NIM811 downregulates transforming growth factor-β signal transduction in vivo and in vitro

Liver fibrosis is the common histological feature of a number of chronic liver diseases, and leads to cirrhosis and hepatocellular carcinoma (HCC). It has been demonstrated that N-methyl-4-isoleucine cyclosporine (NIM811) attenuates CCl4-induced liver fibrosis and inflammation in rats. The present study investigated whether NIM811 downregulated transforming growth factor (TGF)-beta signaling in rats with CCl4-induced liver fibrosis and in HSC-T6 cells. Liver tissues were obtained from rats with CCl4-induced liver fibrosis, with or without NIM811 treatment. HSC-T6 cells were cultured with or without NIM811 for 18 h under serum-free conditions. Expression of collagen I, alpha-smooth muscle actin (alpha-SMA), TGF-beta 1, TGF-beta receptor I (T beta R-I) and TGF-beta pathway downstream signaling molecules were measured by reverse transcription-quantitative polymerase chain reaction and/or western blotting. Collagen I and TGF-beta 1 content in the cell supernatant was measured by ELISA. NIM811 profoundly inhibited collagen I, alpha-SMA, TGF-beta 1 and T beta R-I expression in the liver of CCl4-treated rats. Phosphorylation of Smad2, 3 and 1/5/8 was decreased in the liver of NIM811-treated groups, accompanied by increased In addition, Smad7 expression compared with the CCl4-treated rats. NIM811 inhibited collagen I, TGF-beta 1 and T beta R-I expression in HSC-T6 cells. Smad1 mRNA and phospho-Smad1/5/8 protein levels decreased following NIM811 treatment, accompanied by increased Smad7 expression in HSC-T6 cells compared with normal controls. Furthermore, NIM811 also inhibited collagen I mRNA expression in the liver of rats with CCl4-induced liver fibrosis and in HSC-T6 cells. The results suggest that the antifibrotic effect of NIM811 was due to the inhibition of TGF-beta 1 and its downstream signaling molecules.