Bezdecny Steven A, Karmaus Peer, Roth Robert A, Ganey Patricia E
Department of Pharmacology and Toxicology, Center for Integrative Toxicology and National Food Safety and Toxicology Center, Michigan State University, East Lansing, MI 48824, USA.
Toxicol Appl Pharmacol. 2007 Jun 15;221(3):285-94. doi: 10.1016/j.taap.2007.03.019. Epub 2007 Mar 30.
Polychlorinated biphenyls (PCBs) are ubiquitous, persistent environmental contaminants that affect a number of cellular systems, including neutrophils. Among the effects caused by the noncoplanar PCB 2,2',4,4'-tetrachlorobiphenyl (2244-TCB) in granulocytic HL-60 cells are increases in superoxide anion production, activation of phospholipase A(2) with subsequent release of arachidonic acid (AA) and upregulation of the inflammatory gene cyclooxygenase-2 (COX-2). The objective of this study was to determine the signal transduction pathways involved in the upregulation of COX-2 by 2244-TCB. Treatment of HL-60 cells with 2244-TCB led to increased expression of COX-2 mRNA. This increase was prevented by the transcriptional inhibitor actinomycin D in cells pretreated with 2244-TCB for 10 min. The increase in COX-2 mRNA was associated with release of (3)H-AA, phosphorylation of p38 and extracellular signal-regulated kinase (ERK) mitogen-activated protein (MAP) kinases, increased levels of nuclear NF-kappaB and increased superoxide anion production. Bromoenol lactone, an inhibitor of the calcium-independent phospholipase A(2), reduced (3)H-AA release but had no effect on COX-2 mRNA, protein or activity. Pretreatment with SB-202190 or SB-203580, inhibitors of the p38 MAP kinase pathway, prevented the 2244-TCB-mediated induction of COX-2 and phosphorylation of p38 and ERK MAP kinases. These inhibitors did not alter (3)H-AA release. Treatment with PD 98059 or U 0126, inhibitors of the MAP/ERK (MEK) pathway, prevented the 2244-TCB-mediated activation of ERK but had no effect on COX-2 induction or p38 phosphorylation. 2244-TCB treatment did not affect c-Jun N-terminal kinase (JNK) phosphorylation. 2244-TCB exposure increased the amount of nuclear NF-kappaB. This increase was prevented by pretreatment with p38 MAP kinase inhibitors, but not by pretreatment with MEK inhibitors. Pretreatment with inhibitors of NF-kappaB prevented the 2244-TCB-mediated induction of COX-2 mRNA. 2244-TCB-mediated increases in superoxide anion were prevented by the NADPH oxidase inhibitor apocynin or the free radical scavenger 4-hydroxy TEMPO, but neither of these inhibitors affected the 2244-TCB-induced changes in COX-2 mRNA levels or (3)H-AA release. Taken together these data suggest that p38 MAP kinase-dependent activation of NF-kappaB is critical for the 2244-TCB-mediated upregulation of COX-2 mRNA.
多氯联苯（PCBs）是普遍存在且持久的环境污染物，会影响包括中性粒细胞在内的多种细胞系统。在粒细胞性HL-60细胞中，非共面多氯联苯2,2',4,4'-四氯联苯（2244-TCB）所产生的影响包括超氧阴离子生成增加、磷脂酶A2激活并随后释放花生四烯酸（AA）以及炎症基因环氧合酶-2（COX-2）上调。本研究的目的是确定2244-TCB上调COX-2所涉及的信号转导途径。用2244-TCB处理HL-60细胞会导致COX-2 mRNA表达增加。在用2244-TCB预处理10分钟的细胞中，转录抑制剂放线菌素D可阻止这种增加。COX-2 mRNA的增加与（3）H-AA的释放、p38和细胞外信号调节激酶（ERK）丝裂原活化蛋白（MAP）激酶的磷酸化、核NF-κB水平升高以及超氧阴离子生成增加有关。钙非依赖性磷脂酶A2的抑制剂溴烯醇内酯可减少（3）H-AA的释放，但对COX-2 mRNA、蛋白或活性没有影响。用p38 MAP激酶途径的抑制剂SB-202190或SB-203580预处理可阻止2244-TCB介导的COX-2诱导以及p38和ERK MAP激酶的磷酸化。这些抑制剂不会改变（3）H-AA的释放。用MAP/ERK（MEK）途径的抑制剂PD 98059或U 0126处理可阻止2244-TCB介导的ERK激活，但对COX-2诱导或p38磷酸化没有影响。2244-TCB处理不影响c-Jun N端激酶（JNK）的磷酸化。2244-TCB暴露会增加核NF-κB的量。用p38 MAP激酶抑制剂预处理可阻止这种增加，但用MEK抑制剂预处理则不能。用NF-κB抑制剂预处理可阻止2244-TCB介导的COX-2 mRNA诱导。NADPH氧化酶抑制剂夹竹桃麻素或自由基清除剂4-羟基TEMPO可阻止2244-TCB介导的超氧阴离子增加，但这两种抑制剂均不影响2244-TCB诱导的COX-2 mRNA水平变化或（3）H-AA释放。综上所述，这些数据表明p38 MAP激酶依赖性的NF-κB激活对于2244-TCB介导的COX-2 mRNA上调至关重要。
