PROJECT SUMMARY/ABSTRACT Biliary tract disease and inflammation, including pediatric biliary atresia and primary sclerosing cholangitis, are leading causes of liver failure. Chronic biliary inflammation, which can be associated with the presence of gallstones, is a risk factor for development of cholangiocarcinoma, a rare but deadly malignancy. Acute inflammation of the gallbladder, or cholecystitis, accounts for 20% of all biliary tract disease hospitalizations, usually requires surgical intervention and has significant mortality if left untreated. Together, biliary tract disease, including common gallstones, represents an enormous human health burden. Biliary tract inflammation plays clear roles in progression of various common and rare biliary diseases, however our understanding of inflammation and immune processes within the biliary tract are exceedingly limited. The resident immune cell populations in the extrahepatic biliary tree at homeostasis have never been characterized, and whether host or environmental factors can influence the development of the biliary niche has never been explored. It is unknown if resident biliary immune cells could impact development and progression of biliary disease. During my postdoctoral training, I developed a reproducible tissue digest method that yields highly viable immune, epithelial, and stromal cell populations. Single cell RNA sequencing revealed that there is a diversity of resident immune cells present at homeostasis in the mouse gallbladder/extrahepatic bile ducts, including innate lymphoid cells, adaptive lymphocytes, neutrophils, macrophages, and dendritic cells. In the course of studying biliary tuft cells—rare, chemosensory epithelial cells with known immunomodulating properties—I found that the biliary immune niche is altered in the absence of tuft cells, and is also sensitive to the microbiome-status of the host. My data suggest that the microbiome and biliary epithelial cells (including tuft cells) regulate the establishment of the biliary immune niche. In this proposal, I will test the role of the microbiome and microbial metabolites in setting biliary immune “tone,” and will define a role for tuft cells in regulating the biliary immune cell make-up. I will test whether alterations in the biliary immune niche at homeostasis, specifically the presence or absence of neutrophils, can impact the progression of cholesterol gallstone disease. To accomplish these aims, I have formed collaborations with experts in liver/biliary biology and microbiome manipulations, and have established a career development plan that will facilitate increasing computational independence. These studies will reveal new links between host factors and biliary inflammation, with the potential for therapeutic targeting of biliary epithelial cells and the microbiome to impact biliary health and disease.

PROJECT NARRATIVE Biliary tract inflammation contributes to the development and progression of many common human biliary and liver pathologies, but our understanding of immune processes within the biliary tract is exceedingly limited. I have characterized resident immune cell populations in the extrahepatic biliary tree at high resolution, and found that the make-up of this biliary immune niche is impacted by biliary epithelial cells and the microbiome-status of the host. This project will test how the microbiome, microbial metabolites, and biliary epithelial cells impact the biliary immune niche, and will test whether alterations in the biliary immune niche at homeostasis, specifically the presence or absence of neutrophils, impacts the progression of cholesterol gallstone disease.