Effect Of Shh Signal On Neurorepair Of Cerebral Ischemic/reperfusion Injury In Rats For Resveratrol Pretreatment

Objective: To examinate the effect of Shh signal on brain repair of cerebral ischemic/reperfusion injury in rats for resveratrol pretreatment.Methods: Middle cerebral artery occlusion/reperfusion(MACO/R) model for 80 male Sprague-Dawley rats in vivo and oxygen-glucose deprivation/reoxygenation(OGD/R) model for neurons in vitro were made. There were sham(sham) in vivo or normal(Nor) in vitro, ischemic control(Con), resveratrol pretreatment(Res), resveratrol+cyclopamine(Res+Cyc) and cyclopamine(Cyc) groups. Each group is 16 rats in vivo study. The rats were treated with 30mg/kg resveratrol or 10mg/kg cyclopamine or ethyl alcohol(intraperitoneally) once a day for 7 days and 30 min before MCAO/R. In vitro study, Neurons were maintained in neuronal culture medium containing 40 μmol/L resveratrol or 5 μmol/L cyclopamine or ethyl alcohol for 24 h before OGD/R. Neurological deficit scores at 24 h,7d and 14 d after MCAO/R were assessed with Longa Score, Bederson Score and modified Neurological Severity Score(mNSS), respectively. Neurogenesis of the SVZ was detected with BrdU immunostaining. The expressions of BrdU+/DCX+,BrdU+/Nestin+,BrdU+/NG2+,BrdU+/MAP2+,BrdU+/CNPas+,Brd U+/TUNEL+,NeuN+/Shh+,GFAP+/Shh+ proteins in penumbra and Gli-1 protein in vitro were investigated with immunofluorescence staining, respectively. The protein expressions of GFAP and SY in penumbra were detected with immunohistochemistry at 14 d after MCAO/R. The levels of Shh、Ptc-1、Smo and Gli1 mRNA in vivo were analysed with RT-PCR and the protein expression of the Gli-1 in vitro were detected by Western blotting analysis.Results: Resveratrol pretreatment significantly improved neurological function recovery, promoted proliferation, differentiation, migration, and survival of neural stem cells, inhibitted astrocyte activation, strengthened synaptophysin expression than those in the control group in vivo after stroke(P <0.05). At the same time, resveratrol strengthened the activation of the Shh signaling pathway(P <0.05). However, inhibition of the Shh signaling pathway with cyclopamine, a Smo inhibitor, completely canceled the above effects of resveratrol(P <0.05).Conclusion: These results suggest that promoted neurorepair of cerebral ischemic injury and improved neurological function by resveratrol may, at least in part, be mediated by the Shh signaling pathway.