Long-term culture and expansion of primary human hepatocytes.
Hepatocytes have a critical role in metabolism, but their study is limited by the inability to expand primary hepatocytes in vitro while maintaining proliferative capacity and metabolic function. Here we describe the oncostatin M (OSM)-dependent expansion of primary human hepatocytes by low expression of the human papilloma virus (HPV) genes E6 and E7 coupled with inhibition of epithelial-to-mesenchymal transition. We show that E6 and E7 expression upregulates the OSM receptor gp130 and that OSM stimulation induces hepatocytes to expand for up to 40 population doublings, producing 1013to 1016cells from a single human hepatocyte isolate. OSM removal induces differentiation into metabolically functional, polarized hepatocytes with functional bile canaliculi. Differentiated hepatocytes show transcriptional and toxicity profiles and cytochrome P450 induction similar to those of primary human hepatocytes. Replication and infectivity of hepatitis C virus (HCV) in differentiated hepatocytes are similar to those of Huh7.5.1 human hepatoma cells. These results offer a means of expanding human hepatocytes of different genetic backgrounds for research, clinical applications and pharmaceutical development.
Alexander Grass Center for Bioengineering, The Hebrew University of Jerusalem, Jerusalem, Israel.
Gahl
G
Levy
Alexander Grass Center for Bioengineering, The Hebrew University of Jerusalem, Jerusalem, Israel.
David
D
Bomze
Upcyte Technologies GmbH, Hamburg, Germany.
Stefan
S
Heinz
Medicyte GmbH, Heidelberg, Germany.
Sarada Devi
SD
Ramachandran
Upcyte Technologies GmbH, Hamburg, Germany.
Astrid
A
Noerenberg
Alexander Grass Center for Bioengineering, The Hebrew University of Jerusalem, Jerusalem, Israel.|Department of Cell and Developmental Biology, The Hebrew University of Jerusalem, Jerusalem, Israel.
Merav
M
Cohen
Liver Unit, Department of Gastroenterology, Tel Aviv Medical Center and Tel Aviv University, Tel Aviv, Israel.
Oren
O
Shibolet
Department of Clinical Microbiology and Immunology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.
Ella
E
Sklan
Medicyte GmbH, Heidelberg, Germany.|Fraunhofer Translational Center, Würzburg, Germany.
Joris
J
Braspenning
Alexander Grass Center for Bioengineering, The Hebrew University of Jerusalem, Jerusalem, Israel.|Department of Cell and Developmental Biology, The Hebrew University of Jerusalem, Jerusalem, Israel.
Yaakov
Y
Nahmias