Apoptosis Induced by Transforming Growth Factor-β in Fetal Hepatocyte Primary Cultures

Abstract

Transforming growth factor-β (TGF-β), a growth regulator of fetal hepatocytes in primary culture, also regulates death of these cells. Dose-response analysis showed that the TGF-β concentration needed to induce hepatocyte death (2.5 ng/ml) was 5 times that needed to inhibit growth in these cells (0.5 ng/ml). In response to TGF-β, hepatocytes induced DNA fragmentation and the appearance of nuclei with a DNA content lower than 2C (diploid content), typical of a programmed cell death model. TGF-β-induced apoptosis in fetal hepatocytes was preceded by an induction of reactive oxygen species production and a decrease in the glutathione intracellular content, indicating that this factor induces oxidative stress in fetal hepatocytes. Studies performed to analyze levels of c- fos mRNA, a gene whose expression is modulated by redox state, demonstrated that only high, apoptotic concentrations of TGF-β (2.5 ng/ml) produced an increase in the mRNA levels of this gene, the level of induction being similar to that found when cells were incubated in the presence of tert -butyl hydroperoxide. Gel mobility shift assays showed that the c- fos -induced expression was coincident with an increase in AP-1 activity. Finally, cell death induced by TGF-β in fetal hepatocytes was partially blocked by radical scavengers, which decreased the percentage of apoptotic cells, whereas these agents did not modify the growth-inhibitory effect elicited by TGF-β in these cells. In summary, the results presented in this paper provide evidence for the involvement of an oxidative process in the apoptosis elicited by TGF-β in fetal hepatocytes.