Table_3_MicroRNA Expression Differences in Blood-Derived CD19+ B Cells of Methotrexate Treated Rheumatoid Arthritis Patients.xlsx

Rheumatoid arthritis (RA) is a complex disease with a wide range of underlying susceptibility factors. Recently, dysregulation of microRNAs (miRNAs) in RA have been reported in several immune cell types from blood. However, B cells have not been studied in detail yet. Given the autoimmune nature of RA with the presence of autoantibodies, CD19+ B cells are a key cell type in RA pathogenesis and alterations in CD19+ B cell subpopulations have been observed in patient blood. Therefore, we aimed to reveal the global miRNA repertoire and to analyze miRNA expression profile differences in homogenous RA patient phenotypes in blood-derived CD19+ B cells. Small RNA sequencing was performed on CD19+ B cells of newly diagnosed untreated RA patients (n=10), successfully methotrexate (MTX) treated RA patients in remission (MTX treated RA patients, n=18) and healthy controls (n=9). The majority of miRNAs was detected across all phenotypes. However, significant expression differences between MTX treated RA patients and controls were observed for 27 miRNAs, while no significant differences were seen between the newly diagnosed patients and controls. Several of the differentially expressed miRNAs were previously found to be dysregulated in RA including miR-223-3p, miR-486-3p and miR-23a-3p. MiRNA target enrichment analysis, using the differentially expressed miRNAs and miRNA-target interactions from miRTarBase as input, revealed enriched target genes known to play important roles in B cell activation, differentiation and B cell receptor signaling, such as STAT3, PRDM1 and PTEN. Interestingly, many of those genes showed a high degree of correlated expression in CD19+ B cells in contrast to other immune cell types. Our results suggest important regulatory functions of miRNAs in blood-derived CD19+ B cells of MTX treated RA patients and motivate for future studies investigating the interactive mechanisms between miRNA and gene targets, as well as the possible predictive power of miRNAs for RA treatment response.

类风湿关节炎（Rheumatoid arthritis, RA）是一种复杂疾病，其潜在易感因素范围广泛。近年来，多项研究已在血液来源的多种免疫细胞中报道了RA患者体内微小RNA（microRNAs, miRNAs）的表达失调，但尚未对B细胞展开详细研究。鉴于RA具有自身免疫特性且存在自身抗体，CD19+ B细胞是RA发病机制中的关键细胞类型，且已有研究在患者血液中观察到CD19+ B细胞亚群的异常改变。因此，本研究旨在揭示血液来源CD19+ B细胞的全局miRNA表达谱，并分析同质RA患者表型间的miRNA表达谱差异。

本研究对三组样本的CD19+ B细胞进行了小RNA测序：初诊未治疗的RA患者（n=10）、经甲氨蝶呤（methotrexate, MTX）治疗后达到缓解的RA患者（n=18）以及健康对照（n=9）。多数miRNA在所有表型样本中均可被检测到。然而，与健康对照相比，经MTX治疗的RA患者中有27个miRNA存在显著表达差异，而初诊患者与健康对照之间未观察到显著表达差异。其中多个差异表达miRNA此前已被报道在RA中存在失调，包括miR-223-3p、miR-486-3p及miR-23a-3p。

本研究以差异表达miRNAs及miRTarBase数据库中的miRNA-靶基因相互作用作为输入数据进行miRNA靶基因富集分析，结果显示富集得到的靶基因多在B细胞活化、分化及B细胞受体信号通路中发挥重要作用，例如STAT3、PRDM1与PTEN。值得注意的是，与其他免疫细胞类型相比，这些基因在CD19+ B细胞中呈现出高度的共表达特征。

本研究结果表明，miRNAs在经MTX治疗的RA患者血液来源CD19+ B细胞中发挥重要的调控功能，这为后续研究miRNA与靶基因间的相互作用机制，以及miRNA作为RA治疗应答预测标志物的潜在价值提供了依据。