diagnostics

Article
Non-Alcoholic Fatty Liver Disease in Patients with
Type 2 Diabetes: Evaluation of Hepatic Fibrosis and
Steatosis Using Fibroscan
Tran Thi Khanh Tuong 1,† , Dang Khoa Tran 2,† , Pham Quang Thien Phu 1 ,
Tong Nguyen Diem Hong 3 , Thien Chu Dinh 4, * and Dinh Toi Chu 5, *
1 Department of Internal Medicine, Pham Ngoc Thach University of Medicine, Ho Chi Minh City 700000,
Vietnam; drkhanhtuong@gmail.com (T.T.K.T.); phu2981992@gmail.com (P.Q.T.P.)
2 Department of Anatomy, Pham Ngoc Thach University of Medicine (PNTU), Ho Chi Minh City 700000,
Vietnam; khoatrandr@gmail.com
3 Dai Phuoc Polyclinic of Ho Chi Minh, Ho Chi Minh City 700000, Vietnam; diemhong1702@gmail.com
4 Institute for Research and Development, Duy Tan University, 03 Quang Trung, Danang 550000, Vietnam
5 Department of Human and Animal Physiology, Faculty of Biology, Hanoi National University of Education,
Hanoi 100000, Vietnam
* Correspondence: chudinhthien@duytan.edu.vn (T.C.D.); chudinhtoi.hnue@gmail.com (D.T.C.)
† These authors contributed equally to this work.




Received: 19 February 2020; Accepted: 12 March 2020; Published: 14 March 2020


Abstract: Patients with type 2 diabetes mellitus (T2DM) are at increased risk of non-alcoholic fatty liver
disease (NAFLD) and might eventually progress to advanced fibrosis, cirrhosis and hepatocellular
carcinoma (HCC). Recommendations on whether to screen for NAFLD in diabetic patients remains
conflicted between major guidelines. Transient elastography using FibroScan with CAP (controlled
attenuation parameter) can assess both liver steatosis and fibrosis simultaneously. This paper took a
new look at the prevalence of NAFLD and the severity of fibrosis among T2DM patients in Vietnam.
The study was conducted using a cross-sectional design in T2DM adults who attended Dai Phuoc
Ho Chi Minh Polyclinic and Polyclinic of Pham Ngoc Thach University of Medicine. Liver steatosis
and fibrosis was assessed by FibroScan. NAFLD was diagnosed if CAP > 233 dB/m (steatosis > 5%).
Data were analyzed using STATA 12 software program. We found that a total of 307 type 2 diabetic
patients qualified for the study’s criteria. The prevalence of NAFLD in T2DM patients based on
FibroScan was 73.3%. Rates of mild, moderate and severe steatosis were 20.5%, 21.8% and 30.9%,
respectively. The prevalence of significant fibrosis (≥ F2), advanced fibrosis (≥ F3) and cirrhosis (F4)
was 13.0%, 5.9% and 3.6%, respectively. On multivariate analysis, aspartate aminotransferase (AST)
(OR: 1.067; 95% CI: 1.017–1.119; p = 0.008) and platelet levels (OR: 0.985; 95% CI: 0.972–0.999; p =
0.034) were independent of risk factors of advanced fibrosis. Thus, our study supports screening for
NAFLD and for evaluating the severity of liver fibrosis in T2DM patients.

Keywords: NAFLD; NASH; diabetes; fibrosis; steatosis; FibroScan; CAP

1. Introduction
Estimated incidences of non-alcoholic fatty liver disease (NAFLD) worldwide have increased
twice in the last two decades, while the incidences of other chronic liver diseases (CLD) have remained
unchanged or are on downward trends [1]. Traditionally, NAFLD has been reported as a burden
condition only in the United States (US) or Western countries. However, nowadays, urbanization has
brought about sedentary lifestyles and overnutrition in many Asian countries, leading to the increase of
obesity and metabolic disorders. As the result, NAFLD has been recognized as the most common CLD

Diagnostics 2020, 10, 159; doi:10.3390/diagnostics10030159 www.mdpi.com/journal/diagnostics

Diagnostics 2020, 10, 159 2 of 10

in Asia [2,3]. Currently, the population prevalence of NAFLD in the US and Europe is approximately
30%, whereas in Asia the prevalence is around 25%, like many Western countries [3].
Although almost all NAFLD patients have simple steatosis only, 10%–20% of patients represent
the active form: non-alcoholic steatohepatits (NASH) [1]. NASH can develop into liver fibrosis,
progressing to cirrhosis, heptatocellular carcinoma (HCC), and finally end-stage liver failure. Moreover,
NAFLD patients are at 64 times higher risk of cardiovascular disease (CVD) including coronary artery
disease and stroke than patients without NAFLD [4]. Mortality in NAFLD patients is mostly due to
CVD events, markedly exceeding the common population [5].
Type 2 diabetes mellitus (T2DM) is the main risk factor of NAFLD. Patients with T2DM are at a
greater risk of NAFLD and have a higher rate of death and progression to cirrhosis than non-diabetic
individuals [6]. Therefore, screening for NAFLD and evaluating liver fibrosis in the diabetic population
is extremely essential for early detection and management, preventing the progression to advanced
fibrosis, cirrhosis and HCC.
NAFLD is diagnosed when there is evidence of ≥ 5% hepatic steatosis either by histology or
imaging and absence of secondary causes of fatty accumulation [7,8]. FibroScan with controlled
attenuation parameter (CAP) measurement can assess hepatic steatosis levels. Nevertheless, transient
elastography (TE) performed by FibroScan is recommended by the American Association for the
Study of Liver Diseases (AASLD) and European Association for the Study of the Liver (EASL) to
evaluate liver fibrosis in patients with NAFLD [7,8]. This is a non-invasive, simple-to-perform imaging
modality with high accuracy to assess liver stiffness and hepatic fat deposition. Thus far there has been
little knowledge on the prevalence of NAFLD and liver fibrosis in diabetic populations in Vietnam.
Therefore, our study proposed to estimate the prevalence of CAP-defined NAFLD and the severity of
liver fibrosis by TE performance in T2DM patients.

