baked 100
Potatoes, boiled 84
Carrots, cooked 80
Beets, cooked 64
Corn 55
Peas 44
Coconut 35
Tomato sauce 35
Carrots, raw 30
Asparagus 15
Cucumbers 15
Lettuce 15
Mushrooms 15
Olives 15
Spinach 15
Tomatoes 15
Zucchini 15
Avocados 10
Fruits
Watermelons 90
Pineapples 66
Apricots 57
Strawberries 56
Mangos 55
Bananas 52
Grapes 50
Oranges 46
Apples 39
Peaches 30
Raspberries 25
(continued)
96 6 PCOS Therapy
Table 6.2 (continued) Foods’ glycemic index

Cherries 25
Others
Beer 110
Rice milk 84
Mango juice 55
Orange juice 45
Coconut milk 40
Soy milk 36
Yogurt, low-fat fruit 33
Almond milk 30
Dark chocolate 25
Lemon juice 20
Pesto 15
Vinegar 5
Water 0
6.1.3 Glycemic Load (GL)
Some authors believe that the glycemic load (GL) is a more useful measure of food
value than the glycemic index alone.
Glycemic load accounts for how much carbohydrate is in the food and how much
each gram of carbohydrate in the food raises blood glucose levels.
GL is a GI-weighted measure of carbohydrate content [39].
GL = GI ´ carbohydrate ( grams ) / 100

For instance, watermelon has a high GI, but a typical portion of watermelon does
not contain many carbohydrates, so the glycemic load of eating it is low. Whereas
glycemic index is defined for each type of food, glycemic load can be calculated for
any size serving of a food, an entire meal, or an entire day’s meals.
GL cutoffs are listed in Table 6.3.
Foods that have a low GL in a typical serving size have usually a low GI. Foods
with an intermediate or high GL in a typical serving size range from a very low to
very high GI (Table 6.4).
For detailed information about all the foods, visit the website www.glycemicin-
dex.com
Table 6.3 Glycemic load Glycemic load cutoffs

cutoffs
High ≥20
Intermediate 11–19
Low <10
Table 6.4 Foods’ glycemic Glycemic load (per 100 g serving)

load list
Baguette 15
Banana 16
Potato 20
Carrots 2
Rice 30
Watermelon 4
6.1.4 Insulin Index
The insulin index is a measure used to quantify the typical insulin response to vari-
ous foods. The index is similar to the glycemic index and glycemic load, but rather
than relying on glycemia levels, the insulin index is based upon insulinemia. This
measure can be more useful than either the glycemic index or the glycemic load
because certain foods (e.g., lean meats and proteins) cause an insulin response
despite there being no carbohydrates present, and some foods cause a disproportio-
nate insulin response relative to their carbohydrate load [40].
Holt et al. have noted that the glucose and insulin scores of most foods are highly
correlated, but high-protein foods and bakery products that are rich in fat and refined
carbohydrates “elicit insulin responses that were disproportionately higher than
their glycemic responses” [40]. They also conclude that insulin indices may be use-
ful for dietary management and avoidance of non-insulin-dependent diabetes mel-
litus and hyperlipidemia.
Glycemic Index (GI) considers each food relative to eating 100 % glucose, while
the insulin index is relative to eating white bread (GI of ~70 to 75) (Table 6.5).
6.1.5 Exercise
Exercise reduces insulin resistance by two mechanisms. It induces a reduction in

visceral fat even if it results in moderate weight loss and BMI reduction [41].
Visceral fat is more metabolically active than subcutaneous fat and central adiposity
is more closely related to IR [32].
Exercise, besides, increases muscle cell metabolism: it modulates the expres-
sion or the activity of proteins mediating insulin signaling in the skeletal muscles
[41, 42].
It has been shown that exercise improves menstrual abnormalities and restores
ovulation in obese patients with PCOS [43], and its benefit on reproductive function
is greater than the benefit of low-calories diet only [44].
Exercise exerts its beneficial effects on body composition with a 45 % greater
reduction in fat mass and a 60 % better preservation of fat-free mass [45].
In fact, it is important to clarify that improved abdominal obesity and insulin
sensitivity may occur without a total change in body weight: body composition of
98 6 PCOS Therapy
Table 6.5 Foods’ glycemic Glycemic index Insulin index

and insulin index list
Porridge 60 ± 12 40 ± 4
Muesli 43 ± 7 46 ± 5
Cornflakes 76 ± 11 75 ± 8
Average: 59 ± 3 57 ± 3
White bread (baseline) 71 ± 0 100 ± 0
White pasta 46 ± 10 40 ± 5
Brown pasta 68 ± 10 40 ± 5
Brown rice 104 ± 18 62 ± 11
French fries 71 ± 16 74 ± 12
White rice 110 ± 15 79 ± 12
Whole-meal bread 97 ± 17 96 ± 12
Potatoes 141 ± 35 121 ± 11
Eggs 42 ± 16 31 ± 6
Cheese 55 ± 18 45 ± 13
Beef 21 ± 8 51 ± 16
Lentils 62 ± 22 58 ± 12
patients who exercise regularly may change with increased lean body mass and
decreased fat mass, but no overall change in weight [8].
At the moment, there are no guidelines for the type, intensity, frequency, and
duration of exercise in patients with PCOS [45, 46].
6.1.5.1 PCOS Exercise Recommendations

1. Moderate-intensity aerobic physical activity (e.g., brisk walking) for at least
30 min and for at least 5 days per week should be recommended in all PCOS
patients [32].
2. Vigorous-intensity aerobic activity (e.g., jogging) for at least 20 min and for at
least 3 days per week or combinations of moderate- and vigorous-intensity exer-
cise can also be recommended [32].
3. Resistance training for at least two nonconsecutive days per week [32].
4. Endurance exercise: for patients who cannot manage high-intensity exercise,
prolonged lower-level activity is an appropriate way to gain fitness and to
increase energy expenditure [8].
6.2 Insulin-Sensitizing Agents and Statins
Examining scientific literature, studies are very conflicting to each other, and a
unanimous opinion on the effectiveness of insulin-sensitizing drugs has not yet been
reached.
According to the ASRM Committee of 2008, insulin-sensitizing agents should
be considered in patients with impaired glucose tolerance (IGT) and PCOS [47].
6.2 Insulin-Sensitizing Agents and Statins 99
Particularly, in 2010, AE-PCOS Society consensus treatment emphasized that

metformin should be used in women with PCOS who have already started lifestyle
treatment (diet and exercise) and do not have improvement in IGT or in those who
have normal weight but still having IGT [48].
When administered to insulin-resistant patients, these drugs act to increase target
tissue responsiveness in order to reduce hyperinsulinemia [49].
In the past, limited studies on the use of diazoxide, acarbose, and somatostatin
for PCOS women were conducted; then, thiazolidinediones aroused more interest,
while, to date, metformin is the most worldwide studied insulin-sensitizing agent.
Moreover, statins have also been used to improve lipid profile in PCOS women.
6.2.1 Thiazolidinediones
Thiazolidinediones (TZDs) include pioglitazone, rosiglitazone, and troglitazone:

during the past, they have been used in PCOS women to reduce insulin resistance.
TZDs are selective ligands of the nuclear transcription factor peroxisome
proliferator-activated receptor-γ (PPAR-γ) [50].
They exert their insulin-sensitizing actions by two mechanisms:
• Promoting fatty acid uptake and storage in adipose tissue

