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

ABSTRACT: 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.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.Sevoflurane treatment induced apoptosis and markedly increased the LC3-II level and GFP-LC3 puncta number, decreased p62 expression in H4 cells. Activation of autophagy by rapamycin (1 μmol/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 μmol/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.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 neurotoxicity.

SUBMITTER: Zhou YF

PROVIDER: S-EPMC4857550 | biostudies-other | 2016 May

REPOSITORIES: biostudies-other

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Project description:Aims/introductionAs a common complication in elderly patients after surgery/anesthesia, postoperative cognitive dysfunction (POCD) is mainly characterized by memory, attention, motor, and intellectual retardation. Neuroinflammation is one of the most uncontroversial views in POCD. The sevoflurane-induced neurotoxicity has attracted widespread attention in recent years. However, its mechanism has not been determined. This study aimed to observe the effects of sevoflurane on cognitive function and the changes in inflammatory indices and autophagy protein expression in the prefrontal cortex in aged rats.MethodBefore the experiment, D-galactose was diluted with normal saline into a liquid with a concentration of 125 mg/kg and injected subcutaneously into the neck and back of rats for 42 days to establish the aging rat model. Morris water maze experiments were performed, including positioning navigation (5 days) and space exploration (1 day). The POCD model was established by 3.2% sevoflurane inhalation. The rats were treated with or without MCC950, a potent and selective nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) inhibitor, followed by autophagy agonists and autophagy inhibitors. The expression levels of inflammasome-related protein NLRP3 and autophagy-related proteins LC3B and P62 were detected to test the behavior of rats with a water maze.ResultsWe found that sevoflurane exposure affected learning and working memory ability in aged rats. We also observed microglia activation in the prefrontal cortex. NLRP3 protein expression was significantly upregulated after sevoflurane inhalation. NLRP3 inflammasome activation induced increased expression and mRNA expression of cleaved Caspase-1 and inflammatory cytokines IL-1β and IL-18, and increased secretion of peripheral proinflammatory cytokines. The inhibitor MCC950 was used to improve cognitive ability and inflammation in rats and inhibit the secretion of cytokines. In addition, we demonstrated that significant inhibition of autophagy (decreased LC3-II/I and increased P62) was accompanied by increased activation of NLRP3 inflammasomes and more severe neural cell damage. However, autophagy inhibitor rapamycin administration to activate autophagy resulted in the inhibition of NLRP3 inflammasomes, ultimately attenuating neuronal injury.ConclusionsThe activation of autophagy suppressed the formation of NLRP3 inflammasomes. It also alleviated cognitive impairment in aged rats.

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

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