Experimental Study Of PI3K/Akt/mTOR Regulating Autophagy Of Vascular Endothelial Cells In Steroid-induced Avascular Necrosis Of Femoral Head

Objective: Corticosteroids can cause steroid-induced avascular necrosis of the femoral head(SANFH),and its mechanism is complex.Current studies have shown that vascular endothelial cell injury,regulation of differentially expressed genes and osteocyte autophagy are involved in the pathogenesis of SANFH.However,the relationship between vascular endothelial cells and autophagy and the regulatory mechanism are not clear.The purpose of this study was to investigate the role of vascular endothelial cells autophagy in SANFH,and to explore the relationship between autophagy and SANFH by studying PI3K/Akt/mTOR signaling pathway and autophagy related genes.At the same time,the possible repair effect of mTOR-shRNA on vascular endothelial cell injury was studied.Methods: To study the method of culturing rat femoral head vascular endothelial cells in vitro and transfection of mTOR-shRNA adenovirus at the same time.The autophagy formation of each group was observed by transmission electron microscope(TEM)at 4 time points(12,24,48,72h).The mRNA expression of PI3 K,Akt,mTOR,Beclin1 and MAP1LC3 was detected by RT-PCR,the protein expression of the above factors was detected by Western blot and immunofluorescence,and the expression of 6-keto-PGF1?,a marker of vascular endothelial cell injury,was detected by ELISA.Finally,statistical methods were used to analyze whether there was a statistical difference between the experimental group and the control group at each time point.Results:(1)Adrenocortical hormone can inhibit physiological autophagy of vascular endothelial cells,and mTOR-shRNA can enhance autophagy level.(2)Adrenocortical hormone can increase the expression of PI3K/Akt/mTOR signaling pathway related factors,but decrease the expression of Beclin1 and MAP1LC3 in femoral head vascular endothelial cells.(3)Adrenocortical hormone can lead to VEC damage of femoral head,and mTOR-shRNA has a certain repair effect on VEC injury.(4)mTOR-shRNA can block PI3K/Akt/mTOR signaling pathway,enhance physiological autophagy and repair VEC damage.Conclusion:(1)The abnormal autophagy of vascular endothelial cells induced by adrenocortical hormone is related to the pathogenesis of SANFH.(2)In the process of SANFH,with the increase of action time,hormone increased the expression of PI3K/Akt/mTOR signaling pathway related factors and decreased the expression of Beclin1 and MAP1LC3 in femoral head vascular endothelial cells.It inhibits the protective effect of physiological autophagy on vascular endothelial cells,leads to the injury of vascular endothelial cells,and may eventually lead to SANFH.(3)In the process of SANFH,mTOR-shRNA can enhance autophagy,repair hormone damage to vascular endothelial cells by inhibiting mTOR and blocking PI3K/Akt/mTOR signaling pathway,and then repair or delay avascular necrosis of femoral head.