Autophagy activation prevents sevoflurane-induced neurotoxicity in H4 human neuroglioma cells  

作　　者：You-fa ZHOU Qing-xia WANG Hai-yan ZHOU Gang CHEN 

机构地区：[1]Department of Anesthesiology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou 310020, China [2]Department of Anesthesiology, the Second Hospital of Jiaxing, Jiaxing 314000, China

出　　处：《Acta Pharmacologica Sinica》2016年第5期580-588,共9页中国药理学报（英文版）

基　　金：This study was supported by grants from the National Natu- ral Science Foundation of China （No 81371214） and Zhejiang Provincial Natural Science Foundation of China （No LY12H09005）.

摘　　要：Aim： The inhaled anesthetic sevoflurane may induce cognitive impairment in both animals and humans. Previous study has shown that sevoflurane triggers ER stress and may lead to apoptosis in rat hippocampal neurons. In this study, we examined whether sevoflurane caused autophagy and its contributions to sevoflurane induced neuronal cell injury. Methods： H4 human neuroglioma cells were exposed to 4.1% sevoflurane for 6 h. Cell viability and apoptosis ratio were assessed using a CCK8 kit and flow cytometry, respectively. Autophagosomes in the cells were detected using GFP-LC3 plasmid transfection or transmission electronic microscopy. The expression of LC3B, p62/SQSTM, C/EBP homologous protein （CHOP） and glucose-related protein 78 （GRP78） was assessed with Western blotting. Results： Sevoflurane treatment induced apoptosis and markedly increased the LC3-11 level and GFP-LC3 puncta number, decreased p62 expression in H4 cells. Activation of autophagy by rapamycin （1 pmol/L） significantly reduced sevoflurane-induced apoptosis and increased cell viability, whereas inhibition of autophagy with 3-MA （5 mmol/L） caused the opposite effects. Furthermore, sevoflurane treatment markedly increased the expression of CHOP and GRP78, two hallmark proteins of ER stress. Inhibition of ER stress by 4-phenylbutyrate （500 pmol/L） abrogated sevoflurane-induced autophagy and apoptosis, and improved the viability. Moreover, sevoflurane-stimulated expression of CHOP and GRP78 was inhibited by rapamycin, but further enhanced by 3-MA. Conclusion： Sevoflurane treatment induces ER stress and activates autophagy, which antagonizes sevoflurane-induced apoptosis in H4 human neuroglioma cells. The results suggest that autophagy may be a potential therapeutic target in preventing sevoflurane-induced neu rotoxicity.Aim： The inhaled anesthetic sevoflurane may induce cognitive impairment in both animals and humans. Previous study has shown that sevoflurane triggers ER stress and may lead to apoptosis in rat hippocampal neurons. In this study, we examined whether sevoflurane caused autophagy and its contributions to sevoflurane induced neuronal cell injury. Methods： H4 human neuroglioma cells were exposed to 4.1% sevoflurane for 6 h. Cell viability and apoptosis ratio were assessed using a CCK8 kit and flow cytometry, respectively. Autophagosomes in the cells were detected using GFP-LC3 plasmid transfection or transmission electronic microscopy. The expression of LC3B, p62/SQSTM, C/EBP homologous protein （CHOP） and glucose-related protein 78 （GRP78） was assessed with Western blotting. Results： Sevoflurane treatment induced apoptosis and markedly increased the LC3-11 level and GFP-LC3 puncta number, decreased p62 expression in H4 cells. Activation of autophagy by rapamycin （1 pmol/L） significantly reduced sevoflurane-induced apoptosis and increased cell viability, whereas inhibition of autophagy with 3-MA （5 mmol/L） caused the opposite effects. Furthermore, sevoflurane treatment markedly increased the expression of CHOP and GRP78, two hallmark proteins of ER stress. Inhibition of ER stress by 4-phenylbutyrate （500 pmol/L） abrogated sevoflurane-induced autophagy and apoptosis, and improved the viability. Moreover, sevoflurane-stimulated expression of CHOP and GRP78 was inhibited by rapamycin, but further enhanced by 3-MA. Conclusion： Sevoflurane treatment induces ER stress and activates autophagy, which antagonizes sevoflurane-induced apoptosis in H4 human neuroglioma cells. The results suggest that autophagy may be a potential therapeutic target in preventing sevoflurane-induced neu rotoxicity.

关 键 词：SEVOFLURANE inhalation anesthetics H4 human neuroglioma cells AUTOPHAGY apoptosis ER stress rapamycin 3-MA 4-phenylbutyrate neuroprotection 

分 类 号：Q786[生物学—分子生物学] X503.1[环境科学与工程—环境工程]