Lu S C, Ge J L, Huang H Y, Kuhlenkamp J, Kaplowitz N
Department of Medicine, University of Southern California School of Medicine, Los Angeles 90033.
J Clin Invest. 1993 Sep;92(3):1188-97. doi: 10.1172/JCI116689.
In cultured rat hepatocytes, cystine led to an inhibition of GSH efflux by lowering the Vmax by approximately 35% without affecting the Km. The cystine-mediated inhibition of GSH efflux was rapid in onset (< 1 h), with near maximum effect at 0.1 mM. Inhibition was still observed when cystine uptake was prevented. Cystine and sulfobromophthalein-GSH, a selective inhibitor of sinusoidal transport of GSH, did not exhibit additive inhibitory effects on GSH efflux. Depletion of ATP or membrane depolarization after cystine treatment were excluded as potential mechanisms. DTT not only reversed the cystine-mediated inhibition of GSH efflux, it stimulated GSH efflux up to 400-500%. The DTT effect was immediate in onset, reaching maximum after 30 min, and was partially reversed by cystine, suggesting that the two share a common site(s) of action. DTT treatment did not alter cellular ATP levels or change the membrane potential. In cultured hepatocytes, DTT treatment increased the Vmax of GSH efflux by approximately 500% without affecting the Km. Inhibition of microtubular function and vesicular acidification did not affect basal or DTT stimulated efflux. Both cystine and DTT effects on sinusoidal GSH efflux were confirmed in perfused livers. In summary, the capacity of the sinusoidal GSH transporter is markedly influenced by thiol-disulfide status.
在培养的大鼠肝细胞中，胱氨酸通过将最大反应速度（Vmax）降低约35%而不影响米氏常数（Km），从而抑制谷胱甘肽（GSH）外排。胱氨酸介导的GSH外排抑制起效迅速（<1小时），在0.1 mM时接近最大效应。当胱氨酸摄取被阻止时，仍可观察到抑制作用。胱氨酸和磺溴酞钠-GSH（一种GSH窦状转运的选择性抑制剂）对GSH外排未表现出相加抑制作用。排除了胱氨酸处理后ATP耗竭或膜去极化作为潜在机制。二硫苏糖醇（DTT）不仅逆转了胱氨酸介导的GSH外排抑制，还将GSH外排刺激高达400 - 500%。DTT的作用起效迅速，30分钟后达到最大值，且部分被胱氨酸逆转，这表明二者共享一个共同的作用位点。DTT处理未改变细胞ATP水平或膜电位。在培养的肝细胞中，DTT处理使GSH外排的Vmax增加约500%而不影响Km。微管功能抑制和囊泡酸化抑制均不影响基础或DTT刺激的外排。胱氨酸和DTT对窦状GSH外排的作用在灌注肝脏中得到证实。总之，窦状GSH转运体的能力受硫醇-二硫键状态的显著影响。
