基于VEGF-B/AMPKα通路研究香青兰总黄酮对阿霉素诱导内皮细胞损伤的保护作用

Study of the protective effect of total flavonoids of Dracocephalum moldavica L.on doxorubicin induced injury in HUVECs based on VEGF-B/AMPKαpathway

目的探究香青兰总黄酮(total flavonoids of Dracocephalum moldavica L.,TFDM)对阿霉素诱导的内皮细胞损伤的保护作用及机制。方法CCK-8法检测细胞活力;显微镜法观察细胞形态;试剂盒法检测LDH、SOD和线粒体膜电位变化;Transwell法检测细胞迁移情况;Western blot法检测内皮功能障碍和VEGF-B/AMPKα通路相关蛋白表达情况。结果与模型组相比,TFDM可明显提高细胞活力,改善阿霉素诱导的人脐静脉内皮细胞(HUVEC)细胞形态学变化,降低LDH漏出量,升高SOD活力,升高线粒体膜电位,促进内皮细胞迁移,抑制内皮细胞损伤。Western blot结果显示,与模型组比较,TFDM升高非受体酪氨酸激酶(Src)和黏着斑激酶(FAK)表达水平,升高舒血管因子eNOS磷酸化水平,降低内皮素-1(ET-1)蛋白表达水平,从而抑制内皮功能障碍。TFDM明显上调VEGF-B、NRP1、VEGFR1蛋白表达水平明显升高、明显升高p-AMPKα/AMPKα比例。结论TFDM抑制阿霉素诱导的内皮细胞损伤的机制可能与激活VEGF-B/AMPKα通路有关。

Aim To investigate the protective effect of TFDM on doxorubicin-induced endothelial cell injury and its mechanism.Methods Cell viability was detected by CCK-8 assay.Cell morphology was observed by microscope.The changes of LDH,SOD and mitochondrial membrane potential were detected by kit method.Cell migration was detected by Transwell assay;Endothelial dysfunction and VEGF-B/AMPKαpathway related protein expression were detected by Western blot.Results Compared with model group,TFDM significantly increased cell viability,improved the morphologic changes of HUVEC induced by DOX,decreased LDH leakage,increased SOD activity,increased mitochondrial membrane potential,promoted endothelial cell migration,and inhibited endothelial cell injury.The results of Western blot showed that compared with control group TFDM increased the expression levels of non-receptor tyrosine kinase(Src)and focal adhesion kinase(FAK),increased the phosphorylation level of eNOS,and decreased the expression level of ET-1 protein,thereby inhibiting endothelial dysfunction.The protein expression levels of VEGF-B,NRP1,VEGFR1 and the ratio of p-AMPKα/AMPKαsignificantly increased in the administration group.Conclusion TFDM may inhibit doxorubicin-induced endothelial cell injury by activating VEGF-B/AMPKαpathway.