The inhibitors used including DPI, SB, SP, U0126, or LY. 1% DMSO was used as a negative control.
Figure 2A. DPI blocked JFH1 HCV mediated ROS generation. ROS level was normalized by cell viability to calculate the ROS/Cell Viability Arbitrary Unit. DPI completely blocked the HCV induced ROS production when compared to JFH1 in DMSO (P<0.001, n=4).
Figure 2B. DPI inhibited JFH1 HCV mediated TGF-β1 expression. TGF-β1 level was normalized to GAPDH level to calculate the TGF-β1/GAPDH arbitrary unit. DPI completely blocked the HCV stimulated TGF-β1 expression (P=0.002). SB, SP, or U0126 partially reduced TGF-β1 production by 29.5% (P=0.08), 32.2% (P=0.07), and 27.5% (P=0.1), respectively compare to JFH1 in DMSO (n=4). In contrast, LY had no effect on TGF-β1 expression.
Figure 2C. The ROS inhibitor DPI blocked phosphorylation of p38 MAPK, JNK, ERK. JFH1 HCV activated the phosphorylation of p38 MAPK, p42 ERK, and JNK (lane #2). ROS inhibitor DPI blocks phosphorylation of p38 MAPK, JNK, ERK to levels comparable to those seen with their specific inhibitors (Lane #3). Antibody to PI3K was directed against unphosphorylated protein. We found that HCV does not activate PI3K phosphorylation (data not shown). Lane#1: Huh7.5.1+DMSO; Lane#2 JFH1+DMSO; Lane#3 JFH1+DPI; Lane#4 JFH1+SB; #5 JFH1+SP; #6 JFH1+U0126; #7 JFH1+LY.