2016 Article 929
2016 Article 929
2016 Article 929
DOI 10.1186/s13075-016-0929-x
miRNAs use this seed region to determine the possible that has emerged as a master regulator appears to be
cognate targets of miRNAs by conserved binding sites. miRNA-29. This is a family of miRNA29s and includes
Such databases include TargetScan (www.targetscan.org). a, b, and c and differs in one, two, or three bases. These
Although these programs are useful, they are not abso- are clearly enmeshed in the pathogenesis of fibrosis. First
lutely correct and undoubtedly have false positives in described in fibrosis of the heart by van Rooij et al. in a
there. There can be large differences in targets using dif- landmark study, it has now been demonstrated to play a
ferent target prediction software. Furthermore, it has been major role in fibrosis [3].
found that miRNAs can target 5′ UTRs and it is now ap- Decreased levels of miRNA-29 are associated with fi-
preciated that they may actually be more promiscuous brosis in multiple organs, including the heart, liver [4],
than first thought, underscoring their importance in post- kidney [5] and skin, and in SSc [6], suggesting that this
transcriptional regulation. MiRs often target mRNAs that is a core ‘fibromiRNA’ [7]. In the heart, antagonism of
govern the same pathway that results in modulation of the miRNA-29 in vivo resulted in enhanced collagen in the
pathway. tissue, whereas introduction of miRNA-29 mimics after
heart damage reduced the collagen content [3]. Zhu et al.
Fibrosis signalling have also found downregulation of miRNA-29 in SSc
The ultimate target cells in fibrosis are the fibroblasts fibroblasts and this correlated with the levels of its tar-
where it produces a surfeit of ECM that ultimately im- get gene collagen1A1 [8]. Whatever downregulates the
pairs normal functioning of the tissue and organ. In- miRNA-29a, this results in a de-repression of its tar-
flammation is a key component of fibrosis and many gets that to date include the major structural collagens.
cytokines and chemokines are involved in orchestrating We have found TAB1 to be a bone fide target of
this. Although there are many diverse proteins involved, miRNA-29a in dermal fibroblasts [9]. This gene regu-
transforming growth factor-beta (TGF-β) is considered lates TIMP-1, which blocks the action of matrix metal-
the most important (see below). Other cytokines such loproteinases, the enzymes that degrade ECM, and
as interleukin (IL)-6, IL-1, IL-13 and inflammasome thus alters the balance of ECM turnover. Dysregulated
components are also considered important mediators TIMP-1 has been previously reported in SSc, and
in disease pathogenesis. Also, molecules such as the TIMP-1 is critical in hepatic fibrosis. Indeed, chemical
neurotransmitter serotonin are important in mediating inhibition of TAB1 also reduced ECM deposition [9],
fibrosis but these may also lie upstream of TGF-β and and we found downregulation of miRNA-29a in SSc fi-
converge on this molecule. Clearly, all these molecules broblasts [9]. MiRNA-29a has also been shown to be
may themselves regulate the expression of miRNAs downregulated in SSc and is reduced by Toll-like re-
involved in fibrosis. Table 1 demonstrates miRNAs in ceptor (TLR) 4 stimulation [10]. TAB is activated by
fibrosis and their confirmed targets. TGF-β via the non-canonical pathway [11]. Furthermore,
miRNA-29 has been shown to target platelet-derived
miRNA29 as a ‘master fibromiRNA’ regulator growth factor receptor (PDGFR) [12], and platelet-derived
The exact relative contribution of miRNAs to fibrosis growth factor signalling is known to be important in fibro-
and the bona fide targets are not fully known. MiRNAs sis [13]. Interestingly, the miRNA-29 family has been
can be either pro- or anti-fibrotic. However, one miRNA shown to target the DNA methyltransferases (DNMTs)
Table 1 MicroRNAs in fibrosis
MicroRNA Target gene Pro- or anti-fibrotic Tissue References
17-5p Smad7 Pro Liver [34]
21 Sprouty homolog-1, PTEN, SMAD7, STAT3, PPAR-α Pro Heart, skin, kidney [22, 23, 63]
29, a,b,c Collagen1A1, 1A2, 4A5, FBN, ELN1, PDGFR, TAB1, ADAM Anti Heart, skin, liver [9, 14–17, 20]
33a PPAR-α Pro Liver [29]
122 P4HA1 Anti Liver [59]
129-5p Collagen1A1 Anti Skin [39]
132 MeCP2 Anti Liver [41]
133a Collagens Anti Liver [28]
192 ZEB1 Pro Kidney [31]
199b DYRK1A Pro Heart [51]
199a-5p CAV1 Pro Lungs, skin [47]
214 CTGF Anti Liver [70]
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and this results in methylation abnormalities [14]. Thus, systemically injected ameliorated hepatic fibrosis in-
reduced miRNA-29 could be targeting DNMTs, thus duced by carbon tetrachloride [21].
