FoxO1 Promotes Liver Fibrosis through TGF‐β1 mediated Hepatic Stellate Cell Activation

The O-class of the forkhead transcription factor FoxO1 is a crucial factor mediating the action of insulin→PI3K→Akt and governs diverse cellular processes. To investigate the role of FoxO1 in control of liver fibrosis, we generated liver-specific deletion of FoxO1 gene in mice (L-F1KO). Using the mouse model, we injected intraperitoneally (i.p.) corn oil or 20% solution of carbon tetrachloride (CCL4) twice per week for 4 weeks (with 2.5 ul/g body weight) in WT and L-F1KO mice. H&E staining and Sirius Red staining revealed that CCl4-induced liver injury and fibrosis were significantly appeared, but the effects were largely attenuated in L-F1KO mice. Moreover, we found that both mRNA and protein levels of TGF-β1, a cytokine that activates hepatic stellate cell (HSC) for fibrogenesis, significantly decreased in the liver L-F1KO mice compared to WT mice in response to CCL4 treatment. In primary hepatocytes, with FoxO1-loss of function and gain-of function, we confirmed the regulation of TGF-β1 expression by FoxO1. Moreover, conditional medium (CM) collected from WT hepatocytes treated with CCL4 activated human HSC cell line (LX-2), such effect was attenuated by FoxO1 deletion in primary hepatocytes or neutralization of TGF-β1 in the CM using TGF-β1 antibody. These results indicate a crosstalk between hepatocytes and HSC via FoxO1-regualted TGF-β1. To further confirm this concept in vivo, we overexpressed FoxO1 in the liver of WT and liver-specific TGF-β1 deficient (L-TGF-β1KO) mice with adenovirus expressing FoxO1 (adFoxO1), and injected (i.p.) these mice with CCL4. We found that FoxO1 overexpression in WT mice promoted CCL4-induced liver fibrosis and HSC activation, and such effect was blocked in L-TGF-β1KO mice. Taken together, our data indicate that hepatic FoxO1 promotes CCL4-inducled liver fibrosis via upregulating TGF-β1 expression in hepatocytes and stimulating TGF-β1-mediated HSC activation. Hepatic FoxO1 may be a therapeutic target for prevention and treatment of liver fibrosis.