MicroRNAs in systemic rheumatic diseases

MicroRNAs (miRNAs) are endogenous, non-coding, single-stranded RNAs about 21 nucleotides in length. miRNAs have been shown to regulate gene expression and thus influence a wide range of physiological and pathological processes. Moreover, they are detected in a variety of sources, including tissues, serum, and other body fluids, such as saliva. The role of miRNAs is evident in various malignant and nonmalignant diseases, and there is accumulating evidence also for an important role of miRNAs in systemic rheumatic diseases. Abnormal expression of miRNAs has been reported in autoimmune diseases, mainly in systemic lupus erythematosus and rheumatoid arthritis. miRNAs can be aberrantly expressed even in the different stages of disease progression, allowing miRNAs to be important biomarkers, to help understand the pathogenesis of the disease, and to monitor disease activity and effects of treatment. Different groups have demonstrated a link between miRNA expression and disease activity, as in the case of renal flares in lupus patients. Moreover, miRNAs are emerging as potential targets for new therapeutic strategies of autoimmune disorders. Taken together, recent data demonstrate that miRNAs can influence mechanisms involved in the pathogenesis, relapse, and specific organ involvement of autoimmune diseases. The ultimate goal is the identification of a miRNA target or targets that could be manipulated through specific therapies, aiming at activation or inhibition of specific miRNAs responsible for the development of disease.


Introduction
Since their initial discovery in 1993 [1], microRNAs (miRNAs) have been studied extensively due to their role in the regulation of almost every cellular process thus far investigated. miRNAs are non-coding RNAs about 21 nucleotides in length that function as post-transcriptional regulators of gene expression [2]. Th ey can infl uence the activity of about 50% or more of all protein-coding genes in mammals [2], and their change in expression is associated with human diseases, including infectious diseases, cancer, and rheumatic diseases [3][4][5]. Over 800 human miRNAs have been identifi ed so far [2], and they have been shown to negatively regulate protein expression through the inhibition of translation and/or decrease in mRNA stability [6][7][8]. It is now apparent that miRNAs can potentially regulate every aspect of cellular activity, from diff erentiation and proliferation to apoptosis, and they can also modulate a large range of physiological and pathological processes [6].

Biogenesis and function of miRNAs
Th e fi rst step in the biogenesis of mammalian miRNAs is the generation of primary miRNA transcripts (pri-miRNAs) in the nucleus [2]. Th e pri-miRNAs fold into hairpins and act as substrate for Drosha, which is one of the two members of the RNase III family involved in the miRNA maturation process. Th e product of Drosha cleavage, an approximately 70-nucleotide precursor miRNA (pre-miRNA), is exported to the cytoplasm where Dicer, the second RNase III family member, processes the pre-miRNA to a 20-to 23-nucleotide miRNA/miRNA* duplex [2]. Th e preferential loading of one miRNA strand (the guiding strand or mature miRNA) onto the RNA-induced silencing complex (RISC) over the other strand (passenger strand, miRNA*) apparently is based on the thermodynamic stability of the miRNA duplex. Th e mature miRNA with RISC will bind to and silence its target mRNA based on seed sequence complementarity, generally at the 3' UTR. Th e miRNA* may be discarded and eventually degraded [8,9], but recent reports are showing that some miRNA* are stably expressed and they are implicated in important functions as well [9]. An interesting example is the report of Zhou and colleagues [10] demonstrating upregulation of miR-155 and miR-155* in human plasmacytoid dendritic cells to coordinate function in stimulating type I IFN production.
