Woolbright Benjamin L, Dorko Kenneth, Antoine Daniel J, Clarke Joanna I, Gholami Parviz, Li Feng, Kumer Sean C, Schmitt Timothy M, Forster Jameson, Fan Fang, Jenkins Rosalind E, Park B Kevin, Hagenbuch Bruno, Olyaee Mojtaba, Jaeschke Hartmut
Department of Pharmacology, Toxicology & Therapeutics, University of Kansas Medical Center, Kansas City, KS, USA.
MRC Centre for Drug Safety Science, Department of Molecular and Clinical Pharmacology, Institute of Translational Medicine, University of Liverpool, Liverpool, UK.
Toxicol Appl Pharmacol. 2015 Mar 15;283(3):168-77. doi: 10.1016/j.taap.2015.01.015. Epub 2015 Jan 28.
Accumulation of bile acids is a major mediator of cholestatic liver injury. Recent studies indicate bile acid composition between humans and rodents is dramatically different, as humans have a higher percent of glycine conjugated bile acids and increased chenodeoxycholate content, which increases the hydrophobicity index of bile acids. This increase may lead to direct toxicity that kills hepatocytes, and promotes inflammation. To address this issue, this study assessed how pathophysiological concentrations of bile acids measured in cholestatic patients affected primary human hepatocytes. Individual bile acid levels were determined in serum and bile by UPLC/QTOFMS in patients with extrahepatic cholestasis with, or without, concurrent increases in serum transaminases. Bile acid levels increased in serum of patients with liver injury, while biliary levels decreased, implicating infarction of the biliary tracts. To assess bile acid-induced toxicity in man, primary human hepatocytes were treated with relevant concentrations, derived from patient data, of the model bile acid glycochenodeoxycholic acid (GCDC). Treatment with GCDC resulted in necrosis with no increase in apoptotic parameters. This was recapitulated by treatment with biliary bile acid concentrations, but not serum concentrations. Marked elevations in serum full-length cytokeratin-18, high mobility group box 1 protein (HMGB1), and acetylated HMGB1 confirmed inflammatory necrosis in injured patients; only modest elevations in caspase-cleaved cytokeratin-18 were observed. These data suggest human hepatocytes are more resistant to human-relevant bile acids than rodent hepatocytes, and die through necrosis when exposed to bile acids. These mechanisms of cholestasis in humans are fundamentally different to mechanisms observed in rodent models.
胆汁酸的蓄积是胆汁淤积性肝损伤的主要介质。最近的研究表明，人类和啮齿动物之间的胆汁酸组成存在显著差异，因为人类中甘氨酸结合胆汁酸的比例更高，鹅去氧胆酸盐含量增加，这增加了胆汁酸的疏水性指数。这种增加可能导致直接毒性，杀死肝细胞，并促进炎症。为了解决这个问题，本研究评估了胆汁淤积患者中测量的胆汁酸病理生理浓度如何影响原代人肝细胞。通过超高效液相色谱/四极杆飞行时间质谱法（UPLC/QTOFMS）测定了肝外胆汁淤积患者血清和胆汁中的个体胆汁酸水平，这些患者伴有或不伴有血清转氨酶同时升高。肝损伤患者血清中的胆汁酸水平升高，而胆汁中的水平降低，提示胆道梗死。为了评估人胆汁酸诱导的毒性，用源自患者数据的模型胆汁酸甘氨鹅去氧胆酸（GCDC）的相关浓度处理原代人肝细胞。用GCDC处理导致坏死，凋亡参数无增加。用胆汁胆汁酸浓度处理可重现这一结果，但血清浓度处理则不能。血清全长细胞角蛋白-18、高迁移率族蛋白B1（HMGB1）和乙酰化HMGB1的显著升高证实了受损患者存在炎症性坏死；仅观察到半胱天冬酶切割的细胞角蛋白-18有适度升高。这些数据表明，人肝细胞比啮齿动物肝细胞对与人类相关的胆汁酸更具抗性，并且在暴露于胆汁酸时通过坏死死亡。人类胆汁淤积的这些机制与在啮齿动物模型中观察到的机制根本不同。
