Interleukin-1β regulation of inducible nitric oxide synthase and cyclooxygenase-2 involves the p42/44 and p38 MAPK signaling pathways in cardiac myocytes
Interleukin-1β regulation of inducible nitric oxide synthase and cyclooxygenase-2 involves the p42/44 and p38 MAPK signaling pathways in cardiac myocytes
复制标题
DOI：
10.1161/01.hyp.33.1.276
复制
发表时间：
1999-01-01
期刊：
影响因子：
8.3
通讯作者：
Isenovic, E
中科院分区：
文献类型：
--
作者：
LaPointe, MC;Isenovic, E
编码诱导型一氧化氮合酶（iNOS）和环氧合酶-2（考克斯-2，也称为胰蛋白酶-内过氧化物合酶-2）的基因在许多类型的细胞中响应于促炎细胞因子而被诱导。我们先前已经表明白细胞介素-1 β（IL）刺激心肌细胞中的iNOS和考克斯-2 mRNA。由于IL在许多不同的细胞中激活丝裂原活化蛋白激酶（MAPK）信号通路，我们使用p42/44激活的特异性抑制剂PD 98059（PD）和p38抑制剂SB 205380（SB）测试p42/44和p38 MAPK通路是否参与IL刺激iNOS和考克斯-2。使用Griess试剂测量亚硝酸盐，使用酶免疫法测量前列腺素PGE（2），使用Western印迹分析测量iNOS和考克斯-2蛋白，使用北方印迹分析测量iNOS mRNA：单独测试，p38激酶和MAPK抑制剂部分降低IL对亚硝酸盐、iNOS蛋白和iNOS mRNA的刺激;一起使用，它们完全消除IL的作用。SE和PD分别抑制IL-刺激的考克斯-2蛋白的60%和80%，IL-刺激的考克斯-2蛋白被抑制剂的组合完全阻止。单独使用任何一种药物均可抑制PGE（2）的产生超过99%，这表明对酶活性存在翻译后效应。为了测试这种翻译后效应是否涉及胞浆磷脂酶A（2）（cPLA（2））亚型，对cPLA（2）蛋白进行了蛋白质印迹检测。结果表明，IL刺激cPLA（2）的活性和合成，SE可抑制cPLA（2）的活性和合成，PD则不抑制cPLA（2）的活性和合成。这些结果表明：（1）IL诱导iNOS合成依赖于p42/44和p38两条信号通路，主要在转录水平起作用;和（2）IL对考克斯-2合成的调节涉及p42/44和p38信号传导途径，另外的调节水平发生在产后，可能在cPLA（2）同种型的活化水平，其可能参与细胞内信号传导以及花生四烯酸释放对考克斯-2活性的调节。
The genes encoding inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2, also known as prostaglandin-endoperoxide synthase-2) are induced in many types of cells in response to proinflammatory cytokines, We have previously shown that interleukin-1 beta (IL) stimulates iNOS and COX-2 mRNA in cardiac myocytes. Because IL has been shown to activate mitogen-activated protein kinase (MAPK) signaling pathways in many different cells, we tested whether the p42/44 and p38 MAPK pathways were involved in IL stimulation of iNOS and COX-2, using a specific inhibitor of p42/44 activation, PD98059 (PD), and the p38 inhibitor SB205380 (SB). Nitrites were measured using the Griess reagent, prostaglandin PGE(2) by an enzyme immunoassay, iNOS and COX-2 protein by Western blot analysis, and iNOS mRNA by Northern blot analysis: Tested separately, the p38 kinase and MAPK inhibitors partially reduced IL stimulation of nitrite, iNOS protein, and iNOS mRNA; used together, they completely abolished the effect of IL. SE and PD inhibited IL-stimulated COX-2 protein by 60% and 80%, respectively, and IL-stimulated COX-2 protein was totally prevented by the combination of inhibitors. PGE(2) production was inhibited more than 99% by either drug alone, suggesting a posttranslational effect on enzyme activity, To test whether this posttranslational effect involved the cytosolic phospholipase A(2) (cPLA(2)) isoform, Western blots were probed for cPLA(2) protein. Results indicated that IL stimulated cPLA(2) activity and synthesis, which was inhibited by SE but not PD, These data indicate that (1) IL induction of iNOS synthesis depends on both the p42/44 and p38 signaling pathways, acting primarily at the level of transcriptional regulation; and (2) IL regulation of COX-2 synthesis involves the p42/44 and p38 signaling pathways, with an additional level of regulation occurring posttranslationally, perhaps at the level of activation of the cPLA(2) isoform, which may be involved in intracellular signaling, as well as regulation of arachidonic acid release for COX-2 activity.