Since the fi rst miRNAs discovered (lin-4 and let-7) were shown to bind the 3' UTR of target mRNAs, it has been widely believed that miRNAs exert their eff ects Abstract MicroRNAs (miRNAs) are endogenous, non-coding, single-stranded RNAs about 21 nucleotides in length. miRNAs have been shown to regulate gene expression and thus infl uence a wide range of physiological and pathological processes. Moreover, they are detected in a variety of sources, including tissues, serum, and other body fl uids, such as saliva. The role of miRNAs is evident in various malignant and nonmalignant diseases, and there is accumulating evidence also for an important role of miRNAs in systemic rheumatic diseases. Abnormal expression of miRNAs has been reported in autoimmune diseases, mainly in systemic lupus erythematosus and rheumatoid arthritis. miRNAs can be aberrantly expressed even in the diff erent stages of disease progression, allowing miRNAs to be important biomarkers, to help understand the pathogenesis of the disease, and to monitor disease activity and eff ects of treatment. Diff erent groups have demonstrated a link between miRNA expression and disease activity, as in the case of renal fl ares in lupus patients. Moreover, miRNAs are emerging as potential targets for new therapeutic strategies of autoimmune disorders. Taken together, recent data demonstrate that miRNAs can infl uence mechanisms involved in the pathogenesis, relapse, and specifi c organ involvement of autoimmune diseases. The ultimate goal is the identifi cation of a miRNA target or targets that could be manipulated through specifi c therapies, aiming at activation or inhibition of specifi c miRNAs responsible for the development of disease. through a perfect or imperfect complementarity with sequences in the 3' UTR only. Th e imperfect complementarity still requires perfect target matching of the second through the seventh nucleotides ('seed sequence') starting from the 5' end of the miRNA [3]. However, it has been recently shown that miRNAs can also bind to the 5' UTR region and to protein coding sequences, albeit causing relatively weak repression [3]. Another very recent study challenged the traditional seed match principle by demonstrating a novel centered pairing between miRNA and mRNA, the 'centered sites' , which consist of a class of miRNA target sites that lack both perfect seed pairing and 3' compensatory pairing and have 11 to 12 contiguous Watson-Crick pairs of miRNA nucleotides 4 to 15 [11]. Th is leads to more versatility in miRNA regulation of specifi c targets and, more importantly, may fail to be predicted by the most common algorithms designed to detect miRNA binding sites in the 3' UTR [11]. Other recent studies have also shown the ability of certain miRNAs in translational activation [12], which suggests that our knowledge of overall biological function for miRNAs remains somewhat incomplete.

Key macromolecules of RISC are targets of human autoantibodies
Th e two best characterized protein families in the RISC complex, the Argonaute family and GW182 (glycinetryptophan dipeptide-rich protein of 182 kDa), are known to play a central role in silencing mRNA translation as well as triggering mRNA degradation. Th ey are essential components of the GW bodies (also known as mammalian processing bodies, or P bodies). Interestingly, both are known autoantigens recognized by autoantibodies in various disease states [13][14][15]. Th e Argonaute family comprises four Argonaute (Ago) proteins (Ago1 to 4) in mammals, and they have all been shown to interact with miRNAs [16,17] and repress protein translation when artifi cially tethered to the 3' UTR of reporter mRNAs [18,19]. However, Ago-mediated repression requires them to interact with another protein, GW182, which is the key silencer downstream of Ago2 [19]. GW182 (also known as TNRC6A) is a 182-kDa protein characterized by multiple glycine (G) and tryptophan (W) motifs and is a very important constituent of GW bodies [20,21]. Th e GW182 family includes three paralogues of TNRC6 (GW182-related) proteins, GW182/ TNGW1, TNRC6B (containing three isoforms), and TNRC6C, in mammals [22,23]. A number of diff erent models have been proposed for the GW182 silencing mechanism in the miRNA pathway, including its interference with translational initiation and 80S complex assembly as well as post-initiation steps, but the detailed molecular process remains to be explored [8,9]. Recent studies also demonstrated that GW182 interacts with Poly-A binding protein (PABP) and further recruits deadenylase complex to promote miRNA-targeted mRNA decay [24,25]. GW182 was originally identifi ed and cloned in 2002 as a novel protein recognized by an autoimmune serum from a patient with motor and sensory neuropathy [15,26]. In 2006, Jakymiw and colleagues [13] showed that the Ago2 protein corresponds to the 100-kDa component of the so-called 'anti-Su antibodies' , and for this reason we now call these antibodies 'anti-Ago2/Su' . Since their identifi cation [27,28], anti-Ago2/Su antibodies have been detected in various diseases, including autoimmune and infectious disease [13,15,[28][29][30]. However, the clinical signifi cance of anti-Ago2/Su antibodies has not been established yet [15,30].

