Anthony C. Huggett

Growth modulatory effects of a liver-derived growth inhibitor, transforming growth factor β1, and recombinant tumor necrosis factor α, in normal and neoplastic cells

Abstract The growth modulatory effects of a rat liver-derived growth inhibitor (LDGI), transforming growth factor β1 (TGF-β1), and recombinant tumor necrosis factor (rTNF-α) were examined in a variety of liver-derived and nonliver-derived normal and neoplastic cell culture systems. Normal rat liver epithelial (RLE) cells were highly sensitive to the growth inhibitory effects of LDGI (ID50 = 0.2 ng/ml) and TGF-β1 (ID50 = 0.25 ng/ml) but were less sensitive to rTNF-α (ID40 = 5000 Units/ml). Aflatoxin B1-transformed RLE cells showed sensitivity to the cytostatic effects of LDGI (ID50 = 1.5 ng/ml); however, these cells were completely resistant to the antiproliferative effects of TGF-β1 and rTNF-α. Clones isolated from these transformed cells, exhibited a wide range of sensitivities to LDGI but all of the clones were resistant to the growth inhibitory effects of both TGF-β1 and rTNF-α. Rat hepatoma Reuber cells were extremely sensitive to the antiproliferative effects of rTNF-α (ID50 = 10 Units/ml) but exhibited sensitivity to LDGI only at concentrations above 1.5 ng/ml and were resistant to the antiproliferative effects of TGF-β1. Rat hepatoma UVM 7777 cells and human hepatoma HepG2 cells, however, were insensitive to the growth inhibitory effects of all three factors. Among the nonliver-derived cells, human breast carcinoma (MCF-7) cells were extremely sensitive to rTNF-α (ID50 = 20 Units/ml, exhibited some sensitivity to LDGI (ID50 = 1 ng/ml), and were resistant to the antiproliferative effects of TGF-β1. In contrast, the rate of DNA synthesis is rat kidney fibroblasts and human foreskin fibroblasts was significantly stimulated in response to TGF-β1, LDGI, and rTNF-α. These data demonstrate that LDGI, TGF-β1, and rTNF-α exert positive and negative modulations of growth in different cell systems and that the growth regulatory effects of LDGI differ from those of TGF-β1 and rTNF-α in some cell types.

Effects of Interleukin-6 on the Growth of Normal and Transformed Rat Liver Cells in Culture

Recombinant human interleukin-6 produced a dose-dependent inhibition of DNA synthesis in both growing and mitogen-stimulated cultures of normal rat liver epithelial cells and also in primary rat hepatocytes. A significant inhibition of DNA synthesis (P less than 0.001) was obtained with 1 ng/ml (10 Units/ml) interleukin-6 in normal rat liver epithelial cells. The ID50 for inhibition of DNA synthesis in primary rat hepatocytes was 1 ng/ml. In contrast to the effects of transforming growth factor beta (Type I), where an almost complete inhibition of DNA synthesis could be achieved with either cell type, the maximal inhibition observed with interleukin-6 for both of these cell types was about 45%. Thus distinct mechanisms are involved in the inhibition of liver cell growth by these growth modulators. Transformed liver-derived cell lines were relatively resistant to the growth inhibitory effects of both interleukin-6 and TGF-beta 1 compared with the normal cells. However, human Hep G2 cells, which were completely resistant to the growth inhibitory effects of TGF-beta 1, were moderately inhibited by interleukin-6, indicating that the mechanisms responsible for the acquired resistance to growth inhibition is different for these growth inhibitors. The ability of interleukin-6 to function as a growth inhibitor in vitro was confirmed using normal rat liver epithelial cells. Interleukin-6 at a concentration of 10 ng/ml produced a significant decrease (P less than 0.05) in the proliferation of these cells. These data demonstrate that interleukin-6 may have the capability of functioning as a growth regulatory polypeptide for liver cells in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

Dexamethasone prevents the growth inhibitory effects of recombinant tumor necrosis factor in a rat hepatoma cell line reuber-RC-3: An association with the changes in the messenger RNA levels for multidrug resistance gene

Two days exposure to recombinant tumor necrosis factor (rTNF-alpha) produced a dose-dependent reduction in (methyl-3H) thymidine incorporation in RC-3 cells (ID50 = 25 units/ml). Prolonged treatment with rTNF-alpha further resulted in a significant reduction in colony formation (ID50 = 200 units/ml), which was reversed upon removal of the agent. Interferon levels were undetectable in the supernatants of the rTNF-alpha treated cells. Simultaneous exposure to dexamethasone prevented the growth inhibition in rTNF-alpha-treated RC-3 cells. Significant dose-dependent increase in the steady state levels of the mRNA for multidrug resistance (MDR1) gene was observed after rTNF-alpha treatment while simultaneous exposure to dexamethasone produced a substantial reduction in the mRNA levels for MDR1 gene. These data suggest that growth inhibitory effects of TNF are regulated by dexamethasone and are associated with changes in MDR1 mRNA levels in hepatoma-derived cells.

Immunochemical and catalytical characterization of the human liver NADPH-cytochrome P450 reductase.

1. The NADPH‐cytochrome P450 reductases (EC 1.6.2.4) from human and rabbit liver have been purified to electrophoretic homogeneity. The human reductase had an apparent monomeric molecular weight of 77 500 and the rabbit enzyme of 76 500.