Isolation and culture of primary human hepatocytes.

As our knowledge of the species differences in drug metabolism and drug-induced hepatotoxicity has expanded significantly, the need for human-relevant in vitro hepatic model systems has become more apparent than ever before. Human hepatocytes have become the "gold standard" for evaluating hepatic metabolism and toxicity of drugs and other xenobiotics in vitro. In addition, they are becoming utilized more extensively for many kinds of biomedical research, including a variety of biological, pharmacological, and toxicological studies. This chapter describes methods for the isolation of primary human hepatocytes from liver tissue obtained from an encapsulated end wedge removed from patients undergoing resection for removal of liver tumors or resected segments from whole livers obtained from multiorgan donors. The maintenance of normal cellular physiology and intercellular contacts in vitro is of particular importance for optimal phenotypic gene expression and response to drugs and other xenobiotics. As such, methods are described for culturing primary hepatocytes under various matrix compositions and geometries. Differential expression of liver-selective properties occurs over time in primary hepatocytes dependent on the culture and study conditions. Overall, improved isolation and cultivation methods have allowed for exciting advances in our understanding of the pathology, biochemistry, and cellular and molecular biology of human hepatocytes.