苦参碱对体外培养大鼠视网膜微血管内皮细胞增殖及VEGF表达影响.pdfVIP

过了50%。细胞计数绘制生长曲线结果与MTT 结果相一致。2、 AO/EB 荧光双染 色法观察:在 Mat 浓度达到40mg/L 后处理RRMECs 呈现不同程度的细胞凋亡形态 学特征。3、免疫细胞化学结果显示：VEGF 表达定位于胞膜和胞浆,阳性部分为 棕黄色颗粒。血管抑素（130mg/L）和Mat(40mg/L)处理后其阳性表达明显降低， 灰度值越少（P0.05）且两者作用无明显差别；高糖组的 VEGF 阳性表达最高， 灰度值也最大，Mat(40mg/L)同样能下调其表达（P0.05）。4、Western-Blotting 半定量检测VEGF 蛋白的表达与免疫细胞化学的结果一致。 结论：1、 Mat 能有效地抑制体外培养的RRMECs 的增殖，在一定的浓度和时间 范围内其抑制作用呈明显的剂量和时间依赖性；2、Mat 能有效地诱导 RRMECs 凋亡；3、通过免疫细胞化学和 Western-Blotting 证实其抑制主要是通过下调 VEGF 表达而发生作用的，高糖能上调 VEGF 的表达，Mat 对高糖条件下的 VEGF 表达同样也有抑制作用，提示Mat 对糖尿病视网膜病变等引起的视网膜新生血管 防治具有一定的应用前景。 关键词：苦参碱；视网膜微血管内皮细胞；血管内皮生长因子；视网膜新生血 管；免疫印迹方法；免疫细胞化学 4 The effects of matrine on cell proliferation and expression of intercellular VEGF in microvascular endothelial cells from rat retina by cultured in vitro Abstract Objective ：To evaluate the effects of matrine on cell proliferation and expression of intercellular VEGF in RRMECs in vitro and to provide a new clue and theory basis for developing a drug of preventing and curing retinal neovascularization in the future. Methods ： RRMECs were primary cultured, the third generation of RRMECs were treated. The effect of Mat on the proliferation of RRMECs was measured by MTT assay. The influence of Mat on the growth of RRMECs was determined by cell counting method in trypan blue staining. AO/EB double fluorescence staining method was used to observe the morphological changes of RRMECs apoptosis after treatment with Mat. RRMECs were divided into 5 groups: the control group, angiostatin (130mg/L) group, Mat (40mg/L) group, high glucose (25mmol/L)group, and Mat(40mg/L)+high glucose(25mmol/L) group. After 48 hour, the localization of VEGF protein was examined by immunocytochemistry. Western-blotting was used to measure the