NITRIC-OXIDE SYNTHESIS IN ENDOTHELIAL-CELLS - EVIDENCE FOR A PATHWAY INDUCIBLE BY TNF-ALPHA
NITRIC-OXIDE SYNTHESIS IN ENDOTHELIAL-CELLS - EVIDENCE FOR A PATHWAY INDUCIBLE BY TNF-ALPHA
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DOI：
10.1152/ajpcell.1991.261.4.c634
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发表时间：
1991-10-01
影响因子：
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通讯作者：
MARSDEN, PA
中科院分区：
文献类型：
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作者：
LAMAS, S;MICHEL, T;MARSDEN, PA
一氧化氮（NO）的释放解释了内皮源性舒张因子的生物活性。鉴于肿瘤坏死因子-α（TNF-α）已被认为是感染性休克的重要介质，我们探讨了TNF-α是否能增强内皮细胞中NO和L-瓜氨酸的L-精氨酸依赖性合成。在共孵育生物测定中检测NO的释放，其中测量报告单细胞层（即肾小球系膜细胞或胎儿肺成纤维细胞）中鸟苷3 '，5'-环一磷酸（cGMP）的产生，反映可溶性鸟苷酸环化酶的活化。在TNF-α处理的牛主动脉和肾动脉内皮细胞的存在下，报告细胞单层cGMP含量大于在溶剂处理的内皮细胞的存在下。TNF-α刺激的报告细胞单层cGMP含量的内皮依赖性增加在8 h首次明显，在16-24 h达到最大。此外，TNF-α刺激的内皮依赖性报告细胞单层cGMP含量的增加被血红蛋白和亚甲蓝消除，被N-ω-硝基-L-精氨酸钝化，并被超氧化物歧化酶和钙激动剂缓激肽增强。这些观察结果表明，TNF-α增强NO的释放。此外，通过定量阳离子交换色谱和薄层色谱测定，TNF-α以时间和浓度依赖性方式增强L-[C-14]精氨酸形成L-[C-14]瓜氨酸。因此，很明显，内皮细胞响应于TNF-α而释放NO持续较长时间，并且当用钙激动剂刺激时短暂释放NO。从TNF-α刺激的内皮细胞释放NO的延长可能与感染性休克的发病机制有关。
Nitric oxide (NO) release accounts for the biological activity of endothelium-derived relaxing factor. Given that tumor necrosis factor-alpha (TNF-alpha) has been implicated as an important mediator in septic shock, we explored whether TNF-alpha enhances L-arginine-dependent synthesis of NO and L-citrulline in endothelial cells. The release of NO was detected in a coincubation bioassay where measurement of guanosine 3',5'-cyclic monophosphate (cGMP) production in reporter monolayers, namely glomerular mesangial cells or fetal lung fibroblasts, reflected activation of soluble guanylate cyclase. Reporter monolayer cGMP content was greater in the presence of TNF-alpha-treated bovine aortic and renal artery endothelial cells than in the presence of vehicle-treated endothelial cells. TNF-alpha-stimulated endothelium-dependent increases in reporter monolayer cGMP content were first evident at 8 h and maximal at 16-24 h. In addition, TNF-alpha-stimulated endothelium-dependent increases in reporter monolayer cGMP content were abrogated by hemoglobin and methylene blue, blunted by N-omega-nitro-L-arginine and augmented by superoxide dismutase and the calcium agonist bradykinin. These observations suggested that TNF-alpha enhanced release of NO. Furthermore, the formation of L-[C-14]citrulline from L-[C-14]arginine, as determined by quantitative cation-exchange chromatography and thin-layer chromatography, was enhanced by TNF-alpha in a time- and concentration-dependent manner. Thus it is evident that endothelial cells release NO for a prolonged period in response to TNF-alpha and transiently when stimulated with calcium agonists. The prolonged release of NO from TNF-alpha-stimulated endothelial cells may be implicated in the pathogenesis of septic shock.