MicroRNAs in rheumatic diseases
As miRNAs emerge to play important roles in many biological processes, they have been referred to as master regulators of gene expression, with a concept where a single miRNA may regulate an entire pathway or even multiple pathways [6]. Regulation of the immune system is vital to prevent many pathogenic disorders and mammals have developed a complex system of checks and balances for immune regulation in order to maintain self-tolerance while allowing immune responses to foreign pathogens [5]. Only in recent years has more evidence emerged to support a central role for miRNAs also in abnormal immune processes and in rheumatic diseases. In fact, the potential of miRNAs as biomarkers in rheumatic diseases is a new and growing area of research [3,5]. Th e identifi cation of candidate miRNAs that target genes implicated in rheumatic disorders and the evaluation of the consequences of mutations in their target sites coupled to phenotypic and gene expression studies should improve our understanding of the mole cular mechanisms responsible for rheumatic diseases [31]. Increased knowledge of miRNAs has led to the development of mouse models for studying in vivo therapeutic approaches using specifi c miRNAs [32]. In particular, Nagata and colleagues [32] have performed the intraarticular injection of double-stranded miR-15a in the synovium of mice with autoantibody-mediated arthritis. Th rough this experiment, they have shown that this miRNA is capable of cell entry and induces cell apoptosis through targeting Bcl-2, which is known normally to suppress apoptotic processes [32].
miR-146a appears to be an interesting example of a master regulator in several aspects of immunity. Specifically, it contributes to controlling the overproduction of cytokines, such as TNF-α, and it functions as a negative feedback control of innate immunity in toll-like receptor (TLR) signaling during recurrent bacterial infection by establishing endotoxin tolerance [33] and cross-tolerance [34]. Lu and colleagues [35] recently demonstrated that miR-146a is critical for the suppressor functions of regulatory T (Treg) cells. In fact, a miR-146a knockout mouse showed some loss of immunological tolerance, responsible for fatal IFNγ-dependent immune-mediated lesions in diff erent organs [35]. Th is is an example of how specifi c cellular aspects can also be controlled by a single miRNA, where the lack of function of miRNAs can be responsible for the onset of autoimmune disease. In another study, Curtale and colleagues [36] showed that miR-146a is involved in T-cell activation and is highly expressed in mature human memory T cells. miR-146a can modulate activation-induced cell death processes, thus acting as an anti-apoptotic factor in T cells, and it is also able to reduce the expression of cytokines, such as IL-2, induced by T-cell receptor engagement in the adaptive immune response [36].
Another miRNA widely studied for its key role in autoimmunity is miR-155. It functions in the hematopoietic compartment to promote the development of infl ammatory T cells, including the T helper (Th )17 and Th 1 cell subset [37]. Divekar and colleagues have investigated the infl uence of miR-155 on Treg cells in a mouse model (MRL/lpr) of systemic lupus erythematosus (SLE) [38]. Th ese investigators have shown an increase in CD4+CD25+Foxp3+ Treg cells that have an altered phenotype and reduced suppressive capacity. Searching for the reason for this alteration, they detected a signifi cant reduction of Dicer expression and the overexpression of some miRNAs in MRL/lpr Treg cells, including miR-155, which is able to target CD62L in Treg cells. Th e results of this study show that elevated miR-155 expression together with a reduced level of Dicer can be responsible for the Treg cell phenotype in MRL/lpr mice [38]. Th is study also introduces a new concept that some miRNAs may be produced in this SLE model independently of Dicer, as described recently in mouse embryonic stem cells [39]. miR-155 also plays an important role in mouse models of collagen-induced arthritis and K/BxN serum transfer arthritis [40]. In fact, miR-155 knockout mice do not develop collagen-induced arthritis. In the K/BxN serum transfer arthritis model, the miR-155 -/mice show a reduction in pathogenic autoreactive B and T cells and cytokine production (IL-6, IL-17 and IL-22) and local bone destruction is reduced because of a decreased generation of osteoclasts [40]. Th ese results support a possible therapeutic role for miRNAs in rheuma toid arthritis (RA).
