The Role Of Clock Gene In Autophagy And Apoptosis Of Vascular Endothelial Cells Induced By Angiotensin ?

Objective:To investigate the role of core clock gene Clock in autophagy and apoptosis of human umbilical vein endothelial cell(HUVEC)induced by angiotensin ?(Ang ?).Methods:In this study,HUVEC was stimulated by Ang ? with different concentrations and at different times.Western blot and Q-PCR were used to detect the expression of autophagy-related proteins and apoptosis-related proteins.SiRNA was transfected into endothelial cells by Liposome 2000 for 6 hours(down-regulation of Clock expression),then replace medium.Endothelial cells were treated with 10-7 mol/L Ang ? for 24 hours.MTT assay was used to detect the cell viability after 24 hours with given-concentrated Ang ?;Q-PCR was used to detect the knockdown efficiency of Clock and the expression of autophagy-related genes Atg5 and Atg7;Western blot and Q-PCR were used to detect the expression of autophagy-related proteins[LC3,Beclin-1(BECN-1),p62]and apoptosis-related proteins[Bax,Caspase-3,Bcl-2].Results:1.Autophagy results:1.1 Ang ?(10-7 mol/L)could increase the expression of LC3-?/LC3-? and BECN 1 in HUVEC but decrease the expression of p62 at 24 hours,which suggest that the given concentration of Ang ? could induce autophagy of HUVEC.1.2 Clock knockdown at both protein and transcription levels could reduce the level of autophagy induced by angiotensin ? in vascular endothelial cells.2.Apoptosis results:2.1 Ang ?(10-7 mol/L)could rise up the expression of Bax and Caspase-3 and decrease the expression of Bcl-2 in HUVEC after 24 hours,and the given concentration of Ang ? could induce apoptosis of HUVEC.2.2 Clock knockdown at both protein and transcription levels could reduce the apoptotic level of vascular endothelial cells induced by angiotensin ?.3.SiRNA transfection into endothelial cells by Liposome 2000 for 6 hours had no effects on the viability of vascular endothelial cellsConclusion:1.Angiotensin ? induces autophagy and apoptosis of vascular endothelial cells.2.Clock knockdown do not affect the viability of vascular endothelial cells.3.Clock knockdown can down-regulate autophagy and apoptosis of vascular endothelial cells induced by angiotensin ?.