标题：METTL3-m6A途径抑制肺动脉平滑肌细胞增殖的实验研究

      作者：陈悦，唐竞桐，罗仕蓉    遵义医药高等专科学校，贵州 遵义 563006

      卷次： 2021年32卷8期

      【摘要】 目的 探索甲基转移样酶3 (METTL3)依赖的6-甲基腺嘌呤(m6A)修饰在体外调控肺动脉平滑肌细胞增殖中的功能。方法 培养 SD大鼠肺动脉平滑肌细胞(PASMCs)，采用 siRNA敲低METTL1的表达。根据转染siRNA的不同分为 siNC组(对照 siRNA转染组)与 siMETTL1组(METTL3 siRNA转染组)。两组PASMCs分别经过siNC及 siMETTL1转染后加入含20%FBS的DMEM完全培养液诱导细胞发生增殖。通过试剂盒比色法检测两组PASMCs的m6A总体修饰差异、通过CCK-8细胞活性检测及EdU细胞增殖实验检测两组 PASMCs增殖能力改变。结果 与 siNC组比较，siMETTL3组METTL1的基因表达[(1.0±0.1) vs (0.4±0.1)]及蛋白表达[(1.81±0.05) vs(1.35±0.09)]发生显著下调，差异均有统计学意义(P<0.05)；且与siNC组比较，siMETTL3组PASMCs的m6A修饰发生显著下调[(0.10±0.01) vs (0.03±0.01)]、细胞活力显著下降[(0.35±0.01) vs (0.11±0.01)]、EdU+细胞数显著降低[(48.22±2.92) vs(30.29±1.78)]，差异均有统计学意义(P<0.05)。结论 敲低METTL3的表达可下调m6A修饰并抑制PASMCs的增殖功能。
      【关键词】 肺动脉高压；肺动脉平滑肌细胞；6-甲基腺苷；甲基转移样酶3；细胞增殖
      【中图分类号】 R543.2 【文献标识码】 A 【文章编号】 1003—6350（2021）08—0953—04

Role of METTL3-dependent m6A in inhibiting proliferation of pulmonary artery smooth muscle cells.
CHEN Yue,TANG Jing-tong, LUO Shi-rong. Zunyi Medical and Pharmaceutical College, Zunyi 563006, Guizhou, CHINA
【Abstract】 Objective To investigate the role of methyltransferase-like 3 (METTL3) dependent N6-methylade-nine (m6A) modification regulates the proliferation of pulmonary artery smooth muscle cells. Methods SD rat pulmo-nary artery smooth muscle cells (PASMCs) were cultured, and siRNAs were used to knock down METTL1 expression.The PASMCs were divided into siNC group (control siRNA transfected group) and siMETTL1 group (METTL3 siRNAtransfected group) according to the different siRNAs. After transfection, the DMEM cultured medium containing 20%FBS were added to induce PASMCs proliferation. The differences in the global m6A levels of PASMCs in the two groupswere detected by the colorimetric assay kit, and the changes in the proliferation ability of PASMCs in the two groups weredetected by CCK-8 assay and EdU assay. Results Compared with the siNC group, significant down-regulation of MET-TL1 gene expression and protein expression were observed in the siMETTL3 group: (1.0±0.1) vs (0.4±0.1), (1.81±0.05) vs(1.35±0.09). Significant downregulation of global m6A level with (0.10±0.01) vs (0.03±0.01), decreased cell viability with(0.35±0.01) vs (0.11±0.01), and decreased EdU+ cells with (48.22±2.92) vs (30.29±1.78) were observed in the siMETTL3group. Conclusion Knockdown of METTL3 can downregulate m6A and inhibit the proliferation of PASMCs.
      【Key words】 Pulmonary artery hypertension; Pulmonary artery smooth muscle cell (PSAMC); N6-methylade-nine (m6A); Methyltransferase like 3 (METTL3); Cell proliferation