黄芩苷对缺氧/复氧小胶质细胞TLR4通路的影响

Effect of Baicalin on TLR4 pathway in cultured rat's microglia with hypoxia/reoxygenation

目的:模拟脑梗塞时脑组织内的缺氧/再灌注病理过程,探索缺氧/复氧对原代培养的小胶质细胞Toll样受体4(TLR4)通路的影响和黄芩苷的干预作用;方法:原代培养小胶质细胞,采用缺氧/复氧的方式制备模型,用黄芩苷25μmol/L、50μmol/L和100μmol/L3个浓度梯度进行干预,用逆转录聚合酶链反应检测小胶质细胞TLR4 mRNA的表达,Western Blot检测TLR4、抑制因子(I-кB)、p38、ERK蛋白,免疫荧光双染观察髓性分化因子88(MyD88)和核因子кB(NF-кB)。结果:缺氧/复氧激活了TLR4通路,并通过MyD88依赖途径激活了TLR4和下游蛋白MyD88、NF-кB、ERK、I-кB和p38;黄芩苷高剂量组明显抑制了通路中TLR4、MyD88、NF-кB、ERK与p38蛋白的活性和I-кB蛋白的降解。结论:缺氧/复氧激活了小胶质细胞的TLR4通路,黄芩苷通过对TLR4通路蛋白的抑制降低了小胶质细胞的活性,发挥抗炎效应,促进脑缺血的恢复。

Objective： To simulate ischemic-reperfusion in cerebral congestion,and explore the effect of hypoxia/reoxygenation and intervention of baicalin on TLR4 pathway in primary cultured microglia;Methods： To culture primary microglia,and prepare the model of hypoxia/ reoxygenation,and intervene with baicalin 25mmol/L,50mmol/L and 100mmol/L.Then we detected the expression of TLR4 mRNA in microglia with the reverse transcription polymerase chain reaction,and detected the TLR4,I-кB,p38,ERK protein with Western Blot,observed MyD88 and NF-кB with immunofluorescence double staining;Results： The hypoxia/ reoxygenation activated the TLR4 pathway,and activated TLR4,MyD88,NF-кB,ERK,p38 and I-кB protein on the TLR4 pathway through the MyD88-dependent signaling,and the high dose group inhibited the activation of TLR4,MyD88,NF-кB,ERK,p38 protein and blocked the degradation of I-кB protein.Conclusion： The hypoxia/reoxygenation activated the protein on the TLR4 pathway.Baicalin played anti-inflammatory through inhibiting the TLR4 pathway,and promoted the recovery of cerebral congestion.