Serum-Free, Long-Term Cultures of Human Hepatocytes: Maintenance of Cell Morphology, Transcription Factors, and Liver-Specific Functions

• Published in: Biochemical and biophysical research communications Vol. 269; no. 1; pp. 46 - 53
• Main Authors: Runge, Dieter, Runge, Dorothee M., Jäger, Dana, Lubecki, Kimberly A., Beer Stolz, Donna, Karathanasis, Sotirios, Kietzmann, Thomas, Strom, Stephen C., Jungermann, Kurt, Fleig, Wolfgang E., Michalopoulos, George K.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 05.03.2000
• Subjects: Blood Proteins - genetics; Blood Proteins - metabolism; C/EBP^a protein; Cell Culture Techniques - methods; Cell Differentiation; cell morphology; Cells, Cultured; Culture Media, Serum-Free; cytokeratin 18; cytokeratin 19; cytokeratins; Gene Expression; HNF-4 protein; human hepatocytes; Humans; Liver - cytology; Liver - metabolism; Liver - physiology; Microscopy, Electron; phosphoenolpyruvate carboxykinase; RNA, Messenger - genetics; RNA, Messenger - metabolism; serum proteins; Time Factors; transcription factors; Transcription Factors - genetics; Transcription Factors - metabolism; human hepatocytes; cytokeratins; cell morphology; phosphoenolpyruvate carboxykinase; transcription factors; serum proteins
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1006/bbrc.2000.2215

Since human hepatocytes are available only in limited number, the development of a serum-free culture system for long-term cultivation of differentiated and functional hepatocytes is of great importance. Here we describe the culture of human hepatocytes in a chemically defined serum-free medium for up to 5 weeks. Cell morphology was assayed by light and electron microscopy and revealed a well-preserved cellular morphology. Marker proteins for epithelial and bile duct cells, cytokeratin (CK) 18 and 19, and liver-specific proteins, like phosphoenolpyruvate carboxykinase-2 (PCK2) and serum proteins, were expressed. Liver-enriched transcription factors CCAAT/enhancer binding protein α (C/EBPα) and hepatocyte nuclear factor-4 (HNF-4), cytokine and mitogen activated factors (nuclear factor kappa B) NFκB, and activator protein-1 (AP-1) were maintained and active for several weeks in our cultures. In summary, our serum-free culture system allows the culture of differentiated human hepatocytes for several weeks. It may serve as a model system for metabolic, pharmacologic–toxicologic studies, and studies on human pathogens under defined chemical conditions.