The Mechanism Of MicroRNA Involved In The Protective Effect Of Postconditioning Following Cerebral Ischemia/reperfusion

Objective:To explore the regulatory pathway involving microRNA with the protective effect of Postconditioning following cerebral ischemia/reperfusion (I/R); then further study in vivo and in vitro models in order to determine the exact mechanism of miRNA in the ischemia postconditioning (I-PostC) and cerebral ischemia/reperfusion.Methods:In mice with cerebral ischemia/reperfusion injury model, miRNAprofile on the brain tissue of cortex and hippocampus were determined from isolated RNA using miRNA microarrays, followed by validation with quantitative RT-PCR (qRT-PCR). In vivo, the mouse were treated by intracerebroventricular injection of miRNA-124 agomir and miRNA-124 antagomir, cerebral infarction volume, apoptosis, PI3K/Akt signaling pathway and the expression of apoptosis related proteins such as BcI-2, Bax, caspase 3 were detected using TTC, TUNEL, NeuN staining, Western blot and qRT-PCR. In vitro, PC 12 cells induced by oxygen glucose deprivation/reperfusion model, and prestained cells with miRNA-124 agomir and miRNA-124 antagomir. Apoptosis, PI3K/Akt2 signaling pathway and the expression of apoptosis related proteins such as Bcl-2, Bax, caspase 3 were detected using Dihydroethidium, Cell Counting Kit-8, Annexin V-FITC/PI Apoptosis Detecting Kit, Immunofluorescence staining, Western blot and qRT-PCR.Results:The data showed that miRNA-1, let-7, miR19a and miRNA-124 were differently modulated between I/R and IPostC treated groups in cerebral cortex and hippocampus of mice (P<0.05). The results were then evaluated by RT-qPCR. In vivo, downregulated miRNA-124 expresssion level enhancing the neuroprotection induced by I-PostC, further reduced the volume of cerebral infarction and cell apoptosis, promoted cell survival and enhanced the PI3K/Akt signal pathway and expression, the expression of the anti apoptotic protein Bcl-2 up regulation and apoptosis related protein Bax, caspase 3 expression decreased (P<0.05), overexpressed miRNA-124 otherwise weakened the neuroprotection induced by I-PostC. The above results were confirmed in vitro experiments.Conclusions:The results showed that miRNA expressional alteration in cortex and hippocampus of mice following I/R was associated with the neuroprotection induced by I-PostC, miRNA-1, let-7, miR19a and miRNA-124 either alone or in combination with other miRNAs, was associated with this recovery process. miRNA-124 is involved in the regulation of the pathophysiological process of cerebral ischemia reperfusion injury and the anti-apoptotic effect of I-PostC by targeting PI3K/Akt2 pathway.