2. Materials and Methods

2.1. Study Design: This Study Was A Cross-Sectional Analysis

2.2. Populations

2.2.1. Inclusion Criteria

Patients older than 18 years with known T2DM or previously unknown diabetes but a displaying
fasting glucose of ≥ 126 mg/dL (7 mmol/L) or HbA1c ≥ 6.5% [9] and who were admitted for medical
check-up at Dai Phuoc Ho Chi Minh Polyclinic and the Polyclinic of Pham Ngoc Thach University of
Medicine between 07/2018 and 06/2019 were included in the study. All procedures performed in the
studies involving human participants were in accordance with the ethical standards of the local Pham
Ngoc Thach University of Medicine’s Ethical Committee (No.05/HDDD), and with the 1964 Helsinki
declaration and its later amendments or comparable ethical standards.

2.2.2. Exclusion Criteria

(1) Excessive daily alcohol intake ≥ 30 g in men and ≥ 20 g in women. (2) Causes of secondary
hepatic steatosis: pregnancy, abuse of steatogenic drugs (aminodarone, tamoxifen, methotrexate,
corticosteroid, estrogen), severe malnutrition. (3) Positive hepatitis B surface antigen or hepatitis C viral
antibody. (4) Other chronic liver diseases: autoimmune hepatitis, hemochromatosis, Wilson’s disease,
primary biliary cirrhosis, drug-induced hepatitis. (5) Measurement failure or unreliable results on TE
(ascites, cholestasis, acute exacerbation or hepatitis flare with ALT > 5 times baseline, interquartile
range to median ratio (IQR/med) > 30% or success rate < 60%).
Diagnostics 2020, 10, 159 3 of 10

2.3. Sample Size

With a predicted prevalence of NAFLD defined on FibroScan as 72.8% in a previous report [10],
our study needed a minimum sample size of 305 patients with T2DM to evaluate the prevalence with a
95% confidence interval and 5% precision.

2.4. Performance

2.4.1. Clinical Assessment

Patients who met all study criteria were enrolled. Participating subjects who had given their written
informed consent were checked for medical histories, physical examinations and laboratory tests.

2.4.2. FibroScan Examination

Participants were required to fast for at least 3 h before the procedure. TE was performed by a
single experienced operator using FibroScan 530 compact (Echosens, Paris, France) with a standard M
probe. Liver stiffness measurement (LSM) and CAP were described by the median of 10 successful
measurements. LSM was considered reliable only if IQR/med < 30% and success rate > 60%. CAP
score was considered reliable if 10 valid acquisitions were obtained.
Significant liver fibrosis (≥F2), advanced fibrosis (≥F3) and cirrhosis (F4) based on TE were defined
as LSM ≥ 7 kPa, ≥ 8.7 kPa and ≥ 11.5 kPa, respectively, according to the previous study of LSM using
TE in NAFLD patients [11]. Patients were considered to have NAFLD based on FibroScan if CAP > 233
dB/m [12] with mild, moderate, severe steatosis defined as CAP 234–269 dB/m (S1), 270–300 dB/m (S2)
and ≥ 301 dB/m (S3), respectively.
Obesity was established as a body mass index (BMI) ≥ 25 kg/m2 for adult Asians, as proposed by
the World Health Organization (WHO) [13]. Waist circumference ≥ 80 cm in females and ≥ 90 cm in
males indicated central obesity for adult Asians [14]. Hypertension was considered if the patient had a
history of hypertension and was taking antihypertensive drugs or if their systolic blood pressure (SBP)
was ≥ 140 mmHg or diastolic blood pressure (DBP) ≥ 90 mmHg [15]. The presence of dyslipidemia was
ascertained in patients who were previously diagnosed and were taking lipid-lowering drugs or who
displayed at least 1 of 4 features: cholesterol > 200mg/dL (>5.2 mmol/L), triglyceride > 150 mg/dL (>1.7
mmol/L), HDL-C < 40mg/dL and LDL-C > 130 mg/dL [16]. Patients with metabolic syndrome (MS)
were diagnosed following the standard proposed by the International Diabetes Federation (IDF) [14]:
central obesity with at least 2 of the following points: triglyceride ≥ 150 mg/dL (≥1.7 mmol/L); HDL-C
< 40 mg/dL (<1.06 mmol/L) in males or < 50 mg/dL (<1.3 mmol/L) in females, blood pressure ≥ 130/80
mmHg or on antihypertensive medication with fasting glucose plasma ≥ 100 mg/dL or the presence of
T2DM. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were determined to be
elevated when results were ≥ 33 IU/L in males and ≥ 25 IU/L in females [17]. Patients with NAFLD
with increased ALT or at least with significant fibrosis were diagnosed with NASH [18].

