Peroxiredoxin ? Inhibits Alcohol-induced Apoptosis Of HT22 Cells Via Reaction Of Mitochondrial Membrane Potential By ROS/GSK3? Signaling Pathway

After entering the nerve cells,alcohol metabolism will produce a large number of reactive oxygen species(ROS).Mitochondria are highly sensitive organelle to ROS.Excessive accumulation of ROS will cause mitochondrial damage and lead to neuronal apoptosis.The previous study found that Peroxiredoxin ?(Prx ?)can protect the integrity of mitochondrial function,but its mechanism remains to be further studied.Therefore,in this study,we used the mouse hippocampal neuron cell line(HT22)to establish an acute alcoholism cell model.To explore the protective effect of Prx ? on the mitochondrial damage in HT22 induced by alcohol,and Prx ? mediated ROS/GSK3b signal to inhibit alcohol induced apoptosis of HT22 cells.Constructed mock HT22(control),sh Prx ? HT22(Prx ? knockdown)and wt Prx ? HT22(re-expression of Prx ?)by lentivirus transduction.Treat mock HT22 and sh Prx ? HT22 with different concentrations of alcohol.The MTT assay was used to detect cell viability.Fluorescence microscopy and Flow cytometry were used to detect ROS level,apoptosis level,and mitochondrial membrane potential change.Morphological changes of mitochondria observed by TEM.Western blot was used to detect the expression level of apoptosis proteins and GSK3b/b-catenin proteins.Alcohol treatment and related testing were performed after pretreatment with ROS scavenger NAC and re-expression of Prx ? in sh Prx ? HT22.Alcohol treatment produces the following results.1.The levels of ROS and apoptosis in sh Prx ? HT22 were significantly higher than mock HT22.The expression levels of pro-apoptotic proteins were significantly increased and anti-apoptotic proteins were significantly decreased.2.The mitochondrial membrane potential of sh Prx ? HT22 decreased significantly and the morphology of mitochondria changed significantly.Lots of cytochrome C was released to activated caspase 3 signaling,leading to mitochondria-dependent apoptosis.3.The expression level of p-GSK3b(ser9)decreased significantly,p-b-catenin increase and the b-catenin decreased in sh Prx ? HT22 cells.4.NAC pretreatment and re-expression of Prx ? in sh Prx ? HT22 significantly inhibited alcohol-induced mitochondrial damage and apoptosis in HT22.The results suggested that Prx ? mediates ROS to regulate GSK3b signaling pathway(11)maintain mitochondrial membrane potential stability,protect mitochondria and inhibited alcohol-induced apoptosis of HT22.The results may provide a potential therapeutic target for alcohol-induced mitochondrial damage and apoptosis of neuronal cells,and provide a new theoretical basis for the prevention and treatment of related neurodegenerative diseases.