Rojanasakul Y, Ye J, Chen F, Wang L, Cheng N, Castranova V, Vallyathan V, Shi X
Department of Basic Pharmaceutical Sciences, West Virginia University, Health Sciences Center, Morgantown 26506, USA.
Mol Cell Biochem. 1999 Oct;200(1-2):119-25. doi: 10.1023/a:1007051402840.
Tumor necrosis factor alpha (TNFalpha) plays an important role in the pathogenesis of silicosis and other chronic inflammatory lung diseases. The present study investigates the role nuclear transcription factor kappaB (NF-kappaB) and oxygen free radicals in silica-induced TNFalpha production in primary alveolar macrophages and RAW 264.7 cells. Using electrophoretic mobility shift assay (EMSA) and enzyme-linked immunoadsorbent assay (ELISA), we have demonstrated that silica can induce NF-kappaB activation and TNFalpha expression in a dose-dependent manner. Transient transfection assays with a plasmid construct containing NF-kappaB binding sites linked to a reporter gene further show that silica is able to induce the transcriptional activation of NF-kappaB-dependent gene. Inhibition of NF-kappaB activation by SN50, a specific NF-kappaB blocker, abolishes silica-induced TNFalpha production. Pretreatment of the cells with catalase (H2O2 scavenger) or deferoxamine (*OH scavenger) effectively inhibits NF-kappaB and TNFalpha activation, whereas superoxide dismutase (O2 scavenger) has an opposite effect. These results indicate that silica-mediated free radical generation and NF-kappaB activation play important roles in silica-induced TNFalpha gene expression.
肿瘤坏死因子α（TNFα）在矽肺和其他慢性炎症性肺病的发病机制中起重要作用。本研究调查了核转录因子κB（NF-κB）和氧自由基在二氧化硅诱导原代肺泡巨噬细胞和RAW 264.7细胞产生TNFα中的作用。使用电泳迁移率变动分析（EMSA）和酶联免疫吸附测定（ELISA），我们已证明二氧化硅能以剂量依赖性方式诱导NF-κB激活和TNFα表达。用含有与报告基因相连的NF-κB结合位点的质粒构建体进行瞬时转染分析进一步表明，二氧化硅能够诱导NF-κB依赖性基因的转录激活。用特异性NF-κB阻滞剂SN50抑制NF-κB激活可消除二氧化硅诱导的TNFα产生。用过氧化氢酶（H2O2清除剂）或去铁胺（*OH清除剂）预处理细胞可有效抑制NF-κB和TNFα激活，而超氧化物歧化酶（O2清除剂）则有相反作用。这些结果表明，二氧化硅介导的自由基生成和NF-κB激活在二氧化硅诱导的TNFα基因表达中起重要作用。