2.5. Data Analysis

Data were summarized and examined using STATA 12 software. Continuous variables with normal
distribution were expressed as mean and standard deviation (SD), whereas median with IQR (25 th–75
th) were used to illustrate results with asymmetric distribution. In addition, categorical elements were
represented as numbers and percentages. Comparison between continuous variables was performed
by t-test or Mann–Whitney test where appropriate. Chi-square test was performed to deal with the
categorical factors. Logistic regression was used to investigate patients’ characteristics independently
associated with advanced fibrosis. Variables proven to be significant on univariate analysis were
selected for multivariate analysis. p-value < 0.05 was assumed to be statistically significant.
Diagnostics 2020, 10, 159 4 of 10

3. Results

3.1. Population Characteristic

From 7/2018 to 6/2019, a total of 311 patients qualified for the study’s inclusions. Four patients
were eliminated due to unreliable FibroScan results (n = 3) and failed examination (n = 1). Data from
165 women and 142 men were analyzed. The average age was 56.8 ± 11.3. The BMI of the sample was
normally distributed, with a mean of 24.9 ± 2.9 kg/m2 . Approximately one-half of the study population
had a BMI greater than 25 kg/m2 . The mean waist circumference was significantly higher (p < 0.05) in
men (90.4 ± 8.9 cm) compared to women (82 ± 9.1 cm). However, there was no statistical difference in
the prevalence of central obesity between men and women, which consisted of 63.5% for both sexes.
The patient characteristics are shown in Table 1.

Table 1. Features of patients with and without non-alcoholic fatty liver disease (NAFLD) based on
transient elastography.

Patients without
Patients with NAFLD
NAFLD p
(n = 225)
(n = 82)
Age (year) 56.5 ± 10.5 57.6 ± 13.1 0.508
Male (%) 46.7 45.1 0.810
Duration of DM (year) 3 3 0.439
SBP (mmHg) 125.2 ± 13.9 122.7 ± 14.0 0.164
DBP (mmHg) 77.7 ± 9.0 74.7 ± 9.3 0.010
Hypertension (%) 65.8 52.4 0.033
BMI (kg/m2 ) 25.4 ± 2.8 23.4 ± 2.9 <0.001
Waist circumfenrence (cm) 87.9 ± 9.3 80.5 ± 9.4 <0.001
Obesity (%) 54.7 31.7 <0.001
Central obesity (%) 72.4 39.0 <0.001
Dyslipidemia (%) 93.8 73.2 <0.001
MS (%) 68.9 36.6 <0.001
Glucose (mg/dL) 136 (120–168) 125 (112–153) 0.016
Cholesterol (mg/dL) 186.7 ± 52.1 173.5 ± 51.6 0.050
Triglyceride (mg/dL) 199.1 (136.4–264) 127.3 (87–193.2) <0.001
HDL (mg/dL) 50.6 ± 13.0 56.6 ± 15.0 0.002
LDL (mg/dL) 114.2 ± 39.0 100.3 ± 41.5 0.009
ALT (U/L) 34 (23.4–54.1) 20.8 (15.9–29.8) <0.001
AST (U/L) 24.4 (20.1–33.9) 20.2 (15.8–23.7) <0.001
GGT (U/L) 46 (36.7–77.6) 37.5 (21.4–46.8) <0.001
Platelet (K/µL) 224 ± 53 229 ± 61 0.538
The diagnosis of NAFLD was based on CAP ≥ 233 dB/m. DM, diabetes mellitus; SBP, systolic blood pressure;
DBP, diastolic blood pressure. BMI, body mass index. MS, metabolic syndrome. HDL, high density lipoprotein.
LDL, low density lipoprotein. p-values were determined by t-test, and Mann-Whitney test or Chi-square test,
where appropriate.

3.2. Prevalence of NAFLD in Type 2 Diabetic Patients

Among 307 patients having reliable results, the prevalence of NAFLD based on TE was 73.3%,
with mild, moderate and severe hepatic steatosis consisting of 20.5%, 21.8% and 31.0%, respectively
(Figure 1). Moreover, the prevalence of NAFLD increased with the increasing BMI (Figure 2).
for both sexes. The
Duration patient characteristics
of DM are shown
1.092 1.000–1.193 in Table 1.
0.050
BMI 1.191 1.006–1.411 0.043 1.152 0.837–1.586 0.385
3.2. Prevalence of NAFLD in Type 2 Diabetic Patients
Obesity 2.219 0.811–6.073 0.121
Waist
Among circumference 1.055 reliable
307 patients having 1.004–1.110 0.036
results, the prevalence0.964
of NAFLD0.870–1.069 0.48673.3%,
based on TE was
Central
Diagnostics
with mild, obesityand severe
10,
moderate
2020, 159 4.916 hepatic
1.109–21.792 0.036
steatosis consisting of 3.220
20.5%, 21.8%0.460–22.527 0.239 5 of 10
and 31.0%, respectively
SBP
(Figure 1). Moreover, 0.993 0.959–1.029
the prevalence 0.713 with the increasing BMI (Figure 2).
of NAFLD increased
DBP 0.997 0.947–1.051 0.924
35.0%
Hypertension 1.621 0.563–4.672 0.371 31.0%

