Discussion
RA represents a chronic inflammatory autoimmune disease, which leads to irreversible destruction of the joints. The role of Treg cells in the pathogenesis is not entirely clear, as the decrease in the number of Treg cells in synovium and peripheral blood has been described in the past few years[33-35]. Our FCM data also demonstrate a significant decrease of Treg cells in PBMCs and spleen from CIA mice. The prevailing view in the functions of Treg cells is that they use different pathways to inhibit effector T cell proliferation and immune responses, including immunomodulatory cytokines such as IL-10, TGF-β and IL-35. For instance, IL-10 reporter (IL-10R) depletion results in the selective dysregulation of Th17 immune responses[36]. And several studies reported that mice with IL-10 deficient Treg cells could develop autoimmune diseases, such as spontaneous colitis[37]. Our data have validated the inverse correlation between plasma IL-10 and DAS28 scores in the RA patient cohort, suggesting that Treg-targeted might be an effective and potential strategy for RA treatment.
Previous studies have found that miRNAs play a key regulatory role in the pathogenesis of RA[38-40], and their regulatory role in Th cell differentiation and function has also been confirmed. Interestingly, results from emerging studies of miR-143-3p expression in RA patients appear to be conflicting, which has been reported to show different expression in different individuals with RA[17,27,28]. Hence, We conducted the clinical research and analysis of the correlation between miR-143-3p and Th in RA patients, and explored that miR-143-3p, with negatively correlated with DAS28, was actively associated with IL-10 secreted by Treg cells. The results in CIA mice were consistent with the clinical study. Meanwhile, a recent study reported that miR-143-3p was negatively associated with plasma inflammatory cytokines IL-6 in RA and sepsis[41,42], and IL-6 functions as a critical switch in inhibiting development of TGF-β-induced Treg cells[43]. Based on the above analyses, it is rational to hypothesize that miR-143-3p promoted the differentiation of Treg cells and treated Treg-deficiency diseases such as RA.
To further investigate the potential mechanisms associated with the protective effect of miR-143-3p in RA, we verified Treg cells as the critical node of miR-143-3p in this function. The elevated expression of miR-143-3p was verified to inhibit the mRNA levels of Foxp3 in vitro, which was the master transcription factor of Treg cells, indicating that miR-143-3p is capable to facilitate Treg cells differentiation. To further illuminate the role of miR-143-3p on the differentiation of Treg cells during the RA progress, the overexpression of miR-143-3p was conducted with synthetic miR-143-3p mimics in vivo. Our data corroborated that miR-143-3p mimic treatment resulted in a higher proportion of Treg cells in PBMC and spleen, along with the higher levels of IL-10 in plasma and Foxp3 mRNA in lymph nodes and spleen. These data demonstrate that miR-143-3p could upregulate Foxp3 expression to promote Treg cells differentiation.
Nevertheless, the molecular mechanism of the effect of miR-143-3p on the Treg cells differentiation is still unclear. In order to understand potential mechanism of miR-143-3p in Treg-deficiency of RA, we explored the microRNA database (miRBase at http://www.mirbase.org), which is the high confidence microRNA data set to identify potential target of the miR‐143-3p[44,45]. With the TargetScan and miRDB miRNA target prediction tool, we have identified that Kras, c-maf, Mapk7, Bmp5, Etv6 may be the miR-143-3p targets. It is known that c-maf can affect the differentiation and function of CD4+T cells through its unique regulatory mechanism, especially Treg cells[46]. It has been recent reported that c-maf was highly expressed in intestinal suppressive effector Treg cells (eTreg cells)[47], which were critical enforcers of the immune equilibrium. Moreover, Treg-derived IL-10 production was c-maf dependent[48]. These evidence indicate that the effect of miR-143-3p on Treg cells may be medicated by c-maf, which requires further studies for verification.
In conclusion, our results provide the evidence that the expression of miR-143-3p in PBMCs is negatively correlated with RA disease activity. And miR-143-3p could act as a key regulator of Treg cells differentiation to ameliorate CIA symptoms. These results provide a novel strategy to treat Treg-deficiency autoimmune diseases such as RA.