Persistent biliary hypoxia and lack of regeneration are key mechanisms in the pathogenesis of posttransplant nonanastomotic strictures

Background and Aims: Nonanastomotic biliary strictures (NAS) are a major cause of morbidity after orthotopic liver transplantation (OLT). Although ischemic injury of peribiliary glands (PBGs) and peribiliary vascular plexus during OLT has been associated with the later development of NAS, the exact underlying mechanisms remain unclear. We hypothesized that bile ducts of patients with NAS suffer from ongoing biliary hypoxia and lack of regeneration from PBG stem/progenitor cells. Approach and Results: Forty-two patients, requiring retransplantation for either NAS (n = 18), hepatic artery thrombosis (HAT; n = 13), or nonbiliary graft failure (controls; n = 11), were included in this study. Histomorphological analysis of perihilar bile ducts was performed to assess differences in markers of cell proliferation and differentiation in PBGs, microvascular density (MVD), and hypoxia. In addition, isolated human biliary tree stem cells (hBTSCs) were used to examine exo-metabolomics during in vitro differentiation toward mature cholangiocytes. Bile ducts of patients with NAS or HAT had significantly reduced indices of PBG mass, cellular proliferation and differentiation (mucus production, secretin receptor expression, and primary cilia), reduced MVD, and increased PBG apoptosis and hypoxia marker expression, compared to controls. Metabolomics of hBTSCs during in vitro differentiation toward cholangiocytes revealed a switch from a glycolytic to oxidative metabolism, indicating the need for oxygen. Conclusions: NAS are characterized by a microscopic phenotype of chronic biliary hypoxia attributed to loss of microvasculature, resulting in reduced proliferation and differentiation of PBG stem/progenitor cells into mature cholangiocytes. These findings suggest that persistent biliary hypoxia is a key mechanism underlying the development of NAS after OLT.