Optimal erythropoietin expression in human hepatoma cell lines requires activation of multiple signalling pathways.
Hypoxia is thought to be a common precursor of coronary artery disease and malignant tumors, both diseases representing the leading causes of death in industrial nations. So far, investigations of oxygen-regulated erythropoietin (EPO) gene expression in the human hepatoma cell lines Hep3B and HepG2 allowed many important insights into the mechanisms of oxygen-sensing, signalling and regulation of an increasing number of oxygen-responsive genes. To differentiate the various signalling pathways involved in EPO production by these two cell lines, we examined several factors that positively influenced EPO expression. The results demonstrate a keen differential effect of cell density and oxygen concentration on EPO induction in Hep3B compared to HepG2 cells. Using optimized cell culture conditions, EPO production rates as high as 1 U EPO per 10(6) Hep3B cells in 24 h could be achieved. We also found a moderate but reproducible positive effect of CoCl2 on hypoxia-induced EPO expression in Hep3B but a negative CoCl2 effect on hypoxic induction in HepG2 cells. CoCl2 inhibited cell growth in a concentration-dependent manner. Interleukin-6 was synergistic with hypoxia on EPO induction in Hep3B as well as HepG2 cells, and dexamethasone enhanced this effect in Hep3B but not in HepG2 cells. The moderate CoCl2-dependent increase of EPO production observed in hypoxic Hep3B cells might indicate that CoCl2 and hypoxia do not necessarily act via, identical signalling pathways.
Wenger
R H
RH
Institute of Physiology, University of Zürich-Irchel, CH-8057 Zürich, Switzerland.
Marti
H H
HH
Bauer
C
C
Gassmann
M
M
D006528:Carcinoma, Hepatocellular; D002452:Cell Count; D015687:Cell Hypoxia; D003035:Cobalt / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D004921:Erythropoietin / Q000096:biosynthesis*; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology; D008113:Liver Neoplasms; D010100:Oxygen / Q000378:metabolism; D015398:Signal Transduction
D015850:Interleukin-6; D004921:Erythropoietin; D003035:Cobalt; D003907:Dexamethasone; C018021:cobaltous chloride; D010100:Oxygen