Beside immune and autoimmune mechanisms, the study of miRNAs as biomarkers is most advanced in oncology [3]. Initial reports showed that cancer cells and tissues have diff erent miRNA profi les from normal cells and tissues, suggesting that they could be used for diagnosis, prognosis and therapeutic outcome [3]. By the regulation of gene expression at the post-transcriptional level, they aff ect various signaling cascades during the progression of neoplastic diseases [41]. Sustained angiogenesis is one of the mechanisms leading to cancer progres sion. Recently, a role of the secreted protein epidermal growth factor-like domain 7 (EGFL7) in the control of vascular tubulogenesis has been suggested. Interestingly, the two biologically active miRNAs miR-126 and its complement miR-126*, which are encoded by intron 7 of the EGFL7 gene, have been shown to mediate vascular functions [41], promoting blood vessel growth and infl am mation. Complex networks of reprogramming of miRNAs have been detected in cancer and leukemia and, given the critical role that miRNAs play in tumorigenesis processes and their disease-specifi c expression, they have the potential to become therapeutic targets and specifi c cancer biomarkers [42,43].
In the present review, we will focus our attention on recent developments in understanding the role of miRNAs in autoimmune rheumatic diseases, such as SLE, RA, systemic sclerosis (SSc; scleroderma), Sjögren's syndrome (SS) and polymyositis/dermatomyositis (PM/DM).

Systemic lupus erythematosus
SLE is a systemic infl ammatory autoimmune disease characterized by the presence of autoantibodies against a large number of self-antigens, including chromatin, ribonucleoproteins, and phospholipids. Clinical manifestations are heterogeneous and include malar rash, photosensitivity, arthritis, glomerulonephritis, and neurological disorders [5,44,45]. Since 2007, diff erent groups have reported altered miRNA expression in tissues and peripheral blood mononuclear cells (PBMCs) from SLE patients [46,47], but these fi rst reports mainly identifi ed groups of miRNAs that were aberrantly expressed through microarray chip analysis, without defi ning poten tial pathways they participate in. Table 1 summarizes studies that are more focused on the identifi cation of specifi c aberrant miRNAs in SLE and other diseases. For example, Tang and colleagues [48] have studied the role of miR-146a, showing that it is down-regulated in SLE. Th ey have evaluated the IFN score through the expression levels of the IFN signature genes OAS1, MX1, and LY6E [48]. Since it is known that miR-146a targets adaptors TRAF6 (TNF receptor-associated factor 6) and IRAK1 (IL-1 receptor-associated kinase 1) in the pathway to NF-κB activation (Figure 1a), they have postulated that the lower expression levels of miR-146a in lupus PBMCs is inversely correlated with the IFN score and may be responsible for IFN overproduction in SLE [48]. Th ey have also demonstrated that low miR-146a and high IFN expression correlate with SLE disease activity, in particular with renal disease [48].
Th e same group studied another miRNA, miR-125a, reporting that the miR-125a level is reduced in PBMCs from SLE patients, and the expression of the predicted target of miR-125a, KLF13, was increased [49]. Th e fi nal result is the signifi cant over-expression of the infl ammatory chemokine RANTES (Regulated upon activation, normal T-cell expressed, and secreted; also called CCL5), which is known to be highly expressed and have detrimental eff ects in infl ammatory processes, including arthritis and nephritis (Figure 1b). Th is study demonstrated that miR-125a negatively regulates RANTES expression by targeting KLF13, as shown by manipulation studies of activated T cells from lupus patients [49]. CDK2, cyclin-dependent kinase 2; DNMT1, DNA methyltransferase 1; IRAK1, IL-1 receptor-associated kinase 1; LPS, lipopolysaccharide; MCP-1, monocyte chemoattractant protein 1; MMP, matrix metalloproteinase; PBMC, peripheral blood mononuclear cell; PDGF, platelet-derived growth factor; RA, rheumatoid arthritis; RANTES, Regulated upon activation, normal T-cell expressed, and secreted; RASGRP1, RAS guanyl-releasing protein 1; SS, Sjögren's syndrome; TGF, transforming growth factor; TRAF6, TNF receptor-associated factor 6.
