Interferon α‐induced apoptosis on rat preneoplastic liver is mediated by hepatocytic transforming growth factor β 1

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 β 1 (TGF‐β 1 ) 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‐β 1 levels were increased in IFN‐α2b–treated rats. Immunohistochemical studies showed that IFN‐α2b significantly increased the quantity of TGF‐β 1 –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‐β 1 , 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‐β 1 secretion from hepatocytes. Neither Kupffer cells nor peritoneal macrophages secreted detectable TGF‐β 1 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‐β 1 signaling. When anti–TGF‐β 1 was added to the culture media, TGF‐β 1 activation and apoptosis induced by IFN‐α2b were blocked. In conclusion , IFN‐α2b–induced production of TGF‐β 1 by hepatocytes from preneoplastic liver is involved in the apoptotic elimination of altered hepatic foci. (H EPATOLOGY 2004;40:394–402.)