leading to hypermethylation and silencing of genes im-
portant in ECM regulation. Cushing et al. also showed MiRNAs that modulate TGF-β signalling
that miRNA-29 regulates a variety of ECM genes that Amongst the many cytokines fuelling fibrosis, TGF-β is
include the standard ECM proteins such as collagens one of the most pivotal cytokines mediating fibrosis in
and lamins but also integrins that themselves are in- both organ-specific disease and SSc. TGF-β is critical in
volved in ECM regulation by activating latent TGF-β fibrosis but also in the development of the organism. It
[15]. Furthermore, miRNA-29 is reduced in kidney fi- is elevated in many diseases and animal models of fibro-
brosis and is modulated by TGF-β, and the target of sis and affects the activation of quiescent fibroblasts to
miRNA-29 was found to be disintegrin metalloprotease ‘myofibroblasts’: the effector cells in fibrosis. The myofi-
(a disintegrin and metalloproteinase, or ADAM) [16]. It broblast expresses high amounts of alpha-smooth muscle
was also found that miRNA-29 is reduced in lung fibro- actin, secretes copious amounts of ECM and endows the
sis in both a TGF-β- and Smad3-dependent manner cell with enhanced contractile force. TGF-β is secreted as
[17]. Smad3 genetically reduced mice were protected a latent protein non-covalently linked to latency-activating
from fibrosis induced by bleomycin, confirming its regula- protein (LAP) and through enzymatic proteolytic cleavage
tion by TGF-β. Importantly, sleeping beauty transponson- becomes ‘active’ after dissociating from LAP to bind to its
mediated gene transfer of miRNA-29 to replace the cognate receptors. TGF-β then mediates gene expression
reduced miRNA resulted in an attenuation of fibrosis via activation of the canonical Smad signalling pathway
[17]. This is an important step as this circumvents the from transcriptional activation. MiRNAs that regulate
need for the use of viral vectors that have problems TGF-β would therefore be of prime importance. Only re-
associated with them, such as triggering host immune cently have we begun to understand that miRNAs can be
responses to the vector and labour costs. There is a fur- regulated by TGF-β and TGF-β itself can be regulated by
ther problem with the transduction, and the efficiency miRNAs. One such miRNA is miRNA-21, which has been
of viral vectors depends on the tropism of the virus for found to be upregulated by TGF-β stimulation. Further-
a particular tissue. A further study showed that diabetic more, miRNA-21 is overexpressed in heart disease, and in
kidney disease-associated inflammation and fibrosis are a pressure overload model it was found that miRNA-21
aggravated by genetic renal loss of miRNA-29b in mice, overexpression modulates cardiac myocyte hypertrophy
further underpinning the role of miRNA-29 in fibrosis. and interstitial fibrosis through reduction of sprouty
Gene replacement therapy of lost miRNA-29b expression homologue-1, a potent inhibitor of the extracellular
in the mouse kidney reduced the fibrosis and inflamma- regulated kinase/mitogen-activated protein kinase (ERK/
tion in the diabetic kidney disease model, and the mech- MAPK) pathway, and thus elevates this signalling cascade
anism appears to include the TGF-β/Smad3 axis by [22]. In the bleomycin model of fibrosis, this increased
modulating this [18]. Upregulation of miRNA-29a either miRNA-21 levels and the addition of antisense oligonucle-
by enforced overexpression or by the use of carveldilol, a otides to miRNA-21 reduced the severity of lung fibrosis
beta-adrenoreceptor antagonist, was shown to reduce even after a lag time of many days after lung injury [23].