While miR-146a and miR-125a are down-regulated in SLE patients, other miRNAs can be up-regulated, as is the case for miR-21, miR-148, and miR-126 (Figure 1c) [50,51]. In contrast, miR-21 and miR-148 are overexpressed in PBMCs of SLE patients, and it has been demonstrated that they can target the DNA-methylation pathway, causing DNA hypomethylation and overexpres sion of autoimmune-associated methylationsensitive genes, such as CD70 and LFA-1 (CD11a) [50]. Th e same targets are also infl uenced by another miRNA, miR-126, which is also directed to the EGFL7 gene. In this case, the fi nal result is the DNA hypomethylation and overexpression of autoimmune-associated genes, leading to the autoimmune response in SLE [51].

Rheumatoid arthritis
RA is a systemic autoimmune disorder characterized by chronic infl ammation of synovial tissue that results in irreversible joint damage [52]. Infl ammatory cytokines, especially TNF-α, IL-1β, and IL-6, are known to play an important role in the pathogenesis of RA, as the inhi bition of these cytokines can ameliorate disease symp toms in patients [53]. In recent years, many studies have focused on the identifi cation of altered miRNA expression in RA patients compared to healthy controls or patients aff ected by osteoarthritis [54][55][56]. Some studies mainly considered miRNA expression in plasma and serum, while others mainly focused on tissue analysis (Table 1) [57]. Two of these studies examined miRNA expression in RA synovial tissue and fi broblasts. Stanczyk and colleagues [56] reported an increase of miR-155 and miR-146a expression in both RA synovial fi broblasts (RASFs) and RA synovial tissue compared to osteoarthritis patients. Th ese investigators concluded that the infl ammatory milieu of RA may alter miRNA expression profi les in resident cells of the rheumatoid joints. peripheral blood mononuclear cells (PBMCs) and this may amplify the activation of NF-kB through its direct regulation of NF-kB upstream regulators IRAK1 (IL-1 receptor-associated kinase 1) and TRAF6 (TNF receptor-associated factor 6). Activation of NF-kB leads to increased type I IFN production and thus increased expression of 'IFN signature genes' , including LY6E, OAS1, and MX1 [48]. (B) miR-125a is down-regulated in PBMCs from SLE patients, which leads to elevated expression of its target transcriptional factor KLF13. KLF13 can trigger the expression of the pro-infl ammatory chemokine RANTES (Regulated upon activation, normal T-cell expressed, and secreted), which is known to enhance infl ammatory processes such as arthritis and nephritis [49]. (C) Up-regulation of miR-21, miR-148, and miR-126 can either directly or indirectly inhibit DNA methyltransferase 1 (DNMT1) levels. This inhibition in turn reduces the CpG methylation level and causes over-expression of autoimmune-associated genes in SLE, such as those encoding CD70, LFA-1 (CD11a) and EGFL7 (epidermal growth factor-like domain 7) [50,51]. A n , poly-A tail; CH3, methyl groups; RASGRP1, RAS guanyl-releasing protein 1.

NF-kB
Considering that miR-155 had a repressive eff ect on the expression of two matrix metalloproteinases (MMP-3 and MMP-1) in RASFs, Stanczyk and colleagues [56] hypothesize that miR-155 may be involved in the modulation of joint destructive properties of RASFs, and in the control of the excessive tissue destruction due to infl ammation. Th e same group has recently identifi ed another miRNA, miR-203, highly expressed in RASFs and they demonstrated methylation-dependent regulation of miR-203 expression. Moreover, high expression of miR-203 led to increased secretion of MMP-1 and IL-6 via the NF-kB pathway, contributing to the activated phenotype of synovial fi broblasts in RA [58].
