CONSTITUTIVE PRODUCTION OF COLONY-STIMULATING FACTORS BY HUMAN HEPATOMA-CELL LINES - POSSIBLE CORRELATION WITH CELL-DIFFERENTIATION

A panel of two poorly differentiated (HA22T/VGH and SK-Hep-1) and six well-differentiated (HuH-6-cl 5, HuH-7, PLC/PRF/5, Hep G2, Hep 3B, and Tong) human hepatocellular carcinoma (HCC) cell lines were studied fo r the production of colony-stimulating factors (CSFs) using the granul ocyte and macrophage colony formation (CFU-GM) assay, immunocytochemic al staining, and Northern blotting. Medium conditioned by untreated HA 22T/VGH cells contained a high level of CSFs that could stimulate the in vitro colony formation of human myeloid progenitor cells. The HA22T /VGH cell-derived CSF had an apparent molecular weight of 23 kD. Its a ctivity could be effectively neutralized by antiserum against granuloc yte-macrophage CSF (GM-CSF) but not by antibodies to other hematopoiet ic growth factors, including G-CSF, M-CSF, interleukin-3 (IL-3), and I L-6. Correspondingly, immunocytochemical studies using monoclonal anti -GM-CSF showed a strong positive reaction in the cytoplasm of the HA22 T/VGH cells. Northern blot analysis revealed that untreated HA22T/VGH cells expressed a considerable amount of GM-CSF mRNA, confirming that GM-CSF production was constitutive. At optimal concentrations, lipopol ysaccharide (LPS), IL-1 beta, interferon-gamma (IFN-gamma), and tumor- promoting phorbol diester (TPA) could all stimulate HA22T/VGH cells to secrete GM-CSF. In addition to HA22T/VGH, SK-Hep-1 cells could also p roduce GM-CSF, although less effectively, whereas all the well-differe ntiated HCC cell lines tested were negative for CSF production. Morpho logic, cytochemical, and immunocytochemical examinations demonstrated that both poorly differentiated CSF-producing HCC cell lines (HA22T/VG H and SK-Hep-1) were macrophage-like in morphology, possessed nonspeci fic esterase (NSE) activity, and expressed CD14, CD68, and HLA-DR on t heir surface, while all the well-differentiated HCC cell lines were ep ithelioid and lacked myeloid differentiation antigens. These results s uggest that monocytoid features and CSP production may be differentiat ion markers of hepatocytes at the immature stages, and that the HA22T/ VGH and SK-Hep-1 cell lines may be valuable tools for the study of hep atic function and differentiation.