myocardial fibrosis mediated by experimental myocardial Knockdown of miRNA-21 reduced pro-fibrotic responses
infarction in a small animal model [19]. Thus, agents that of TGF-β stimulation, whereas increasing miRNA-21 en-
increase the levels of miRNA-29 would be predicted to hanced the pro-fibrotic response [23]. It appears that the
reduce fibrosis. Although many cytokines that decrease target mRNA of miRNA-21 is Smad7, which is an inhibi-
the levels of miRNA-29 have been identified and these tory Smad, attenuating the Smad-dependent signalling
have been found to correlate in vivo, few, if any, cyto- pathway. There is, of course, cross-talk between the Smad
kines have been found to increase the miRNA. In the and other pathways, adding another layer of complexity.
bleomycin model of skin fibrosis, a mimic of SSc, it Smad3 itself regulates the expression of miRNA-21 by
was found that miRNA-29a is supressed; however, res- increasing this after stimulation with TGF-β; however,
toration of miRNA-29a levels with the use of the tyro- Smad2 inhibits this. Inhibition of miRNA-21 by ultra-
sine kinase inhibitor imatinib attenuated this fibrosis sound microbubble-mediated gene transfer of a nega-
[6], suggesting that miRNA-29a is regulated by tyro- tive miRNA-21 plasmid attenuated kidney fibrosis in an
sine kinase activation. It has recently been shown that obstructive nephropathy model in mice, underscoring
miRNA-29a is also regulated by the alarmin IL-33 [20]. the importance of this miR [24]. MiRNA-21 has also
IL-33 is a danger-associated molecule that is released been demonstrated to be elevated in SSc skin whole tissue
in tendon disease and this leads to downregulation of and isolated fibroblasts [8]. Interestingly, miRNA-21 has a
miRNA-29a and a subsequent increase in collagen signal transducer and activator of transcription 3 (STAT3)
levels. Recently, adenoviral overexpression of miR29a binding site and this transcriptionally regulates the
O’Reilly Arthritis Research & Therapy (2016) 18:11 Page 4 of 10
expression of miRNA-21, altering its targets [25]. STAT3 it- but this cluster also is important in fibrosis. It is critical in
self is an important molecule activated by IL-6 and mediates epithelial lung development, and mice that lack this cluster
fibrosis in SSc fibroblasts [26]. A possible target of miRNA- do not have many lung epithelial cells and die shortly after
21 is the gene phosphatase and tensin homolog (PTEN) and birth. Mice in which transgenic overexpression in the lungs
this abrogates a cell signalling pathway, resulting in in- of this miRNA cluster results in lung hyperproliferation and
creased ECM deposition [27]. It is known that the main blocks differentiation of lung progenitor cells were also
substrate of PTEN is inositol-3,4,5-triphosphate and this ac- demonstrated with transgenic technology [35]. It was found
tivates Akt important in wound repair. It is also known that in idiopathic pulmonary fibrosis samples that miRNA-17-92
PTEN is reduced in SSc skin and that PTEN-ablated mice cluster was reduced and this was confirmed by both poly-
have exacerbated fibrosis compared with wild-type controls. merase chain reaction and in situ hybridisation techniques.