• Increasing the expression of adiponectin, an adipocytokine with an insulin sensi-
tivity effect [51]
Obese women with PCOS who were administered troglitazone demonstrated

benefit in insulin sensitivity, glucose tolerance, and hyperandrogenemia [52–56].
It was demonstrated that even pioglitazone, in doses of 30 mg/day for
3 months, significantly improved insulin sensitivity, hyperandrogenism, and ovu-
lation rates [56].
TZDs were shown to be more effective than metformin in reducing levels of free
testosterone and DHEAS after 3 months of treatment, but this benefit was not evi-
dent after 6 months of therapy [57].
Pioglitazone is able to produce a significant reduction in the incidence of
impaired glucose tolerance and 40 % reversion of previous IGT to normal in PCOS
patients treated with 45 mg daily for 6 months [58]. Significant improvements of
insulin effectiveness in the liver and skeletal muscle, with substantial increase of
circulating adiponectin levels, were also reported [59].
Moreover, some studies demonstrated a clear capacity of pioglitazone to reduce
free fatty acid level in PCOS patients, by decreasing lipolysis and increasing lipo-
genesis [60]; conversely, other studies failed to show any improvement in lipid pro-
file [61].
Additionally, some studies indicate a reduction of inflammatory markers in
pioglitazone-treated PCOS women [62], while others do not [58].
A randomized study using treatment with pioglitazone showed that the latter
increased ovulation frequency [63]. The regulation of ovulation could in turn restore
100 6 PCOS Therapy
normal feedback effects of luteal steroids, normalize serum LH levels, and improve
ovarian steroidogenesis [64]. Additionally, pioglitazone was shown to ameliorate
GnRH-stimulated LH secretion [56].
Administration of pioglitazone during ovarian stimulation period seems to
improve ovarian response to controlled ovarian stimulation in PCOS patients, in
terms of clinical pregnancy rate, as well as risks of ovarian hyperstimulation syn-
drome and multiple pregnancies [64].
Previously, TZDs have been accused of inducing weight gain and water reten-
tion, but recent studies have disconfirmed this supposition [65].
However, the primary concern with TZDs is the liver toxicity: a significant num-
ber of cases of hepatic necrosis were reported in patients using troglitazone, which
was withdrawn from the market in 2000.
Pioglitazone safety in women under 18 is not yet established, so it is not recom-
mended in this female PCOS subgroup.
However, in clinical practice, neither pioglitazone nor rosiglitazone is routinely
used in PCOS women, especially with infertility issues, because TZDs are classified
as pregnancy category C by the FDA, due to the fact that studies in animals have
shown adverse fetal effects such as IUGR [64].
6.2.2 Metformin
Despite there is no universal consensus on metformin benefits in PCOS, in this

chapter all the beneficial effects of metformin therapy in patients with PCOS are
highlighted.
The positive effects of metformin have been demonstrated in nondiabetic women
with PCOS, and they are associated with increased menstrual cyclicity, improved
ovulation, and reduction in circulating androgen levels [66].
To date neither in Europe nor in the United States metformin has been approved for
the treatment of insulin resistance associated with PCOS: its use should be restricted to
those patients with IGT [67]; however, it is largely prescribed as an “off-label” drug.
For “off-label” use of any medication, it is extremely important to fulfill several
criteria for safe use:
• The condition should have health consequences significant enough to warrant

treatment.
• The treatment should have demonstrated safety and efficacy.
• The proposed treatment should be superior to the presently available alternatives
[68].
6.2.2.1 Mechanism of Action

Metformin is a second-generation biguanide used as an oral antihyperglycemic
agent, and it is approved by the US Food and Drug Administration (FDA) as treat-
ment for type II diabetes mellitus.
It is considered an insulin-sensitizing agent because it lowers glucose levels

without increasing insulin secretion, but improving insulin sensitivity.
Metformin causes [69, 70]:
• Increased peripheral insulin sensitivity, by activating glucose transporters

(GLUTs) which allows passage of glucose into hepatic and muscle cells
• Inhibition of hepatic glucose production
• Reduction of circulating free fatty acid concentrations, which helps in reducing
gluconeogenesis
Metformin activates the adenosine monophosphate (AMP)-activated protein kinase

pathway (AMPK) [71, 72]: phosphorylation of threonine in AMPK is necessary for
metformin action, resulting in decreased glucose production and increased fatty acid
oxidation in hepatocytes, skeletal muscle cells [73], and mouse ovarian tissue [74].
Furthermore, metformin inhibits hepatic gluconeogenesis through an AMP-
activated protein kinase-dependent regulation of the orphan nuclear receptor small
heterodimer partner (SHP) [75, 76].
Importantly, the actions of metformin are not associated with an increase in insu-
lin secretion and, consequently, with hypoglycemia.
Metformin affects ovarian function in a dual mode:
• Alleviation of systemic insulin excess acting upon the ovary, particularly on ste-
roidogenesis and follicular growth
• Direct ovarian effect
Furthermore, metformin acts at the hypothalamic level on AMPK pathway: the

latter is essential in the modulation of LH secretion [77].
During the last two decades, some studies demonstrated that metformin inhibits
androstenedione and testosterone production from theca cells through inhibition of
the steroidogenic acute regulatory protein and 17α-hydroxylase expression [78].
At the ovarian level, hyperandrogenic intrafollicular pattern is improved by a
decrease in IGF-1 availability that has an important role in controlling granulosa
cell aromatase levels [79].
It has been shown that granulosa cells from women with PCOS have higher levels of
FSH receptor (FSHR) expression compared with those from normal ovaries [80, 81].
Metformin reduces FSH-stimulated aromatase expression and activity in granu-
losa cells; it exerts this action by reducing FSHR mRNA and consequently the
activity of FSH (as measured by aromatase expression and E2), without altering
cAMP levels. This involves blocking activation of CRE on promoter II of CYP19
via inhibition of pCREB and possible disruption of the formation of the CREB-
CRTC2 co-activator complex. This is via an AMPK-independent mechanism [82].
6.2.2.2 Dosage and Side Effects

Metformin is available as 500, 850, and 1,000 mg tablets with a target dose of
1,500–2,550 mg/day.
102 6 PCOS Therapy
Metformin has a dose-dependent absorption in humans [83], and its bioavailabi-

lity is limited to 50–60 % because the amount available may result from pre-systemic
clearance or binding to the intestinal wall [83].
Therapeutic regimens of metformin administration are not well standardized,
and its dose should probably be adjusted according to the patient’s BMI and insulin
resistance [84].
For example, it was demonstrated that nonobese women with PCOS respond better
than obese women to metformin treatment at a dosage of 1,500 mg/day for 6 months.
Nonobese women, in fact, showed a statistically significant decrease in serum andro-
gen level and fasting insulin level and also an improvement in menstrual cyclicity
[85, 86]. Moreover, it is possible that women who did not respond to metformin 1,5 g
dose per day might show clinical changes if the dose is increased to 2 g [76].
Common side effects are gastrointestinal, such as diarrhea, nausea, vomiting,
bloating, abdominal discomfort, flatulence, and unpleasant metallic taste in the
mouth.
Lactic acidosis and hypoglycemia are very rare.
To reduce these side effects, it is recommended to start metformin with a low
dose (e.g., 250–500 mg/day) and then gradually increase within a period of
4–6 weeks [76].
Metformin may cause vitamin B12 malabsorption, and so every patient should
be monitored for signs and symptoms of vitamin B12 deficiency: numbness, pares-
thesia, macroglossia, behavioral changes, and pernicious anemia [66].
Metformin prescription should be avoided in women with renal insufficiency,
congestive heart failure, sepsis, or hepatic dysfunction [66].
Therefore, testing of hepatic and renal function is necessary in advance of pre-
scription, and thereafter yearly testing is indicated.
However, it has been demonstrated that metformin use for up to 6 months does
not adversely affect renal or liver function in a large sample of PCOS women, even
those with mildly abnormal baseline hepatic parameters [87, 88].
The length of metformin treatment in PCOS patients is not standardized, but data
present in literature [89] showed that, after a long-term metformin treatment, drug
suspension is related to a quick reversion of its beneficial effect on peripheral insu-
lin sensitivity.
6.2.2.3 Metformin and Menstrual Disorders

The main complaint about menstrual disorders from PCOS patients is the absence
or infrequency of menstrual bleeding.
Few studies noticed the regularization of menstrual cycle after 3–6 months of
therapy with metformin alone in 60–70 % of PCOS insulin-resistant patients [90–
93] with an important improvement of LH/FSH ratio [90].
The response to the treatment usually depends on the degree of insulin
resistance.
The positive effect of metformin on menstrual cycle is commonly attributed to its
effectiveness on ovulatory function. However, it is not uncommon to observe dis-
cordance between menstrual and ovulatory cycles.
The presence of ovulation should be confirmed through the measurement of

luteal phase progesterone levels (usually, levels > 4 ng/mL indicate a previous ovu-
lation) [76].
An Italian study revealed that only 79 % of PCOS women had ovulatory cycles
after reaching normal menstrual cycle with metformin treatment [93].
This observation may indicate that the effectiveness of metformin on menstrual
cyclicity is probably secondary to a direct effect on the endometrium and not only
to an effect on the ovary [67].
Ovulation may be a result of a direct action of metformin on the ovary that leads
to normal steroid production and steroid feedback effects that include a lowering of
LH and androgen levels [67].
6.2.2.4 Metformin and Endometrium

