Disruption of redox homeostasis in tumor necrosis factor-induced apoptosis in a murine hepatocyte cell line
Disruption of redox homeostasis in tumor necrosis factor-induced apoptosis in a murine hepatocyte cell line
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DOI：
10.1016/s0002-9440(10)64533-6
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发表时间：
2000-07-01
影响因子：
6
通讯作者：
Fausto, N
中科院分区：
文献类型：
--
作者：
Pierce, RH;Campbell, JS;Fausto, N
肿瘤坏死因子（TNF）是肝脏急性期反应的介质，可以启动肝细胞增殖并导致细胞死亡。我们研究了TNF引起肝细胞凋亡的机制，重点是氧化应激，抗氧化防御和线粒体损伤的作用。研究在培养的AML 12细胞（一种分化的鼠肝细胞系）中进行。与体内肝细胞的情况一样，AML 12细胞对单独TNF引起的细胞死亡不敏感，但当暴露于TNF和小剂量的放线菌素D（Act D）时，细胞凋亡而死亡。直到处理后G小时才检测到细胞凋亡的形态学迹象，到18小时，类似于50%的细胞已经死亡。细胞暴露于TNF+Act D并不阻断NF κ B B核转位、DNA结合或其整体反式激活能力。凋亡诱导的特征在于氧化应激，其由NAD（P）H和谷胱甘肽的损失指示，随后是线粒体损伤，包括线粒体膜电位的损失、内膜结构损伤和线粒体凝聚。这些变化与细胞色素C的释放和caspase-8、-9和-3的激活相一致，TNF诱导的细胞凋亡依赖于谷胱甘肽水平。在谷胱甘肽水平降低的细胞中，TNF本身在缺乏转录阻断的情况下充当凋亡剂。相反，抗氧化剂铜-硫辛酸（其防止暴露于TNF+Act D的细胞中谷胱甘肽的损失）完全防止线粒体损伤、半胱天冬酶活化、细胞色素C释放和凋亡。结果表明，TNF+ ActD诱导的AML 12细胞凋亡涉及氧化损伤和线粒体损伤。由于损伤在更大程度上受细胞谷胱甘肽含量的调节，我们认为TNF+Act D的组合会导致细胞凋亡，因为Act D阻断了抗氧化防御所需基因的转录。
Tumor necrosis factor (TNF) is a mediator of the acute phase response in the liver and can initiate proliferation and cause cell death in hepatocytes, We investigated the mechanisms by which TNF causes apoptosis in hepatocytes focusing on the role of oxidative stress, antioxidant defenses, and mitochondrial damage. The studies were conducted in cultured AML12 cells, a line of differentiated murine hepatocytes. As is the case for hepatocytes in vivo AML12 cells were not sensitive to cell death by TNF alone, but died by apoptosis when exposed to TNF and a small dose of actinomycin D (Act D), Morphological signs of apoptosis were not detected until G hours after the treatment and by 18 hours similar to 50% of the cells had died. Exposure of the cells to TNF+Act D did not block NF kappa B nuclear translocation, DNA binding, or its overall transactivation capacity. Induction of apoptosis was characterized by oxidative stress indicated by the loss of NAD(P)H and glutathione followed by mitochondrial damage that included loss of mitochondrial membrane potential, inner membrane structural damage, and mitochondrial condensation. These changes coincided with cytochrome C release and the activation of caspases-8, -9, and -3, TNF-induced apoptosis was dependent on glutathione levels. In cells with decreased levels of glutathione, TNF by itself ill the absence of transcriptional blocking acted as an apoptotic agent, Conversely, the antioxidant cu-lipoic acid, that protected against the loss of glutathione in cells exposed to TNF+Act D completely prevented mitochondrial damage, caspase activation, cytochrome C release, and apoptosis, The results demonstrate that apoptosis induced by TNF+Act D ill AML12 cells involves oxidative injury and mitochondrial damage. As injury was regulated to a larger extent by the glutathione content of the cells, we suggest that the combination of TNF+Act D causes apoptosis because Act D blocks the transcription of genes required for antioxidant defenses.