Percentage of steatosis
Cholesterol 30.0% 0.987–1.006
0.997 26.7% 0.485
HDL 0.981
25.0% 0.945–1.019 0.322 21.8%
20.5%
LDL 0.997
20.0% 0.984–1.009 0.583
Triglyceride 0.999 0.996–1.003 0.771
15.0%
Glucose 1.003 0.995–1.011 0.429
10.0%
ALT 1.022 1.009–1.035 0.001 0.983 0.954–1.013 0.271
AST 5.0% 1.032–1.077 <0.001
1.055 1.067 1.017–1.119 0.008
GGT 1.007
0.0% 1.003–1.010 <0.001 0.999 0.995–1.005 0.933
Platelet S0
0.981 0.970–0.993 S1
0.002 S2 0.985 S3 0.972–0.999 0.034
CAP classification
DM, diabetes mellitus; SBP, systolic blood pressure; DBP, diastolic blood pressure; BMI, body mass
index. HDL, high density lipoprotein; LDL, low density lipoprotein; CI, confidence interval; OR, odd
Figure 1. Hepatic steatosis on FibroScan with controlled attenuation parameter (CAP). S0, CAP ≤ 233
ratio.
dB/m; S1, CAP 234–269 dB/m; S2, CAP 270–300 dB/m; S3, CAP ≥ 301 dB/m.
Figure 1. Hepatic steatosis on FibroScan with controlled attenuation parameter (CAP). S0, CAP ≤ 233
dB/m; S1, CAP 234–269 dB/m; S2, CAP 270–300 dB/m; S3, CAP ≥ 301 dB/m.
100.0%
100.0%
90.0%with NAFLD had greater BMI and waist circumference than patients without
T2DM patients 80.5%
NAFLD, and tended80.0% to be obese and centrally obese. As far as we know, these patients also more
easily suffered from
70.0%metabolic64.6%
disorders such as hypertension, dyslipidemia or metabolic syndrome.
percentage

ALT, AST and gamma-glutamyltransferase (GGT) were 56.3%

Higher levels of60.0% also found in NAFLD
patients.
50.0%
40.0%
30.0%
20.0% 15.0%
7.0%
10.0%
0.0%
NAFLD Significant fibrosis

BMI <25 BMI 25 - 29.9 BMI ≥ 30

Figure 2. Proportion of patients with NAFLD and significant fibrosis according to BMI. (BMI, body
mass index.
Figure NAFLD,ofnon-alcoholic
2. Proportion patients withfatty liver and
NAFLD disease).
significant fibrosis according to BMI. (BMI, body
mass index. NAFLD, non-alcoholic fatty liver disease).
T2DM patients with NAFLD had greater BMI and waist circumference than patients without
NAFLD, and tended to be obese and centrally obese. As far as we know, these patients also more easily
4. Discussion
suffered from metabolic disorders such as hypertension, dyslipidemia or metabolic syndrome. Higher
In of
levels our study
ALT, AST ofand
307 gamma-glutamyltransferase
T2DM patients who underwent (GGT)TE,were
we found a highinprevalence
also found of NAFLD
NAFLD patients.
of 73.3%, including more than one-third of severe steatosis. The proportion of significant fibrosis,
advanced fibrosis
3.3. Prevalence and cirrhosis
of Hepatic Fibrosiswas 13.0%;
Stages 5.9%2and
in Type 3.6%,Patients
Diabetic respectively. Forty-eight-point five percent
The proportion of at least significant fibrosis, advanced fibrosis and cirrhosis was 13%, 5.9%
and 3.6%. Significant fibrosis was found to be increased with increasing BMI (Figure 2). Patients
with advanced fibrosis were more likely to be centrally obese as they had greater BMI and waist
circumference and probably sustained MS. AST, ALT and GGT were higher and platelet levels were
lower in individuals with advanced fibrosis (Table 2).
Diagnostics 2020, 10, 159 6 of 10

Table 2. Features of patients with and without advanced fibrosis on transient elastography.

