Human hepatocyte culture.

Primary culture of human hepatocytes is an in vitro model widely used to investigate numerous aspects of liver physiology and pathology. The technique used to isolate human hepatocytes is based on two-step collagenase perfusion. Originally performed in situ for obtaining hepatocytes from the adult rat, this technique has been adapted to the ex vivo treatment of human liver from organ donors or from lobectomy resection for medical purposes. This chapter describes experimental protocols for the isolation of hepatocytes from human liver tissue and for the preparation of short- and long-term cultures in which cells retain a differentiated phenotype for at least 1 mo. The various aspects emphasized here include the conditions for obtaining tissue, quality control of tissue for efficient perfusion, collagenase perfusion parameters, solutions for perfusion and culture media, cell substrate, cell plating, specific equipment, and safety conditions.

[1]  D. Friend,et al.  HIGH-YIELD PREPARATION OF ISOLATED RAT LIVER PARENCHYMAL CELLS , 1969, The Journal of cell biology.

[2]  J. Pascussi,et al.  Expression of CYP3A4, CYP2B6, andCYP2C9 Is Regulated by the Vitamin D Receptor Pathway in Primary Human Hepatocytes* , 2002, The Journal of Biological Chemistry.

[3]  R. Gebhardt,et al.  Perifusion of co-cultured hepatocytes: optimization of studies on drug metabolism and cytotoxicity in vitro , 1996, Cell Biology and Toxicology.

[4]  I. Georgoff,et al.  Quantitative assay for albumin-producing liver cells after simian virus 40 transformation of rat hepatocytes maintained in chemically defined medium. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[5]  F. Lasserre,et al.  The small heterodimer partner interacts with the pregnane X receptor and represses its transcriptional activity. , 2003, Molecular endocrinology.

[6]  P. Blanc,et al.  Expression and DNA-binding activity of C/EBPα and C/EBPβ in human liver and differentiated primary hepatocytes , 2001 .

[7]  P. Maurel,et al.  Cyclosporin A drug interactions. Screening for inducers and inhibitors of cytochrome P-450 (cyclosporin A oxidase) in primary cultures of human hepatocytes and in liver microsomes. , 1990, Drug metabolism and disposition: the biological fate of chemicals.

[8]  P. Beaune,et al.  Expression of cytochrome P-450 enzymes in cultured human hepatocytes. , 1990, European journal of biochemistry.

[9]  J. Neuberger,et al.  Growth of normal human hepatocytes in primary culture: Effect of hormones and growth factors on DNA synthesis , 1991, Hepatology.

[10]  C. Bonfils,et al.  Induction protocols for cytochromes P450IIIA in vivo and in primary cultures of animal and human hepatocytes. , 1991, Methods in enzymology.

[11]  J. Pascussi,et al.  Dexamethasone enhances constitutive androstane receptor expression in human hepatocytes: consequences on cytochrome P450 gene regulation. , 2000, Molecular pharmacology.

[12]  P. Maurel,et al.  Omeprazole is an aryl hydrocarbon-like inducer of human hepatic cytochrome P450. , 1990, Gastroenterology.

[13]  R. Lanford,et al.  Analysis of plasma protein and lipoprotein synthesis in long-term primary cultures of baboon hepatocytes maintained in serum-free medium , 1989, In Vitro Cellular & Developmental Biology.

[14]  P. Maurel,et al.  Differential effects of cytokines on the inducible expression of CYP1A1, CYP1A2, and CYP3A4 in human hepatocytes in primary culture , 1995, Hepatology.

[15]  M. Wright,et al.  Induction of cytochrome P450 3A by metyrapone in human hepatocyte culture , 1996, Human & experimental toxicology.

[16]  G. Scarfe,et al.  The olivacine derivative s 16020 (9-hydroxy-5,6-dimethyl-N-[2-(dimethylamino)ethyl)-6H-pyrido(4,3-B)-carbazole-1-carboxamide) induces CYP1A and its own metabolism in human hepatocytes in primary culture. , 2004, Drug metabolism and disposition: the biological fate of chemicals.

[17]  C. Sureau,et al.  In vitro infection of adult normal human hepatocytes in primary culture by hepatitis C virus. , 1998, The Journal of general virology.

[18]  J. Domergue,et al.  Omeprazole and lansoprazole are mixed inducers of CYP1A and CYP3A in human hepatocytes in primary culture. , 1994, The Journal of pharmacology and experimental therapeutics.

[19]  J. Pascussi,et al.  Dexamethasone induces pregnane X receptor and retinoid X receptor-alpha expression in human hepatocytes: synergistic increase of CYP3A4 induction by pregnane X receptor activators. , 2000, Molecular pharmacology.

[20]  A. Li,et al.  Rifampicin induction of lidocaine metabolism in cultured human hepatocytes. , 1995, The Journal of pharmacology and experimental therapeutics.

[21]  P. Maurel,et al.  Long-term primary cultures of adult human hepatocytes. , 1997, Chemico-biological interactions.

[22]  P. Maurel.  The use of adult human hepatocytes in primary culture and other in vitro systems to investigate drug metabolism in man , 1996 .

[23]  J. Pascussi,et al.  Dual effect of dexamethasone on CYP3A4 gene expression in human hepatocytes. Sequential role of glucocorticoid receptor and pregnane X receptor. , 2001, European journal of biochemistry.

[24]  S. Wrighton,et al.  Ethanol and isopentanol increase CYP3A and CYP2E in primary cultures of human hepatocytes. , 1995, Archives of biochemistry and biophysics.

[25]  P. Maurel,et al.  Effects of imidazole derivatives on cytochromes P450 from human hepatocytes in primary culture , 1992, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[26]  P. Seglen.  Preparation of isolated rat liver cells. , 1976, Methods in cell biology.

[27]  T. Nakamura,et al.  Cell cycle progression proteins (cyclins), oncogene expression, and signal transduction during the proliferative response of human hepatocytes to hepatocyte growth factor , 1996, Hepatology.

[28]  J. Saric,et al.  Secretion of functional plasma haemostasis proteins in long‐term primary cultures of human hepatocytes , 2004, British journal of haematology.

[29]  Toshikazu Nakamura,et al.  Effects of hepatocyte growth factor on the growth and metabolism of human hepatocytes in primary culture , 1995, Hepatology.

[30]  P. Maurel,et al.  Effect of corticosteroids on the expression of cytochromes P450 and on cyclosporin A oxidase activity in primary cultures of human hepatocytes. , 1992, Molecular pharmacology.

[31]  J. Pawlotsky,et al.  Alpha Interferon Inhibits Hepatitis C Virus Replication in Primary Human Hepatocytes Infected In Vitro , 2002, Journal of Virology.

[32]  J. Daurès,et al.  Effect of anti‐ulcer drugs on DNA synthesis in adult normal human hepatocytes in culture , 1995, Hepatology.

[33]  J. Pascussi,et al.  Transcriptional Regulation of CYP2C9 Gene , 2002, The Journal of Biological Chemistry.

[34]  F. Zindy,et al.  Mitotic responsiveness of cultured adult human hepatocytes to epidermal growth factor, transforming growth factor alpha, and human serum. , 1992, Gastroenterology.