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钟 林 尢家马 录 肖献忠 孙去病
摘 要 目的：探讨核转录因子-κB(NF-κB)在热休克预处理减轻肿瘤坏死因子-α(TNF-α)所致内皮细胞凋亡中的作用。方法：将培养牛肺动脉内皮细胞(BPAEC)于42℃孵育1 h，37℃再培养16 h后，用TNF-α(2 500 U/mL)损伤24 h，观察BPAEC形态学变化、DNA断裂百分率，抽提DNA作琼脂糖电泳，以确定细胞凋亡情况。Western blot分析70 KD-热休克蛋白(HSP70)合成、NF-κB抑制蛋白I-κBα量的改变以及诱导型一氧化氮合酶(iNOS)的生成。凝胶滞留法检测NFkB活性，免疫组织化学染色法分析NFkB的移位。结果：热休克预处理可明显减轻TNF-α所致BPAEC凋亡，表现为与TNF-α损伤组相比，凋亡细胞减少，DNA电泳未出现明显“梯状”条带，DNA断裂百分率降低。进一步实验发现热休克预处理可诱导BPAEC合成HSP70，并可抑制TNF-α引起的BPAEC的I-κBα的降解、NF-κB向细胞核的移位、NF-κB与DNA的结合活性，以及iNOS的生成。结论：热休克预处理可减轻TNF-α所致BPAEC凋亡，其机制与HSP70阻断I-κBα降解，从而抑制NF-κB活化，抑制iNOS转隶合成有关。
主题词 NF-κB；热休克蛋白质类；内皮；一氧化氮；肿瘤坏死因子
Role of NF-κB in protection of heat shock
pretreatment against TNF-α-induced endothelial cell apoptosis
ZHONG Lin, YOU Jia-Lu XIAO Xian-Zhong, SUN Qu-Bing
Department of Pathophysiology, Hunan Medical University, Changsha (410078)
Abstract AIM:To explore the role of nuclear factor kappa B (NF-κB) in protection of heat shock pretreatment against TNF-α induced endothelial cell apoptosis.METHODS： Cultured bovine pulmonary artery endothelial cells (BPAECs) were exposed to TNF-α(2 500 U/mL, 24 h) 16h after heat shock pretreatment(43℃,1h). The apoptotic morphological changes, DNA ladder pattern on agarose gel electrophoresis, and percentage of DNA fragmentation of BPAEC were analyzed. Inducible heat shock protein-70 kD(HSP70), inhibitor κBα(I-κBα) levels, expression of inducible nitric oxide synthase(iNOS) and NF-κB activity, NF-κB translocation to nuclei were detected using Western blot analysis, electrophoretic mobility shift assays (EMSA), and immunohistochemical analysis respectively.RESULTS：Heat shock pretreatment significantly attenuated TNF-α induced apoptosis in BPAEC. To compared with TNF-α treated BPAEC[1] [2] [3] 下一页
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