A Single-Cell Woodchuck Liver Atlas Identifies Healthy and Disease-Related Cellular Programs Conserved in Human

Abstract

BackgroundModel organisms allowing for longitudinal examinations of liver disease pathogenesis are pivotal for the development of new therapeutic modalities. The eastern North American woodchuck develops chronic hepatitis and liver cancer after woodchuck hepatitis virus (WHV) infection, mirroring aspects of the natural history of the human hepatitis B virus (HBV). However, the cellular landscape of the woodchuck liver and the cell-level relevance of WHV infection to HBV infection is currently uncharacterized. MethodsWe employed single-cell RNA sequencing (RNA-seq) to generate an atlas of healthy woodchuck liver (63,389 cells, n=8) and peripheral blood mononuclear cells (PBMCs) (26,972 cells, n=7). Cell-specific and hepatic zonation gene signatures were validated using spatial transcriptomics (n=1). We employed our atlas to examine immune activation in stimulated precision cut-liver slices (PCLS) and disease-related pathway activation in chronic WHV infection (11,797 cells, n=3). We further employed our atlas to examine shared disease pathways between WHV infection and human HBV infection. ResultsOur atlas revealed woodchuck hepatic cellular diversity comparable to human and murine livers. Applying single-nucleus RNA-seq to PMA/ionomycin-stimulated precision cut liver slices revealed inflammation-associated activation signatures in T cell, myeloid and endothelial cell compartment. Finally, we describe intrahepatic T cells in chronic WHV hepatitis with both exhaustion and activation-associated signatures that resemble intrahepatic T cell genes signatures described in human chronic HBV. ConclusionsWe present a multi-omic atlas of healthy, diseased and ex vivo stimulated woodchuck liver. By identifying shared pathological processes between WHV and HBV infections, our findings reinforce the value of this preclinical model in translational research. This resource aims to advance studies on HBV pathogenesis and oncogenesis to speed the development of novel therapeutic strategies. Impact and Implications/Lay summaryThe liver plays important roles in metabolism, detoxification, and immune processes; liver transplantation is often the only treatment option for severe chronic liver diseases. Therefore, developing animal models that reflect human liver disease and can be studied throughout the disease course is crucial for the discovery of new treatment options. The woodchuck is an animal that develops chronic hepatitis and liver cancer after infection with woodchuck hepatitis virus, which models the human hepatitis type B virus infection (HBV) and associated hepatic carcinoma. However, our understanding of the cells that compose the woodchuck liver is limited, making it challenging to design and test cell-based therapeutics. In this study, we atlased the healthy and chronically infected woodchuck liver, found that liver cell types in woodchuck resemble those in humans, and employed the atlas to show similarities between WHV and HBV at the cell level, reinforcing the potential of WHV-infected woodchuck as a model for human HBV disease. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=196 SRC="FIGDIR/small/641192v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@18fdd40org.highwire.dtl.DTLVardef@6ccf2org.highwire.dtl.DTLVardef@7923e1org.highwire.dtl.DTLVardef@122a8d2_HPS_FORMAT_FIGEXP M_FIG C_FIG