Excessive insulin levels stimulate endometrial growth [94], and most anovulatory
PCOS patients have endometrial vascularization and pattern and thickness abnor-
malities [95, 96]: PI (pulsatility index) and RI (resistance index) are higher than
controls, probably due to the vasoconstrictive effect of androgens on vascular tis-
sues [97].
Metformin may have a positive impact on the endometrium thanks to:
• Indirect effect: androgen decrease, which leads to the reduction of their vasocon-
strictive effects on vascular tissue.
• Direct effect: insulin stimulates glucose oxidation activity in the late luteal phase
in human endometrium; insulin receptors are present at the endometrial level,
reaching their maximal expression in the secretory phase. GLUT-4 is an insulin-
dependent transporter expressed in the endometrium and involved in endome-
trium metabolism; GLUT-4 is reduced in PCOS patients, suggesting that in these
subjects both insulin resistance and hyperinsulinemia induce an inadequate
GLUT-4 expression and so a decreased glucose supply. Thus, by improving
hyperinsulinemia, metformin could be effective in restoring endometrial recep-
tivity through a direct effect.
PCOS women who ovulated under metformin treatment showed a triple-line

endometrial pattern in a percentage of cases similar to those observed in healthy
controls [95], and a triple-line pattern is associated with a significantly higher preg-
nancy rate.
Another aim of metformin treatment is to reduce the long-term risks of unchal-
lenged endometrial proliferation: hyperplasia and carcinoma.
Their main pathogenic mechanism assumed was hyper-estrogenic stimulation of
endometrial growth, unopposed by progesterone. In fact, estrogens act by genetic
and epigenetic mechanisms on cancer cells, and a close relationship between estro-
gens, growth factors, and oncogenes is important in the development of several
human cancer [98].
The second hypothesis taken in consideration was the known mitogenic effect
exerted by insulin [99].
104 6 PCOS Therapy
6.2.2.5 Metformin and Hyperandrogenism

Metformin determines a great improvement on the hyperandrogenism symp-
toms of patients with PCOS, ameliorating hyperandrogenemia and reducing
circulating insulin levels [92, 101–103]. Moreover, as insulin acts as an ana-
bolic growth factor in hair [104], it is possible that the suppression of circula-
ting insulin levels alone may be sufficient to improve the rate of terminal hair
growth [76].
A 20–30 % reduction of total and free testosterone, increased SHBG levels,
a 30 % decline of androstenedione levels, a modest decrease of FG hirsutism score,
and an improvement of acanthosis nigricans were shown [92, 100].
Poor effects on the acne score of young PCOS women were recorded [105].
Several data suggest that metformin could act on hyperandrogenism by interfe-
ring both with direct and specific mechanisms on peripheral androgen-secreting
organs and with free androgen fraction-regulating systems [67]: in fact, a reduced
ovarian and adrenal secretion of androgens, a reduced pituitary secretion of LH, and
an increased liver SHBG production seem to be the mechanisms that mediate met-
formin effect on hyperandrogenism [69].
On the other hand, other studies compared metformin effects to those obtained
from oral contraceptives or antiandrogen drugs: the latter achieved a more effective
results on hyperandrogenism than metformin alone [106–108].
According to our clinical experience, in overweight/insulin-resistant/hirsute
PCOS women, metformin should be considered a first-line treatment, to be associa-
ted in combination with antiandrogen therapy.
Moreover, a case reported by an English study group demonstrated how impor-
tant is the metformin administration even in underweight PCOS patients with men-
strual disorder and hirsutism, underlying the essential role of insulin resistance in
PCOS pathogenesis, sometimes independent of fat mass or distribution [109].
6.2.2.6 Metformin and Fertility

Metformin reduces insulin levels and alters its effects on ovarian androgen biosyn-
thesis, theca cell proliferation, and endometrial growth; it inhibits ovarian gluconeo-
genesis, reducing ovarian androgen production [110–112]: all these actions lead to
an improved ovulation induction in PCOS patients.
According to the ESHRE and ASRM guidelines issued in 2007, the use of met-
formin should be limited to patients with impaired glucose tolerance and should be
interrupted before the administration of clomiphene citrate, thus restricting the use
of metformin to a minority of PCOS patients [113].
However, more recent data suggest that these guidelines should be
reconsidered.
Metformin alone has a significant benefit on inducing ovulation in PCOS women,
but there is limited evidence that it improves pregnancy rate [101]. According to a
multicenter study, metformin alone is not as effective as clomiphene citrate (CC)
alone for the treatment of infertility: 55.3 vs. 75.1 % in cumulative ovulation and
7.2 % vs. 22.5 % of life birth [101].
On the contrary, an Italian study stated that the cumulative ovulation rate was
similar in women treated with CC or metformin, whereas the pregnancy rate was
significantly higher in women treated with metformin [114].
However, metformin is more effective than placebo alone, and it is associated
with a significantly lower multiple pregnancy and ovarian hyperstimulation syn-
drome (OHSS) rate [76].
Because of the lack of evidence, metformin should not be used as first-line
monotherapy, but only in those patients who:
1. Want to improve both metabolic and reproductive functions, but they do not want
to immediately get pregnant.
2. Absolutely wish to avoid multiple gestations.
3. Do not tolerate CC or are resistant to CC [76].
Clomiphene resistance is defined as the inability to achieve ovulation after
two cycles of clomiphene administration at a dose of 150 mg/day for 5 days
[115].
4. Do not achieve a pregnancy (CC failure): metformin could be administered as
pretreatment [67].
In CC-resistant women, a combined therapy with CC + metformin (contempo-

rarily or as pretreatment) is suggested: in a meta-analysis, this combination signifi-
cantly improved ovulation and pregnancy rates, decreasing OHSS rate, when
compared with CC alone [116].
The percentage of patients with PCOS and clomiphene resistance ranges in the
different studies between 15 and 40 % [117, 118]. In these patients, metformin/
clomiphene combination induces ovulation in 62.5–77.7 % of cases [116,
119–122].
This result is probably secondary to various mechanisms:
• Changes in intrafollicular steroidogenesis resulting from the effect of metformin

on granulosa cells through an increase in insulin-like growth factor 1 [120]
• Inhibition of androgen synthesis by the direct action of metformin on the interna
theca cells [78]
• Metformin-induced decrease of adrenal responsiveness to adrenocorticotropic
hormone, resulting in reduced adrenal steroidogenesis [123]
• Reduction in serum LH and prolactin levels resulting from the effects of metfor-
min on the hypothalamic–pituitary axis [124]
Thus, it is possible to state that metformin administration, decreasing insulin

secretion, facilitates the induction of ovulation by using clomiphene citrate [125] in
patients with PCOS.
The beneficial effects of metformin coadministration during gonadotropin ovula-
tion induction and/or IVF cycles are unclear, and therapy with metformin should
depend on the degree of IR.
106 6 PCOS Therapy
It is well known that the response of PCOS women to gonadotropin stimulation

differs significantly from that of normal ovaries: it is defined “explosive” and it is
responsible for the higher risk of canceled cycles and/or for OHSS [126, 127].
In fact, it was shown that during ovarian stimulation, E2 production and E2-to-A
ratio are higher in patients with PCOS who have elevated insulin levels than in
normo-insulinemic women [126]. Increased insulin levels involve greater ovarian
endocrine and morphologic responses to FSH-induced ovulation, which predispose
to OHSS.
Therefore, it seems that the typical response of the polycystic ovary to exogenous
gonadotropin therapy is related to increased plasma concentrations of insulin [128].
6.2.2.7 Metformin and Pregnancy Loss

Few observational studies have shown that metformin could play an important role
in reducing the risk of pregnancy loss [129–131].
In particular, metformin exerts systemic actions by reducing body weight, insu-
lin and PAI-1 levels [131–133], and plasmatic endothelin-1 (ET-1), androgen, and
LH concentrations [135] and by increasing IGFBP-1 and glycodelin levels [136].
Moreover, metformin improved the uterine artery blood flow [95, 136] and
several endometrial receptivity surrogate markers, as well as endometrial vascular-
ization and pattern [95]. It was hypothesized that metformin might improve perifol-
licular and peri-corpus luteum vascularization too [95].
Furthermore, in the past, an experimental study [137] demonstrated that metfor-
min also induced AMPK activation within the blastocyst, leading to improved insu-
lin signaling and pregnancy outcomes. In fact, the preimplantation blastocyst stage
embryo is an insulin-sensitive tissue, responsive to insulin or IGF-1 via the IGF-1
receptor/translocation of GLUT-4, with an increased glucose uptake [138]. High
insulin or IGF-1 concentrations induced a downregulation of IGF-1 receptor [139]
with consequent insulin-stimulated glucose uptake reduction, intraembryonic glu-
cose level dropping, and apoptosis triggering [138].
On the contrary, other studies did not confirm these beneficial effects of metfor-
min in preventing abortion [140, 141].
6.2.2.8 Metformin Administration During Pregnancy

