Sugishita K, Kinugawa K, Shimizu T, Harada K, Matsui H, Takahashi T, Serizawa T, Kohmoto O
Internal Medicine II, Faculty of Medicine, Japan.
J Mol Cell Cardiol. 1999 Aug;31(8):1457-67. doi: 10.1006/jmcc.1999.0989.
There is controversy over whether nitric oxide (NO) mediates acute negative inotropic actions of cytokines including tumor necrosis factor-alpha (TNF- alpha). The reports from established laboratories have appeared inconsistent, which could be due to species differences. Thus, we tried to elucidate the mechanisms underlying negative inotropic actions of interleukin-6 (IL-6) and TNF- alpha in the same model. We studied the effects of cytokines on Ca(2+)transients (using indo-1), cell shortening (CS) (using a video motion detector) and the L-type Ca(2+)channel current (I(Ca)) (using the whole cell perforated patch clamp technique) in isolated guinea-pig ventricular myocytes. IL-6 (1000 U/ml) or TNF- alpha (500 U/ml) decreased both peak systolic Ca(2+)(IL-6: 0.43+/-0.01 to 0.40+/-0.01, n=5, P<0.05; TNF- alpha : 0.42+/-0.02 to 0.39+/-0.02, n=5, P<0.05) and the amplitude of CS (IL-6: 7.5+/-0.9 to 6.2+/-0.5 micrometers, n=5, P<0.05; TNF- alpha : 6.7+/-0.7 to 5.8+/-0.7 micrometers, n=5, P<0.05) without detectable reductions in I(Ca)(IL-6: 0.9+/-0.1 to 0.9+/-0.1 nA, n=4, N.S.; TNF- alpha : 1.1+/-0.3 to 1.1+/-0.2 nA, n=4, N.S.) within 5 min. The nitric oxide synthase (NOS) inhibitor, N(G)-monomethyl- L arginine (300 micromol/l), blocked the effects of IL-6 but not of TNF- alpha. When pretreated with 20 nmol/l isoproterenol, exposure to IL-6 decreased both I(Ca)(2.8+/-0.5 to 2. 0+/-0.3 nA) and the amplitude of CS (10.4+/-2.4 to 7.5+/-1.9 micrometer) within 5 min. TNF- alpha also clearly depressed I(Ca)(2.9+/-0.9 to 2.3+/-0.7 nA) and the amplitude of CS (7.0+/-1.4 to 5.5+/-1.3 micrometer) in beta -adrenergic stimulated cells. TNF- alpha significantly increased the content of sphingosine (product of sphingomyelin pathway) in isolated heart. The effects of low dose sphingosine (5 micromol/l) mimicked those of TNF- alpha on cardiac myocytes. IL-6 produced an acute negative inotropic effect through a NO-dependent pathway while TNF- alpha did so via a sphingomyelin-dependent pathway in isolated guinea-pig ventricular myocytes.
关于一氧化氮（NO）是否介导包括肿瘤坏死因子-α（TNF-α）在内的细胞因子的急性负性肌力作用存在争议。知名实验室的报告似乎并不一致，这可能是由于物种差异所致。因此，我们试图在同一模型中阐明白细胞介素-6（IL-6）和TNF-α负性肌力作用的潜在机制。我们研究了细胞因子对豚鼠离体心室肌细胞中[Ca(2+)]i瞬变（使用indo-1）、细胞缩短（CS）（使用视频运动检测器）和L型Ca(2+)通道电流（I(Ca)）（使用全细胞穿孔膜片钳技术）的影响。IL-6（1000 U/ml）或TNF-α（500 U/ml）在5分钟内可降低收缩期峰值[Ca(2+)]i（IL-6：从0.43±0.01降至0.40±0.01，n = 5，P<0.05；TNF-α：从0.42±0.02降至0.39±0.02，n = 5，P<0.05）以及CS幅度（IL-6：从7.5±0.9降至6.2±0.5微米，n = 5，P<0.05；TNF-α：从6.7±0.7降至5.8±0.7微米，n = 5，P<0.05），而I(Ca)无明显降低（IL-6：从0.9±0.1降至0.9±0.1 nA，n = 4，无显著性差异；TNF-α：从1.1±0.3降至1.1±0.2 nA，n = 4，无显著性差异）。一氧化氮合酶（NOS）抑制剂N(G)-单甲基-L-精氨酸（300 μmol/l）可阻断IL-6的作用，但不能阻断TNF-α的作用。用20 nmol/l异丙肾上腺素预处理后，暴露于IL-6可在5分钟内降低I(Ca)（从2.8±0.5降至2.0±0.3 nA）和CS幅度（从10.4±2.4降至7.5±1.9微米）。TNF-α也可明显降低β-肾上腺素能刺激细胞中的I(Ca)（从2.9±0.9降至2.3±0.7 nA）和CS幅度（从7.0±1.4降至5.5±1.3微米）。TNF-α可显著增加离体心脏中鞘氨醇（鞘磷脂途径的产物）的含量。低剂量鞘氨醇（5 μmol/l）对心肌细胞的作用与TNF-α相似。在豚鼠离体心室肌细胞中，IL-6通过NO依赖途径产生急性负性肌力作用，而TNF-α则通过鞘磷脂依赖途径产生该作用。
