Role Of Anion Exchanger-2(ae2) In High Concentration Glucose-induced Apoptosis Of Endothelial Cells By Ae2 Gene Silencing

Objective: The purpose of this study is to investigate whether transfection of AE2-shRNA to human umbilical vein cells (HUVECs) could down-regulate high glucose–induced apoptosis.Methods: Cultured HUVECs(CRL-2480)were inoculate in a 6-well culture plate uniformly and were randomly divided into four groups,namely, Control group, HG group, Non-silencing shRNA group,and RNAi group,respectively.Control group were only treated with DMEM containing glucose (5.5 mmol/L) for 48h, while other groups were treated with DMEM containing glucose(35.6 mmol/L) for 48h post transfection. Cell viability, expression levels of AE2 mRNA and protein, apoptosis of HUVECs, activity of Caspase-3, intracellular levels of ROS, change of mitochondrial membrane potential and NO level were detected.Results: HUVECs were incubated with media containing 35.6mmol/L of glucose for 48h:the cell viability was significantly decreased, and the expression levels of AE2 mRNA and its protein were upregulated,showing significant difference between HG group and Control group(P<0.01); it was also found that the interleukin 1?–converting enzyme (ICE) / CED-3 family protease(caspase-3)was activated, the levels of ROS and apoptosis were increased ; however, the levels of NO and mitochondrial membrane potential were decreased, which had significant difference compared with Control group(P<0.05). Downregulation of AE2 expression by its gene-silencing may antagonize these changes in all above-mentioned parameters induced by high concentration glucose and there were significant difference between RNAi group and HG group(P<0.05).However, the results from the Non-silencing shRNA group were the similar as those from the High group and there was no significant difference between the two groups (P>0.05). All these results suggest that AE2 gene silencing could raise cell viability, decrease ROS level and enhance mitochondrial membrane potential, reduce caspase-3 activity and suppress the apoptosis of HUVECs induced by high glucose.Conclusion: AE2 can mediate apoptosis of HUVECs induced by high concentration glucose and underlying mechanisms were likely to activate the pathway of ROS/mPTP/Caspase-3; AE2 gene-silencing can protect HUVECs against the injury or/and apoptosis induced by high concentration glucose.