Cultured bacteria isolated from primary sclerosing cholangitis patient bile induce inflammation and cell death

Primary sclerosing cholangitis (PSC) is a chronic liver disease characterized by inflammation and progressive fibrosis of the biliary tree. PSC pathogenesis remains poorly understood, and there are no effective therapies. Previous studies have observed associations between colonic and biliary microbiome alterations and PSC. We aimed to determine whether bacterial isolates cultured from PSC patient bile induce disease-associated phenotypes in cells, specifically cell death, epithelial permeability, inflammation, and changes in host-protective pathways. Bile was collected from PSC patients by endoscopic retrograde cholangiography and from non-PSC controls undergoing cholecystectomies. Biliary bacteria were cultured anaerobically, and 50 colonies per sample were identified by 16S sequencing. No bacteria were isolated from non-PSC controls, while bacteria were cultured from most PSC patients. The PSC bile microbiomes exhibited reduced diversity compared to the gut or oral cavity, with one or two species predominating. The effects of supernatants from seven PSC-associated bacterial isolates on cellular phenotypes were characterized using human colonic (Caco-2), hepatic (HepG2), and biliary (EGI-1) cells. Overall, PSC-associated bacteria produced factors cytotoxic to hepatic and biliary cells. An <i>Enterococcus faecalis</i> isolate, and to a lesser extent a <i>Veillonella parvula</i> isolate, induced epithelial permeability, while <i>Escherichia coli</i>, <i>Fusobacterium necrophorum</i>, and <i>Klebsiella pneumoniae</i> isolates induced inflammatory cytokines in biliary cells. Our data suggest that bacteria cultured from PSC bile induce cellular changes characteristic of PSC pathogenesis, with different isolates inducing distinct cellular responses. Our work provides a starting point for future research into bacterial contributions to PSC with the eventual goal of developing therapies for this disease.IMPORTANCEPrimary sclerosing cholangitis (PSC) is a chronic liver disease in which inflammation and scarring of the bile ducts cause bile to build up in the liver, leading to liver damage and eventually liver failure. The causes of this disease are poorly understood, and the only current treatment is a liver transplant. To develop new treatments, we must first better understand what leads to this disease. We examined whether bacteria isolated from PSC patient bile can cause disease-related responses in human biliary, liver, and intestinal cells. We observed that different PSC-associated bacteria can induce distinct disease-related cellular changes, including inflammation and cell death. These data suggest that the microbial community in PSC patients may indeed be linked to disease development. Our findings provide new starting points for further exploration into the poorly understood origins of PSC.