Thus, elevated miRNA-21 reduces the expression of PTEN It was found that there was hypermethylation of DNA in
and this leads to altered Akt signalling, leading to increased the promoter region of miRNA-17-92 and that this was
ECM deposition. Interestingly, unsaturated fatty acids ap- leading to repressed expression of the cluster; the enzyme
pear to upregulate miRNA-21 levels, inducing the suppres- DMNT-1 is altered in pulmonary fibrosis and this is also a
sion of PTEN in hepatocytes [27], suggesting a link between target gene of some of the miRNAs in the cluster. Thus, a
the metabolic syndrome, miRNAs and fibrosis. complicated feed-forward loop exists in which miRNAs
TGF-β has been shown to downregulate miRNA-133a from the gene cluster target DNMT-1; thus, the reduction
levels in hepatic stellate cells and during hepatic myofi- of the miRNAs leads to enhanced DNMT-1 levels. Transfec-
broblast development without decreasing stimuli miRNA- tion of miRNAs reduced DNMT-1 levels and thus globally
133a levels [28]. Targets of miRNA-133a appear to be methylated DNA levels. Also, in vitro and in vivo adminis-
collagens, the major component of the fibrotic scar. tration of 5′aza′2-deoxycytidine, a global demethylation
Interestingly, serum expression of miRNA-133a was agent, restores the reduction of the miRNA cluster and
found to be elevated in patients with liver fibrosis and reductions in their targets such as collagen1A1 and
indicated the progression of liver fibrosis [28], indicat- CTGF. In vivo treatment of bleomycin-treated mice
ing that this could be a valid biomarker where today after commencement of fibrosis attenuated the fibrosis
none exists. For confirmation of liver fibrosis, often a in the lungs and also elevated levels of the miRNA clus-
biopsy is the only option but this, of course, is invasive. ter under examination. This was accompanied by a re-
MiRNA-33a has also been shown to be modulated by duction in DNMT-1 enzyme levels as this is targeted by
TGF-β in stellate cells, and the potential targets of the miRNA and also a reduction in pro-fibrotic genes
miRNA-33a are peroxisome proliferator activator receptor driving the fibrosis [36]. This cluster has been impli-
(PPAR) alpha and Akt in these cells. This is important as cated in liver fibrosis with the bona fide target identi-
PPAR is critical in the shift from a quiescent to a myofi- fied as CTGF mediated via p53, a tumour-related gene
broblast in hepatic stellate cells [29]. [37]. CTGF is important as a fibrotic molecule in its
MiRNA-192 is also upregulated by the addition of own right and is often associated with TGF-β.
TGF-β1 and this upregulation promotes collagen depos-
ition in a kidney fibrosis model [30]. Use of antagomirs MiRNA-129-5p
to miRNA-192 reduced kidney fibrosis [30]. It appears that In the prototypic fibrotic disease SSc, there is an in-
miRNA-192 targets the E-Box repressor Zinc finger E-box crease in leucocytes in the skin, including primarily T
binding homeobox 1 (ZEB1) modulating E-cadherin levels, cells. These T cells that are residing in the skin are in
enhancing kidney fibrosis [31]. ZEB1 is a critical transcrip- close proximity to the myofibroblasts, suggesting that
tion factor in morphogenesis and epithelial-to-mesenchymal they are governing their transdifferentiation [38] and may
transition [32]. MiRNA-145 is also upregulated by TGF-β activate other immune cells in the inflammatory foci. It has
and this leads to increased fibrosis by increasing alpha- been described in SSc fibroblasts that miRNA-129-5p is re-
smooth muscle actin levels via activating latent TGF-β [33]. pressed compared with healthy control fibroblasts [39].
Recently, miR17-5p was found to be induced in hep- They also show that the T-cell cytokine IL-17 can increase
atic stellate cells by TGF-β1 induction and activation miRNA-129-5p levels, and using siRNA to knock down
and was also upregulated in the carbon tetrachloride IL-17 receptors in dermal fibroblasts reduced miRNA-129-
model of liver fibrosis [34]. In vitro inhibition of miR17- 5p levels. The actual target mRNA of miRNA-129-5p ap-
5p led to reduction of cell activation and proliferation pears to be collagen alpha-1 [39]. This all suggests that the
but no alteration in apoptosis levels [34]. Th17 cells reduce collagen expression via the upregulation
of the negative regulator miRNA-129-5p; however, in SSc
MiRNA-17-92 cluster fibroblasts, the dysregulated TGF-β levels alter the cells’
The miRNA-17-92 cluster which encodes seven miRNAs is sensitivity to IL-17, possibly through downregulation of
important in oncogenesis (sometimes called OncomiRNA1), the TGF-β receptors, thus leading to reduced IL-17
O’Reilly Arthritis Research & Therapy (2016) 18:11 Page 5 of 10
Fig. 1 miRNA-132 feed-forward loop. Various factors such as IL-6 and dietary factors, including alcohol, reduce the expression of miRNA-132 and
thus lead to increases in its target mRNA meCP2, and meCP2 interacts with the methylase Ezh2 that methylates lysine27 on H3 and interacts with
PRC1; this all leads to repression of the master regulator PPARγ and myofibroblast activation and extracellular matrix expression. Blockade of Ezh2
with DZnep may be beneficial in fibrosis through blocking histone methylation. Ezh2 enhancer of Zeste homologue 2, IL interleukin, MeCP2 methyl cap-
binding protein 2, miRNA microRNA, PPARγ peroxisome proliferator activator receptor-gamma, PRC1 polycomb recessive complex 1, TGF-β transforming
growth factor-beta
O’Reilly Arthritis Research & Therapy (2016) 18:11 Page 6 of 10
Fig. 2 miRNA modulation. Two methods are employed for miRNA therapeutics restoration of miRNA or inhibition. Restoring miRNA is through the
use of double-stranded RNA, which is composed of a guide and passenger strand that is chemically modified usually by a cholesterol modification.
