| Background:Ischemic stroke,which is caused by an interruption of blood supply,is a leading cause of mortality and morbidity worldwide.Despite the development of new therapeutic drugs and endovascular techniques,the annual worldwide incidence and prevalence of stroke has continued to increase.Cerebral tissue damage that occurs after ischemia significantly threatens the survival and quality of life of stroke patients.At present,there is no specific therapeutic drug for ischemia-reperfusion injury after ischemic stroke,and stem cell transplantation may be the most promising treatment.Previous studies have reported that long non-coding RNA(LncRNA)plays a critical role in regulating the function of Bone marrow mesenchymal stem cells(BMSCs),and it has been reported that the expression of long non-coding RNA small nucleolus RNA host gene 12(LncRNA SNHG12)is significantly abnormal in the brain tissue of rat cerebral ischemia model.However,the effect of IncRNA SNHG12 on BMSCs in injured brain tissue has rarely been reported.This study mainly explored whether BMSCs after silencing SNHG12 has a neuroprotective effect on cerebral ischemia-reperfusion injury by activating PI3K/AKT-mTOR signal pathway and its corresponding mechanism.Methods:Rats brain mcrovascular endothelial cells(BMECs)were treated with oxygen glucose deprivation/reperfusion(OGD/R),and the expression level of SNHG12 at different time points in BMECs was detected by qPCR.OGD/R treated rat brain microvascular endothelial cells(BMECs)were co-cultured with BMSCs or OGD/R pretreated BMSCs.Next,the expression level of SNHG12 after co-cultuer was detected by qPCR.BMECs proliferation was detected by using CCK-8 and EdU assays,and cell apoptosis was determined by using flow cytometry and the Hoechst staining method.Autophagy of BMECs was determined using immunofluorescence and expression of the PI3K/AKT-mTOR pathway proteins was measured by western blot.Moreover,BMECs proliferation,apoptosis,and autophagy were also determined after the BMECs had been co-cultured with shSNHG12-BMSCs.Finally,BMSCs or shSNHG12-BMSCs were transplanted into rat models of middle cerebral artery occlusion(MCAO)to further confirm the findings of cell experiments.Moreover,neurobehavioral assessment was performed on MCAO model rats,TTC staining and HE staining were used to understand the changes in cerebral infarction volume,and Tunel assay was used to detect brain tissue apoptosis.Results:In vitro,BMECs after OGD/R treatment significantly upregulated the expression of SNHG12 in BMECs,inhibited the proliferation of BMECs,promoted the expression of apoptosis protein Caspase-3,promoted the expression of autophagy protein LC3B,inhibited the expression of autophagy protein P62,and inhibited the phosphorylation of PI3K,AKT and mTOR proteins in BMECs.However,the co-cultured of BMSCs and BMECs treated with OGD/R significantly inhibited the up-regulation of SNHG12 expression in BMECs,significantly improved the inhibition of BMECs proliferation,and significantly inhibited the increase of BMECs apoptosis and autophagy,and also reversed the inhibition of PI3K,AKT and mTOR protein phosphorylation in BMECs.It was also confirmed that silencing SNHG12 significantly enhanced the effect of BMSCs.In vivo experiments further confirmed that the transplantation of BMSCs significantly reduced the cerebral infarction area,decreased the neurological deficit score,inhibited the increase of apoptosis and autophagy,and reversed the inhibitory effect of PI3K,AKT and mTOR protein phosphorylation in MCAO rats.It was also confirmed that silencing SNHG12 significantly enhanced the therapeutic effect of BMSCs in MCAO rats.Conclusion:The results of in vivo and in vitro experiments showed that silencing of SNHG12 in BMSCs significantly enhanced the effectiveness of BMSCs in promoting cell proliferation and reducing cell apoptosis and autophagy by activating the PI3K/AKT-mTOR signaling pathway,and alleviated neurological dysfunction after cerebral ischemia reperfusion injury. |
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