【摘要】 目的:研究反义寡核苷酸(antisense oligodeoxynucleotides,AS-ODN)抑制早期生长反应基因1(early growth response gene-1,Egr-1)表达对内皮细胞缺氧复氧(anoxia/reoxygenation,A/R)损伤的影响。方法:采用新生大鼠进行心脏微血管内皮细胞(cardiac microvascular endothelial cells,CMECs)原代培养,取3~4代的CMECs建立缺氧复氧模型。细胞随机分为6组:对照(Con)组、缺氧复氧组(A/R)、溶剂组(LIP)、反义寡核苷酸转染组(AS)、正义寡核苷酸转染组(S)和错配寡核苷酸转染组(Sc)。通过测定细胞培养上清液中乳酸脱氢酶(LDH)及内皮细胞中超氧化物歧化酶(SOD)、丙二醛(MDA)含量,观察内皮细胞损伤程度;应用ELISA法测定细胞培养上清液肿瘤坏死因子-α(TNF-α)的含量,观察内皮细胞炎症反应水平;Western-blot法检测培养内皮细胞中Egr-1的蛋白表达水平;显微镜下观察细胞形态学改变并计算存活率。结果:A/R造成内皮细胞内MDA升高,SOD下降,上清液中LDH、TNF-α含量升高,A/R刺激下细胞Egr-1的蛋白表达水平明显升高;A/R刺激前给予AS-ODN可抑制Egr-1蛋白的表达,减轻内皮细胞的损伤及炎症反应程度,提高细胞存活率。结论:AS-ODN抑制培养内皮细胞Egr-1的表达,并降低A/R损伤,提示Egr-1与缺氧复氧所致的心脏微血管内皮细胞损伤密切相关。更多还原

【Abstract】 AIM:To investigate the effects of antisense oligodeoxynucleotides(AS-ODN)on Egr-1protein expression in cultured endothelial cells after anoxia/reoxygenation(A/R).METHODS:The cultured cardiac microvascular endothelial cells(CMECs)A/R model were established.The cells were randomly divided into one of six groups:Con,A/R,Lip,AS,S,and Sc.Levels of lactate dehydrogenase(LDH),superoxide dismutase(SOD), malondialdehyde(MDA),and tumor necrosis factor-α(TNF-α)were measured to assess the degree of injury and inflammation of endothelial cells.Egr-1protein expression was examined by Western-blot analyses.Cell morphology and cell viability were observed to assess the degree of injury.RESULTS:Treatment with Egr-1AS-ODN significantly reduced Egr-1 protein expression and attenuated injury and inflammation of endothelial cells caused by A/R evidenced by the the decrease in leakage of LDH,the increase in SOD activity,the decrease in MDA generation,and release of TNF-αfrom cultured CMECs.CONCLUSION:AS-ODN can protect cultured CMECs from A/R injury,by inhibiting the overexpression of Egr-1.Egr-1 was related to the A/R injury of CMECs.更多还原