METTL3通过调节MAPK和NF-κB信号通路参与脂多糖诱导小鼠小胶质细胞的细胞炎症反应

Methyltransferase Regulates Lipopolysaccharide-induced Inflammation in BV-2 Cells by Inhibiting the MAPK/NF-κB Signaling Pathway

目的探讨甲基转移酶3(METTL3)在神经炎症中的作用及可能机制。方法将培育的小鼠小胶质(BV-2)细胞随机分为4组:对照组、脂多糖(LPS)组、siCtrl+LPS组和siMETTL3+LPS组。利用CCK-8实验确定LPS的最佳作用浓度和细胞活性测定;q-PCR和Western blot法检测LPS诱导BV-2细胞中METTL3表达;采用酶联免疫吸附法(ELISA)检测BV-2细胞培养液中白介素-6(IL-6)、肿瘤坏死因子α(TNF-α)炎症因子表达水平以及RNA m6A修饰水平。结果与对照组比较,LPS组BV-2细胞中METTL3表达水平和生物活性均升高,差异有统计学意义(P<0.01);与siCtrl+LPS组比较,下调METTL3可显著降低IL-6(P<0.05)和TNF-α(P<0.01)水平,抑制LPS导致的细胞凋亡(P<0.01)。此外,沉默METTL3可显著提高丝裂原活化蛋白激酶(MAPK)以及核因子(NF-κB)的磷酸化水平(均P<0.01)。结论下调METTL3通过抑制MAPK和NF-κB信号通路的激活从而抑制神经炎症的发生。

Aim To elucidate the role of methyltransferase(METTL3)modification in neuroinflammation and its possible mechanisms.Methods The optimal concentration of ipopolysaccharide(LPS)was determined by CCK-8.The expression of METTL3 in BV-2 cell induced by lipopolysaccharide(LPS)was detected by q-PCR and Western blot.The expression of IL-6 and TNF-αinflammatory factors in the supernatant were detected by ELISA.Results Compared with the control group,the expression and bioactivity of METTL3 were increased in LPS-induced BV-2 cells(P<0.01).Down-regulated METTL3 can significantly reduce the production and secretion of inflammatory factors and inhibit the apoptosis induced by LPS.The knockdown of METTL3 could increase conspicuously the phosphorylation levels of mitogen-activated protein kinase(MAPK)and nuclear factor kappa-B(NF-κB).(P<0.01)Conclusion METTL3 catalyzes the formation of N6-methyladenosine(m6 A)and mediates the occurrence of neuroinflammation by activating MAPK/NF-κB signaling pathway.