L Camphor Target MicroRNA-125-3p On The Mechanism Of Cerebral Ischemia Reperfusion Injury

ObjectiveIt is well known that cerebral ischemia and reperfusion can cause neuronal damage in specific areas of the brain,but this mechanism is not clear.Early experimental study of our group showed that L-camphor can protect cerebral ischemia-reperfusion injury,however,there are few studies on this protective mechanism.Studies have shown that cerebral ischemia and reperfusion may lead to neuronal cell injury,and ischemic injury and necrosis of the material stimulation will lead to autophagy,apoptosis over-activation,making the cell content degradation caused by too much cell death,leading to tissue organs Serious injury.Protecting neuronal processes and inhibiting apoptosis and autophagy overgrowth may serve as a new target for ischemic brain injury and become a new hotspot in basic research and potential clinical application of cerebral ischemia reperfusion.The aim of this study was to investigate the mechanism of L-camphorum on cerebral ischemia-reperfusion injury,and to explore the protective effect of L-camphor in cerebral ischemic injury.In the previous group,it was proved that L-camphor-protected nerve cells could inhibit the apoptosis and inhibit the autophagy after cerebral ischemia-reperfusion injury.The mechanism may be that the GABAA receptor targets P53.It has also been proved that some microRNAs may play an important role in the pathophysiology of cerebral ischemia and reperfusion after regulating the development and function of the central nervous system by regulating their target genes.Methods1.The rat model of cerebral ischemia was made by right carotid artery insertion.The rats were subjected to neurological score after modeling.And to observe whether the role of L-camphor 24h can affect the neurological score of cerebral ischemia-reperfusion animal model.And then take the brain,take the fresh rat cortex,MiroRNA gene chip analysis.The expression of miR-125a was detected by qRT-PCR,and the expression of miR-125a was studied by using miR-125a and so on.2.Construction of specific expression of rno-miR-125a-3p mimics and inhibitors in miR-125a predicted by bioinformatics software,transfection into PC12 cells,observation of related target protein expression by immunofluorescence assay The detection of miR-125a was associated with the expression of protein Cadm2 compared with the normal and model groups.3.Transfection of rno-miR-125a-3p mimics and inhibitors and the corresponding negative control into PC12 cells were performed by Western Blot.The target gene-3’UTR region was rr The double luciferase reporter gene of the binding site of-miR-125a-3p further validates the relationship between Cadm2 and miR-125a.4.Transfection of rno-miR-125a-3p mimics and inhibitors and the corresponding negative control to PC12 cells were detected by Imag J.The ratio of spindle cells to miR-125a-3p and PC12 cells was detected by Imag J.And whether there is an association between average lengths.5.rno-miR-125a-3p mimetic and inhibitor and negative control were transfected into PC12 cells.Serum-free treatment was added to the left camphor.The expression of caspase-3 was detected by Western blot.(LC3,P62)expression.Results1.In this study,based on the animal model of cerebral ischemia and reperfusion,the behavioral score was performed before and after administration.The results showed that L-camphor can reduce the behavioral score of animal model of cerebral ischemia-reperfusion injury.The expression of miR-125a in the model group was significantly higher than that in the control group by gene chip analysis.QRT-PCR results showed that L-camphorine could inhibit the expression of miR-125a in the animal model of cerebral ischemia-reperfusion compared with the model group,and P<0.05 was statistically significant.2.Bioinformatics The potential target for predicting miR-125a may be Cadm2.Compared with the control group and the model group,the expression of Cadm2 in the miR-125a mimetic group decreased,and the expression of Cadm2 in the inhibitor group increased(P<0.05 was statistically significant).3.The expression of Cadm2 in the mock group was decreased compared with that in the normal group and the negative control group.The expression of Cadm2 in the inhibitor group was significantly higher than that in the control group(P<0.05).The results of gene validation were also validated by Western Blot.4.Imag J detected the spindle cell ratio,differentiation rate and average length of PC12 cells.It was found that the indexes of miR-125a mimetic group were inhibited compared with the normal group and the negative control group,but the indexes of each group(P<0.05 was statistically significant).5.rno-miR-125a-3p mimetic can inhibit the expression of Cadm2 and P62 induced by serum starvation and promote the expression of caspase-3 and LC3.The L-camphor and rno-miR-125a-3p inhibitors can promote the induction of serum starvation Cadm2 and P62 expression,inhibition of caspase-3 and LC3 expression,and when the L-camphor and rno-miR-125a-3p inhibitors in combination,the results are more obvious(P<0.05 was statistically significant).ConclusionIn the animal model of ischemia-reperfusion and cell experiments,we found that L-camphor can protect the expression of miR-125a by regulating the expression of miR-125a by interfering with the expression of miR-125a,Protein up-regulation protects neurite outgrowth.L-camphor can regulate the apoptosis and autophagy of cerebral ischemia-reperfusion injury by regulating miR-125a.This study provides theoretical basis for the prevention and treatment of cerebral ischemia in L-camphor.