Ellouk-Achard S, Mawet E, Thibault N, Dutertre-Catella H, Thevenin M, Claude J R
Université René Descartes--Paris V, Faculté de Pharmacie, Laboratoire de Toxicologie.
Drug Chem Toxicol. 1995 May-Aug;18(2-3):105-17. doi: 10.3109/01480549509014315.
Alteration of calcium homeostasis has been proposed to play a major role in cell necrosis induced by a variety of chemical agents such as acetaminophen (APAP). In this study, a potential protective effect of the dihydropyridine calcium channel blocking agent, nifedipine, was investigated in vitro on acetaminophen-induced hepatocyte damage. Rat hepatocytes were exposed during 20 hours to various concentrations of APAP (0.50 to 4.00 mM). The following metabolic and functional parameters were investigated: -lactate dehydrogenase (LDH) release as an indicator of plasma membrane integrity, -cell viability evaluated by the colorimetric MTT assay, and intracellular calcium concentration as evaluated by two fluorimetric methods: a scanning laser cytometer using indo-1-AM as fluorescent probe and a fluorescence plate reader using fluo-3-AM as calcium indicator. Incubation of hepatocytes with APAP alone in the range 0.50 to 4.00mM resulted in a dose-response relationship with regard to LDH release (243% to 750% of control) and to the loss of cell viability (0 to 67% of control). Moreover these results were correlated with a significant increase in cytosolic calcium content (189 to 406 nM). Nifedipine treatment prior to APAP exposure, partially prevented LDH release, the plasma membrane blebbing, and thereby the loss of viability. In addition, intracellular calcium level progressively returned within the limits of the control values with increasing concentrations of nifedipine. It can be concluded that, in vitro conditions, nifedipine pretreatment exhibits a preventive effect against acetaminophen hepatocyte injury.
钙稳态改变被认为在对乙酰氨基酚（APAP）等多种化学物质诱导的细胞坏死中起主要作用。在本研究中，体外研究了二氢吡啶类钙通道阻滞剂硝苯地平对APAP诱导的肝细胞损伤的潜在保护作用。大鼠肝细胞暴露于不同浓度的APAP（0.50至4.00 mM）中20小时。研究了以下代谢和功能参数：作为质膜完整性指标的乳酸脱氢酶（LDH）释放；通过比色MTT法评估的细胞活力；以及通过两种荧光法评估的细胞内钙浓度：一种使用indo-1-AM作为荧光探针的扫描激光细胞仪，另一种使用fluo-3-AM作为钙指示剂的荧光酶标仪。单独用0.50至4.00 mM范围内的APAP孵育肝细胞，导致LDH释放（对照的243%至750%）和细胞活力丧失（对照的0至67%）呈现剂量反应关系。此外，这些结果与胞质钙含量显著增加（189至406 nM）相关。在APAP暴露前用硝苯地平处理，部分预防了LDH释放、质膜泡状化，从而预防了细胞活力丧失。此外，随着硝苯地平浓度增加，细胞内钙水平逐渐恢复到对照值范围内。可以得出结论，在体外条件下，硝苯地平预处理对APAP诱导的肝细胞损伤具有预防作用。