The safety of metformin in pregnancy has not yet been established. It crosses the
human placenta [142, 143], and it has been detected in umbilical cord blood at
levels equal to or higher than the ones in maternal venous blood [144–146]: in fact,
except for the first hours after metformin intake, the fetus is exposed to higher con-
centrations of metformin than the mother [144]. The knowledge on metformin
metabolism in the fetus is scarce: it has been hypothesized that part of the metfor-
min is excreted to the amniotic fluid [147] and reabsorbed to the fetal circulation by
swallowing. Metformin is then eliminated from the fetus by passage through the
placenta into the maternal circulation [144]. Fetal insulin concentrations are not
affected by maternal metformin treatment.
A recent study demonstrated that intrauterine metformin exposure seems to
result in elevated SHBG levels in newborns [148]. Metformin exposure throughout
pregnancy exerts no major effects on maternal or neonatal androgens or estrogens at

birth [148].
Metformin is classified as pregnancy category B [149]: a meta-analysis con-
cluded that there was no evidence of an increased risk for major malformations
[150].
Additionally, a study demonstrated that metformin did not adversely affect birth
length, birth weight, growth, or motor–social development in the first 18 months of
life [151].
However, current conservative practice would be to stop treatment once pre-
gnancy has been established, but considering the adverse impact of insulin resistance
on the pregnancy, continued metformin treatment after conception in women with
PCOS may be beneficial [152].
The rationale of using metformin during pregnancy in PCOS women is the
attempt to reduce the risk of developing gestational diabetes and other pregnancy
complications associated with insulin resistance, such as preeclampsia.
Metformin seems to reduce the risk of gestational diabetes (GD) [153, 154] that
complicates 5–40 % of pregnancy in women with PCOS.
Continued metformin treatment throughout pregnancy appeared to significantly
reduce the rate of GDM requiring insulin therapy [155].
The mechanisms recognized in reducing GD incidence were the reduction of
preconception weight, insulin, insulin resistance, insulin secretion, and testosterone
levels and the persistence of these effects during pregnancy [156].
A less weight gain in women treated with metformin, compared with those
treated with insulin, has been reported, and also the incidence of neonatal hypogly-
cemia was reduced [157, 158].
Furthermore, during the first trimester of pregnancy, metformin seems to influ-
ence the trophoblastic invasion of the maternal decidua, myometrium, and blood
vessels, allowing a successful placentation with consequent pregnancy outcome
improvement, such as prevention of pregnancy-induced hypertension (PIH) and
preeclampsia [67].
Increased placental insulin resistance directly impairs nutrient supply to the fetus
and leads to fetal growth restriction [159, 160].
Unfortunately, there are only a few studies in literature confirming these prelimi-
nary data.
Generally, it is important to note that the beneficial role of metformin in
pregnancy-related parameters may be accomplished through a continuum of
effects that starts from preconception and lasts throughout pregnancy [152]. In
fact, preconception weight loss and IR reduction promoted by the combination of
metformin and diet may reduce the likelihood of gestational diabetes in PCOS
women [156].
Despite these favorable effects and reassuring clinical data, no definite guide-
lines recommending metformin use in pregnant women exist: further research is
necessary [157, 161].
Finally, it is important to know that metformin is transferred into breast milk in
amounts that appear to be clinically insignificant [162–165]. Thus, metformin use
108 6 PCOS Therapy
by breastfeeding mothers is considered safe. Nevertheless, each decision to breast-

feed should be made after conducting a risk/benefit analysis for each mother and her
infant [163].
6.2.2.9 Metformin and Metabolic Syndrome

As explained before, metformin increases insulin sensitivity [89, 100, 103, 166–
169] and decreases weight, waist circumference, and BMI [100, 102, 167, 170],
particularly if associated with diet and physical exercise.
Some authors state that, without metformin, weight loss (through caloric
restriction and increased exercise) is difficult to achieve and maintain [171, 172],
due to the weight-preserving and anabolic effects of high insulin [173] and
androgens [91].
It was demonstrated that reduction of body weight, BMI, and visceral fat was
greater than placebo, and the combination of metformin plus lifestyle intervention
was more effective than placebo plus lifestyle intervention [174].
Metformin could act to improve body weight in obese PCOS patients both
directly and indirectly:
• Direct effect: on the central nervous system, by modulating appetite in the hypo-
thalamus [175]
• Indirect effect: via adipocytokine modification
Visfatin is the most recently identified adipocytokine, which seems to be prefe-
rentially produced by visceral adipose tissue and has insulin-mimetic action
[176]. Circulating visfatin levels are higher in patients with PCOS than healthy
controls, and it was demonstrated that metformin treatment significantly reduced
visfatin levels after 3 months of therapy [177].
It has been suggested that weight loss may be a dose-related response with
increased weight loss at higher dose [170]. In fact, comparing two different doses, a
significant drop in BMI and waist circumference was seen in those patients using
the higher dose [178].
Investigators have reported a greater weight, BMI, and WC reduction in obese
patients receiving 2,550 mg/day and concluded that the long-term effect of metfor-
min is better with greater dose [170, 179].
Additionally, metformin may slow the progression to type II diabetes melli-
tus [66].
This protective effect might be associated with the preservation of pancreatic
beta-cell function and appeared to be mediated by a reduction in the secretory
demands placed on beta cells by chronic insulin resistance [180].
A recent position statement from the AES (Androgen Excess Society) recom-
mended that women with PCOS, regardless of weight, should be screened for IGT
or type II diabetes mellitus by an oral glucose tolerance test at their initial presenta-
tion and every 2 years thereafter [181].
However, this statement noted that the use of metformin to treat or prevent the
progression of IGT could be considered but should not be mandated at this point in
time because well-designed RCTs demonstrating efficacy have yet to be conducted
[67]. Moreover, it is important to underline that metformin does not maintain its ben-
efits at a biochemical and clinical level after a 12-month treatment suspension [89].
It is widely known that insulin resistance and consequent metabolic syndrome
increase the risk of cardiovascular disease: for this reason, it is very important to
consider long-term health when selecting a medical treatment in overweight women
with PCOS [182].
PCOS young patients usually do not manifest increased blood pressure values
[183], but at menopause women with PCOS have a risk of developing hypertension
2.5-fold higher than age-matched controls [184]: metformin could prevent struc-
tural changes that precede hypertension [67]. In fact, it has been shown that metfor-
min improve endothelial function, coronary microvascular function, and coronary
flow rate [185].
As explained in previous chapters, dyslipidemia is a typical feature of metabolic
syndrome: metformin improves hepatic fatty acid metabolism from lipogenesis
toward oxidation.
Different beneficial effects are reported on dyslipidemia in PCOS women [130,
173, 186–192]:
• Decreased total and LDL cholesterol levels

• Decreased triglyceride levels
• Increased HDL cholesterol levels
To prevent vascular consequences, LDL particles should be normalized.

