Rapamycin mitigates warm ischemia-induced peribiliary fibrosis: A non-transplant experimental model with implications for ischemic cholangiopathy

Background: Warm ischemia is a major contributor to ischemic cholangiopathy and non-anastomotic biliary strictures (NAS) after liver transplantation, particularly in donation-after-circulatory-death grafts.However, the isolated impact of warm ischemia on peribiliary fibrosis is difficult to delineate because clinical settings involve overlapping effects of cold ischemia, reperfusion injury, and alloimmunity.This study aimed to establish a non-transplant rat model that isolates warm ischemic biliary injury and to compare the antifibrotic effects of rapamycin and tacrolimus.Methods: Warm ischemia was induced in Sprague-Dawley rats by ligating both ends of the peribiliary vascular plexus and the hepatic artery, followed by 30 minutes of portal vein clamping.Rats were randomly assigned to control, tacrolimus (1 mg/kg/day), or rapamycin (1 mg/kg/day) groups (n=15 per group).Serum alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase, and total bilirubin were measured serially.Hematoxylin and eosin (H&E) and Sirius red staining were performed at 1, 3, and 6 weeks.Peribiliary fibrosis was quantified using digital image analysis of collagen area fraction.Results: Warm ischemia induced acute hepatobiliary injury with transient enzyme elevations, but no significant intergroup differences were observed.Histologically, biliary epithelial proliferation and collagen deposition increased progressively and became prominent at 6 weeks.At this time point, fibrosis ratios differed significantly (overall p=0.002): controls showed the highest fibrosis (4.8%), followed by tacrolimus (2.4%) and rapamycin (1.7%).Both immunosuppressants significantly reduced fibrosis compared with controls (p<0.05),whereas the difference between tacrolimus and rapamycin was not significant.Conclusion: This warm ischemia model demonstrates that isolated ischemic injury alone can induce progressive peribiliary fibrosis.Rapamycin and tacrolimus attenuated fibrosis, with rapamycin producing the lowest collagen deposition.These findings provide mechanistic insight into ischemic cholangiopathy after liver transplantation-particularly in donation after circulatory death grafts-and underscore the need for further studies using models that incorporate cold ischemia, reperfusion, and alloimmune factors.