基于Ras/Raf-1/ERK信号转导通路调控VEGF的表达探讨活血解毒方对糖尿病大鼠视网膜的影响

Effects of Huoxue Jiedu Recipe on Retinopathy in Diabetic Rats via Down-regulation of VEGF Mediated by Ras / Raf-1 / ERK Signaling Pathway

目的:观察活血解毒方对糖尿病大鼠视网膜血管形态的影响并探讨其可能的作用机制。方法:一次性腹腔注射链脲佐菌素(Streptozotocin,STZ,65 mg/kg)诱导糖尿病大鼠模型,造模后随机分为模型组和活血解毒方组。另设正常对照组。药物干预12周后,应用视网膜消化铺片法观察视网膜血管形态;应用Western blot法检测视网膜VEGF蛋白的表达,应用Real-Time PCR法检测视网膜VEGF、Ras、Raf-1与ERK mRNA的表达。结果:模型组视网膜毛细血管面积密度及内皮细胞/周细胞比例与较正常组升高(P<0.001),VEGF蛋白及VEGF、Ras、ERKmRNA表达升高(P<0.05),Raf-1mRNA表达升高(P>0.05)。与模型组比较,活血解毒方组视网膜毛细血管面积密度和内皮细胞/周细胞比例降低(P<0.01和P<0.001),VEGF蛋白与VEGF、Ras、Raf-1、ERK mRNA表达下降(P<0.05)。结论:活血解毒方能明显抑制视网膜毛细血管和内皮细胞的增生,其机制可能是通过干预Ras/Raf-1/ERK信号转导通路,下调VEGF在视网膜中的表达,抑制血管新生,从而改善DR。

Objective：To observe the effect of Huoxue Jiedu Recipe （ HJR） on the morphology of retinal blood vessels of diabetic rats, and to explore its underlying mechanism .Methods：Streptozotocin（STZ, 65 mg/kg） was intraperitoneally injected to rats to induce diabetes .The diabetic rats were randomly divided into the model group and the HJR group .Other rats were recruited as the normal control group .After 12 weeks intervention ,the retinal vascular morphology was observed by using trypsin digestion .The ex-pression of VEGF protein was examined by Western-blot.The expression of VEGF, Ras, Raf-1, ERK mRNA in retinas of rats was examined by Real-time PCR.Results：Compared with the normal group , capillary density and the ratio of endothelial cells to peri-cytes increased in the model group （ P〈0.001 ） .VEGF protein and VEGF mRNA , Ras mRNA, ERK mRNA expression increased （ P〈0.05 ） , Raf-1 mRNA expression increased （ P〉0.05 ） .Compared with the model group , capillary density decreased in the HJR group （ P〈0.01 ） , and the ratio of endothelial cells to pericytes decreased （ P〈0.001 ） .VEGF protein and VEGF mRNA , Ras mRNA, Raf-1 mRNA, ERK mRNA expression decreased（P〈0.05）.Conclusion：HJR could obviously restrain the prolifera-tion of capillary and endothelial cells .The effect is mediated by down-regulating expression of VEGF which is achieved by inhibi-ting Ras/Raf-1/ERK signaling pathway .