Despite metformin has been shown to improve metabolic alteration, it cannot be
considered as first-line therapy [193], but it should be used as an adjunct to lifestyle
modification.
Besides ameliorating the metabolic syndrome already present, metformin appears
to be also effective in preventing the onset of the metabolic syndrome [194]; a study
reported that PCOS women treated with a combination of metformin and controlled
diet had significant and sustained improvements in all parameters of the metabolic
syndrome over 4 years [195]. Conversely, another study showed that beneficial
effects of metformin on the metabolic syndrome, without a specific lifestyle modifi-
cation regimen, could be sustained over 3 years of routine clinic follow-up [194].
Furthermore, chronic inflammation is one of the PCOS features. Metformin alone
reduces circulating levels of CRP (inflammation marker that is usually higher in PCOS
women) [196]. It exerts a direct vascular anti-inflammatory effect by dose dependently
inhibiting IL-1β-induced release of the pro-inflammatory cytokines IL-6 and IL-8 in
endothelial cells, human vascular smooth muscle cells, and macrophages [67, 197].
Endothelial dysfunction, assessed by reduced flow-mediated dilatation, has shown
promising results in cardiovascular risk stratification and prognosis [198, 199].
Metformin administration for 6 months in women with PCOS induced a significant
increase in flow-mediated dilatation that was restored to normal values [200].
A recent study suggests that metformin decreases serum levels of asymmetric
dimethylarginine (ADMA) levels, an endogenous inhibitor of NOS, by concomitant
effects on insulin action and androgen levels [201].
110 6 PCOS Therapy
Metformin seems to be effective even in decreasing AGE levels, which are oxi-
dative mediators of endothelial dysfunction [134].
Plasminogen activator inhibitor-1 is a pro-thrombotic factor produced by the
endothelium that inhibits fibrinolysis and regulates vascular smooth muscle proli-
feration [202]. Insulin upregulates PAI-1 gene transcription [203] and stimulates
hepatic [204] and endothelial PAI-1 production [205]. It has been demonstrated that
metformin reduces PAI-1 levels [131–133].
6.2.2.10 Metformin and Hypothyroidism

A recent study stated that in overweight PCOS patients with primary sub-
hypothyroidism, treatment with metformin (1,500 mg/day) resulted in a significant
fall in TSH and in some cases improvement of hypothyroidism [206].
This is an important finding because hypothyroidism occurs in more than 10 %
of PCOS patients [207].
Several mechanisms have been hypothesized:
• A slight increase in the gastrointestinal absorption of levothyroxine (in patients

already in treatment with L-thyroxine) [208].
• Influence of changes in body weight, associated with metformin therapy, on TSH
levels [209].
• Increase of dopamine in the hypothalamus [210]. Previous studies, in fact,
have suggested that there was a disruption of the neuroendocrine mecha-
nisms in women with PCOS, mainly due to a deficiency in hypothalamic
dopamine [211].
Further studies are needed to confirm these findings, but some authors suggest
starting to treat obese PCOS patients with subclinical hypothyroidism with metfor-
min and to reevaluate their thyroid function after 6 months [206].
6.2.2.11 Metformin Use in Lean PCOS Women

It was revealed that metformin decreases ovarian cytochrome P450c17α activity:
this mechanism leads to a reduction of free testosterone serum levels even in lean
PCOS women, with a consequent improvement of hyperandrogenism [212].
In fact, it has been demonstrated that women with PCOS who have normal
weight or are thin responded to a reduction in insulin release with decreased ovarian
androgen production and serum ovarian androgens.
As Nestler highlighted several years ago, metformin treatment of nonobese
women leads to [213]:
• Decreased fasting and glucose-stimulated insulin levels

• Decreased basal and GnRH-stimulated LH release
• Decreased ovarian androgen production
• Decreased both serum total and free testosterone concentrations
• Increased serum SHBG concentrations
• Decreased androstenedione and DHEAS levels
Six months of metformin therapy clinically results in beneficial effects in lean

PCOS women in terms of resumption of menses, without any remarkable effect on
metabolic and cardiovascular risk factors [214].
Moreover, a very recent study suggests that treatment with metformin, for at
least 12 weeks prior to, and during, IVF/ICSI, is worth considering as a manage-
ment approach for nonobese women with PCOS [215].
6.2.2.12 Metformin Use in PCOS Adolescents

Metformin is indicated in patients older than 10 years, 2 years after their
menarche.
Few studies demonstrated that metformin improved ovulatory function even in
PCOS adolescents [93, 216], as well as hyperandrogenism [217].
As in adult population, metformin is effective in reducing hyperinsulinemia and
lipid abnormalities.
A recent study reported that PCOS was the main indication for metformin pre-
scription in UK general practice, even if it is off-label [218].
In the FDA approval process, several studies that demonstrated the safety of
metformin use in the adolescent population were conducted.
Contraindications and side effects are the same described for adults.
Even if the literature on metformin in adolescents is limited and the number of
studies inconsistent, it is possible that early intervention might prevent the complete
spectrum of the syndrome in young overweight girls [219].
However, to date, in adolescent population, the first-step treatment is always the
lifestyle modification.
In obese adolescents with PCOS, EP combination pills are the standard of care
when lifestyle modification is not effective [220]. EP pills treat hyperandrogenism
by increasing SHBG and so decreasing free testosterone; it reduces LH and FSH
secretion and decreases ovarian stimulation and androgen production. Progesterone
induces menstrual cyclicity and prevents endometrial hyperplasia [221]. However,
EP pills do not treat IR or components of the metabolic syndrome [222] and are
instead associated with glucose intolerance, decreased insulin sensitivity, abnormal
lipid profiles, and CV disease [223, 224].
A recent study compared metformin monotherapy vs. estrogen–progesterone +
metformin in the treatment of overweight and obese PCOS adolescents [220]: it was
shown that the use of metformin alone was associated with greater decrease in total
cholesterol and triglycerides and with a better improvement in weight loss.
These findings suggest that metformin monotherapy is more effective in redu-
cing cardiovascular risk in overweight and obese adolescents with PCOS than the
combination with EP pill [220].
6.2.2.13 Metformin + Pioglitazone in PCOS Treatment

Pioglitazone as add-on therapy in metformin-resistant PCOS women (e.g., in
women who after 6 months’ 1,500–2,500 mg daily metformin treatment fail to
improve their metabolic and hyperandrogenemia-related clinical signs) may exert
beneficial metabolic (further reduction of IR and glucose levels, improved lipid
112 6 PCOS Therapy
metabolism, and lowering of carotid intima–media thickness) and antiandrogenic

(improved menstrual regularity, significant drop in testosterone and DHEAS levels,
increased SHBG, and improved hirsutism score) effects [64, 225, 226].
Moreover, the addition of pioglitazone was not associated with any adverse side
effects, such as hepatotoxicity and hypoglycemia [226], providing another valid
option for the management of NAFLD and NASH in PCOS women [64].
Pioglitazone safety in women under 18 is not yet established; thus, pioglitazone
is not recommended in this female PCOS subgroup. Its safety in pregnancy and
lactation is not entirely clear, but it is classified as category C drugs by the FDA
due to the fact that studies in animals have shown adverse fetal effects such as
IUGR [64].
However, long-term and large-sampled clinical trials are necessary before stating
definitive conclusions.
6.2.3 Statins
Statins are competitive inhibitors of HMG-CoA reductase, the rate-limiting enzyme

of the cholesterol biosynthesis [227]: inhibition of this enzyme decreases chole-
sterol synthesis with a compensatory increase in the expression of LDL receptors in
the liver. In the general population, statins decrease total cholesterol and LDL cho-
lesterol, and they have antiproliferative and antioxidant features on endothelial cells
[228].
Statins reduce plasma triglycerides in a dose-dependent manner, and they also
have a modest HDL-raising effect, which is not dose dependent [229, 230].
As dyslipidemia is a component of metabolic syndrome, atorvastatin and sim-
vastatin have been used in PCOS women to investigate their effects on this common
syndrome.
To date, there are limited data on the use of statins in PCOS, but short-term
use of statins alone or in combination with metformin appears to improve lipid
levels in PCOS. In a meta-analysis, statins were more effective than placebo in
reducing total cholesterol, LDL, and triglycerides; lipid profile improvement
occurred within the first 3 months of treatment, with no further significant change
thereafter [231].
A combination of metformin with statins was more successful than metformin
alone in lowering fasting glucose, fasting insulin, LDL cholesterol, and triglyce-
rides [232].
Moreover, in the presence of simvastatin, metformin is much more effective in
reducing testosterone, DHEAS, hirsutism, and LH and reversing the LH/FSH ratio
in patients with PCOS [233].
The mechanisms of action of simvastatin on inhibition of T levels are likely
related to the inhibition of the mevalonate pathway [233, 234]. Statins might also
decrease the expression of several key enzymes involved in T production: chole-
sterol side chain cleavage (P450SCC), 17α-hydroxylase/17,20-lyase (P450c17), and
3β-hydroxysteroid dehydrogenase (3βHSD). Such effects of statins were noted in
6.3 Inositol and Other Supplements 113
adrenocortical cells [235, 236]. The mechanisms of these actions might be due to
the inhibitory effects of statins on isoprenylation [237], leading to decreased func-
tion of small guanosine triphosphatases, such as Ras: statin might abrogate Ras-
induced steroidogenesis [236].
Additionally, statins induce inhibition of proliferation of theca interstitial cells
and might reduce T output of the ovary by reducing the size of the theca interstitial
compartment [233].
Thus, although simvastatin plus metformin could successfully reduce hyperan-
drogenism, insulin resistance, and lipid profile, its clinical significance is yet to be
characterized [233].
However, statins are considered pregnancy category X drugs, and so it is always
required to avoid contraception: this represents a very important restriction of use,
and it is not a good option of treatment for all PCOS patients who want to get
pregnant.
Finally, statins should be reserved only for women with PCOS who have
increased LDL cholesterol [238].
6.3 Inositol and Other Supplements
In recent years, more attention has been paid to some supplements, which seem to
have an important role in the therapy of PCOS, such as inositol and antioxidant
molecules.
6.3.1 Inositol and Its Isomers
Several inositol isomers, and in particular myoinositol (MI) and D-chiro-inositol

