Isolation of Human Hepatocytes by a Two-step Collagenase Perfusion Procedure
Isolation of Human Hepatocytes by a Two-step Collagenase Perfusion Procedure
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DOI：
10.3791/50615
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发表时间：
2013-09-01
影响因子：
1.2
通讯作者：
Thasler, Wolfgang E.
中科院分区：
文献类型：
--
作者：
Lee, Serene M. L.;Schelcher, Celine;Thasler, Wolfgang E.
肝脏是一个具有特殊再生能力的器官，具有广泛的功能，如解毒，代谢和体内平衡。因此，肝细胞是各种研究问题的重要模型。特别是，人肝细胞的使用在药代动力学、毒理学、肝再生和转化研究领域尤其重要。因此，该方法提供了Seglen（1）所述的两步胶原酶灌注程序的改良版本，以分离肝细胞。然而，酶促方法已被证明是上级的，因为肝细胞在分离后保留其结构完整性和功能。本文介绍的方法将先前为大鼠肝脏设计的方法适用于人肝片，并产生大量肝细胞，其活力为77+/-10%。该手术的主要区别在于血管插管的过程。此外，本文所述的方法也可以应用于来自具有可比肝脏或血管大小的其他物种的肝脏。
The liver, an organ with an exceptional regeneration capacity, carries out a wide range of functions, such as detoxification, metabolism and homeostasis. As such, hepatocytes are an important model for a large variety of research questions. In particular, the use of human hepatocytes is especially important in the fields of pharmacokinetics, toxicology, liver regeneration and translational research. Thus, this method presents a modified version of a two-step collagenase perfusion procedure to isolate hepatocytes as described by Seglen(1).Previously, hepatocytes have been isolated by mechanical methods. However, enzymatic methods have been shown to be superior as hepatocytes retain their structural integrity and function after isolation. This method presented here adapts the method designed previously for rat livers to human liver pieces and results in a large yield of hepatocytes with a viability of 77+/-10%. The main difference in this procedure is the process of cannulization of the blood vessels. Further, the method described here can also be applied to livers from other species with comparable liver or blood vessel sizes.