Patients with Advanced Patients without

Fibrosis Advanced Fibrosis p
(n = 18) (n = 289)
Age (year) 58.7 ± 11.3 56.7 ± 11.2 0.725
Male (%) 38.9 46.7 0.518
Duration of DM (year) 6.5 (3–10) 3 (2–6) 0.056
SBP (mmHg) 123.3 ± 12.6 124.6 ± 14.0 0.690
DBP (mmHg) 76.7 ± 8.6 76.9 ± 9.2 0.920
Hypertension (%) 72.2 61.6 0.367
BMI (kg/m2 ) 26.3 ± 3.1 24.8 ± 2.9 0.039
Waist circumference (cm) 90.7 ± 9.0 85.6 ± 9.9 0.034
Dyslipidemia (%) 94.4 87.9 0.402
Obesity (%) 66.7 47.4 0.113
Central obesity (%) 88.9 61.9 0.021
MS (%) 88.9 58.5 0.011
Glucose (mg/dL) 147.5 (132–170) 133 (117–165) 0.113
Cholesterol (mg/dL) 174.5 ± 50.2 183.7 ± 52.4 0.469
Triglyceride (mg/dL) 177.1(142.3–244.9) 182.2 (109.9–250.7) 0.547
HDL–C (mg/dL) 48.9 ± 11.9 52.4 ± 13.9 0.248
LDL–C (mg/dL) 105.2 ± 41.2 110.8 ± 40.1 0.582
ALT (U/L) 60.6 (40.8–80.6) 29.7 (19.5–42.2) <0.001
AST (U/L) 45.3 (33.2–67.9) 22.6 (18.2–29.2) <0.001
GGT (U/L) 87.4 (62.4–173.9) 46 (30.4–63.4) <0.001
Platelet (K/µL) 186 ± 41 228 ± 55 0.005
The diagnosis of advanced fibrosis was based on LSM ≥ 8.7 kPa. DM, diabetes mellitus; SBP, systolic blood
pressure; DBP, diastolic blood pressure; BMI, body mass index; MS, metabolic syndrome; HDL, high density
lipoprotein; LDL, low density lipoprotein; p values were determined by t-test, Mann-Whitney test or Chi-square
test, where appropriate.

FibroScan revealed that hepatic fibrosis was mostly found in patients with NAFLD. Majority of
patients with at least significant fibrosis (≥F2) had NAFLD (38/40) while all of patients with advanced
fibrosis (≥F3) and cirrhosis were identified with NAFLD based on FibroScan.

3.4. NASH in Diabetic Patients

Forty-eight-point five percent of diabetic patients were identified with NASH after excluding
other causes of CLD. ALT was observed to be elevated in a great number of those patients (93.3%)
while 10 individuals had normal ALT but significant fibrosis (≥F2) on FibroScan (6.7%).

3.5. Factors Predicting Advanced Fibrosis in Type 2 Diabetic Patients

It can be seen in Table 3 that on univariate analysis, BMI, waist circumference, ALT, AST, GGT,
platelet and the presence of central obesity were significantly related to advanced fibrosis in T2DM
patients. However, on multivariate analysis, independent points associated with advanced fibrosis
were AST (OR: 1.067; 95%CI: 1.017–1.119; p = 0.008) and platelets (OR: 0.985; 95% CI: 0.972–0.999; p =
0.034)
Diagnostics 2020, 10, 159 7 of 10

Table 3. Associations between the presence of advanced fibrosis on transient elastography and T2DM
patients’ characteristics by univariate and multivariate logistic regression analyses.

Univariate Analysis Multivariate Analysis

OR 95% CI p Adjusted OR 95% CI p
Age 1.016 0.974–1.061 0.454
Gender 0.726 0.273–1.925 0.520
Duration of DM 1.092 1.000–1.193 0.050
BMI 1.191 1.006–1.411 0.043 1.152 0.837–1.586 0.385
Obesity 2.219 0.811–6.073 0.121
Waist circumference 1.055 1.004–1.110 0.036 0.964 0.870–1.069 0.486
Central obesity 4.916 1.109–21.792 0.036 3.220 0.460–22.527 0.239
SBP 0.993 0.959–1.029 0.713
DBP 0.997 0.947–1.051 0.924
Hypertension 1.621 0.563–4.672 0.371
Cholesterol 0.997 0.987–1.006 0.485
HDL 0.981 0.945–1.019 0.322
LDL 0.997 0.984–1.009 0.583
Triglyceride 0.999 0.996–1.003 0.771
Glucose 1.003 0.995–1.011 0.429
ALT 1.022 1.009–1.035 0.001 0.983 0.954–1.013 0.271
AST 1.055 1.032–1.077 <0.001 1.067 1.017–1.119 0.008
GGT 1.007 1.003–1.010 <0.001 0.999 0.995–1.005 0.933
Platelet 0.981 0.970–0.993 0.002 0.985 0.972–0.999 0.034
DM, diabetes mellitus; SBP, systolic blood pressure; DBP, diastolic blood pressure; BMI, body mass index. HDL,
high density lipoprotein; LDL, low density lipoprotein; CI, confidence interval; OR, odd ratio.

