Oligo-peptide I-C-F-6 is a Carapax trionycis extract component that has an effect on hepatic fibrosis, however, its mechanism of action is still unclear. This study investigated whether oligo-peptide I-C-F-6 could inhibit liver fibrosis by suppressing NF-κB and Wnt/β-catenin signaling, which are important in liver fibrosis. HSC-T6 cells were treated with oligo-peptide I-C-F-6, and rats were divided randomly into five groups: control (saline), CCl, CCl plus oligo-peptide I-C-F-6 (0.12 and 0.24 mg/kg), and CCl plus colchicine (0.11 mg/kg). Here, we demonstrated that oligo-peptide I-C-F-6 ameliorated liver injury, inflammation, and hepatic fibrogenesis induced by CCl. Oligo-peptide I-C-F-6 also inhibited the activation of hepatic stellate cells (HSCs) in vivo and in vitro, as evaluated by the expression of transforming growth factor-β1 (TGF-β1) and α-smooth muscle actin (α-SMA), which is a specific marker of HSC activation. Moreover, oligo-peptide I-C-F-6 significantly reduced the expression and distribution of β-catenin, P-AKT, phospho (P)-GSK-3β, nuclear factor κB (NF-κB) P65, phospho-P65, and IκB kinase α/β (IKK-α/β) levels; additionally, IκB-α level was elevated both in vivo and in vitro. Together, these results indicate that oligo-peptide I-C-F-6 has hepatoprotective and anti-fibrotic effects in animal models of liver fibrosis, the mechanism of which may be related to modulating NF-κB and Wnt/β-catenin signaling.