Supplementary Material for: Exploring the Associations and Molecular Impacts of miR-146a/rs2910164 and miR-196a2/rs185070757 with Rheumatoid Arthritis in a Pakistani Population

<b><i>Introduction:</i></b> MicroRNAs (miRNAs) are a new class of molecules that participate in post-transcriptional regulation of gene expression and hence have been reported to have a crucial role in the pathogenesis of rheumatoid arthritis (RA). We aimed to investigate the association of miR-146a rs2910164 (G/C) and miR-196a2 rs185070757 (T/G) with RA susceptibility in Pakistani patients and to bioinformatically predict the molecular function of these miRNAs. <b><i>Methods:</i></b> A case-control study on 600 individuals was conducted, including 300 RA patients and 300 matching healthy controls. Genotyping was performed by tetra-primer amplification of refractory mutation system-polymerase chain reaction, and the association between variants and RA was statistically determined using different models. <b><i>Results:</i></b> For the variant rs2910164 (G/C) in miR-146a, no difference in genotype distribution was observed between RA cases and controls, as determined using co-dominant (χ² = 4.33; <i>p</i> = 0.114), homozygous dominant (C/C vs. G/G + C/G) (OR = 0.740 [0.531–1.032]; <i>p</i> = 0.091), homozygous recessive (G/G vs. C/C + G/C) (odds ratio [OR] = 01.432 [0.930–2.206]; <i>p</i> = 0.126), heterozygous (G/C vs. C/C + G/G) (OR = 1.084 [0.786–1.494]; <i>p</i> = 0.682), and additive (OR 0.778 [0.617–0.981]; <i>p</i> = 0.039) models. Similarly, the GT genotype in the rs185070757 (T/G) miR-196a2 variant did not differ between cases and controls with any models (<i>p</i> &gt; 0.05). For the first time, we report no association of miR-146a rs2910164 (G/C) and miR-196a2 rs185070757 (T/G) with RA in a Pakistani population. A subsequent bioinformatic analysis revealed that the CC genotype of miR-146a rs2910164 might have a protective role against RA pathogenesis, with no effect observed with the miR-196a2 rs185070757. <b><i>Conclusion:</i></b> Our findings suggest that these miRNAs might have little-to-no impact on the RA pathogenesis in the Pakistani population.

<b><i>引言：</i></b> 微小RNA（miRNAs）是一类参与基因表达转录后调控的新型分子，既往研究表明其在类风湿关节炎（RA）的发病机制中发挥关键作用。本研究旨在探讨miR-146a rs2910164（G/C）与miR-196a2 rs185070757（T/G）基因多态性与巴基斯坦人群RA易感性的关联，并通过生物信息学手段预测这两种微小RNA的分子功能。<b><i>方法：</i></b> 本研究纳入600名受试者开展病例对照研究，其中包括300例RA患者及300例匹配的健康对照。采用四引物扩增阻滞突变系统-聚合酶链反应（tetra-primer amplification refractory mutation system-polymerase chain reaction）进行基因分型，通过多种统计学模型分析基因变异与RA的关联。<b><i>结果：</i></b> 针对miR-146a的rs2910164（G/C）位点，共显性模型（χ²=4.33；p=0.114）、纯合显性模型（C/C vs. G/G + C/G）（OR=0.740 [0.531~1.032]；p=0.091）、纯合隐性模型（G/G vs. C/C + G/C）（比值比[odds ratio, OR]=1.432 [0.930~2.206]；p=0.126）、杂合模型（G/C vs. C/C + G/G）（OR=1.084 [0.786~1.494]；p=0.682）及加性模型（OR=0.778 [0.617~0.981]；p=0.039）的分析结果均显示，RA患者与健康对照的基因型分布无显著差异。同样，rs185070757（T/G）位点的GT基因型在各模型下的病例组与对照组间均无显著差异（p>0.05）。本研究首次报道，在巴基斯坦人群中，miR-146a rs2910164（G/C）与miR-196a2 rs185070757（T/G）与RA易感性无显著关联。后续生物信息学分析显示，miR-146a rs2910164的CC基因型可能对RA发病具有保护作用，而miR-196a2 rs185070757位点未观察到此类效应。<b><i>结论：</i></b> 本研究结果提示，上述微小RNA可能对巴基斯坦人群的RA发病几乎无影响。