TUMOR NECROSIS FACTOR-ALPHA-MEDIATED DECREASE IN GLUTATHIONE INCREASES THE SENSITIVITY OF PULMONARY VASCULAR ENDOTHELIAL-CELLS TO H2O2
TUMOR NECROSIS FACTOR-ALPHA-MEDIATED DECREASE IN GLUTATHIONE INCREASES THE SENSITIVITY OF PULMONARY VASCULAR ENDOTHELIAL-CELLS TO H2O2
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DOI：
10.1172/jci115658
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发表时间：
1992-03-01
影响因子：
15.9
通讯作者：
MALIK, AB
中科院分区：
文献类型：
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作者：
ISHII, Y;PARTRIDGE, CA;MALIK, AB
我们研究了肿瘤坏死因子-α（TNF-α）刺激内皮细胞对H2O2诱导的内皮通透性增加的影响。将牛肺微血管内皮细胞（BPMVEC）在微孔滤膜上生长至汇合，并测定I-125-白蛋白穿过单层的清除率。用TNF-α（100 U/ml）预处理6 h对跨内皮I-125-白蛋白渗透性没有直接影响。然而，TNF-α预处理增强了BPMVEC对H2O2的敏感性;也就是说，H2O2（10 μ M）单独没有直接影响，而H2O2增加I-125-白蛋白渗透性超过三倍时，加入到用TNF-α预处理6小时的单层细胞中。乳酸脱氢酶释放的测定表明，增加的渗透性不是由于细胞溶解。我们测量了细胞内谷胱甘肽和过氧化氢酶的含量，以确定它们在介导对H2 O2敏感性增加中可能发挥的作用。TNF-α治疗（100 U/ml，6小时）减少总GSH含量，同时增加氧化型GSH含量，但不改变细胞过氧化氢酶活性。GSH的作用进行了研究，通过预处理内皮细胞与2 mM GSH为3小时，产生了80%的细胞内GSH含量增加。GSH补充抑制了TNF-α处理的内皮细胞对H2O2的敏感性增加。我们测试了黄嘌呤氧化酶（XO）抑制的影响，因为XO激活可能是导致细胞GSH含量降低的氧化剂的来源。用0.5 mM羟嘌呤醇预处理可减弱TNF-α和H2 O2对内皮渗透性的协同作用。结果表明，降低氧化剂缓冲能力继发于TNF-α诱导的细胞内GSH含量减少介导的内皮细胞对H2O2的敏感性增加。这一机制可能有助于与TNF-α释放相关的败血症中的氧化剂依赖性血管内皮损伤。
We examined the effects of tumor necrosis factor-alpha (TNF-alpha) stimulation of endothelial cells on the increase in endothelial permeability induced by H2O2. Bovine pulmonary microvascular endothelial cells (BPMVEC) were grown to confluence on a microporous filter and the I-125-albumin clearance rate across the monolayer was determined. Pretreatment with TNF-alpha (100 U/ml) for 6 h had no direct effect on transendothelial I-125-albumin permeability. However, TNF-alpha pretreatment enhanced the susceptibility of BPMVEC to H2O2; that is, H2O2 (10-mu-M) alone had no direct effect, whereas H2O2 increased I-125-albumin permeability more than threefold when added to monolayers pretreated for 6 h with TNF-alpha. Determination of lactate dehydrogenase release indicated that increased permeability was not due to cytolysis. We measured the intracellular contents of GSH and catalase to determine their possible role in mediating the increased susceptibility to H2O2. TNF-alpha treatment (100 U/ml for 6 h) decreased total GSH content and concomitantly increased the oxidized GSH content, but did not alter the cellular catalase activity. The role of GSH was examined by pretreating endothelial cells with 2 mM GSH for 3 h, which produced an 80% increase in intracellular GSH content. GSH repletion inhibited the increased sensitivity of the TNF-alpha-treated endothelial cells to H2O2. We tested the effects of xanthine oxidase (XO) inhibition since XO activation may be a source of oxidants responsible for the decrease in cellular GSH content. Pretreatment with 0.5 mM oxypurinol attenuated the synergistic effect of TNF-alpha and H2O2 on endothelial permeability. The results indicate that decreased oxidant buffering capacity secondary to TNF-alpha-induced reduction in intracellular GSH content mediates the increased susceptibility of endothelial cells to H2O2. This mechanism may contribute to oxidant-dependent vascular endothelial injury in septicemia associated with TNF-alpha release.