Zhou Zhu, Gengaro Patricia, Wang Wei, Wang Xue-qing, Li Chunling, Faubel Sarah, Rivard Christopher, Schrier Robert W
Department of Medicine, University of Colorado at Denver and Health Sciences Center, Box B173, 4200 E 9th Ave., Denver, CO 80262, USA.
Am J Physiol Renal Physiol. 2008 Oct;295(4):F932-41. doi: 10.1152/ajprenal.00066.2008. Epub 2008 Jul 16.
The interaction of tumor necrosis factor (TNF)-alpha with the endothelium is a pivotal factor during endotoxemia. Inflammatory conditions are characterized by the activation of the transcription factor NF-kappaB and the expression of inflammatory mediators. Previous reports indicate that inhibition of NF-kappaB activation during sepsis may be beneficial to the microvasculature. In addition, the phosphatidylinositol-3-kinase/Akt signaling pathway (PI3-kinase/Akt) has been shown to be cytoprotective. In this study, we examined the effect of inhibition of NF-kappaB and PI3-kinase/Akt on cell viability, cytokine production, inducible nitric oxide synthase (iNOS) expression, and nitric oxide (NO) generation by TNF-alpha-treated cultured microvascular endothelial cells. TNF-alpha induced significant cytotoxicity and was associated with increased inflammatory cytokines and NO and increased expression of iNOS. The NF-kappaB inhibitor, pyrrolidine dithiocarbamate (PDTC), prevented these increases and significantly attenuated the TNF-alpha-induced cytotoxicity. TNF-alpha also caused PI3-kinase/Akt activation, which was further increased by PDTC and prevented by the PI3-kinase inhibitor, LY294002. Inhibition of PI3-kinase/Akt also significantly potentiated TNF-alpha-mediated cytotoxicity. LY294002 treatment resulted in the appearance of increased apoptosis, compatible with the known anti-apoptotic properties of PI3-kinase/Akt. The present results therefore demonstrate a cytotoxic effect of TNF-alpha in microvascular endothelial cells which can be attenuated by NF-kappaB inhibition. In addition, PI3-kinase/Akt activation during TNF-alpha exposure may represent a compensatory anti-necrotic and anti-apoptotic pathway. The cytoprotective effects of NF-kappaB inhibition and PI3-kinase/Akt activation may have potential implications in the treatment of endotoxemia and septic shock.
肿瘤坏死因子（TNF）-α与内皮细胞的相互作用是内毒素血症期间的一个关键因素。炎症状态的特征是转录因子NF-κB的激活和炎症介质的表达。先前的报道表明，在脓毒症期间抑制NF-κB激活可能对微血管系统有益。此外，磷脂酰肌醇-3-激酶/蛋白激酶B信号通路（PI3-激酶/蛋白激酶B）已被证明具有细胞保护作用。在本研究中，我们检测了抑制NF-κB和PI3-激酶/蛋白激酶B对TNF-α处理的培养微血管内皮细胞的细胞活力、细胞因子产生、诱导型一氧化氮合酶（iNOS）表达和一氧化氮（NO）生成的影响。TNF-α诱导了显著的细胞毒性，并与炎症细胞因子和NO增加以及iNOS表达增加有关。NF-κB抑制剂吡咯烷二硫代氨基甲酸盐（PDTC）可阻止这些增加，并显著减轻TNF-α诱导的细胞毒性。TNF-α还导致PI3-激酶/蛋白激酶B激活，PDTC可进一步增强这种激活，而PI3-激酶抑制剂LY294002可阻止这种激活。抑制PI3-激酶/蛋白激酶B也显著增强了TNF-α介导的细胞毒性。LY294002处理导致凋亡增加，这与PI3-激酶/蛋白激酶B已知的抗凋亡特性相符。因此，本研究结果表明TNF-α对微血管内皮细胞具有细胞毒性作用，而抑制NF-κB可减轻这种作用。此外，TNF-α暴露期间PI3-激酶/蛋白激酶B激活可能代表一种代偿性的抗坏死和抗凋亡途径。抑制NF-κB和激活PI3-激酶/蛋白激酶B的细胞保护作用可能在内毒素血症和脓毒性休克的治疗中具有潜在意义。
