The Investigation Of Autophagy’s Role In Human Umbilical Vein Endothelial Cells Senescence Induced By Homocysteine

Objective: To investigate the role of autophagy in the senescence of HUVECs induced by homocysteine.Method: The HUVECs was isolated from newborn’s umbilical cord,and was sub-cultured in vitro.The experiment was divided into three parts: 1)control,HCY(25uM,100 uM,500uM),5g/L D-galactose,interfered 24h;2)control,25 uM HCY treated 24 h,36h,48h;3)control,500 uM HCY,500 uM HCY+100uM 3-MA,500 uM HCY+5.5nM rapamycin.The CCK-8 kit,PI kit and flow cytometry,ROS kit,SA-β-Gal staining kit,StubRFP-Sens-GFP-LC3 autophagy related lentivirus were used to test cell proliferation rate,cell cycle suppression rate,intracellular ROS,SA-β-Gal,autophagy flux respectively.Result: in vitro,HCY inhibits the proliferation rate and cell cycle of HUVECs,increases the intracellular ROS,the number of SA-β-Gal staining cells and autophagy dots in doseand time-dependent way(P<0.05).The D-galactose results the same(P<0.05).The 3-MA inhibited autophagy and rapamycin induced autophagy in HUVECs treated with HCY(P<0.05).The inhibition of proliferation rate and cell cycle,increase of intracellular ROS,SA-β-Gal staining cells in HCY treated HUVECs was aggravated by 3-MA(P<0.05),and was ameliorated by rapamycin(P<0.05).Conclusion: In this study,HCY and D-galactose accelerates HUVECs senescence.HCY and D-galactose induced autophagy,and may through oxidative stress mechanism.Properly adjusted autophagy is vital to delay HUVECs senescence,and the clearly identification of conditions which autophagy could play its protection role and the precisely test of autophagy degree are the prerequisites.