电刺激糖尿病模型大鼠创面的愈合及血管生成

Electrical stimulation promotes wound healing and angiogenesis in diabetic rats

背景:糖尿病性创面愈合障碍是糖尿病的常见并发症之一,然其发病机制目前尚未阐明。电刺激疗法是临床治疗创面损伤的常用手段之一,能有效促进损伤皮肤愈合。目的:探讨电刺激疗法对糖尿病大鼠创面愈合及血管生成的影响及潜在机制。方法:SPF级雄性SD大鼠42只,随机分为空白对照组和糖尿病模型组。模型组采用一次性尾静脉注射链脲佐菌素(50 mg/kg)联合背部皮肤创伤的方法建立糖尿病性难愈创面大鼠模型,再将造模成功的24只大鼠随机分为模型组和电刺激治疗组;空白对照组为正常大鼠造成背部创面,治疗组予以电刺激治疗21 d,对照组和模型组正常饲养,于治疗第3,7,14,21天分别评价创面愈合情况。治疗完成后,取大鼠血清和创面组织用于指标检测,采用苏木精-伊红染色法观察大鼠创面病理形态,酶联免疫吸附法检测血清内皮型一氧化氮合酶、血管生成素1和血管内皮生长因子水平,免疫组织化学法和Western-blotting法定位定量观察创面血管生成素受体2和血管内皮细胞生长因子受体2蛋白的表达。实验方案经西南医科大学附属医院动物实验伦理委员会批准。结果与结论:(1)经电刺激治疗后,糖尿病大鼠创面愈合率于第14天时即接近90%,而模型组不到60%,差异有显著性意义(P<0.01);血清内皮型一氧化氮合酶、血管生成素1和血管内皮生长因子水平较模型组均显著升高(P<0.05或P<0.01);(2)与模型组比较,治疗组创面新生血管多,血管管腔较大,血管周围及创面组织的血管生成素受体2和血管内皮细胞生长因子受体2表达均显著升高(P<0.05或P<0.01);(3)电刺激疗法能明显促进糖尿病大鼠创面愈合及新血管生成,其机制与升高内皮型一氧化氮合酶、血管生成素1和血管内皮生长因子水平,上调血管生成素受体2和血管内皮细胞生长因子受体2水平有关。

BACKGROUND:Diabetic wound healing disorder is one of the common complications of diabetes,but its pathogenesis has not yet been elucidated.Electrical stimulation therapy is one of the commonly used methods in the clinical treatment of wound injury,which can effectively promote the healing of injured skin.OBJECTIVE:To investigate the effects of electrical stimulation therapy on wound healing and angiogenesis in diabetic rats and its potential mechanism.METHODS:Forty-two SPF male Sprague-Dawley rats were enrolled in this study.A rat model of diabetic refractory wounds was established in 24 rats by tail vein injection of streptozotocin(50 mg/kg)combined with back skin wounds,and model rats were randomly divided into model group and electrical stimulation group.Meanwhile,normal rats were taken to cause back wounds as a blank control group.The electrical stimulation group was treated with electrical stimulation for 21 days.The blank control group and the model group were fed normally.The wound healing was evaluated on the 3rd,7th,14th and 21 st days after treatment.After treatment,the rat serum and wound tissue were taken for index detection.The pathological morphology of rat wounds was observed by hematoxylin-eosin staining.The serum endothelial nitric oxide synthase,angiopoietin 1 and vascular endothelial growth factor levels were detected by enzyme-linked immunosorbent assay.The expression levels of angiopoietin receptor 2 and vascular endothelial growth factor receptor 2 proteins were detected by immunohistochemistry and western blot.An ethical approval was obtained from the Animal Experimental Ethics Committee of the Affiliated Hospital of Southwest Medical University.RESULTS AND CONCLUSION:The wound healing rate of diabetic rats was close to 90%on the 14 th day after electrical stimulation,while the healing rate in the model group was less than 60%(P<0.01).The serum endothelial nitric oxide synthase,angiopoietin 1 and vascular endothelial growth factor levels were significantly higher in the electrical stimulation group than the model group(P<0.05 or P<0.01).Meanwhile,compared with the model group,the electrical stimulation group had more neovascularization,larger vascular lumen,higher expression of angiopoietin receptor 2 and vascular endothelial growth factor receptor 2 around the blood vessels(P<0.05 or P<0.01).These findings indicate that electrical stimulation therapy can significantly promote wound healing and neovascularization in diabetic rats,and its mechanism is related to the increase of endothelial nitric oxide synthase,angiopoietin 1 and vascular endothelial growth factor levels and up-regulation of angiopoietin receptor 2 and vascular endothelial growth factor receptor 2 levels.