摘要
目的观察复元醒脑汤对糖尿病脑梗死大鼠内皮祖细胞（EPCs）功能的干预作用。方法将120健康雄性sD大鼠按随机数字表法分为对照组、假手术组、模型组、治疗组，每组30只。采用腹腔注射链脲佐菌素（STZ）50mg／kg后给予高能量高脂肪饲料喂养2周的方法复制糖尿病大鼠模型；制模后由颈内动脉注入自体血栓复制糖尿脑梗死模型；假手术组注射生理盐水。制模后治疗组给予复元醒脑汤[人参10g（单煎）、三七10鼽石菖蒲12g、水蛭10g、益母草30g、生南星15g、制大黄9g（后下），浓度388g／L]10．4mL·kg^-1·d。灌胃，每日2次，连续7d；假手术组、模型组给予等量生理盐水灌胃，连续7d。取大鼠外周血10mL离心，吸取界面层的单核细胞，传代培养后用于实验。用流式细胞仪检测各组细胞CD31、CD34、血管内皮生长因子受体-1（FLK-1）和血管性血友病因子（vWF）的表达量，将具有双标阳性的细胞作为EPCs的阳性表达，用四甲基偶氮唑盐（MTY）比色法测定细胞增殖能力，并测定细胞迁移能力。结果假手术组EPCs表达量较对照组显著降低【FLK-1为（34．64±1．92）％比（41．68±1．53）％，vWF为（40．92±1．16）％比（5i．98±1．06）％]，EPCs增殖率培养1d起即较对照组显著降低[（0．014±0＋001）％比（0．021±0．001）％]，持续到培养30d[（0．084±0．009）％比（0．121±0．003）％]，EPCs迁移率亦较对照组显著降低[吸光度（A）值：32．80±4．15比62．20±3．27，均P〈0．05]；模型组EPCs表达量较假手术组显著降低（FLK-1为（30．19±2．00）％比（34．64±1．92）％，vWF为（38．54±2．19）％比（40．92±1．16）％]，细胞增殖率培养1d起即较假手术组显著升高[（0．009±0.001）％比（0．014±0．001）％]，持续到培养30d[（0．057±0．005）％比（0．084±0．009）％]，EPCs细胞迁移率均显著低于假手术组[（18．00±1．58）％比（32．80±4．15）％，均P〈0．05]；治疗组EPCs表达量较模型组升高[FLK-1：（35．90±1．57）％，vWF：（42．05±0．85）％]，但差异无统计学意义（均P〉0．05），培养1d治疗组细胞增殖率与模型组比较差异无统计学意义，随着培养时间延长，两组逐渐显现出统计学差异，培养2d起治疗组EPCs增殖率（A值：0．042±0．003）显著高于模型组（P〈0．05或P〈0．01），持续到培养30d（A值：0．098±0．004），细胞迁移率（A值：43．20±3．27）较模型组明显升高（P〈0．01）。结论复元醒脑汤可提高糖尿病脑梗死大鼠EPCs的增殖、迁移能力，从而使其受损的血管内皮功能得到修复，进而促进局部血管再生和侧支循环的建立。
Objective To observe the interference effect of Fuyuan Xingnao decoction on the function of endothelial progenitor cells （EPCs） in rats with diabetic cerebral infarction. Methods 120 healthy male Sprague- Dawley （SD） rats were divided into control group, sham-operation group, model group and treatment group by random digits table, each n = 30. The rat diabetes mellitus model was reproduced by intraperitoneal injection of urea with streptozotocin （STZ） 50 mg/kg and afterward the animal was fed by high energy high-fat diet for 2 weeks; after the diabetic modeling, the diabetic cerebral infarction model was reproduced by the injection of autologous thrombus into the internal carotid artery; in the sham-operation group, an ophthalmic surgical tweezers was used to ligate the left carotid vein and artery, afterward a clip was applied to close the artery in the neck and normal saline was injected into the internal carotid artery. After diabetic cerebral infarction modeling, in treatment group, 10.4 mL · kg ^-1· d^-1 of Fuyuan Xingnao decoction [ingredients： ginseng 10 g （decocted separately）, notoginseng 10 g, acori graminei 12 g, leech 10 g, motherwort 30 g, south star 15 g, prepared rhubarb 9 g （added later in the boiling decoction） and the concentration was 388 g/L] was given by gastric lavage, twice a day for consecutive 7 days. In the sham-operation group and model group, a same amount of normal saline was given by gastric lavage for consecutive 7 days. The rat peripheral blood 10 mL was collected and centrifuged, the mononuclear cells at the interface layer were absorbed for subculture to harvest the EPCs ready to be used in this experiment. The expressions of CD31, CD34, vascular