Muñoz C, Pascual-Salcedo D, Castellanos M C, Alfranca A, Aragonés J, Vara A, Redondo J M, de Landázuri M O
Servicio de Inmunología, Hospital de la Princesa, Universidad Autónoma de Madrid, Spain.
Blood. 1996 Nov 1;88(9):3482-90.
Endothelial cells (EC) play a key role in the inflammatory response, both by the production of proinflammatory cytokines and by their interaction with leukocytes. Molecular genetic analysis has demonstrated that functional NF-kappa B sites are involved in the transcription of interleukin-6 (IL-6), IL-8, and granulocyte-macrophage colony-stimulating factor (GM-CSF) genes in response to inflammatory mediators. Thus, we have explored the effect of two inhibitors of the NF-kappa B activation, pyrrolidine dithiocarbamate (PDTC) and N-acetylcysteine (NAC), on the production of these cytokines by EC. Both PDTC and NAC inhibited, in a dose-dependent manner, the synthesis of IL-6, IL-8, and GM-CSF induced by tumor necrosis factor (TNF)-alpha or bacterial lipopolysaccharides (LPS) in human umbilical vein endothelial cells (HUVEC). PDTC appeared to prevent IL-6, IL-8, and GM-CSF gene transcription, as it blocked the induction of specific mRNA by TNF-alpha or LPS. The TNF-alpha mediated transcriptional activation of a chloramphenicol acetyltransferase (CAT) plasmid containing three copies of the -72 kappa B binding site from the IL-6 promoter was abrogated by PDTC. According to transfection experiments, electrophoretic mobility shift assays (EMSA) demonstrated that the antioxidant prevented the induction of NF-kappa B DNA-binding activity by TNF-alpha. Under the same conditions, PDTC by itself or in combination with TNF-alpha, enhanced the DNA-binding activity of AP-1, as well as c-fos and c-jun mRNA levels. Altogether, these results indicate that the antioxidant PDTC specifically inhibits the transcription of IL-6, IL-8, and GM-CSF genes through the inhibition of the NF-kappa B activation, while increasing the expression of AP-1. Our data make evident the antiinflammatory and immunoregulatory potential of the pharmacological inhibition of the NF-kappa B activation. In addition, PDTC and related molecules may be a useful tool to explore the expression of genes involved in the inflammatory response.
内皮细胞（EC）在炎症反应中起关键作用，这既体现在促炎细胞因子的产生上，也体现在它们与白细胞的相互作用上。分子遗传学分析表明，功能性核因子-κB（NF-κB）位点参与白细胞介素-6（IL-6）、IL-8和粒细胞-巨噬细胞集落刺激因子（GM-CSF）基因对炎症介质的转录反应。因此，我们探讨了两种NF-κB激活抑制剂，吡咯烷二硫代氨基甲酸盐（PDTC）和N-乙酰半胱氨酸（NAC），对内皮细胞产生这些细胞因子的影响。PDTC和NAC均以剂量依赖性方式抑制人脐静脉内皮细胞（HUVEC）中由肿瘤坏死因子（TNF）-α或细菌脂多糖（LPS）诱导的IL-6、IL-8和GM-CSF的合成。PDTC似乎能阻止IL-6、IL-8和GM-CSF基因的转录，因为它能阻断TNF-α或LPS对特异性mRNA的诱导。PDTC消除了TNF-α介导的含有来自IL-6启动子的三个-72 κB结合位点拷贝的氯霉素乙酰转移酶（CAT）质粒的转录激活。根据转染实验，电泳迁移率变动分析（EMSA）表明，这种抗氧化剂可阻止TNF-α诱导的NF-κB DNA结合活性。在相同条件下，PDTC单独或与TNF-α联合使用时，可增强活化蛋白-1（AP-1）的DNA结合活性以及c-fos和c-jun mRNA水平。总之，这些结果表明，抗氧化剂PDTC通过抑制NF-κB激活特异性抑制IL-6、IL-8和GM-CSF基因的转录，同时增加AP-1的表达。我们的数据表明了对NF-κB激活进行药理学抑制的抗炎和免疫调节潜力。此外，PDTC及相关分子可能是探索参与炎症反应的基因表达的有用工具。
