PI3K/AKT通路在电针促进局灶脑缺血再灌注大鼠脑内血管再生中的作用

Role of phosphoinositide 3-kinase/AKT signaling pathway in promoting angiogenesis in rats with focal cerebral ischemia/reperfusion using electrical acupuncture

目的观察电针对PI3K/AKT信号转导通路激活的影响,探讨电针促进脑内缺血区血管再生的可能机制。方法采用线栓法制备SD大鼠局灶脑缺血再灌注模型。取双侧"合谷"穴(LI4)为电针穴位,将80只SD大鼠完全随机分为正常组(n=10)、模型组(n=30)、电针组(n=30)、电针+LY294002组(n=10)。模型组和电针组分别包括再灌注1、3、7d3个时相点,每个时相点10只。采用免疫组化法检测各时间点大脑缺血半影区p-AKT1、AC133蛋白的表达;逆转录聚合酶链法检测AC133mRNA的表达;免疫印迹法定量检测总AKT1、p-AKT1蛋白的表达。结果与正常组比较,模型组和电针组大鼠脑缺血组织p-AKT1蛋白表达增加(P<0.05),AKT1蛋白表达无明显差异(P>0.05),AC133蛋白和mRNA表达增加(P<0.05)。与模型组比较,电针组p-AKT1蛋白表达于再灌注3、7d有明显增高(P<0.05),再灌注3、7dAC133蛋白和mRNA表达增加(P<0.05)。在PI3K/AKT信号转导通路特异性阻断剂LY294002作用下,大鼠p-AKT1蛋白的表达较电针组显著减少(P<0.05),AC133蛋白表达阴性,AC133mRNA表达减少,与正常组比较无明显差异(P>0.05)。结论电针可能加强PI3K/AKT信号转导通路的激活,促进EPCs归巢至缺血组织区,从而促进血管再生。

Objective To study the role of electrical acupuncture （EA） in activating the PI3K/AKT signaling pathway and its mechanism underlying the angiogenesis in rats with focal cerebral ischemia/reperfusion. Methods A focal cerebral ischemia/reperfusion model was established by filament occlusion. Heku point （LI 4） was selected for electrical acupuncture. Eighty SD rats were randomly divided into control group（n=10）, model group（n=30）, EA group （n=30）, and EA＋LY294002 group（n=10）. Rats in model and EA groups accepted reperfusion on days 1, 3 and 7. Expression of p-AKT1 and AC133 was detected by immunohistochemistry （IHC）. Expression of AC133 mRNA was detected by PCR. Expression of p-AKT1 and AKT1 was detected by Western blotting. Results The expression level of p-AKT1, AC133 and AC133 mRNA was higher in model and EA groups than in control group（P0.05）with no significant difference in expression of AKT1 between the 2 groups. The expression level of p-AKT1, AC133 and AC133 mRNA was significantly higher in EA group than in control group in 3 and 7 d after reperfusion（P0.05）. The expression level of p-AKT1, AC133 and AC133 mRNA was significantly lower in EA＋LY294002 group than in EA group （P0.05）, with no difference between EA＋LY294002 group and control group. Conclusion EA can activate the PI3K/AKT signaling pathway, promote EPC homing to the ischemic zone , and thus lead to angiogenesis.