Colchicine (CNE) is used to treat various inflammatory conditions, but its use is limited by its hepatotoxic potential. This study assessed the protective effect of pumpkin seed oil (PPN) against CNE-induced liver injury in rats. Forty male Sprague-Dawley rats were allocated into four groups: control, PPN (4 mL/kg b.wt), CNE (0.6 mg/kg b.wt), and CNE+PPN, and received oral treatments for 56 days. CNE significantly elevated liver enzymes (ALP, GGT, AST, and ALT) and bilirubin levels, and induced dyslipidemia, while PPN co-treatment restored these parameters toward normal. Antioxidant defenses, including SOD, CAT, and GSH were significantly suppressed by CNE, accompanied by increased MDA, protein carbonyls, 8-OHdG, and caspase-3 activity. PPN treatment improved redox status and reduced apoptotic markers. Histologically, CNE caused ductular reactions, bile duct hyperplasia, and collagen fiber accumulation around the portal triads. Masson's trichrome staining confirmed significant fibrosis, which was significantly reduced with PPN co-administration. Immunohistochemistry revealed a significant reduction in PCNA in CNE-treated livers, partially restored by PPN. BAX expression was significantly increased by CNE and notably reduced in the CNE+PPN group. These findings highlight the hepatoprotective potential of PPN through antioxidant, anti-apoptotic, anti-fibrotic, and lipid-regulating mechanisms, supporting its use as a natural beneficial agent against CNE-induced liver injury.