Sielaff T D, Hu M Y, Rao S, Groehler K, Olson D, Mann H J, Remmel R P, Shatford R A, Amiot B, Hu W S
Department of Surgery, University of Minnesota, Minneapolis, USA.
Transplantation. 1995 May 27;59(10):1459-63. doi: 10.1097/00007890-199505270-00017.
Current bioartificial liver devices are based on the use of a large mass of hepatocytes exhibiting differentiated metabolic function. The pig has become a source of interest for the acquisition of such cells-however, harvesting a large mass of highly viable cells has met with difficulty. This study describes a technique for harvesting large quantities of hepatocytes at viabilities greater than 90% and also describes several features documenting differentiated function. Pigs, 6 to 10 kg body weight, underwent in situ two-step whole liver perfusion (ethylene glycol tetraacetic acid and collagenase) and ex vivo cell harvest. Harvests yielded an average of 19.5 billion cells with an average viability of 94.6%. Hepatocytes were then entrapped in type I collagen (3 x 10(5) cells/well) and cultured in serum-free media for 5 days. Pig hepatocytes produced stable amounts of albumin and maintained cytochrome P-450 and glucuronidation activity over 5 days, as shown by the metabolism of lidocaine and 4-methylumbelliferone. These data indicate that pig hepatocytes can be harvested with high yields and can retain viability and differentiated function over at least 5 days of culture, and therefore should prove to be an excellent source of hepatocytes for bioartificial liver devices.
当前的生物人工肝装置基于使用大量具有分化代谢功能的肝细胞。猪已成为获取此类细胞的一个受关注来源——然而，获取大量高活力细胞遇到了困难。本研究描述了一种以高于90%的活力收获大量肝细胞的技术，还描述了一些记录分化功能的特征。体重6至10千克的猪接受原位两步全肝灌注（乙二醇四乙酸和胶原酶）及体外细胞收获。收获的细胞平均为195亿个，平均活力为94.6%。然后将肝细胞包埋于I型胶原中（每孔3×10⁵个细胞），并在无血清培养基中培养5天。如利多卡因和4-甲基伞形酮的代谢所示，猪肝细胞在5天内产生稳定量的白蛋白，并维持细胞色素P-450和葡萄糖醛酸化活性。这些数据表明，猪肝细胞能够以高产率收获，并且在至少5天的培养过程中能够保持活力和分化功能，因此应被证明是生物人工肝装置肝细胞的极佳来源。