(DCI), were shown to have insulin-mimetic properties and to be efficient in the
treatment of PCOS.
Inositol (cyclohexane-1,2,3,4,5,6-hexol) is a polyol existing under nine stereo-
isomeric forms depending on the spatial orientation of its six hydroxyl groups
(Fig. 6.1).
Myoinositol is naturally present in animal and plant cells, as free form, as
inositol-containing phospholipid (phosphoinositides), or as phytic acid (IP6) [239].
The greatest amounts of myoinositol in common foods are found in fresh fruits
and vegetables and in peas, beans, grains, and nuts [240].
Originally, myoinositol was considered one of the B-complex vitamins, but now
it is no more reputed an essential nutrient because it was shown that it is produced
in sufficient amount in the human body from D-glucose [241].
It was shown that myoinositol is indispensable for the growth and survival of
cells [242] and for the development and function of peripheral nerves [243]; it is
essential to bone formation, osteogenesis, and bone mineral density [244], but its
therapeutic implications are mainly related to its important role in glucose
homeostasis.
114 6 PCOS Therapy
A significant part of the ingested myoinositol is consumed in the form of phos-

phatidylinositol (PI) that may be hydrolyzed by a pancreatic phospholipase A in the
intestinal lumen. Ninety-nine percent of the myoinositol ingested is absorbed from
the human gastrointestinal tract, through an active transport system involving a Na+/
K+-ATPase [239].
Cells mainly derive inositol from three sources:
• De novo biosynthesis from glucose-6-phosphate by 1D-myoinositol-phosphate

synthase (MIPS) and inositol monophosphatase (IMPase)
• Dephosphorylation of inositol phosphates derived from breakdown of inositol-
containing membrane phospholipids
• Uptake from the extracellular fluid via specialized myoinositol transporters [245]
In vivo, conversion of myoinositol to D-chiro-inositol can occur in tissue expres-

sing the specific epimerase.
Myoinositol and D-chiro-inositol can also be bound components of glyco-
sylphosphatidylinositol (GPI) anchors and of inositol phosphoglycan (IPG) that
would constitute second messengers of insulin action in the GPI/IPG pathway
[241].
The exact mechanisms of action of MI and DCI with insulin-mimetic activities
are still unclear; a presumed mechanism of action implies inositol phosphoglycans
(IPGs) containing MI or DCI as insulin mediators [241].
A few studies hypothesized that insulin, other growth factors, and classical hor-
mones stimulated the hydrolysis of glycosylphosphatidylinositol (GPI) generating
water-soluble inositol phosphoglycan (IPG) second messenger. The origin of IPG-A
is thought to be myoinositol-containing GPI [246].
OH
HO OH
OH OH OH
HO OH HO OH HO OH
HO OH
OH HO OH HO OH HO OH
OH OH OH
Myo-Inositol
Scyllo-Inositol Epi-Inositol Cis-Inositol
OH
OH OH OH OH
HO OH
HO OH HO OH HO OH HO OH
HO OH HO OH HO OH HO OH HO OH
OH OH OH OH
OH
D-chiro-Inositol L-chiro-Inositol Muco-Inositol Allo-Inositol Neo-Inositol
Fig. 6.1 Inositol isomers

One of the most interesting models is the one elaborated by Larner and cowor-
kers in 2010 [247]. According to this model, insulin binding to its receptor (IR)
causes the autoactivation of the receptor, and the activated IR can transduce the
signal through two parallel signaling pathways, which act together to mediate insu-
lin action in a complementary and synergistic manner [241]:
1. The first one implies the recruitment and activation of substrate of insulin recep-
tor (IRS) by the activated IR. Subsequent protein activations (PI3K, PDK-1)
finally lead to PKB-Akt recruitment and activation at the plasma membrane.
Activated PKB-Akt induces GLUT-4 translocation to the plasma membrane,
improving glucose entry into the cell.
2. When insulin binds to its receptor, the epimerase converts MI molecules to
DCI. In the second pathway, the IR is fixed to a G protein itself attached to a
phospholipase that catalyzes the hydrolysis of a GPI [248, 249]. The insulin-
induced hydrolysis of the GPI releases an inositol phosphoglycan containing
D-chiro-inositol (DCI-IPG), which acts as a probable second messenger of insu-
lin (INS-2) mediating insulin effects on glucose oxidative and non-oxidative
clearance. INS-2 binds and activates two Mg2+-dependent protein phosphatases:
PP2Cα in the cytosol and PDHP in the mitochondria. Activated PP2Cα stimu-
lates glycogen synthase directly and also indirectly through possible activation
of PI3K-Akt and subsequent inhibition of GSK3. In the mitochondria, activated
PDHP stimulates PDH and so glucose oxidative use [241].
The insulin-sensitizing effect of a MI and DCI supplementation is probably due

to their intracellular enhanced availability for the production of membrane IPG pre-
cursors; numerous evidences support the hypothesis of a role of inositol glycan
insulin second messengers in insulin-mimetic properties of some inositol isomers.
Moreover, it is known that part of MI supplementation effect on insulin sensiti-
vity may come from its partial in vivo intracellular epimerization to DCI [241].
MI intracellular concentration is regulated through processes such as extracel-
lular MI uptake, de novo biosynthesis, regeneration, efflux, and degradation.
Alteration of one or several of these processes can lead to inositol intracellular
abnormalities [241] in diabetes mellitus: inhibition of cellular MI uptake, altered
MI biosynthesis, enhanced MI efflux due to sorbitol intracellular accumulation,
and increased MI degradation are putative mechanisms of MI intracellular deple-
tion [250].
Larner et al. noted a decreased urinary excretion of DCI and an increased urinary
excretion of MI in humans and monkeys with type II diabetes (ten times higher
compared to healthy subjects) [251].
The ratio of MI/DCI is regulated by an epimerase that converts MI into DCI
[252], and Larner showed that each organ has a specific MI/DCI ratio [253].
Altered ratios of increased myoinositol to decreased D-chiro-inositol in urine
have even been proposed as an index of insulin resistance in humans [254]: a deficit
in MI to DCI epimerization activity, due to an epimerase-type enzyme, was supposed
[249, 255].
116 6 PCOS Therapy
Excessive urinary MI excretion could reduce MI plasma level and subsequently

emphasize MI intracellular depletion, particularly in tissues heavily dependent on
extracellular MI import [241]. Decreased production of DCI from MI reduces the
availability of intracellular DCI for its incorporation into IPGs (particularly, DCI-
IPG), probable downstream second messengers of insulin.
Furthermore, the decreased DCI content in insulin target tissues could reduce
insulin signal transduction involving IPGs, in order to contribute to insulin resistance
in those tissues. Depleted plasma levels of DCI observed in PCOS patients underline
the correlation between impaired plasma DCI and insulin resistance [241].
Thus, insulin resistance is associated with:
1. Abnormally low levels of DCI in urine, plasma, and insulin target tissues (liver,
muscle, fat)
2. Excessive MI urinary excretion
3. Intracellular MI deficiency in insulin-sensitive tissues (Fig. 6.2)
On the contrary, more recently (in 2006) Nestler proposed that, in a woman with
PCOS, an initial genetic or environmental insult causing insulin resistance leads to
a compensatory hyperinsulinemia. The latter induces a defect that increases renal
clearance of DCI, and this leads to a reduction in circulating DCI and its availability
to tissue. The consequence is an intracellular deficiency of DCI and of DCI-IPG,
a mediator of insulin action.
Diminished release of DCI-IPG in response to stimulation by insulin results in a
further decrease in insulin sensitivity [256] (Fig. 6.3).
In 2010, Baillargeon et al. [257] showed that when plasma glucose is maintained
at stable levels and plasma insulin is acutely raised and maintained at constant
levels, the circulating DCI-IPG insulin mediator is released rapidly and briefly in
normal women. Conversely, this coupling between insulin action and DCI-IPG
release was entirely absent in obese women with PCOS: the release of bioactive
DCI-IPG was significantly lower in obese PCOS women [257].
Possible explanations for these findings are a deficit in intracellular DCI or DCI-
IPG and/or a defect in incorporation of the substrate DCI with membrane phospho-
glycans to generate DCI-IPG mediator [257].
The possibility that a deficit in circulating DCI, or its precursor MI, is responsi-
ble for defective insulin-stimulated release of DCI-IPG mediator in PCOS is sup-
ported by the findings that oral supplementation with DCI [258–260] or MI [261,
262] to both lean and obese PCOS women improved their insulin resistance and
clinical symptoms.
Moreover, defective DCI-IPG release in response to insulin could be due to a
qualitative (rather than quantitative) defect in the insulin signaling mechanism that
activates DCI-IPG mediator release from the membrane: there may be a primary
defect in the union of the insulin receptor β-unit to the G protein or a defect in
G-protein activation of phospholipase C [257].
This observation fits with Cheang et al. data [263]: they showed, in a number of
hyperinsulinemic PCOS patients who did not respond to DCI treatment, the absence
? MI DCI epimerization failure
Myo-Inositol excretion
DCI production
Insulin action mediator

