BILIARY ATRESIA THROUGH TARGETED ENDOTHELIAL DESTRUCTION (BATTED) PRESENTS A NOVEL LARGE ANIMAL MODEL, RECAPITULATING HUMAN DISEASE

Background: Biliary atresia (BA) remains a major cause of obstructive neonatal cholestatic disease. Although Kasai procedure or hepato-portoenterostomy (HPE) is routinely performed for BA, more than half of these cases eventually require liver transplantation. The intricate mechanisms of bile ductular injury in BA driving its pathogenesis remains elusive and not recapitulated in current small animal and rodent models that rely on bile duct ligation. Addressing prevailing lacunae, we hypothesized that extra and intra hepatic bile duct destruction through an endothelial irritant would recapitulate the human condition. We have thus developed and describe here a novel neonatal piglet BA model titled biliary atresia through targeted endothelial destruction (BATTED). Neonatal piglets have liver and gastro-intestinal homology to human infants and share anatomic and physiological processes through similarity in body size, and anatomy, providing a robust platform to perform BATTED and HPE.
Method: 7-10 day old piglets were procured. Two piglets underwent BATTED and the remaining, sham surgery. BATTED included cholecystectomy, isolation of common bile duct and hepatic duct, with subsequent injection of 95% ethanol and a retained ligation for continued ethanol induced intrahepatic injury. Vascular access ports were placed for blood collection and scheduled ultrasound guided liver biopsies were performed. Six-weeks post-BATTED piglets underwent HPE. 8-weeks after initial surgery, animals were euthanized, and samples collected. Serology, histology, gene expression and immunohistochemistry were performed.
Results: Serological evaluation revealed a surge in conjugated bilirubin 6 weeks after BATTED procedure from baseline (mean Δ 0.39mg/dL to 3.88mg/dL). Gamma-glutamyl transferase (GGT) also exhibited a several fold increase (mean: Δ 16.3IU to 89.5IU). Sham did not display these elevations (conjugated bilirubin: Δ 0.39mg/dL to 0.98mg/dL, GGT: Δ 9.2IU to 10.4IU). Sirius red staining demonstrated significant periportal and diffuse liver fibrosis (16-fold increase) and bile duct proliferation marker CK-7 increased 9-fold with BATTED. Piglets in the BATTED group demonstrated enhanced CD-3 (7-fold), alpha-SMA (8.85-fold), COL1A1 (11.7-fold) and CYP7A1 (7-fold), vs sham. Successful HPE was accomplished in piglets with a reduction in conjugated bilirubin (Δ 4.89 mg/dL to 2.11 mg/dL).
Conclusion: BATTED model replicated BA features, including hyperbilirubinemia, GGT elevation, significant hepatic fibrosis, bile duct proliferation, and inflammatory infiltration with subsequent successful HPE. This model offers substantial opportunities to elucidate the mechanism underlying BA and adaptation post HPE, paving the path for the development of diagnostics and therapeutics.