The guide strand is identical to the miRNA that is diminished. This is then incorporated into the RISC complex and the target mRNAs reduced.
Inhibition of miRNA function is achieved through single-stranded chemically modified LNA antagomirs which can be cholesterol-conjugated
and have increased stability. These molecules bind the mature miRNA and stop them from being loaded into the RISC, therefore increasing
the mRNA target(s). In the context of fibrosis, to the right of the illustrations are examples of miRNA as both a mimic and antagomir binding
the target mRNA and altering the protein output. Smad7 is a negative regulator of the fibrotic cascade, so elevated levels reduce fibrosis.
PPAR-α is also a negative regulator of fibrosis and thus its restoration is positive. miRNA, MicroRNA, PPARα peroxisome proliferator activator
receptor-alpha, RISC RNA-induced silencing complex
O’Reilly Arthritis Research & Therapy (2016) 18:11 Page 7 of 10
monomer introduced and increases the binding affinity of lung cancer [61]. It is suggested that the use of a neutral
the nucleic acid to the mRNA [53]. LNA-modified antago- lipid vehicle does not aggregate so much and is less likely
mirs were first used in vivo in non-human primates to to be engulfed by macrophages [61]. As further delivery
target the liver miRNA-122 and resulted in a sustained systems are developed for miRNAs to target them more
and long-lasting decrease in cholesterol [54]. Injection specifically to the target tissue and cells, further wide-
of unmodified miRNAs into the tail vein of mice results spread use will occur. A further issue with the blockade of
in breakdown and renal excretion very quickly [55]. A miRNAs with antagomirs may be that the levels of redun-
different chemical modification is a 2′O-methyl modifi- dancy may restrict the therapeutic benefit.
cation that increases nuclease resistance. Another chem- The vista is optimistic for using miRNAs therapeutic-
ical modification is the addition of a cholesterol moiety at ally to treat fibrotic conditions such as SSc. This is one
the 3′ end. Despite the theoretical rationale for modula- of the fastest growing areas in scientific research and
tion, only one antagomir is currently in clinical use. This offers the potential to revolutionise treatment. It was
is the antagomir for miRNA-122, marketed as Miravirsen shown in an animal model of kidney fibrosis that the use
(Roche, Copenhagen, Denmark), and has shown great of an antagomir to miRNA-214 attenuated kidney fibro-
clinical efficacy [56]. Miravirsen targets miRNA-122, sis [62] and that this is independent of classic TGF-β
which is a liver-specific miRNA and thus is not expressed signalling.