Intracellular deficit of DCI-IPG
Release of DCI-IPG following

insulin stimulation
Insulin sensitivity
Insulin resistance
Fig. 6.2 MI and DCI alteration in insulin resistance, proposed by Larner [251]
of changes in DCI-IPG release suggesting that a functional defect rather than a

simple inositol nutritional deficiency might be present [263, 264].
6.3.1.1 Inositol as Treatment for PCOS

A supplementation with myoinositol or D-chiro-inositol was found to be safe and
effective in improving metabolic and hormonal parameters in PCOS patients: the
main mechanism of action is based on improving insulin sensitivity of target tissues,
118 6 PCOS Therapy
Unknown genetic or environmental

“insult”
Insulin-resistance and
compensatory hyperinsulinemia
DCI urinary clearance
Insulin action mediator

Intracellular deficit of DCI-IPG
Release of DCI-IPG following

insulin stimulation
Insulin sensitivity
Fig. 6.3 DCI alteration in insulin resistance, proposed by Nestler [256]
resulting in the reduction of insulinemia which has a positive effect on the reproduc-
tive axis and metabolism.
One of the first studies was conducted in 1999 by Nestler et al. [258], who found
that the administration of D-chiro-inositol to women with polycystic ovary syndrome
decreased the insulin response to orally administered glucose; simultaneously with
the reduction in insulin secretion, women who received DCI had a significant improve-
ment in ovulatory function and decreased serum androgen concentrations [258].
It was demonstrated in various studies that both DCI and MI are able to:
• Reduce LH levels, LH/FSH ratio, and testosterone levels [258, 265–268].

• Restore spontaneous ovulation and menstrual cycles [258, 260, 262, 265, 269].
• Improve cutaneous disorders of hyperandrogenism, reducing hirsutism and acne
score [266].
• Decrease HOMA index [264, 265, 267].

• Reduce systolic arterial blood pressure [267, 268].
• Reduce leptin, LDL cholesterol levels, and triglycerides [269].
• Increase HDL cholesterol level [260, 269].
In view of all these findings, recently we conducted a research to evaluate the

clinical, endocrine, and metabolic response of young women with PCOS, treated for
12 weeks with DCI.
From a clinical point of view, our study has highlighted a significant retrieval of
menstrual cycle regularity (p < 0.001) in a rate higher than 60 % in patients treated
and a significant improvement of acne score (p < 0.05) in patients with D-chiro-
inositol treatment. Moreover, there was a significant decrease of triglyceride (p < 0.05)
and basal insulin serum levels (p < 0.05) in patients treated with D-chiro-inositol
[270].
Another clinical study of our group has shown that the administration of D-chiro-
inositol in association with estro-progestins (0.03 mg of ethinyl estradiol and 3 mg
of drospirenone) leads to a significant decrease of acne already from the second
month of treatment, amplifying earlier the effects of oral contraceptives [271].
In literature no side effects after MI or DCI administration were reported when
clinical dosage was used (max 1 g of DCI or 4 g of MI). Clinical trial data indicate
that adverse events related to inositol treatment are gastrointestinal symptoms (nau-
sea, flatus, loose stools, diarrhea) at a dose of 12 g/day or higher [272].
Moreover, MI or DCI supplementation was demonstrated to be effective in
reducing the risk of gestational diabetes (GB) in PCOS women [273, 274], even if
more studies are needed to confirm these preliminary data.
Finally, we suggest the use of MI (4 g die) or DCI (1 g die) as first-line treatment
for those lean PCOS patients suffering from oligomenorrhea and mild hyperan-
drogenism. Combined therapy with diet, exercise, and metformin is reserved for
insulin-resistant and overweight PCOS patients with oligomenorrhea and moderate
hyperandrogenism.
6.3.1.2 Inositol and Oocyte Quality

Myoinositol function is also linked to the important role of IP3 in oocyte develop-
ment and maturation [275, 276].
Oocyte cycle is usually arrested at metaphase of the second meiotic division.
Calcium release mechanisms are shown to undergo modification during oogenesis,
and maximal sensitivity of calcium release is acquired during the final stages of
oocyte maturation: after fertilization, an increased level of intracellular Ca++ occurs,
and subsequent conclusion of meiosis [277, 278].
It was experimentally observed that immature oocytes (germinal vesicles or
oocytes undergone in vitro process of maturation) contain a number of IP3 receptors
less than those matured in vivo, leading to a reduction in Ca++ intracytoplasmic rise.
The disposal of Ca++ from intracellular deposits is required for the oocyte’s acti-
vation that is manifested by the exocytosis of cortical granules, the perpetuation of
the second meiotic division, the extrusion of the II polar body, the formation of two
120 6 PCOS Therapy
pronuclei, and the activation of protein synthesis from maternal RNA to prime the
first mitosis.
Inositol depletion dramatically reduces transduction signal mechanisms media-
ted by IP3, altering the dynamics linked to the intracellular Ca++ fluctuations.
Myoinositol supplementation may prevent this block and promote meiotic pro-
gression of the germinal vesicles; in fact, it was demonstrated that follicles contai-
ning high levels of MY, dosed in follicular fluid, present oocytes of good quality,
and this may be related to a close correspondence between MI and inositol phos-
phates, necessary during oocyte maturation PIP2-mediated [279].
In human follicular fluid a greater concentration of myoinositol is a marker of
good oocyte quality.
A recent clinical trial showed that only MI rather than DCI is able to improve
oocyte quality [280]; the reason was explained by the “DCI paradox in the ovary”
[281]: it is explained that “ovaries in PCOS patients likely present an enhanced
MI to DCI epimerization that leads to a MI tissue depletion; this, in turn, could
eventually be responsible for the poor oocyte quality characteristic of these
patients” [282].
However, this hypothesis has yet to be confirmed: in fact, even DCI supplemen-
tation has shown a significant improvement in oocyte quality.
One of our recent studies showed that, in patients with PCOS, treatment with
myoinositol and folic acid, compared to only acid folic treatment, reduces the num-
ber of germinal vesicles and degenerated oocytes at the time of oocytes’ pickup,
without affecting the total number of oocytes retrieved. Moreover, an increased
number of transferred embryos of good quality and a reduced amount of FSHR IU
administered for the ovulation induction were shown [283].
These results were consistent with those found in other studies [284], suggesting
the positive effect that myoinositol plays in the development of mature oocytes.
Furthermore, recent data demonstrate that by providing both MI and DCI in a
physiological ratio (40:1), hormonal and metabolic imbalances are treated much
more quickly compared to MI alone [252], especially in overweight PCOS patients
who need to control insulin levels and increase ovarian MI content, reducing the risk
of developing a metabolic disease [285, 286].
6.3.2 Antioxidants
Polycystic ovary syndrome is also associated with decreased antioxidant concentra-

tions, and it is considered an oxidative state [287].
The decrease in mitochondrial O2 consumption and GSH levels along with
increased ROS production explains the mitochondrial dysfunction in PCOS patients
[288]. The mononuclear cells of women with PCOS are increased in this inflamma-
tory state [289], which occurs mostly in response to hyperglycemia and C-reactive
protein (CRP) [290].
Physiological hyperglycemia generates increased levels of ROS from mononu-
clear cells, which activate the release of TNF-α and increase inflammatory
transcription factor NF-kappa B. As a result, concentrations of TNF-α, a known