4. Discussion
In our study of 307 T2DM patients who underwent TE, we found a high prevalence of NAFLD
of 73.3%, including more than one-third of severe steatosis. The proportion of significant fibrosis,
advanced fibrosis and cirrhosis was 13.0%; 5.9% and 3.6%, respectively. Forty-eight-point five percent
of patients in our study had NASH, which consisted of more than 65% NAFLD patients. AST and
platelets were independent predictive factors of advanced fibrosis.
We compared our results with other prior studies. The studies of Kwok (Hongkong) and Lee-Lee
Lai (Malaysia) showed that the prevalence of NAFLD in diabetic patients based on FibroScan was 72.8%
and 72.4% with the CAP cut-offs of 222 dB/m and 263 dB/m, respectively [10,19]. Overall, our findings
were in line with the previous studies despite different CAP cut-offs. The cut-off in ours was 233 dB/m,
which was the cut-off being using at our center as well. Our study populations had considerably lower
mean BMI, that is, 24.9 kg/m2 versus 28.2 kg/m2 and 26.6 kg/m2 in the study by Kwok and Lee-Lee Lai,
respectively. BMI is known to impact CAP measurements [20]. Furthermore, more recent evidence
highlights that the epidemiology of NAFLD is significantly influenced by ethnicity [21,22], which may
also take a crucial part in designing screening programs of individual countries.
Regarding the percentage of diabetic patients with NAFLD, the range is between 49%–69.4% [6].
We believe that our data is in agreement with the published literature. There are various reported
prevalence rates of NAFLD in diabetes depending on different methods used to evaluate the liver.
Liver biopsy is considered the “gold standard” for both steatosis and fibrosis assessment. However,
this is an invasive procedure with rising risk of complications. Abdominal ultrasonography was
used to detect hepatic steatosis by most of the previous research, which is an inexpensive and widely
available equipment. However, liver ultrasound may be inadequately accurate if there is less than 33%
hepatic fat accumulation [23]. Recently, FibroScan with CAP measurement is a novel non-invasive,
easy-to-perform tool developed to assess both hepatic steatosis and fibrosis simultaneously with high
sensitivity and specificity [24]. Screening for NAFLD in T2DM individuals using FibroScan/CAP is
recommended in the latest Asia–Pacific Working Party on Non–Alcoholic Liver Disease guidelines [25].
Diagnostics 2020, 10, 159 8 of 10

Moreover, we also found increasing rates of MS, hypertension, dyslipidemia and obesity in patients
with NAFLD. Therefore, it is important to evaluate all components of MS in NAFLD patients and vice
versa, and to screen for NAFLD in patients with obesity, MS, hypertension or dyslipidemia.
FibroScan in our center is not equipped with an XL probe, so the accuracy of LSM in individuals
having a BMI greater than 28 kg/m2 may have been influenced [26]. As far as we know, there are two
other studies using only an M probe and the same cut-offs for fibrosis stages like ours. In their study,
Sobhonslidsuk and colleagues pointed out that the prevalence of significant fibrosis and advanced
fibrosis based on TE was 16.1% and 5.8%, respectively [27]. Apparently, these findings concurred
well with our study findings. This is likely explained by the similarities in population characteristics
between Vietnamese and Thai studies. Otherwise, the study of de Lédingen and colleagues discovered
that patients with advanced fibrosis accounted for 12.9% [28], which was remarkably higher compared
to our results. This may be because of the diversity in patient characteristics between two studies, for
instance, BMI and waist circumference in the French study (27.2 kg/m2 and 97.2 cm) was greater than in
ours (24.9 kg/m2 and 85.9 cm). Another possible reason for this is that liver fibrosis related to NAFLD
in Asia is not as common as in Europe. Natural development of NAFLD to end-stage liver disease
usually progresses slowly, lasting for around 10 to 20 years, and the latter appearance of NAFLD
results in a lower proportion of advanced fibrosis or cirrhosis in many Asian surveys compared to
those in Europe [3].
The presence of NAFLD in T2DM patients can lead to substantial risk of liver fibrosis. In our study,
the proportion of fibrosis severities in patients with NAFLD were considerably higher than patients
without NAFLD (p < 0.05). Vice versa, diabetes is associated with increased risk of advanced fibrosis
as well. Another study using FibroScan to evaluate liver fibrosis in both diabetic and non-diabetic
patients revealed that advanced fibrosis in diabetic subjects was 26.9% while only 4.5% of non-diabetic
individuals had advanced fibrosis [29]. In addition, it is important to note that among diabetic patients
with BMI < 25 kg/m2 , 64.6% patients still had NAFLD and 7% had significant fibrosis. In previous
studies about NAFLD, such non-obese patients usually have other components of MS [30]. Obviously,
the presence of diabetes itself is one of the components of MS and the prevalence of MS was high among
NAFLD patients in our study as well. These non-obese populations still face metabolic problems and
the risk of CVD equivalently to obese individuals [31].
NAFLD is believed to be the leading reason of ALT elevation in type 2 diabetic patients [6]. Our
study revealed that elevated liver enzymes in patients with NAFLD were mainly due to increased
ALT related to NASH after dropping out other causes of CLD. Ninety percent of NASH patients had
increased ALT and AST, but in 10% of NASH patients the ALT may be normal [1]. We figured out
that nearly 7% of patients had significant fibrosis but not elevated ALT. In our findings, AST and
platelet levels appeared to be independent predictive factors of advanced fibrosis in T2DM patients.
ALT is mainly located in the cytoplasm of the hepatic cell. By contrast, AST is mostly present in the
mitochondrial membrane (80%), while the remainder is in the cytosol [32]. Chronic liver injuries can
damage the mitochondria leading to the increasing release of AST into the blood, which is commonly
seen in patients with advanced fibrosis. Moreover, hepatic fibrosis progression can also result in
reduced elimination of AST in hepatic sinusoid contributing to the elevation of AST serum [33]. On
the other hand, advanced fibrosis can precede portal hypertension inducing thrombocytopenia due to
increased sequestration and destruction of platelets in the enlarged spleen [34]. Lower platelet levels
predict a more extreme hepatic fibrosis.
Our work has several limitations. First of all, we do not have an XL probe to perform FibroScan
examination on patients with BMI greater than 28 kg/m2 . Secondly, owning to lack of liver biopsy for
diagnostic confirmation of TE findings, we do not have the precise prevalence of NASH. Finally, our
findings may not be generalizable to patients with T2DM in all primary care settings as well as diabetic
inpatients at hospitals. Future studies should concentrate on enhancing the quality of the current topic.
Diagnostics 2020, 10, 159 9 of 10

5. Conclusions
To sum up, we found that the prevalence of NAFLD based on transient electrography is high
among T2DM patients. Patients with NAFLD are at an increased risk of advanced fibrosis than those
without NAFLD. Our study supports screening for NAFLD and evaluating severity of hepatic fibrosis
in T2DM patients.

