Interferon α-induced apoptosis on rat preneoplastic liver is mediated by hepatocytic transforming growth factor β1

Abstract

In previous work we showed that interferon alfa-2b (IFN-α2b) increases apoptosis on rat hepatic preneoplastic foci. The aim of this study was to determine if transforming growth factor β1 (TGF-β1) was involved in the programmed cell death on the foci. Animals were divided into 6 groups: subjected to a 2-phase model (diethylnitrosamine plus 2-acetylaminofluorene) of preneoplasia development (group 1); treated with IFN-α2b during the 2 phases (group 2); treated with IFN-α2b during initiation with diethylnitrosamine (group 3); treated with IFN-α2b during 2-acetylaminofluorene administration (group 4); subjected only to an initiation stage (group 5); and treated with IFN-α2b during the initiation period (group 6). Serum TGF-β1 levels were increased in IFN-α2b-treated rats. Immunohistochemical studies showed that IFN-α2b significantly increased the quantity of TGF- β1-positive hepatocytes in groups 2 to 4. Phosphorylated-Smads- 2/3 (p-Smads-2/3) proteins in liver nuclear extracts were significantly elevated. To determine the source of TGF-β1, isolated hepatocytes, Kupffer cells, and peritoneal macrophages from animals in groups 1 and 5 were cultured with or without IFN-α2b. IFN-α2b stimulus induced several-fold increases of TGF-β1 secretion from hepatocytes. Neither Kupffer cells nor peritoneal macrophages secreted detectable TGF-β1 levels when they were treated with IFN-α2b. IFN-α2b-stimulated cultured hepatocytes from preneoplastic livers showed enhanced apoptosis, measured by fluorescence microscopy and caspase-3 activity. They presented higher nuclear accumulation of p-Smads-2/3, indicating increased TGF-β1 signaling. When anti-TGF- β1 was added to the culture media, TGF-β1 activation and apoptosis induced by IFN-α2b were blocked. In conclusion, IFN-α2b-induced production of TGF-β1 by hepatocytes from preneoplastic liver is involved in the apoptotic elimination of altered hepatic foci.