anywhere else; it uses LNA technology. It works by block- One of the most studied fibrotic miRNAs is miRNA-21,
ing the interaction with the miRNA (122) and its target and use of an antagomir in two mouse models of kidney
RNA in the hepatitis C virus (HCV) 5′ UTR [57]. fibrosis demonstrated great clinical efficacy in reducing fi-
MiRNA-122 is essential for the replication of HCV [58] brosis [63]. Mechanistically, it was found that miRNA-21
and lipid and iron metabolism in the liver [59]. It appears targets the 3′ UTR of PPAR-α, an important receptor in
safe and well tolerated and has no adverse effects. Another involved in lipid metabolism; indeed, deletion of PPAR-α
approach to targeting miRNAs is termed tiny LNAs. abrogated the effect of anti-miRNA-21 anti-fibrotic effects
These are tiny (8-mer) LNA anti-miRNAs that target only [63]. The authors further demonstrated that mitochon-
the conserved seed region [60]. An alternative strategy is drial oxidative regulators are also a target of miRNA-21
the use of miRNA masking (miRNA-Mask); this works by [63], suggesting that metabolic stress is important in fi-
the introduction of single-stranded RNA that targets the brosis generation. Regulus Therapeutics (San Diego, CA,
3′ UTR of the protein coding mRNA and thus ‘covers up’ USA) also has an antagomir against miRNA-21 (RG-012)
the binding site to the miRNA and thereby de-represses in the developmental pipeline and this antagomir showed
the protein target. This technology has been used to con- good efficacy in animal models of Alport syndrome, an
firm targets of miRNAs in vitro. Further study is needed inherited kidney disease in which mutations in collagen
to determine which chemical modification is optimum for lead to excess collagen deposition within the kidney.
stability without undue toxicity. Another way of targeting
miRNAs in fibrosis is to reconstitute reduced levels of the Replacing reduced miRNAs
miRNA with miRNA replacement, thus restoring the sup- Replacement of reduced miRNA expression by using miRNA
pression of the putative target(s). The introduction of such mimics could be one therapeutic strategy in fibrosis.
miRNA mimic replacement can occur through the direct This involves the use of RNA duplexes in which there
introduction of the mimic or the use of viral vectors to ex- is a ‘guide strand’ and a passenger strand that may be
press the miRNA. Viral vectors, however, may induce an chemically modified. Although restitution of miRNAs is
unwanted immune response, and tissue tropism may limit in its infancy, it has been proven in animal models of
their effectiveness. Therapeutically, antagomirs are much cancer and more recently fibrosis. Often restoring the
more advanced than miRNA replacement therapy. One lost miRNA is achieved by viral vectors. This has its
issue that is currently hampering the field is that the own challenges, as the vector size may be large and sys-
introduction of miRNA therapeutics is not cell- or temic delivery may be difficult to achieve and can also
organ-specific and thus being systemic may have major have high immunogenicity. Montgomery et al. recently
side effects. Getting the miRNA therapeutic to the cor- published the use of a miRNA-29 mimic in an animal
rect target tissue remains a challenge, especially if model of fibrosis, the bleomycin lung fibrosis model, and
injected systemically; getting the miRNA out of the cir- showed that introduction of the miRNA mimic in vivo
culatory system and crossing the endothelium will be was effective in reducing this fibrosis [64] and confirmed
challenging if over 5 nm in diameter. One recent study reductions in the miRNAs target proteins. But what is
used a neutral lipid emulsion as a vehicle for the sys- more impressive is that introduction of the mimic was
temic delivery of miRNA-34a, which is downregulated effective in prevention of fibrosis with no observable
in cancer, and this lipid vehicle delivery system was cap- side effects [64]. miRagen Therapeutics (Boulder, CO, USA)
able of reducing lung tumour burden in a mouse model of has an miRNA-29 mimic in pre-clinical development for
O’Reilly Arthritis Research & Therapy (2016) 18:11 Page 8 of 10
fibrotic conditions and uses a non-viral vector for systemic elevated significantly in fibrosis and this correlates with
delivery. miRagen Therapeutics should move into clinical serum alanine aminotransferase (ALT) levels, a marker of
trials soon with this molecule. (Table 2 illustrates miRNA damaged liver [67] and may be a more reliable marker
therapeutics in development.) than ALT in assessing fibrosis [67] and has been found in
Although miR therapeutics is a promising area, chal- other independent cohorts [68]. It has been suggested that
lenges do remain. These include the effective delivery of serum miRNA-122 is elevated in hepatocyte injury regard-
the mimic or antagomirs targeted to the correct tissue less of the aetiology [69] and this may prevent it being
and cell type. Strategies to do so include viral vector de- used specifically for liver fibrosis. miRNA-214 has also
livery, liposomes and nanoparticle delivery. Of course, been found extracellularly and to actually mediate signals
some tissues such as the skin and lung are more access- through downregulation of its target gene CTGF via extra-
ible than others. The use of ligands that are attached to cellular exosomes to target cells mediating fibrosis [70].
