Davila J C, Davis P J, Acosta D
Department of Pharmacology and Toxicology, College of Pharmacy, University of Texas, Austin 78712-1074.
Toxicol Appl Pharmacol. 1991 Mar 15;108(1):28-36. doi: 10.1016/0041-008x(91)90265-g.
The purpose of this study was to elucidate the mechanism of hepatotoxicity of papaverine hydrochloride (papaver) in vitro. To evaluate the role of metabolism in the toxicity of papaver, cells were pretreated with SKF-525A or benzyl imidazole (cytochrome P450 system inhibitors) for 24 hr at 1 x 10(-5) or 1 x 10(-4) M, respectively, or with phenobarbital sodium (cytochrome P450 system inducer) for 3 days at 2 x 10(-3) M. Cells then were exposed to concentrations of papaver ranging from 1 x 10(-5) to 1 x 10(-3) M for 4 to 24 hr. Cytotoxicity was evaluated by enzyme leakage (lactate dehydrogenase) and by energy status of the cells (ATP/ADP). The role of biological reactive intermediates in the toxicity of papaver was investigated by measuring changes in cellular reduced glutathione levels (GSH), by inhibiting GSH synthesis, and by determining the production of lipid peroxidation (LPX). Papaverine produced concentration- and time-dependent increases in enzyme leakage, with significant effects occurring by the 8-hr exposure period. Pretreatment with SKF-525A or benzyl imidazole increased enzyme leakage induced by papaver especially at a later time frame (24 hr), but pretreatment with phenobarbital delayed the onset of cytotoxicity from 8 to 12 hr. Decreases in GSH levels paralleled the time course of enzyme leakage. However, the administration of buthionine sulfoximine to cell cultures dramatically decreased the time by which papaver induced cellular injury (2 hr vs 8 hr). Changes in cellular energy status (ATP/ADP) were also detected earlier than enzyme leakage (4 hr vs 8 hr). In contrast, no significant production of lipid peroxidation was noted in papaver-treated cultures. We suggest that the mechanism of papaver-induced hepatotoxicity may be related to alterations in glutathione balance of the cells and to disruption of energy homeostasis.
本研究的目的是阐明盐酸罂粟碱（罂粟碱）在体外的肝毒性机制。为了评估代谢在罂粟碱毒性中的作用，细胞分别用SKF - 525A或苄基咪唑（细胞色素P450系统抑制剂）以1×10⁻⁵或1×10⁻⁴M预处理24小时，或用苯巴比妥钠（细胞色素P450系统诱导剂）以2×10⁻³M预处理3天。然后将细胞暴露于浓度范围为1×10⁻⁵至1×10⁻³M的罂粟碱中4至24小时。通过酶泄漏（乳酸脱氢酶）和细胞能量状态（ATP/ADP）评估细胞毒性。通过测量细胞内还原型谷胱甘肽水平（GSH）的变化、抑制GSH合成以及测定脂质过氧化（LPX）的产生来研究生物活性中间体在罂粟碱毒性中的作用。罂粟碱导致酶泄漏呈浓度和时间依赖性增加，在暴露8小时时出现显著影响。用SKF - 525A或苄基咪唑预处理可增加罂粟碱诱导的酶泄漏，尤其是在较晚的时间点（24小时），但用苯巴比妥预处理可将细胞毒性的发作从8小时延迟至12小时。GSH水平的降低与酶泄漏的时间进程平行。然而，向细胞培养物中加入丁硫氨酸亚砜胺可显著缩短罂粟碱诱导细胞损伤的时间（2小时对8小时）。细胞能量状态（ATP/ADP）的变化也比酶泄漏更早被检测到（4小时对8小时）。相比之下，在罂粟碱处理的培养物中未观察到显著的脂质过氧化产生。我们认为，罂粟碱诱导的肝毒性机制可能与细胞内谷胱甘肽平衡的改变以及能量稳态的破坏有关。
