五味子醇甲通过调控自噬流减轻大鼠脑缺血再灌注损伤

目的:探究五味子醇甲(Sch A)减轻大鼠脑缺血再灌注损伤的作用及其潜在机制。方法:成年雄性SD大鼠随机分为假手术组(sham组)、模型组大脑中动脉闭塞(MCAO)组]及Sch A低、中、高剂量治疗组,每组6只。制备大鼠左侧MCAO模型,90 min后再灌注,治疗组Sch A的剂量分别为40、80和160μg·kg^?1·d^?1。造模7 d后进行神经功能损伤方面的评价(mNSS评分),并用TTC染色检测MCAO大鼠脑梗死体积。通过Western blot检测MCAO大鼠脑缺血半影区神经元自噬相关蛋白P62、LAMP1、cathepsin B和ubiquitin的表达水平,综合分析自噬流情况;使用免疫荧光双标法检测Sch A处理后MCAO大鼠脑缺血半影区神经元自噬水平诱导情况。结果:相比于MCAO组,Sch A处理后大鼠mNSS评分和脑梗死体积显著降低(P<0.05),并有剂量依赖性,以Sch A高剂量组差异显著。Western blot结果显示,与sham组比较,MCAO组大鼠cathepsin B蛋白表达水平下降,P62及ubiquitin的表达水平升高(P<0.05);与MCAO组比较,Sch A处理显著升高cathepsin B表达水平的同时降低了P62及ubiquitin的表达水平,以Sch A高剂量组差异显著(P<0.05);而LAMP1在各组间表达仅有下调或上调趋势,但无显著差异(P>0.05)。免疫荧光结果显示,与MCAO组比较,Sch A处理提高了MCAO大鼠脑缺血半影区LC3-Ⅱ表达阳性的神经元比例,以Sch A高剂量组升高显著(P<0.01)。结论:Sch A可逆转MCAO大鼠脑缺血半影区cathepsin B、P62及ubiquitin表达水平而发挥抗脑缺血再灌注损伤的作用,其机制可能与神经元自噬有关。

Schisandrin A attenuates cerebral ischemia-reperfusion injury in rats by regulating autophagic flux

AIM:To investigate the protective effect of schisandrin A(Sch A)on the brain of rats with cerebral ischemia-reperfusion injury and its potential mechanism.METHODS:Male SD rats were subjected to left middle cerebral artery occlusion(MCAO)for 90 min before reperfusion.The rats were divided into sham operation group(sham group),model group(MCAO group),low-dose Sch A group,medium-dose Sch A group,and high-dose Sch A group(n=6 in each group).The low,medium and high doses of Sch A were 40,80 and 160μg·kg^?1·d^?1,respectively.The modified neurological severity scores(mNSS)were evaluated after 7 d,and the volume of cerebral infarction was detected by TTC staining.The autophagic flux was assessed by Western blot detection of the expression levels of autophagy-associated proteins P62,LAMP1,cathepsin B and ubiquitin in brain ischemic penumbra neurons.Immunofluorescence double labeling was used to detect the induction of autophagy in the neurons of cerebral ischemic penumbra by Sch A.RESULTS:Compared with MCAO group,the mNSS and cerebral infarction volume of the MCAO rats treated with Sch A were decreased in a dose-dependent manner,with significant difference in high-dose Sch A group(P<0.05).Compared with sham group,the protein expression of cathepsin B in MCAO group was decreased,and the protein expression of P62 and ubiquitin was increased(P<0.05).Compared with MCAO group,the protein expression of cathepsin B was increased significantly after Sch A treatment,while the protein expression of P62 and ubiquitin was decreased,also with significant difference in high-dose Sch A group(P<0.05).The difference of LAMP1 level in each group was not statistically significant(P>0.05).Moreover,the immunofluorescence results showed that compared with MCAO group,the proportion of LC3-Ⅱ positive neurons in the cerebral ischemic penumbra of the rats in Sch A groups was increased,and the increase was significant in high-dose Sch A group(P<0.01).CONCLUSION:Sch A may induce neuronal autophagy in the cerebral ischemic penumbra of SD rats and reverse the expression levels of neuronal autophagy-related proteins cathepsin B,P62 and ubiquitin to protect against cerebral ischemia-reperfusion injury.