Tepperman B L, Soper B D
Department of Physiology, Faculty of Medicine, University of Western Ontario, London, Canada.
Eur J Pharmacol. 1995 Oct 6;293(3):259-66. doi: 10.1016/0926-6917(95)00027-5.
Pertubations in cellular Ca2+ homeostasis can lead to oxidative stress whereas nitric oxide has been shown to inactivate oxygen radicals. Therefore the effects of inhibition of nitric oxide (NO) synthase activity on Ca2+-mediated disruption to rabbit dispersed gastric mucosal cells have been examined. Addition of the Ca2+ ionophore A23187 (3-25 microM) to the incubation medium induced a concentration-dependent increase in cell damage is assessed by trypan blue dye uptake and decreased cellular metabolic activity as estimated by alamar blue absorbance. These responses were exacerbated by inhibition of NO synthase activity with NG-monomethyl-L-arginine (300 microM). The deleterious effects of ionophore A23187 and NG-monomethyl-L-arginine were ameliorated by addition of the NO donor S-nitroso-acetyl-penicillamine to the cell suspension. An increase in cellular Ca2+ in response to ionophore A23187 (12.5 microM) resulted in enhanced 2'7'-dichlorofluorescein fluorescence suggesting an elevation in oxidative stress. Ca2+-mediated cell injury was abolished by the oxygen radical scavengers, catalase and 2',2'-dipyridyl. However, the cytotoxic effect of combined treatment with A23187 and NG-monomethyl-L-arginine was not reduced by administration of oxygen radical scavengers. NG-monomethyl-L-arginine treatment exacerbated the increase in cytosolic Ca2+ in response to ionophore A23187 as assessed by indo-1 fluorescence. Furthermore this increase in cytosolic Ca2+ was reduced by addition of S-nitroso-acetyl-penicillamine to the incubation medium. These data suggest that NO synthase inhibition in gastric mucosal cells exacerbates the damaging actions of the Ca2+ ionophore A23187. The increase in cell damage in response to the NO synthase inhibitor NG-monomethyl-L-arginine does not appear to be mediated by an increase in oxidative stress and may be associated in part with changes in cellular Ca2+ flux.
细胞内钙离子稳态的紊乱可导致氧化应激，而一氧化氮已被证明可使氧自由基失活。因此，研究了抑制一氧化氮（NO）合酶活性对钙离子介导的兔分散胃黏膜细胞破坏的影响。向孵育培养基中添加钙离子载体A23187（3 - 25 microM）会导致细胞损伤呈浓度依赖性增加（通过台盼蓝染料摄取评估），并降低细胞代谢活性（通过alamar蓝吸光度估计）。用NG - 单甲基 - L - 精氨酸（300 microM）抑制NO合酶活性会加剧这些反应。向细胞悬液中添加NO供体S - 亚硝基 - 乙酰青霉胺可改善离子载体A23187和NG - 单甲基 - L - 精氨酸的有害作用。响应离子载体A23187（12.5 microM），细胞内钙离子增加导致2',7' - 二氯荧光素荧光增强，表明氧化应激升高。钙离子介导的细胞损伤被氧自由基清除剂过氧化氢酶和2',2' - 二吡啶消除。然而，氧自由基清除剂的给药并未降低A23187和NG - 单甲基 - L - 精氨酸联合处理的细胞毒性作用。通过indo - 1荧光评估，NG - 单甲基 - L - 精氨酸处理加剧了响应离子载体A23187时胞质钙离子的增加。此外，向孵育培养基中添加S - 亚硝基 - 乙酰青霉胺可降低这种胞质钙离子的增加。这些数据表明，胃黏膜细胞中NO合酶的抑制会加剧钙离子载体A23187的破坏作用。响应NO合酶抑制剂NG - 单甲基 - L - 精氨酸时细胞损伤的增加似乎不是由氧化应激增加介导的，可能部分与细胞钙离子通量的变化有关。