the nucleotides that are specific for certain cell surface This has also been shown in in vitro models and in vivo
receptors that bind and then internalise may be one way [71]. It now appears that exosome mediates the transfer of
of enhancing specific cell uptake. Here, they can release miRNAs to distal cells to trigger intracellular signalling
their ‘cargo’. A major concern is also ‘off target effects’ and that exosomes display cell surface markers that the
of the miR therapeutics. A principle benefit of targeting target can recognise and internalise through receptors,
miRNAs is that they often target mRNAs in the same thereby giving rise to direct mRNA repression. The im-
pathway and so fine-tune the output of a particular path- portant fibrotic miRNA-29a has also been found in the
way; however, this may be a wanted effect in one specific serum to be associated with cardiac fibrosis in cardiac
cell type but not in another cell type. A further consider- hypertrophy [72]. In SSc, a few circulating miRNAs have
ation should be drug resistance, as in cancer treatment been demonstrated to be higher in serum compared with
drug resistance can develop; this itself can be mediated controls, including miRNA-142-3p [73]. Furthermore,
through miRs, altering gene expression of mRNAs critical miRNA-196a has been demonstrated to be reduced in
in driving this effect. It could be speculated that, in miR SSc serum compared with healthy controls [74]. There
therapeutics, antagomir therapy may reduce the miR and was also a correlation between lower serum miRNA-196a
thus increase its targets, but mechanisms to counter this levels and the modified Rodnan skin score, which is a
through upregulation of the miR being targeted by in- measurement of skin thickness due to fibrosis [74]. In-
creased biogenesis may lead to diminishing effectiveness. deed, levels of miRNA-196a have also been found to be
Finally, the TLR system can also respond to RNA and in- differentially expressed in the hair of patients with SSc
duce an antiviral response through downstream adaptor [75]. Although miRNAs in serum have been demonstrated
proteins and this could be theoretically activated by miRs. in association with fibrotic organ-specific diseases or in
SSc, their clinical utility remains to be determined, and
Serum miRNAs as diagnostic/prognostic markers larger studies are needed to validate these reports. Stratifi-
MiRNAs appear remarkably stable in serum and other cation of patients on the basis of specific miRNAs may
bodily fluids such as urine and saliva [65]; this is because allow a more targeted therapeutic approach. Furthermore,
they are enclosed in extracellular membrane-bound vesi- in the case of serum miR, there is no consensus yet on the
cles or combined with high-density lipoproteins. Thus, ‘normalisation’ method used for these studies. Tissue-
they are attractive as a non-invasive diagnostic biomarker based markers of miR expression are of limited value as
of disease. There are associations with various cytokines whole tissue comprises multiple cell types and miRs are
in fibrotic diseases; however, none of these is sufficiently cell-type dependent. The additional effort to isolate single
robust to be diagnostic or guide treatment. The hepatic- cells yields much more informative results.
specific miRNA-122, which is the target of miravirsen, has
been shown to be reduced in hepatic fibrosis in cells [66]; Conclusions
however, the circulating serum levels of miRNA-122 are Since their discovery two decades ago, miRNAs have
been found to be associated with various diseases. There
Table 2 MicroRNA therapeutics in fibrosis is currently good evidence for the role of miRNAs in fi-
Company MicroRNA target Stage brotic diseases, either organ-specific or systemic fibrosis,
Regulus Therapeutics 21 (RG-012)/Antagomir Phase 1 clinical trial such as SSc. Important steps have been made in recent
Alport syndrome
years, including the identification of dysregulated miRNAs
miRagen Therapeutics 29 mimic Pre-clinical and their targets. Whereas the exact targets of these
development
miRNAs are unknown for some, they are known for
miRagen Therapeutics 155a mimic Pre-clinical others, and they are regulating key downstream pathways
Marina Biotech 21 Antagomir Pre-clinical in disease pathogenesis such as miRNA-29, which is a key
a
Although developed for immune modulation could be useful in fibrosis mediator of fibrosis. Gene therapy with the restoration of
O’Reilly Arthritis Research & Therapy (2016) 18:11 Page 9 of 10
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