mediator of insulin resistance, are further increased [290].
Oxidative stress and inflammation promotes hyperandrogenism, which aug-
ments the inflammatory load [289].
Oxidative stress promotes its effects causing damage to follicular proteins by the
marking of free thiol groups [291].
Furthermore, reactive oxygen species (ROS) has been considered to play a criti-
cal role in the success of different IVF techniques. ROS are produced within the
follicle, especially during the ovulatory process [292], and it is believed that oxida-
tive stress may be a cause of poor oocyte quality [293]. In fact, high levels of oxi-
dants, as H2O2, have been found in fragmented embryos [294].
MI and DCI are considered an effective therapy for PCOS women even for its
antioxidant activity.
A recent study demonstrated that MI treatment positively affected the oxidative
status of red blood cells (RBC), as shown by the partial restoration of GSH contents
and the reduction of both band 3 Tyr-P levels and protein glutathionylation [295].
Moreover, there is evidence that melatonin plays an important role in the
regulation of reproductive activity [296], and high levels of melatonin have been
found in human preovulatory follicular fluid in concentrations that are almost
threefold higher than serum levels [297–299]. It is known that melatonin and its
metabolites are potent direct free radical scavengers [300–303] and indirect
antioxidants, due to their ability to modulate gene transcription for antioxidant
enzymes [304].
An Italian study demonstrated that, in patients undergoing IVF, treatment with
melatonin plus myoinositol and folic acid reduced the number of germinal vesicles
and degenerated oocytes and increased the number of top-quality embryos, com-
pared to the therapy with only MI [305].
Other two important antioxidant molecules are SOD (superoxide dismutase) and
ALA (α-lipoic acid).
Therapeutic strategy to reduce the oxidative stress includes diet rich in vegeta-
bles, weight reduction, physical exercise, smoking cessation, alcohol consumption
reduction, and adequate number of sleeping hours.
6.3.3 Vitamin D
Vitamin D has pleiotropic effects on a large spectrum of intracellular regulatory

processes, including insulin metabolism, or intrinsic apoptotic pathway, on both
classical and nonclassical tissues, such as the ovary [306].
Moreover, as explained previously, calcium has an important role in follicle
development, and both calcium and vitamin D deficiencies are considered as poten-
tial risk factors for insulin resistance and obesity [307–310].
Hypovitaminosis D was found in about 80 % of PCOS women [311, 312].
Supplementation of vitamin D (50,000 IU/week) and calcium (1 g/day) seems to
support the positive effect of metformin therapy, with greater results in restoring
122 6 PCOS Therapy
normal menstrual regularity and improving hyperandrogenism symptoms, weight

loss, and follicle maturation compared to metformin treatment alone [311].
Further studies are needed to confirm these data in order to use vitamin D + cal-
cium supplementation as routine PCOS treatment protocol.
6.3.4 Glucomannan
Recently, glucomannan has been introduced as supplement for insulin resistance

treatment.
Glucomannan is a high-molecular-weight polysaccharide obtained from tubers
of Amorphophallus konjac: it consists of molecules of D-glucose and D-mannose,
and it is soluble and absorbs water up to 200 times its weight.
Glucomannan exerts its activity by increasing the viscosity of food bolus during
digestion: it creates a viscous gel that makes the bolus smooth and soft, and it forms
a nondigestible coating around food particles.
This leads to a decreased time of food permanence inside the gastrointestinal
tract: as main consequence, the action of digestive enzymes is partially avoided,
resulting in reduced absorption of nutrients [313, 314].
Thus, glucomannan slows both lipid and carbohydrate absorption, reducing total
and LDL cholesterol [315].
In diabetic patients, it is able to reduce postprandial glycemia and insulinemia [316].
Moreover, glucomannan increases the secretion of glucagon-like peptide 1
(GLP1), cholecystokinin (CCK), and peptide YY (PYY) [317], induces satiation
and satiety [318], and preserves weight loss [319].
Minor adverse effects are normally GI related and include diarrhea, flatulence,
and bloating.
Recently, an Italian study has shown that the association inositol–glucomannan
may represent a good therapeutic strategy in the treatment of PCOS women with
insulin resistance [320].
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Erratum to: Chapter 4 in Psychological
Implications of PCOS
Agata Ando’ and Antonio Maria D’Alessandro
Erratum to:
Chapter 4 in Mariagrazia Stracquadanio, Lilliana Ciotta, Metabolic Aspects of

PCOS: Treatment with Insulin Sensitizers. doi:10.1007/978-3-319-16760-2_4.
The co-author information on the chapter opening page of chapter 4 was missing.
Co-authors for this chapter were:

Agata Ando' and Antonio Maria D'Alessandro
The online version of the original chapter can be found under

doi:10.1007/978-3-319-16760-2_4
Agata Ando’
Department of Psychology, University of Torino, Sydney, Italy
Antonio Maria D’Alessandro
Department of Psychology, University of Catania,
Catania, NSW, Italy
© Springer International Publishing Switzerland 2015 E1

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4.2 PCOS and Mental Disorders. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65

1.1 PCOS Origins

© Springer International Publishing Switzerland 2015 1

1.2 Definition and Epidemiology

Polycystic ovary syndrome (PCOS) is a heterogeneous endocrine and metabolic

1. Oligomenorrhea or anovulatory cycles with menstrual irregularities

The previous criteria processed by NIH (National Institute of Health) in 1992

1. Clinical and/or biochemical hyperandrogenism

1. Clinical and/or biochemical hyperandrogenism

Approximately 85–90 % of women with oligomenorrhea have PCOS, while

2.1 Genetics of PCOS

PCOS is a multifactorial polygenic disease (interaction between several genetic and

© Springer International Publishing Switzerland 2015 5

These environmental factors may cluster in certain families because exercising

hyperandrogenism [18]. Moreover, recent experiments involving a large number

More recently, genes encoding inflammatory cytokines have been identified as

represent a negative feedback cycle in which shortened telomeres, in turn, affect

2.2 PCOS Physiopathology

• Exaggerated Adrenarche: It is possible that PCOS might be established and

• Rosenfield’s Hypothesis: Rosenfield suggested that PCOS results from a hyper-

• Hyperinsulinemia: High level of insulin accelerates the development of granu-

2.3 Role of Insulin in the Pathogenesis of PCOS

Insulin receptor binding Post-binding abnormality

Liver SHBG synthesis Free androgens

Fig. 2.1 Correlation between PCOS, hyperinsulinemia, hyperandrogenism, and ovarian

TNF-α is secreted by adipose tissue macrophages and has pro-inflammatory prop-

3. Kahsar-Miller MD, Nixon C, Boots LR et al (2001) Prevalence of polycystic ovary syndrome

3.1 Endocrine Aspects of PCOS

Polycystic ovary syndrome (PCOS) is a chronic and self-perpetuating endocrine

3.1.1 Endocrine Pattern

© Springer International Publishing Switzerland 2015 21

3.1.1.2 Sex Hormones

3.1.1.3 Estrogens and Progesterone

newborn, AMH is undetectable, but it increases gradually until puberty, remaining

3.1.2 Clinical Endocrine Features

3.1.2.1 Menstrual Disorder

Therefore, ovarian dysfunction usually manifests as oligomenorrhea/amenorrhea

Terminal hair growth requires androgen stimulation, specially testosterone and

3.1.2.4 Acne and Seborrhea

3.1.2.5 Androgenic Alopecia

3.1.2.6 Other Clinical Features

In literature, an unusual case of atypical oral hirsutism secondary to PCOS was

3.1.2.7 PCOS and Thyroid Dysfunction

• Probable common genetic predisposition

3.2 Metabolic Aspects of PCOS

The prevalence of metabolic syndrome is similar across racial backgrounds [78].

3.2.1 The Role of the Adipocyte in Linking PCOS to Metabolic

In PCOS subcutaneous adipocytes, there is a reduced catecholamine-mediated

3.2.2 The Role of Vitamin D in the Development of Metabolic

Moreover, it is not fully understood whether vitamin D insufficiency results from

3.2.3 Metabolic Syndrome and Associated Disorders

3.2.3.1 Visceral Obesity

5.1 Differential Diagnosis

5.2 Risk Factors

5.3 Clinical–Endocrine Features

5.3.1 Oligomenorrhea and Anovulation

5.4 Endocrine Blood Tests

5.5 Ultrasound Features

5.6 Clinical–Metabolic Features