Shindo T, Ikeda U, Ohkawa F, Kawahara Y, Yokoyama M, Shimada K
Department of Cardiology, Jichi Medical School, Minamikawachi, Tochigi, Japan.
Cardiovasc Res. 1995 Jun;29(6):813-9.
The aim was to investigate nitric oxide (NO) synthase activity in cultured neonatal rat cardiac myocytes and fibroblasts upon treatment with inflammatory cytokines interleukin 1 beta (IL-1 beta), tumour necrosis factor alpha (TNF-alpha), IL-2, IL-6, IL-8, transforming growth factor beta (TGF-beta) and gram negative bacterial lipopolysaccharide (LPS).
NO and guanosine 3',5'-cyclic monophosphate (cGMP) synthesis was measured in cultured neonatal rat cardiac myocytes and fibroblasts, using Griess reagent and an enzyme immunoassay kit, respectively. The expression of inducible NO synthase (iNOS) mRNA and protein was assayed by northern and western blotting, respectively.
Incubation of cardiac myocytes for 24 h with IL-1 beta (10 ng.ml-1) or LPS (1 microgram.ml-1) caused significant increases in NO and cGMP production. TNF-alpha, IL-2, IL-6, IL-8, and TGF-beta showed no significant effect on their production. IL-1 beta induced NO and cGMP production in a time and dose dependent manner. IL-1 beta also increased iNOS mRNA and protein accumulation in cardiac myocytes. Simultaneous incubation of IL-1 beta with NG-monomethyl-L-arginine, genistein, calphostin C, cycloheximide, or actinomycin D completely inhibited the IL-1 beta induced NO production by cardiac myocytes. TGF-beta, dexamethasone, or cyclosporin A also dose dependently inhibited the IL-1 beta induced NO production. Exposure to IL-1 beta for 12-24 h decreased the beating rate of cardiac myocytes, but addition of dexamethasone completely overcame this inhibition. In contrast to cardiac myocytes, incubation of cardiac fibroblasts for 24 h with IL-1 beta or LPS showed no significant effect on NO or cGMP production.
These observations suggest that IL-1 beta/LPS responsive iNOS, which is an important regulator of contractile function of the heart, is present in cardiac myocytes but not in cardiac fibroblasts.
研究炎性细胞因子白细胞介素1β（IL-1β）、肿瘤坏死因子α（TNF-α）、IL-2、IL-6、IL-8、转化生长因子β（TGF-β）以及革兰氏阴性菌脂多糖（LPS）处理后，培养的新生大鼠心肌细胞和成纤维细胞中一氧化氮（NO）合酶的活性。
分别使用格里斯试剂和酶免疫分析试剂盒，测定培养的新生大鼠心肌细胞和成纤维细胞中NO和鸟苷3',5'-环磷酸（cGMP）的合成。分别通过Northern印迹和Western印迹检测诱导型NO合酶（iNOS）mRNA和蛋白的表达。
心肌细胞与IL-1β（10 ng.ml-1）或LPS（1 μg.ml-1）孵育24小时后，NO和cGMP的生成显著增加。TNF-α、IL-2、IL-6、IL-8和TGF-β对它们的生成无显著影响。IL-1β以时间和剂量依赖性方式诱导NO和cGMP的生成。IL-1β还增加了心肌细胞中iNOS mRNA和蛋白的积累。IL-1β与NG-单甲基-L-精氨酸、染料木黄酮、钙磷蛋白C、环己酰亚胺或放线菌素D同时孵育，可完全抑制IL-1β诱导的心肌细胞NO生成。TGF-β、地塞米松或环孢素A也以剂量依赖性方式抑制IL-1β诱导的NO生成。暴露于IL-1β 12 - 24小时会降低心肌细胞的搏动率，但加入地塞米松可完全克服这种抑制作用。与心肌细胞不同，心肌成纤维细胞与IL-1β或LPS孵育24小时后，对NO或cGMP的生成无显著影响。
这些观察结果表明，IL-1β/LPS反应性iNOS存在于心肌细胞而非心肌成纤维细胞中，它是心脏收缩功能的重要调节因子。
