目的:类风湿性关节炎(RA)是一种以滑膜增生和进行性骨破坏为特征的自身免疫性疾病。成纤维细胞样滑膜细胞(FLSs)的肿瘤样生长在RA的发病机制中起着至关重要的作用。N6甲基腺嘌呤(m6A)mRNA甲基化修饰,由甲基转移酶(METTL3)和去甲基化酶调节,是RA发展过程中的一种新型表观遗传调节因子。然而,关于RA滑膜炎中m6A甲基化修饰的研究有限,缺乏对RA-FLSs功能影响的机制研究。
方法:本研究使用临床滑膜组织标本和FLS作为研究对象。检测m6A甲基化水平以及甲基转移酶和去甲基化酶的表达。采用RNA干扰和基因过表达方法研究了METTL3在RA-FLSs中的作用机制。该研究还检查了增殖,凋亡,迁移,入侵,和RA-FLSs的细胞因子水平,以及METTL3在RA动物模型中的表达。
结果:在这项研究中,我们发现RA患者的滑膜组织和FLS中m6A甲基化水平升高.免疫组织化学染色显示RA滑膜组织中METTL3和METTL14水平上调,METTL3、METTL14、WTAP、FTO,RA患者滑膜组织和FLSs中ALKBH5明显增高。过表达METTL3可以促进细胞增殖,迁移,IL-6、RANKL的分泌;抑制METTL3的表达可以抑制RA-FLSs的异常增殖,迁移,入侵,和分泌IL-6,RANKL,同时促进OPG的凋亡和分泌,从而抑制RA-FLSs肿瘤样生长。InCIA小鼠,使用MTX和STM2457降低了METTL3的表达,滑膜增生和骨质破坏。
结论:RA患者的滑膜组织和FLS中存在m6A甲基化的异常修饰,抑制METTL3可以减少滑膜炎和骨破坏。我们的研究结果表明,m6A甲基化可能控制FLS介导的肿瘤样表型,并成为RA治疗的新靶点。
OBJECTIVE: Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovial hyperplasia and progressive bone destruction. The tumor-like growth of fibroblast-like synoviocytes (FLSs) plays a crucial role in the pathogenesis of RA. The N6 methyladenine (m6A) mRNA methylation modification, regulated by methyltransferases (METTL3) and demethylation enzymes, is a novel epigenetic regulator in the development of RA. However, there is limited research on m6A methylation modifications in RA synovitis and a lack of mechanistic studies on their impact on the function of RA-FLSs.
METHODS: This study utilized clinical synovial tissue specimens and FLSs as research subjects. The m6A methylation level and the expression of methyltransferases and demethylation enzymes were detected. RNA interference and gene overexpression methods were employed to investigate the mechanism of METTL3 in RA-FLSs. The study also examined the proliferation, apoptosis, migration, invasion, and cytokine levels of RA-FLSs, as well as the expression of METTL3 in RA animal models.
RESULTS: In this study, we found that m6A methylation levels were elevated in synovial tissues and FLSs of RA patients. Immunohistochemical staining showed that METTL3 and METTL14 levels were up-regulated in synovial tissues of RA, the mRNA levels of METTL3, METTL14, WTAP, FTO, and ALKBH5 were significantly higher in synovial tissues and FLSs of RA patients. Overexpression of METTL3 could promote the proliferation, migration, and secretion of IL-6, RANKL of RA-FLSs; inhibition of METTL3 expression could inhibit the abnormal proliferation, migration, invasion, and secretion of IL-6, RANKL, at the same time promoted the apoptosis and secretion of OPG, thus inhibited RA-FLSs tumor-like growth. In CIA mice, the use of MTX and STM2457 reduced METTL3 expression, synovial hyperplasia and bone destruction.
CONCLUSIONS: Abnormal modification of m6A methylation exists in synovial tissues and FLSs of RA patients, and inhibition of METTL3 can reduce synovitis and bone destruction. Our findings suggest that m6A methylation might control FLS-mediated tumor-like phenotype, and be a novel target for RA treatment.