Solá Susana, Brito Maria A, Brites Dora, Moura José J G, Rodrigues Cecília M P
Centro de Patogénese Molecular, Faculdade de Farmácia, University of Lisbon, Lisbon, Av. das Forcas Armadas, Portugal.
Clin Sci (Lond). 2002 Nov;103(5):475-85. doi: 10.1042/cs1030475.
The accumulation of toxic bile salts within the hepatocyte plays a key role in organ injury during liver disease. Deoxycholate (DC) and glycochenodeoxycholate (GCDC) induce apoptosis in vitro and in vivo, perhaps through direct perturbation of mitochondrial membrane structure and function. In contrast, ursodeoxycholate (UDC) and its taurine-conjugated form (TUDC) appear to be protective. We show here that hydrophobic bile salts induced apoptosis in cultured rat hepatocytes, without modulating the expression of pro-apoptotic Bax protein, and caused cytochrome c release in isolated mitochondria. Co-incubation with UDC and TUDC prevented cell death and efflux of mitochondrial factors. Using spin-labelling techniques and EPR spectroscopy analysis of isolated rat liver mitochondria, we found significant structural changes at the membrane-water surface in mitochondria exposed to hydrophobic bile salts, including modified lipid polarity and fluidity, altered protein order and increased oxidative injury. UDC, TUDC and cyclosporin A almost completely abrogated DC- and GCDC-induced membrane perturbations. We conclude that the toxicity of hydrophobic bile salts to hepatocytes is mediated by cytochrome c release, through a mechanism associated with marked direct effects on mitochondrial membrane lipid polarity and fluidity, protein order and redox status, without modulation of pro-apoptotic Bax expression. UDC and TUDC can directly suppress disruption of mitochondrial membrane structure, which may represent an important mechanism of hepatoprotection by these bile salts.
肝细胞内有毒胆盐的蓄积在肝脏疾病期间的器官损伤中起关键作用。脱氧胆酸盐（DC）和甘氨鹅脱氧胆酸盐（GCDC）在体外和体内均可诱导细胞凋亡，可能是通过直接扰乱线粒体膜的结构和功能。相比之下，熊去氧胆酸盐（UDC）及其牛磺酸结合形式（TUDC）似乎具有保护作用。我们在此表明，疏水性胆盐可在培养的大鼠肝细胞中诱导细胞凋亡，而不调节促凋亡蛋白Bax的表达，并导致分离线粒体中细胞色素c的释放。与UDC和TUDC共同孵育可防止细胞死亡和线粒体因子的外流。使用自旋标记技术和对分离的大鼠肝线粒体进行电子顺磁共振光谱分析，我们发现在暴露于疏水性胆盐的线粒体中，膜-水表面发生了显著的结构变化，包括脂质极性和流动性的改变、蛋白质有序性的改变以及氧化损伤的增加。UDC、TUDC和环孢素A几乎完全消除了DC和GCDC诱导的膜扰动。我们得出结论，疏水性胆盐对肝细胞的毒性是通过细胞色素c的释放介导的，其机制与对线粒体膜脂质极性和流动性、蛋白质有序性和氧化还原状态的显著直接影响相关，而不调节促凋亡蛋白Bax的表达。UDC和TUDC可直接抑制线粒体膜结构的破坏，这可能是这些胆盐发挥肝保护作用的重要机制。