Two other miRNAs have been detected at high levels in RA. In particular, miR-223 is up-regulated in CD4+ naïve T lymphocytes of RA patients, and a possible role of this miRNA in the pathogenesis of the disease has been hypothesized [59]. Alsaleh and colleagues [60] studied the overexpression of miR-346 in RA fi broblast-like synoviocytes, showing that miR-346 indirectly regulates the release of the pro-infl ammatory cytokine IL-18.
Nakasa and colleagues [54] have studied the expression pattern of miR-146a in synovial tissue from patients with RA. Th ey reported increased expression of mature miR-146a and primary miR-146a/b in RA synovial tissue, which also expressed TNF-α [54]. Cells positive for miR-146a are primarily CD68+ macrophages, but also some CD3+ T cell subsets and CD79a+ B cells [54]. Th e expression of miR-146a/b is markedly up-regulated in RASFs after stimulation with TNF-α and IL-1β [54] ( Figure 2a). Our group has implemented a diff erent approach to examine miRNA expression in RA patients compared to healthy controls [55]. Pauley and colleagues [55] have shown increased expression of miR-146a, miR-155, miR-132 and miR-16 in RA PBMCs. In addition, two targets of miR-146a, TRAF6 and IRAK1, are similarly expressed between RA patients and control individuals, despite increased expression of miR-146a in patients with RA. Repression of TRAF6 and/or IRAK1 in THP-1 cells resulted in up to an 86% reduction in TNF-α production, implying that normal miR-146a function is critical for the regulation of TNF-α production. Our data thus demonstrate that miRNA expression in RA PBMCs mimics that of synovial tissue/fi broblasts, and our hypothesis is that miR-146a is upregulated but unable to properly regulate TRAF6/IRAK1, leading to prolonged TNF-α production in RA patients [55]. More recently, Nimoto and colleagues [61] confi rmed the upregu lation of miR-146 a/b in PBMCs of RA patients, which seems to be involved in the overexpression of the pro-infl ammatory cytokine IL-17. Other groups have also demonstrated the overexpression of miR-146a in CD4+ T cells from RA patients, which is closely related to TNF-α expression and to regulation of T-cell apoptosis, thus maintaining the pro-infl ammatory milieu typical of RA patients [62]. A recent report by Nakamachi and colleagues [63] has shown another miRNA, miR-124a, is involved in RA. Th ey have found that miR-124a levels are signifi cantly decreased in RA synoviocytes compared to osteoarthritis synoviocytes. Transfection of precursor miR-124a into RA synoviocytes led to the signifi cant suppression of cell proliferation and arrest of the cell cycle at the G1 phase. Th ey identifi ed a putative consensus site for miR-124a binding in the 3' UTR of cyclindependent kinase 2 (CDK2) and monocyte chemoattractant protein 1 (MCP-1) mRNA. In fact, induction of miR-124a in RA synoviocytes signifi cantly suppressed the production of the CDK2 and MCP-1 proteins [63]. Th us, these investigators show that miR-124a is also a key miRNA in the post-transcriptional regulatory mechanism of RA synoviocytes (Figure 2b).

Other autoimmune diseases
Sjögren's syndrome SS is an autoimmune infl ammatory exocrinopathy aff ecting the lacrimal and salivary glands, leading to dry eyes and mouth [64]. It is often associated with positive anti-SSA/Ro and anti-SSB/La antibodies and with other systemic symptoms, such as arthritis, lymphadenopathy, interstitial pneumonia, and renal disease [64]. Th e role of miRNAs in SS has not been widely explored yet (Table 1). Alevizos and colleagues [65] identifi ed miRNA signatures from the minor salivary glands of patients with SS and normal controls. Th is analysis allowed them to distinguish between these two populations, as well as between subsets of SS patients with low-grade or high-grade infl ammation [65]. Michael and colleagues [66] explored the presence of miRNAs in saliva exosomes isolated from parotid and submandibular glands of patients with SS. Th ey have shown that miRNAs can be identifi ed in saliva, which suggests it may be possible to obtain information from these target organs without the need for invasive methods, such as biopsies. Th e same group also identifi ed the miR-17-92 cluster as responsible for the accumulation of pro-B cells and the marked reduction of pre-B and more mature B cells in patients aff ected by SS [67] ( Table 1). Alevizos and colleagues [68] also reported the impor tance of miRNAs as biomarkers in SS, as they identifi ed a specifi c pattern of miRNA expression in infl amed salivary glands from SS patients with diff erent degrees of infl ammation. Th is opens the possibility to use predicted target pathways of diff erentially expressed miRNAs to identify either infl ammation or exocrine gland dysfunction. Recently, Pauley and colleagues [69] reported the altered expression of miR-146a in PBMCs of SS patients and an established mouse model of SS. In this report, miR-146a was signifi cantly overexpressed in SS patients compared with healthy controls, and functional experiments conducted on THP-1 cells have shown the infl uence of miR-146a on increased phagocytic activity and suppressed infl ammation cytokine production. Th is is another example of how altered miRNAs can infl uence pathogenetic mechanisms in autoimmune diseases such as SS.

