1C). Consequently, we investigated the expression of alpha smooth muscle actin (α-SMA), a marker of activated HSCs. As expected, the CCl4-treated SMP30 KO mice group exhibited much lower numbers of α-SMA immunopositive

cells per field compared with that of the CCl4-treated WT mice (Fig. 1D,E). We confirmed identical immunoblot results (Fig. 1F,G). These data indicate that CCl4-induced liver fibrosis is inhibited in SMP30 KO mice and suggest that SMP30 might play an important role in the HSC activation. In the WT mice the CCl4 treatment induced a decreased level of SMP30 expression around the central vein characterized by necrotic hepatocytes and infiltration of inflammatory cells compared with that of the control group. However, in the SMP30 KO mice group we could not detect an SMP30 expression, R428 confirmed with the SMP30 KO mice (Fig. 2A,B). The results, using immunoblotting and RT-PCR for SMP30 Z IETD FMK expression, were observed to be the same as the results obtained using immunohistochemistry (Fig. 2C-E). In serum vitamin C level measurements, the control group of the WT mice indicated normal serum vitamin C levels,

whereas the serum vitamin C level of the CCl4-treated WT mice was significantly decreased. In SMP30 KO mice, the serum vitamin C was undetectable in both the control group and the CCl4-treated groups (Fig. 2F). These data reveal that the SMP30 expression significantly decreased due to CCl4-induced liver injury. It was noticed that the SMP30 KO mice exhibited higher TGF-β expression levels in comparison with those of the WT mice (Fig. 3A,B). To evaluate p-Smad3, downstream of TGF-β1, expression levels in CCl4-treated WT mice and SMP30 KO mice, immunoblotting and immunohistochemistry were performed. selleck screening library In immunoblot results, whole liver tissues of SMP30 KO mice exhibited an elevated total of p-Smad3 expression levels compared with that of WT mice (Fig. 3C,D). However, in immunohistochemistry, CCl4-treated WT mice, exhibited significantly higher numbers of nuclear p-Smad2/3-positive parenchymal cells and nonparenchymal

cells, compared with CCl4-treated SMP30 KO mice (Fig. 3E-G). We observed more critical differences in nonparenchymal cells than in hepatocytes, which means the nuclear translocation of p-Smad2/3 was more severely inhibited in nonparenchymal cells, including HSCs and inflammatory cells. To confirm the immunohistochemistry results, we extracted nuclear proteins from the whole liver tissue for immunoblotting. The cytoplasmic p-Smad3 expression showed the same expression pattern as the total p-Smad3 expression pattern (Fig. 3C,H). Additionally, extracts of nuclear proteins also revealed well-matched results with the immunohistochemical nuclear p-Smad2/3 expression (Fig. 3C,I). Surprisingly, CCl4-treated SMP30 KO mice showed a significantly lower level of ROS generation and lipid peroxidation compared with CCl4-treated WT mice (Fig.