A study of neuro-autophagy mediated by the TRPC3 pathway in a neonatal rat model of hypoxic-ischemic brain damage
Objective:To explore neuro-autophagy mediated by the transient receptor potential channel 3(TRPC3)pathway in a neo-natal rat model of hypoxic-ischemic brain damage(HIBD).Methods:Seven-day-old male Sprague-Dawley(SD)mice were randomly divided into four groups,with 20 rats in each group:sham group,HIBD group,HIBD+vector group,and HIBD+TRPC3 group.At 24 hours before the induction of the HIBD models,vector and TRPC3 were injected into the left ventricle of the young mice in the HIBD+vector and HIBD+TRPC3 groups,respectively.The mice in each group were assessed for brain infarct area,neurological score,cerebral edema volume,and neuronal apoptosis and mitophagy.The in vitro mouse hippocampal neuronal cell line(HT22)was divided into the following six groups:control group,OGD/R group,OGD/R+TRPC3 group,OGD/R+NC group,OGD/R+si-Pink1 group,and OGD/R+TRPC3+si-Pink1 group.The oxygen-glucose deprivation/reperfusion(OGD/R)method was used to establish an in vitro cerebral ischemia-reperfusion model in all groups except the control group.Mitochondrial damage was measured based on the mitochondrial membrane potential.Results:Compared with the sham group,the HIBD and HIBD+vector groups had greater infarct area,brain water content,neurological score,and neuronal apoptosis(P＜0.05).The infarct area,brain water content,neurological score,and neuronal apoptosis were significantly reduced in the HIBD+TRPC3 group compared with the HIBD+vector group(P＜0.05).The expression levels of mitophagy-related proteins PTEN-induced putative kinase 1(Pink1),Parkin RBR E3 ubiquitin-protein ligase(Parkin),and light chain 3(LC3)Ⅱ were significantly higher in the HIBD and HIBD+vector groups than in the sham group(P＜0.05).The expression levels of Pink1,Parkin,and LC3 Ⅱ were further increased in the HIBD+TRPC3 group compared with the HIBD+vector group(P＜0.05).In vitro experiments showed that TRPC3 upregulation signifi-cantly increased Pink1 and Parkin expression and the LC3 Ⅱ/LC3 Ⅰ ratio,and reduced sequestosome-1(p62)expression.Silencing Pink1 partially blocked the effect of TRPC3 on mitophagy.TRPC3 upregulation increased the number of healthy mitochondria,reduced the number of damaged mitochondria,and increased HT-22 cell survival rate.However,silencing Pink1 prevented TRPC3 from restor-ing mitochondrial function and proliferative capacity.Conclusion:TRPC3 plays an important protective role in the early stages of HIBD by activating the Pink1/Parkin pathway-mediated mitophagy.
万方数据
维普
