葛根素对脑梗死大鼠p-AMPK、Egr-1水平的影响

Effects of puerarin on the levels of p-AMPK and Egr-1 in rats with cerebral infarction

目的研究葛根素对脑梗死大鼠梗死周围脑组织磷酸化腺苷酸活化蛋白激酶(p-AMPK)、腺苷酸活化蛋白激酶(AMPK)、早期生长反应因子-1(Egr-1)蛋白水平的影响,探讨葛根素保护作用的分子机制。方法42只健康雄性SD大鼠随机分为假手术组、对照组和葛根素组,每组14只。线栓法建立右侧永久性大脑中动脉闭塞(MCAO)模型。假手术组仅分离劲动脉。葛根素组于MCAO术后立即予葛根素(100 mg/kg,溶于10%乙醇)腹腔注射,对照组予等体积溶剂(10%乙醇)腹腔注射。术后24 h,采用改良的Longa评分法来评价神经功能;TTC染色显示脑梗死体积;采用western blot来观察3组p-AMPK、AMPK、Egr-1蛋白水平的变化。结果与对照组比较,葛根素组大鼠神经功能明显改善,脑梗死体积显著减小(P<0.05)。与假手术组比较,对照组大鼠脑组织Egr-1蛋白表达水平显著升高,差异有统计学意义(P<0.05);而与对照组比较,葛根素组p-AMPK水平以及p-AMPK/AMPK比值明显上调,Egr-1蛋白水平有显著下调,差异均有统计学意义(P<0.05)。3组大鼠AMPK蛋白水平差异无统计学意义(P>0.05)。结论葛根素通过下调过高表达的Egr-1水平、上调p-AMPK蛋白表达和p-AMPK/AMPK比值,抑制过度的炎性反应,且能够显著改善了MCAO术后神经功能,减小了脑梗死体积。通过激活AMPK抑制炎性反应,是葛根素对大鼠脑梗死脑保护作用的分子机制。

Objective To investigate the effects of puerarin on the protein levels of phosphorylated adenylate-activated protein kinase(p-AMPK),adenylate-activated protein kinase(AMPK)and early growth response factor-1(Egr-1)in brain tissue around the infarcted area of rats with cerebral infarction,and to explore the molecular mechanism of the protective effect of puerarin.Methods Forty-two healthy male SD rats were randomly divided into sham operation group,control group and puerarin group,with 14 rats in each group.The suture method was used to establish a model of permanent middle cerebral artery occlusion(MCAO)on the right side.In the sham operation group,only arteries were separated.Puerarin group was intraperitoneally injected with puerarin(100mg/kg,dissolved in 10%ethanol)immediately after MCAO,and the control group was intraperitoneally injected with equal volume of solvent(10%ethanol).At 24h after operation,the improved Longa scoring method was used to evaluate neurological function,and TTC staining was used to reveal cerebral infarction volume.Western blot was used to observe the changes in the levels of p-AMPK,AMPK and Egr-1 proteins in the three groups.Results As compared with that in control group,the nerve function of the rats in puerarin group was significantly improved,and the cerebral infarction volume was significantly reduced(P<0.05).As compared with those in sham operation group,the expression levels of Egr-1 protein in brain tissue in control group were significantly increased(P<0.05).As compared with those in control group,the p-AMPK levels and the ratio of p-AMPK/AMPK puerarin group were significantly up-regulated,while the protein levels of Egr-1 were significantly down-regulated(P<0.05).There were no significant differences in AMPK protein levels among the three groups(P>0.05).Conclusion Puerarin inhibits excessive inflammatory response by down-regulating the over-expressed Egr-1 level,and up-regulating the expression of p-AMPK protein and the ratio of p-AMPK/AMPK.In addition,it can significantly improve the neurological function after MCAO and reduce cerebral infarction volume.The inhibition of inflammatory response by activating AMPK is the molecular mechanism of the protective effect of puerarin on the brain in rats with cerebral infarction.