Inhibition of Sevoflurane-induced Apoptosis of HT22 Cells in Mouse Hippocampal Neurons by Targeted Regulation of PTEN via miR-374b-5p
Objective To investigate the protective effect of miR-374b-5p against sevoflurane(Sev)induced cell damage in mouse hippocampal neuron HT22 cells.Methods HT22 cells were divided into 6 groups:control group,Sev group,miR-con+Sev group,miR-374b-5p+Sev group,miR-374b-5p+pcDNA+Sev group,and miR-374b-5p+pcDNA-PTEN+Sev group.RT-qPCR was used to detect the expression lev-els of miR-374b-5p and PTEN mRNA in each group.Western blot was used to detect the expression lev-els of PTEN,Bax,Bcl-2 and Cleaved caspase-3 in each group.Double Luciferase activity experiment was used to verify the targeting effect of miR-374b-5p on PTEN,and CCK8 was used to detect the activity of HT22 cells;Flow cytometry was used to detect HT22 cell apoptosis;DCFH-DA probe was used to detect ROS levels in HT22 cells.Enzyme-linked immunosorbent assay(ELISA)was used to detect malondialdehyde(MDA)and glutathione(GSH)levels in HT22 cells.Results Compared with the control group,miR-374b-5p expression in the Sev treatment group decreased,while the levels of PTEN mRNA and PTEN protein increased(P＜0.05).And miR-374b-5p inhibited PTEN expression by binding to the 3'UTR of PTEN mRNA in HT22 cells.Compared with the control group,both the levels of PTEN mRNA,PTEN,ROS,MDA,Cleaved caspase-3,Bax,and apoptosis rate in HT22 cells of hippocampal neurons in Sev group increased,while levels of miR-374b-5p,Bcl-2,GSH content,and cell survival rate decreased(P＜0.05).Compared with the miR-con+Sev group,the levels of PTEN mRNA,PTEN,ROS,MDA,Cleaved caspase-3,Bax expression and apoptosis were decreased,while levels of miR-374b-5p,GSH content,Bcl-2 expression and cell survival were elevated in HT22 cells in miR-374b-5p+Sev group(P＜0.05).Compared with the miR-374b-5p+pcDNA+Sev group,the miR-374b-5p+pcDNA PTEN+Sev group showed an increase in levels of PTEN mRNA,PTEN,ROS,MDA content,Cleaved caspase-3,Bax,and cell apoptosis rate in HT22 cells of mouse hippocampal neurons,while showed a decrease in levels of GSH content,Bcl-2 expression level,and cell survival rate decreased(P＜0.05).Conclusion Pretreatment with miR-374b-5p may exert a protective effect on neuronal cells by inhibiting apoptosis in Sev-treated HT22 cells through inhibition of PTEN.
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