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目的从基因水平探究氧化应激在内皮细胞功能障碍中的作用及机制.方法用胶原酶消化结合免疫磁珠分选法纯化大鼠海绵体内皮细胞,通过黄嘌呤/黄嘌呤氧化酶(200μM/L,60mU/L,X/XO)体系氧化损伤48h,应用基因芯片研究损伤前后基因表达的差异,并利用分析软件筛选出与内皮细胞功能障碍相关的细胞信号通路.结果经纯化的海绵体内皮细胞纯度达96%(抗 vWF 法)和92.7%(流式细胞仪法)且生长增殖良好.氧化损伤后,内皮细胞透亮度减低,发生皱缩,释放乳酸脱氢酶(3200±223U/L vs 420±56U/L)显著增加(P <0.05).甲醛变性凝胶电泳中,提取的 RNA 在氧化损伤组与对照组均有清晰的18s、28s 条带电泳图,28s:18s 约为2:1.在对照组,表达基因13090个(占基因总数的31.92%),不表达27918个(占总数68.07%);氧化损伤组表达基因12039个(29.35%),不表达28972个(70.65%),其中,差异表达基因2480个(差异倍数2.0),1454个基因表达上调,1026个基因表达下调,并筛选出黏附功能、内吞作用、ErbB、细胞骨架肌动蛋白调控、蛋白转运等多个与内皮功能障碍关系密切的主要信号路径蛋白.结论氧化应激在内皮细胞功能障碍中有重要作用,并且通过多条信号通路调控相关基因的表达水平以影响内皮细胞功能.“,”Objective To investigate the role of oxidative stress in endothelial dysfunction in gene level. Methods The collection of purified cavernosum endothelial cells were procedured by collagenase digestion combined with magnetic activated cell sorting(MACS), after injured by xanthine (200 μM/L) and xanthine oxidase (60mU/L, X/XO) for 48h, the differentially expressed genes and their pathways related to oxidative stress had been screened by gene chip and analysed by SAS software. Results The purity of endothelial cells attained 96%(anti-vWF) and 92.7%(FC), respectively, and the cells grew well. After injured by X/XO, the cells shrunk and their transparent ability reduced. Furthermore, the releasing of LDH of injuried cells (3200±223)U/L significantly increased than that of normal control group (420±56)U/L, P < 0.05. In formaldehyde- agarose gel electrophoresis, the intact bands(18s,28s) were showed in both groups, and 28s:18s were around 2:1. And in gene microarray, there were 13 090 (31.92% of total) genes expressing in normal control group with 27918 (68.07% of total) genes being not; in injured group, there were 12 039 (29.35%) genes expressing and 28972 (70.65%) genes did not. When the cut-off value was set as 2, there were 2 480 genes differentially expressed between X/XO injured group and normal control group, furthermore, 1 454 genes upregulated and 1026 genes downregulated. What’s more, relevant proteins of certain pathways, such as Adhension juncion, Endocytosis, ErbB signaling pathways, Regulation of actin cytoskeleton and Protein export had been pro-posed to be significant in endothelial dysfunction. Conclusion Oxidative stress may play a crucial role in endothelial dysfunction through some certain cell signal pathways by regulating the expression of oxidative stress-related genes.