Author Contributions: T.T.K.T., P.Q.T.P. and T.N.D.H. designed and performed experiments and collected data
and informed consent. T.T.K.T., D.K.T., P.Q.T.P., T.N.D.H., T.C.D. and D.T.C. analysed and interpreted the results,
and edited and corrected the manuscript. T.T.K.T., D.K.T., P.Q.T.P., T.C.D. and D.T.C. wrote the manuscript. All
authors have read and agreed to the published version of the manuscript.
Funding: The authors declared that this study was funded by the Pham Ngoc Thach University of Medicine
Research Fund.
Acknowledgments: We would like to thank Phuong Linh Nguyen (a freelance English editor, and an English
teacher at the Ban Mai School, Ha Dong, Hanoi 100000, Vietnam) for critical reading and checking to improve
the manuscript.
Conflicts of Interest: The authors declare no conflict of interest.

References
1. LaBrecque, D.; Abass, Z.; Anania, F.; Ferenci, P. Nonalcoholic fatty liver disease and nonalcoholic
steatohepatitis. In World Gastroenterology Organisation Global Guidelines; WGO: Milwaukee, WI, USA,
2012.
2. Ashtari, S.; Pourhoseingholi, M.; Zali, M. Non-alcohol fatty liver disease in Asia: Prevention and planning.
World J. Hepatol. 2015, 7, 1788–1796. [CrossRef] [PubMed]
3. Jian-Gao, F.; Seung-Up, K.; Vincent, W. New trends on obesity and NAFLD in Asia. J. Hepatol. 2017, 67,
862–873.
4. Targher, G.; Byrne, C.; Lonardo, A.; Zoppini, G.; Barbui, C. Non-alcoholic fatty liver disease and risk of
incident cardiovascular disease: A meta-analysis. J. Hepatol. 2016, 65, 589–600. [CrossRef] [PubMed]
5. Ekstedt, M.; Franzen, L.; Mathiesen, U.; Thorelius, L. Long-term follow-up of patients with NAFLD and
elevated liver enzymes. Hepatology 2006, 44, 865–873. [CrossRef] [PubMed]
6. Younossi, Z.; Gramlich, T.; Matteoni, C.; Boparai, N.; Mccullough, A. Nonalcoholic fatty liver disease in
patients with type 2 diabetes. Clin. Gastroenterol. Hepatol. 2004, 2, 262–265. [CrossRef]
7. Chalasani, N.; Younossi, Z.; Lavine, J.; Charlton, M.; Cusi, K.; Rinella, M.; Harrison, S.; Brunt, E.M.; Sanyal, A.J.
The diagnosis and management of nonalcoholic fatty liver disease: Practice guidance from the American
Association for the study of liver diseases. Hepatology 2018, 67, 328–357. [CrossRef]
8. EASL; EASD; EASO. EASL–EASD–EASO clinical practice guidelines for the management of non-alcoholic
fatty liver disease. J. Hepatol. 2016, 59, 1121–1140.
9. Association, A.D. Classification and diagnosis of diabetes: Standards of medical care in diabetes 2018. In
Standards of Medical Care in Diabetes—2018; Riddle, M., Ed.; American Diabetes Association: Arlington
County, VA, USA, 2018; Volume 41, pp. 13–27.
10. Kwok, R.; Choi, K.; Wong, G.; Zhang, Y. Screening diabetic patients for non-alcoholic fatty liver disease with
controlled attenuation parameter and liver stiffness measurements: A prospective cohort study. Gut 2015, 65,
1359–1368. [CrossRef]
11. Wong, V.; Vergniol, J.; Wong, G.; Foucher, J. Diagnosis of fibrosis and cirrhosis using liver stiffness measurement
in nonalcoholic fatty liver disease. Hepatology 2010, 51, 454–462. [CrossRef]
12. Karlas, T.; Petroff, D.; Garnov, N.; Bohm, S. Non-invasive assessment of hepatic steatosis in patients with
NAFLD using controlled attenuation parameter and H-MR spectroscopy. PLoS ONE 2014, 9, e91987.
[CrossRef]
13. World Health Organization. The Asia-Pacific Perspective: Redefining Obesity and Its Treatment; WHO: Geneva,
Switzerland, 2000; pp. 18–20.
14. International Diabetes Federation. The IDF Consensus Worldwide Definition of the Metabolic Syndrome;
International Diabetes Federation: Brussels, Belgium, 2006.
Diagnostics 2020, 10, 159 10 of 10

