Catecholamines decrease nitric oxide production by cytokine-stimulated hepatocytes
Catecholamines decrease nitric oxide production by cytokine-stimulated hepatocytes
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DOI：
10.1067/msy.2001.115900
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发表时间：
2001-08-01
期刊：
影响因子：
3.8
通讯作者：
Billiar, TR
中科院分区：
文献类型：
--
作者：
Collins, JL;Vodovotz, Y;Billiar, TR
背景儿茶酚胺在炎症反应中显著升高，并在脓毒症中起调节作用。一氧化氮（NO）也是脓毒症中的关键炎症介质，由肝脏中的诱导型一氧化氮合酶（iNOS）大量产生。本研究的目的是验证儿茶酚胺在肝细胞NO产生的调节中起作用的假说。从健康雄性Sprague-Dawley大鼠中分离原代肝细胞，并在存在或不存在不同浓度的肾上腺素或去甲肾上腺素的情况下，用正常培养基培养或用cytomix（白细胞介素-1 β、干扰素-γ和肿瘤坏死因子-α）刺激。处理后6小时分离总RNA，并通过北方印迹分析iNOS mRNA。在12小时获得蛋白质提取物，并通过Western免疫印迹分析iNOS。在24小时时分析细胞培养上清液中的NO，确定为稳定的终产物NO2-。肾上腺素和去甲肾上腺素显着降低刺激肝细胞中的NO2-水平，但对iNOS mRNA或蛋白水平没有影响。腺苷酸环化酶刺激剂毛喉素可复制NO2-的减少。蛋白激酶A抑制剂Rp-8-Br-环磷酸腺苷可完全逆转儿茶酚胺诱导的NO2-减少。儿茶酚胺减少肝细胞对细胞因子刺激的NO产生。这种作用似乎是由于翻译后事件，似乎是介导的环磷酸腺苷的一部分。
Background. Catecholamines are significantly elevated in inflammatory responses and play a regulatory role in sepsis. Nitric oxide (NO), also a key inflammatory mediator in sepsis, is produced in large amounts by the inducible nitric oxide synthase (iNOS) in the liver The purpose of this study was to test the hypothesis that catecholamines play a role in the regulation of NO production by hepatocytes.Methods. Primary hepatocytes were isolated from healthy male Sprague-Dawley rats and either cultured with normal medium or stimulated with cytomix (interleukin-1 beta, interferon-gamma, and tumor necrosis factor-alpha) in the presence or absence of epinephrine or norepinephrine at varying concentrations. Total RNA was isolated 6 hours after treatment and analyzed by Northern blotting for iNOS mRNA. Protein extracts were obtained at 12 hours and were analyzed by Western immunoblotting for iNOS. Cell culture supernatants were analyzed for NO, determined as the stable end-product NO2-, at 24 hours.Results. Epinephrine and norepinephrine significantly decreased NO2- levels in stimulated hepatocytes but had no effect on iNOS mRNA or protein levels. Ae decrease in NO2- was reproduced by the adenylate cyclase stimulator, forskolin. The catecholamine-induced decrease in NO2- was completely reversed by the protein kinase A inhibitor Rp-8-Br-cyclic adenosine monophosphate.Conclusions. Catecholamines decrease hepatocyte Production of NO in response to cytokine stimulation. This effect seems to be due to post-translational events and appears to be mediated in part by cyclic adenosine monophosphate.