水蛭素对缺血性脑卒中大鼠微血管新生及HIF-VEGF-Notch通路的影响

Effect of Hirudin on Microangiogenesis and HIF-VEGF-Notch Pathway in Rats with Ischemic Stroke

目的探讨水蛭素对缺血性脑卒中大鼠脑组织微血管新生及缺氧诱导因子(HIF)-血管内皮生长因子(VEGF)-Notch通路的影响。方法将120只SD大鼠随机分为空白组、假手术组、模型组、阳性对照组、水蛭素低剂量组(0.5 mg/kg)和水蛭素高剂量组(1.0 mg/kg),每组20只。采用线栓法制备大鼠局灶性脑缺血模型(MCAO),以尼莫地平为阳性对照药物(1 mg/kg),阳性对照组及水蛭素各剂量组连续腹腔注射7 d,应用Longa法对各组大鼠进行神经功能学评分,干/湿重法测定脑组织含水量,2,3,5-氯化三苯基四氯唑(TTC)染色观察脑组织梗死体积,免疫组化法测定梗死灶周围微血管密度(CD31)表达情况,免疫印迹法(Western Blot)检测脑组织HIF-VEGF-Notch通路相关蛋白的表达。结果与空白组比较,模型组大鼠神经功能评分增高(P<0.01),脑组织含水量及脑梗死体积增大(P<0.01),脑组织中CD31阳性细胞表达及HIF-1α、VEGF、Notch1的蛋白表达增加(P<0.05或P<0.01)。水蛭素低剂量组、水蛭素高剂量组神经功能评分、脑含水量及梗死体积低于模型组(P<0.05或P<0.01),大鼠脑组织CD31阳性细胞表达及HIF-1α、VEGF、Notch1蛋白表达较模型组升高(P<0.05或P<0.01)。结论水蛭素可改善缺血性脑卒中大鼠神经功能,促进脑组织微血管新生,其作用可能与激活HIF-VEGF-Notch信号通路相关。

Objective To investigate the effect of hirudin on microangiogenesis and hypoxia-inducible factor(HIF)-vascular endothelial growth factor(VEGF)-Notch pathway of rats with ischemic stroke.Methods One hundred and twenty SD rats were randomly divided into blank group,sham operation group,model group,positive control group,low-dose hirudin group(0.5 mg/kg),and high-dose hirudin group(1 mg/kg),with 20 rats in each group.The rat model of focal cerebral ischemia model(MCAO)was established by suture method.Nimodipine was used as the positive control drug(1 mg/kg),and the positive control group and the hirudin group were continuously injected intraperitoneally for 7 days.Neurological function scores of the rats in each group were evaluated by Longa method,the water content of brain tissue was detected by dry/wet weight method,the infarct volume of brain tissue was detected by 2,3,5-triphenyltetrazolium chloride(TTC)staining,the expression of microvessel density(CD31)around the infarct was detected by immunohistochemistry,and the expression of HIF-VEGF-Notch pathway related proteins in brain tissue was detected by Western Blot.Results Compared with the blank group,the neurological function scores in the model group were significantly increased(P<0.01),the water content of the brain tissue and the volume of cerebral infarction were significantly increased(P<0.01),Notch1 protein expression was significantly increased(P<0.05 or P<0.01).The neurological function score,brain water content and infarct volume in the low-dose hirudin group and the high-dose hirudin group were significantly lower than those in the model group(P<0.05 or P<0.01).Compared with the model group,the expression was further increased(P<0.05 or P<0.01).Conclusion Hirudin could improve the neurological function and promote microvascular angiogenesis in brain tissue of rats with ischemic stroke,which might be related to the activation of HIF-VEGF-Notch signaling pathway.