Upregulation of the NADPH oxidase NOX4 by TGF-beta in hepatocytes is required for its pro-apoptotic activity

Background/Aims The transforming growth factor-beta (TGF-β) induces apoptosis in hepatocytes through an oxidative stress process. Here, we have analyzed the role of different NADPH oxidase isoforms in the intracellular signalling induced by TGF-β in hepatocytes, to later explore whether this mechanism is altered in liver tumor cells. Methods Primary cultures of rat and human hepatocytes, HepG2 and Hep3B cells were used in in vitro studies to analyze the TGF-β response. Results TGF-β-induced apoptosis in rat hepatocytes does not require Rac-dependent NADPH oxidases. TGF-β upregulates the Rac-independent Nox4, which correlates with its pro-apoptotic activity. Regulation of Nox4 occurs at the transcriptional level and is counteracted by intracellular survival signals. siRNA targeted knock-down of Nox4 attenuates NADPH oxidase activity, caspase activation and cell death in rat hepatocytes. NOX4 upregulation by TGF-β is also observed in human hepatocytes, coincident with apoptosis. In human hepatocellular carcinoma (HCC) cell lines, NOX4 upregulation by TGF-β is only observed in cells that are sensitive to its cytotoxic effect, such as Hep3B cells. siRNA targeted knock-down of NOX4 in these cells impairs TGF-β-induced apoptosis. Conclusions Upregulation of NOX4 by TGF-β is required for its pro-apoptotic activity in hepatocytes. Impairment of this TGF-β-induced response might confer apoptosis resistance in HCC cells.