低氧预处理人羊膜间充质干细胞来源外泌体在改善血管衰老中的作用研究

Study on the role of human amniotic mesenchymal stem cell⁃derived exosomes cultured under hypoxic condition in delaying vascular aging

目的明确低氧预处理人羊膜间充质干细胞来源的外泌体(human amniotic mesenchymal stem cell⁃derived exosomes,hAMSC⁃exos)是否在改善血管衰老中发挥作用。方法D⁃半乳糖(D⁃galactose,D⁃gal)诱导人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVEC)亚急性衰老,收集低氧预处理hAMSC⁃exos作用于D⁃gal诱导衰老的HUVEC,通过衰老相关β⁃半乳糖苷酶(senescence⁃associatedβ⁃galactosidase,SA⁃β⁃Gal)活性染色,P53、P16和γH2AX蛋白表达水平测定HUVEC的衰老水平变化,通过划痕实验、transwell小室迁移实验和成管实验来检测HUVEC的成血管功能变化。结果D⁃gal诱导能够成功构建HUVEC亚急性衰老模型;SA⁃β⁃Gal检测及P53、P16和γH2AX蛋白表达检测显示低氧预处理hAMSC⁃exos可更好地逆转D⁃gal诱导HUVEC导致的SA⁃β⁃Gal、P53、P16和γH2AX上调;划痕实验、transwell小室迁移实验和成管实验结果显示,低氧预处理hAMSC⁃exos可改善D⁃gal诱导的HUVEC的成血管功能下调。结论本研究初步明确了低氧预处理hAMSC⁃exos能改善HUVEC的衰老表型。

Objective To analyze the role of human amniotic mesenchymal stem cell⁃derived exosomes(hAMSC⁃exos)cultured under hypoxic condition in delaying vascular aging.Methods D⁃galactose(D⁃gal)stimulation was used to induce subacute senescence in human umbilical vein endothelial cells(HUVEC).Hypoxia⁃induced hAMSC⁃exos(Hy hAMSC⁃exos)was collected to incubate with D⁃gal⁃induced HUVEC.Senescence of HUVEC was evaluated by senescence⁃associatedβ⁃galactosidase activity(SA⁃β⁃Gal)staining.Protein expression levels of P53,P16 andγH2AX,angiogenic function of HUVEC were detected by scratch assay,transwell migration assay and tube formation assay.Results Subacute senescence model of HUVEC was successfully constructed via D⁃gal stimulation;SA⁃β⁃Gal and detection of protein expression levels of P53,P16 andγH2AX showed that Hy hAMSC⁃exos could reverse the upregula⁃tion ofβ⁃Gal,P53,P16 andγH2AX caused by D⁃gal⁃induced HUVEC;scratch assay,transwell migration assay and tube formation assay showed that Hy hAMSC⁃exos could alleviate the weakened angiogenic function of HUVEC induced by D⁃gal.Conclusion The study indicates that Hy hAMSC⁃exos could alleviate vascular aging.