基于PI3K/AKT/mTOR信号通路研究破淤汤对缺血性糖尿病足溃疡大鼠促血管新生的作用

Approach to effect of Poyu decoction on promoting angiogenesis in rats with ischemic diabetic foot ulcer,based on PI3K/AKT/mTOR signal pathway

目的:基于PI3K/AKT/mTOR信号通路探索破淤汤对缺血性糖尿病足溃疡大鼠促血管新生的作用机制。方法:将造模成功的缺血性糖尿病足溃疡大鼠随机分为非药物组、西洛他唑组、破淤汤低剂量组、破淤汤中剂量组、破淤汤高剂量组,各组给予相应药物或非药物干预后,通过创面肉芽组织HE染色观察组织结构,通过CD31免疫组化染色观察血管新生,通过酶联免疫吸附(ELISA)双抗体夹心法检测血清中VEGFR-2水平,通过Western免疫印迹法检测PI3K/Akt/mTOR信号通路关键蛋白表达情况。结果:HE染色结果显示低剂量破淤汤组与非药物组比较,新生毛细血管明显增多,同样含大量炎细胞;中、高剂量破淤汤组较非药物组改善明显,新生毛细血管多,炎症细胞少,接近于正常对照组。与非药物组比较,西洛他唑组及低、中、高剂量破淤汤组大鼠糖尿病足溃疡肉芽组织微血管数目、P-PI3K、P-AKT、P-mTOR表达水平及大鼠血清VEGFR2表达量均升高(P均﹤0.05)。结论:破淤汤可能通过促进缺血溃疡组织VEGER-2蛋白表达,促进PI3K/AKT/mTOR信号通路关键蛋白的表达来促进缺血溃疡组织的血管新生。

Objective:To approach the effect and mechanism of Poyu decoction on promoting angiogenesis in rats with ischemic diabetic foot ulcer,based on PI3K/AKT/mTOR signaling pathway.Methods:After successful modeling,the rats with ischemic diabetic foot ulcer were randomly divided into non-drug group,cilostazol group,low-,middle-and high-dose Poyu decoction groups.After each group was intervened by corresponding drug or non-drug,the tissue structure was observed by HE staining of wound granulation tissue;the angiogenesis was observed by CD31 immunohistochemical staining,VEGFR-2 level in serum was detected by enzyme-linked immunosorbent assay(ELISA)-double antibody sandwich technique;The expression of key proteins in PI3K/Akt/mTOR signaling pathway was detected by Western blot.Results:The results of HE staining showed that low-dose Poyu decoction group increased the neonatal capillaries significantly compared with non-drug group,and also contained a large number of inflammatory cells;middle-and high-doses Poyu decoction groups were significantly improved compared with non-drug group,increased more neonatal capillaries and contained fewer inflammatory cells,which was close to normal control group.Compared with the non-drug group,the number of microvessels and the expression levels of P-PI3K,P-AKT and P-mTOR in granulation tissue of rats with diabetic foot ulcer as well as the expression level of VEGFR2 in serum of rats all were increased in cilostazol group as well as in low-,medium-and high-dose Poyu decoction groups(P﹤0.05).Conclusion:Poyu decoction may promote the angiogenesis of ischemic ulcer tissue by promoting the expression of VEGER-2 protein in ischemic ulcer tissue and the expression of key proteins in PI3K/AKT/mTOR signal pathway.