15. Williams, B.; Mancia, G.; Spiering, W.; Agabiti Rosei, E.; Azizi, M.; Burnier, M.; Clement, D.L.; Coca, A.; de
Simone, G.; Dominiczak, A.; et al. 2018 ESC/ESH guidelines for the management of arterial hypertension.
Eur. Heart J. 2018, 39, 3021–3104. [CrossRef] [PubMed]
16. National, C.E.P.C.C. National Cholesterol Education Program Expert Panel on Detection, Evaluation, and Treatment
of High Blood Cholesterol in Adults (Adult Treatment Panel III); International Medical publishing, Inc.: Mclean,
VA, USA, 2002; p. 106.
17. Kwo, P.; Cohen, S.; Lim, J. ACG practice guideline: Evaluation of abnormal liver chemistries. Am. J.
Gastroenterol. 2016, 112, 18–35. [CrossRef] [PubMed]
18. Adams, L.; Feldstein, A. Non-invasive diagnosis of nonalcoholic fatty liver and nonalcoholic steatohepatitis.
J. Dig. Dis. 2011, 12, 10–16. [CrossRef] [PubMed]
19. Lai, L.; Yusoff, W.; Vethakkan, S.; Mustapha, N. Screening for non-alcoholic fatty liver disease in patients
with type 2 diabetes mellitus using transient elastography. J. Hepatol. 2019, 34, 1396–1403. [CrossRef]
20. Wang, Y.; Fan, Q.; Wang, T.; Wen, J.; Wang, H.; Zhang, T. Controlled attenuation parameter for assessment of
hepatic steatosis grades: A diagnostic meta-analysis. Int. J. Clin. Exp. Med. 2015, 8, 17654–17663. [PubMed]
21. Bambha, K.; Belt, P.; Abraham, M.; Wilson, L.A. Ethnicity and nonalcoholic fatty liver disease. Hepatology
2012, 55, 769–780. [CrossRef]
22. Rich, N.; Oji, S.; Mufti, A.; Browning, J.D.; Parikh, N.D.; Odewole, M.; Mayo, H.; Singal, A.G. Racial and
ethnic disparities in nonalcoholic fatty liver disease prevalence, severity, and outcomes in the united states: A
systematic review and meta-analysis. Clin. Gastroenterol. Hepatol. 2018, 16, 198–210.e2. [CrossRef] [PubMed]
23. Ryan, C.; Johnson, L.; Germin, B.; Marcos, A. One hundred consecutive hepatic biopsies in the workup of
living donors for right lobe liver transplantation. Liver Transplant. 2002, 8, 1114–1122. [CrossRef]
24. Je´rome, B.; Paul, C. Controlled attenuation parameter (CAP): A new device for fast evaluation of liver fat?
Liver Int. 2012, 32, 875–877.
25. Wong, V.; Chan, W.; Chitturi, S.Y.C. Asia-Pacifc working party on non-alcoholic fatty liver disease guidelines
2017-Part 1: Definition, risk factors and assessment. J. Gastroenterol. Hepatol. 2018, 33, 70–85. [CrossRef]
26. Wong, G.; Wong, V.; Chim, A.; Yiu, K. Factors associated with unreliable liver stiffness measurement and
its failure with transient elastography in the Chinese population. Hepatology 2011, 26, 300–305. [CrossRef]
[PubMed]
27. Sobhonslidsuk, A.; Pulsombat, A.; Kaewdoung, P.; Petraksa, S. Non-alcoholic fatty liver disease (nafld) and
significant hepatic fibrosis defined by non-invasive assessment in patients with type 2 diabetes. Asian Pac. J.
Cancer Prev. 2015, 16, 1789–1794. [CrossRef] [PubMed]
28. Lédinghen, V.; Vergniol, J.; Gonzalez, C.; Foucher, J. Screening for liver fibrosis by using FibroScan®and
FibroTest in patients with diabetes. Dig. Liver Dis. 2012, 44, 413–418. [CrossRef] [PubMed]
29. Hajiani, E.; Alavinejad, P.; Hashemi, S.; Masjedizadeh, A. Comparison of the transient elastography (fibroscan)
results among diabetic and non-diabetic patients with non- alcoholic fatty liver disease. Gastroenterol. Hepatol.
2014, 1, 00021.
30. Das, K.; Das, K.; Mukherjee, P.; Ghosh, A.; Ghosh, S. Nonobese population in a developing country has a high
prevalence of nonalcoholic fatty liver and significant liver disease. Hepatology 2010, 51, 1593–1602. [CrossRef]
31. Sookoian, S.; Pirola, C. Systematic review with meta-analysis: Risk factors for non-alcoholic fatty liver disease
suggest a shared altered metabolic and cardiovascular profile between lean and obese patients. Aliment.
Pharm. 2017, 46, 85–95. [CrossRef]
32. Rej, R. Aspartate aminotransferase activity and isoenzyme proportions in human liver tissues. Clin. Chem.
1978, 24, 1971. [CrossRef]
33. Wai, C.; Marrero, J.A.; Conjeevaram, H.S.; Lok, A.S.; Greenson, J.K.; Fontana, R.J.; Kalbfleisch, J.D. A simple
noninvasive index can predict both significant fibrosis and cirrhosis in patients with chronic hepatitis C.
Hepatology 2003, 38, 518–526. [CrossRef]
34. Witters, P.; Freson, K.; Verslype, C.; Peerlinck, K. Review article: Blood platelet number and function in
chronic liver disease and cirrhosis. Aliment. Pharm. Ther. 2008, 27, 1017–1029. [CrossRef]

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).