萝卜硫素通过Src/PI3K/Akt通路调控人脐静脉内皮细胞NO的生成机制

Sulforaphane regulated nitric oxide production via Src/PI3K/Akt pathway in human umbilical vein endothelial cells

目的:研究萝卜硫素(SFN)促进人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVEC)生成一氧化氮(NO),参与内皮细胞修复的作用机制。方法:采用MTT法检测SFN对HUVEC细胞存活率的影响;检测SFN对HUVEC NO释放量的影响;采用Western blot法检测SFN对NO释放途径相关蛋白表达量的影响。结果:SFN浓度低于50μmol·L-1时,对HUVEC细胞无明显毒性(P<0.05);SFN可显著增加NO释放量,呈剂量依赖性,eNOS特异性抑制剂L-NAME明显抑制NO生成(P<0.05);SFN呈时间和剂量依赖性显著增加eNOS磷酸化(p-eNOS)水平,促使Akt磷酸化(p-Akt)及PI3K上游调节因子Src激酶的活化。结论:SFN通过Src/PI3K/Akt信号通路增强HUVEC细胞中eNOS的活性,促进NO生成参与内皮细胞修复,为SFN在预防内皮功能障碍、动脉粥样硬化和其他心血管疾病方面的应用提供实验依据。

OBJECTIVE To explore the mechanism of action of sulforaphane(SFN)on repairing the endothelial cell by promoting the production of nitric oxide(NO)in human umbilical vein endothelial cells(HUVEC).METHODS Methyl thiazolyl tetrazolium(MTT)method was employed for examining the effect of SFN on cell viability of HUVEC cells.And the effect of sulforaphane on NO production was also examined.The effects of SFN on NO production and signaling pathway were analyzed by Western blot.RESULTS SFN below50μmol·L-1 showed no significant toxicity in HUVEC cells(P<0.05).And SFN significantly increased NO production in a dose-dependent manner under the suppression of L-NAME,a specific inhibitor of eNOS.SFN markedly promoted the phosphorylation of eNOS(p-eNOS)in a time/dose-dependent fashion.The phosphorylation of Akt was also elevated.Furthermore,SFN activated Src kinase,a further upstream regulator of PI3K.CONCLUSION SFN promotes NO production by boosting the activity of eNOS through Src/PI3K/Akt signaling pathway in HUVEC cells.Thus it provides experimental rationales for SFN preventing endothelial dysfunction,atherosclerosis and other cardiovascular diseases.