endothelial growth factor receptor （FLK-1） and yon willebrand factor （vWF） were detected bv the flow cvtometrv and the oositive cells with double labelinz wereconsidered to be EPCs positive expression. The cell proliferation ability was determined by methylthiazolyl tetrazolium （MTT） and the cell migration ability was calculated in the two groups, Results The EPCs expression in sham- operation group was significantly lower than that in the control group [FLK-1 was （34.64 ± 1.92）% vs. （41.68 ± 1.53）%, vWF was （40.92 ± 1.16）% vs. （51.98 ± 1.06）%], the EPCs proliferation ability in sham-operation group was obviously lower than that in the control group on the 1st day [（0.014 ± 0.001）% vs. （0.021 ± 0.001）%], persisting until culture for 30 days [（0.084 ±0.009）% vs. （0.121 ± 0.003）%], migration ability of EPCs in sham-operation group was also markedly lower than that in control group [absorbauce （A） value： 32.80 ± 4.15 vs. 62.20 ±3.27, all P 〈 0.01]; the expression of EPCs in model group was significantly less than that in the control group [FLK-1 was （30.19 ± 2.00）% vs. （34.64 ±1.92）%, vWF was （38.54± 2.19）% vs. （40.92± 1.16）%], the cell proliferation rate in model group was obviously increased compared with that in sham-operation group from the 1st day of culture [（0.009 ± 0.001）% vs. （0.014 ±0.001）%], and the increase persisted for 30 days in culture [（0.057 ± 0.005）% vs. （0.084 ±0.009）%], EPCs migration rate in the model group was remarkably lower than that in the sham-operation group [（18.00 ± 1.58）% vs. （32.80 ± 4.15）%, all P 〈 0.05]. The expression of EPCs in the treatment group was significantly higher than that in the model group [FLK-I： （35.90 ± 1.57）%, vWF： （42.05± 0.85）%], but there was no statistical significant difference （both P 〉 0.05）. There was no statistically significant difference in cell proliferation rate in the model group compared with that of treatment group on the 1st day, but the difference gradually appeared between the two groups as the incubation time extended. The EPCs proliferation ability in the treatment group （A value： 0.042 ± 0.003） was significantly higher than that in model group on the 2nd day of culture （P 〈 0.05 or P 〈 0.05）, and the difference lasted until the 30th day （A value： 0.098 ± 0.004）; the cell migration ability in the treatment group （A value： 43.20 ± 3.27） was higher than that in the model group （P 〈 0.01）. Conclusion Fuyuan Xingnao decoction can elevate the EPCs functions of proliferation and migration in rats with diabetic cerebral infarction, thus the function and repair ability of damaged vascular endothelial cell can be improved to promote the local vascular regeneration and establishment of collateral circulation.
出处
《中国中西医结合急救杂志》
CAS
北大核心
2016年第4期412-416,共5页
Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care
基金
国家自然科学基金（81273725）
关键词
复元醒脑汤
内皮祖细胞
糖尿病脑梗死
Fuyuan Xingnao decoction
Endothelial progenitor cell
Diabetic cerebral infarction
