USDA, ARS, Animal and Natural Resources Institute, Animal Biosciences and Biotechnology Laboratory, Beltsville Agricultural Research Center, Bldg. 200, Rm. 13, BARC-East, Beltsville, MD 20705, USA.
In Vitro Cell Dev Biol Anim. 2010 Jan;46(1):11-9. doi: 10.1007/s11626-009-9244-z.
Liver in vitro models are needed to replace animal models for rapid assessment of drug biotransformation and toxicity. The PICM-19 pig liver stem cell line may fulfill this need since these cells have activities associated with xenobiotic phase I and II metabolism lacking in other liver cell lines. The objective of this study was to characterize phase I and II metabolic functions of a PICM-19 derivative cell line, PICM-19H, compared to the tumor-derived human HepG2 C3A cell line and primary cultures of adult porcine hepatocytes. Following exposure of PICM-19H cells to either 3-methylcholanthrene, rifampicin or phenobarbital, the induced activities of cytochrome P450 (CYP450) isozymes CYP-1A, -2, and-3A were assessed. Relative to adult porcine hepatocytes, PICM- 19H cells exhibited 30% and 43%, respectively, of CYP1A and 3A activities, while HepG2 C3A cells exhibited 7% and 0% of those activities. Fluorescent metabolites were extensively conjugated, i.e., 52% and 96% of CYP450-1A and-3A metabolites were released from medium samples following treatment with β-glucuronidase/arylsulfatase. Rifampicin induction of CYP450 isozyme activities was confirmed by conversion of testosterone to 6β-OH-, 2α-OH- and 2β-OH-testosterone, as determined by mass spectrometry. Susceptibility of PICM-19H cells to acetaminophen toxicity was determined; CD50 was calculated to be 14.9±0.9 mM. Toxicity and bioactivation of aflatoxin B1 was determined in 3-methylcholanthrenetreated cultures and untreated controls; CD50 were 1.59 μM and 31 μM, respectively. These results demonstrate the potential use of PICM-19H cells in drug biotransformation and toxicity testing and further support their use in extracorporeal artificial liver device technology.
需要使用体外肝脏模型来替代动物模型，以便快速评估药物的生物转化和毒性。PICM-19 猪肝干细胞系可能满足这一需求，因为这些细胞具有与异生物质相 I 和 II 代谢相关的活性，而其他肝细系缺乏这些活性。本研究的目的是比较 PICM-19 衍生细胞系 PICM-19H 与肿瘤衍生的人 HepG2 C3A 细胞系和成年猪原代肝细胞的相 I 和 II 代谢功能。在 PICM-19H 细胞暴露于 3-甲基胆蒽、利福平或苯巴比妥后，评估细胞色素 P450（CYP450）同工酶 CYP-1A、-2 和-3A 的诱导活性。与成年猪肝细胞相比，PICM-19H 细胞分别显示 CYP1A 和 3A 活性的 30%和 43%，而 HepG2 C3A 细胞分别显示 7%和 0%的活性。荧光代谢物被广泛共轭，即在用β-葡糖苷酸酶/芳基硫酸酯酶处理后，CYP450-1A 和-3A 代谢物中有 52%和 96%从培养基样品中释放。通过用质谱法测定睾酮转化为 6β-OH-、2α-OH-和 2β-OH-睾酮，证实利福平诱导 CYP450 同工酶活性。测定 PICM-19H 细胞对乙酰氨基酚毒性的敏感性；CD50 计算为 14.9±0.9 mM。在 3-甲基胆蒽处理的培养物和未处理的对照物中测定黄曲霉毒素 B1 的毒性和生物活化；CD50 分别为 1.59 μM 和 31 μM。这些结果表明 PICM-19H 细胞可用于药物生物转化和毒性测试，并进一步支持它们在体外人工肝脏设备技术中的应用。