Scleroderma
Another autoimmune disease in which miRNAs have not been widely studied is SSc, a multisystemic fi brotic disorder with high morbidity and mortality rates [70]. Th e progressive replacement of normal tissue by collagenrich extracellular matrix leads to impairment and, ultimately, to functional failure of aff ected organs. Fibroblasts are activated by profi brotic cytokines and growth factors, such as IL-4, transforming growth factor (TGF)β, and platelet-derived growth factor (PDGF)-B [71]. Maurer and colleagues [71] identifi ed miR-29 as one key regulator of collagen expression in SSc (Table 1). Th is miRNA is strongly downregulated in SSc fi broblasts and skin sections, and transfection experiments showed a possible direct regulation of collagen by miR-29a. Moreover, TGF-β, PDGF-B, and IL-4 reduce the levels of miR-29a in normal fi broblasts to those seen in SSc fi broblasts, while inhibition of PDGF-B and TGF-β pathways by treatment with imatinib restored the levels of miR-29a in vitro [71].

Polymyositis/dermatomyositis
PM/DM is a T-cell mediated infl ammatory myopathy in which the cellular immune response is a key feature in promoting muscle damage [72][73][74]. As in other systemic autoimmune diseases, a strong association of autoantibodies with distinct clinical phenotypes is found in patients with PM/DM [75]. Th e study of miRNAs in this disease is mainly limited to work by Eisenberg and colleagues [73] showing the possible infl uence of miR-146b, miR-221, miR-155, miR-214, and miR-222 on the NF-kB pathway leading to muscle infl ammation (Table 1). from systemic lupus erythematosus patients, miR-146a is up-regulated in rheumatoid arthritis (RA) synoviocytes and PBMCs. miR-146a is a known regulator of IRAK1 (IL-1 receptor-associated kinase 1) and TRAF6 (TNF receptor-associated factor 6) mRNA and this may be responsible for the altered regulation of IRAK1 and TRAF6, both of which act through the NF-kB pathway to prolong the production of proinfl ammatory cytokines and chemokines, including TNF-α and IL-1β [3,55]. (B) miR-124a is down-regulated in synoviocytes from RA patients. Its target proteins, CDK2 (cyclin-dependent kinase 2) and MCP-1 (monocyte chemoattractant protein 1), are up-regulated and this leads to increased synovial proliferation, angiogenesis and chemotaxis [63].

NF-kB CDK2
Conclusion miRNAs play important roles in fundamental cellular processes, and their dysregulated expression is observed in diff erent pathological conditions, including rheumatic diseases, infl ammation, and tumorigenesis [31]. Th e use of miRNAs or miRNA-mimic oligonucleotides has been tested in diff erent cancer cell lines, in mice, and in nonhuman primates [31]. Th ese previous investigations have shown that miRNA-based gene therapies targeting dysregulated miRNAs have the potential to become therapeutic tools. It will be interesting if these miRNAbased gene therapies will be developed to treat patients with rheumatic diseases, such as RA and SLE, in the future. However, further studies in multiple populations and conducted by independent investigators are needed to validate and elucidate these mechanisms and whether or not miRNAs could serve as useful disease markers or therapeutic targets.