Activation of Slit2-Robo1 signaling promotes liver fibrosis

Background & Aims The secretory protein Slit2 and its receptor Robo1 are believed to regulate cell growth and migration. Here, we aimed to determine whether Slit2-Robo1 signaling mediates the pathogenesis of liver fibrosis. Methods Serum levels of Slit2 in patients with liver fibrosis were determined by ELISA. Liver fibrosis was induced in wild-type (WT), Slit2 transgenic ( Slit2 -Tg) and Robo1 +/− Robo2 +/− double heterozygotes ( Robo1/2 +/− ) mice by carbon tetrachloride (CCl 4 ). The functional contributions of Slit2-Robo1 signaling in liver fibrosis and activation of hepatic stellate cells (HSCs) were investigated using primary mouse HSCs and human HSC cell line LX-2. Results Significantly increased serum Slit2 levels and hepatic expression of Slit2 and Robo1 were observed in patients with liver fibrosis. Compared to WT mice, Slit2 -Tg mice were much more vulnerable to CCl 4 -induced liver injury and more readily develop liver fibrosis. Development of hepatic fibrosis in Slit2 -Tg mice was associated with a stronger hepatic expression of collagen I and α-smooth muscle actin (α-SMA). However, liver injury and hepatic expression of collagen I and α-SMA were attenuated in CCl 4 -treated Robo1/2 +/− mice in response to CCl 4 exposure. In vitro , Robo1 neutralizing antibody R5 and Robo1 siRNA downregulated phosphorylation of Smad2, Smad3, PI3K, and AKT in HSCs independent of TGF-β1. R5 and Robo1 siRNA also inhibited the expression of α-SMA by HSCs. Finally, the protective effect of R5 on the CCl 4 -induced liver injury and fibrosis was further verified in mice. Conclusions Slit2-Robo1 signaling promotes liver injury and fibrosis through activation of HSCs.