摘要
目的观察不同浓度的葡萄糖、胰岛素和氧化型低密度脂蛋白(ox-LDL)对内皮细胞的协同损伤作用,建立一种2型糖尿病血管并发症体外内皮细胞损伤模型.方法采用L9(34)正交设计方法,将生长融合状的血管内皮细胞(ECV-304)随机分为9组,分别施加不同浓度的葡萄糖、胰岛素和ox-LDL条件液,培养24 h后观测细胞活性、细胞一般形态学和超微结构改变以及细胞凋亡情况.结果显微镜下见模型组细胞回缩变圆,轮廓分明,细胞内出现粗糙颗粒样变化;线粒体肿胀,嵴丢失,内质网结构模糊、肿胀,呈小泡状;琼脂糖凝胶电泳可见模型组有明显的细胞凋亡典型的阶梯状区带电泳图谱(DNA Ladder).结论三因素对细胞有协同损伤作用,可以诱导细胞发生凋亡和细胞器结构发生变化,含100 mg/L ox-LDL、20 mmol/L葡萄糖和100 mU/L胰岛素的混合条件培养液对细胞的损伤最显著,可作为糖尿病血管并发症体外内皮细胞损伤模型的合适诱导条件液.
Objective To observe the effects of different concentration of glucose, insulin and ox-LDL on the endothelial cells cultured in vitro, and to establish a new in vitro cell-injuring model of diabetic vascular complication. Methods According to the orthogonal design L9 （34）, we randomized the well growing human umbilical vein endothelial cell line ECV-304 into 9 groups, and each group cells were exposed to different concentration of glucose, insulin and ox-LDL according the orthogonal table for 24 hours. Then, cell viability, common morphology changes and microstructure changes of the cells and apoptosis were examined. Results Compared with other group cells, the model group cells showed more pyknocyte, wider intercellular space and more abnormal granule th cytoplasm under optics microscope, and more typical DNA ladders occurred in the agarose gel electrophoresis. Moreover, under electron microscope the model group cells took on abnormal microstructure with swelling mitochondria, losing ridge and clouding endoplasmic reticulum. Conclusion The synergistic damage action of the three risk factors is responsible for more occurrences of apoptosis and alterations in microstructure of the model group cells. A new in vitro cell-injuring model of diabetic vascular complication could be gotten, with the conditional incubation medium consisting of 100 mg/L ox- LDL, 20 mmol/L glacose and 100 mU/L insulin.
出处
《血栓与止血学》
2005年第6期245-249,256,共6页
Chinese Journal of Thrombosis and Hemostasis
基金
湖南省教育厅科研基金重点资助项目(02A031)
