Isogenic pluripotent stem cell(PSC)-derived cholangiocyte and macrophage organoid co-culture for modelling human cytomegalovirus associated biliary atresia [Oral presentation]

Abstract

<p>Purposes: Macrophages are first targets during human cytomegalovirus (HCMV) infection<br>and are crucial for viral persistence and dissemination. Perinatal HCMV infection has been<br>implicated in the pathogenesis of Biliary Atresia (BA), but the pathogenesis of HCMV<br>associated BA remains poorly understood. In this study, we developed an isogenic pluripotent<br>stem cell (PSC)-derived cholangiocyte and macrophage co-culture to investigate the role of<br>CMV infection in the pathogenesis of BA.</p><p><br>Methods: iPSC-derived macrophages were infected with HCMV, and HCMV-infected<br>macrophages were then co-cultured with isogenic PSC-derived cholangiocytes.<br>Morphological/molecular/single-cell RNA-sequencing (sc-RNA-seq) analyses were conducted<br>to investigate the infection/immune responses/cholangiocyte development in the co-culture.</p><p>Results: In contrast to the expanded mono-cystic organoids in the mock culture, multi-cystic,<br>deformed, and poorly expanded organoids were formed in the HCMV infected co-culture. Postinfection<br>day 1 (PD-1) HCMV infected cholangiocytes proliferated, released proinflammatory<br>cytokines and chemokines (IL8, IL1B, CXCL5, and CXCL1), down-regulated cholangiocyte<br>markers KRT19 and EPCAM expression. In PD-5 infected co-culture, we observed (i) viral<br>gene UL22A expression mainly in the BCL2- large cholangiocytes; (ii) an expansion of large<br>cholangiocyte population; (iii) a reduced percentage of large cholangiocyte that expressed<br>cholangiocyte markers (KRT19, EPCAM).</p><p><br>Conclusion: Our data showed that (i) infected macrophages could transfer HCMV to large<br>cholangiocytes; (ii) HCMV infection of cholangiocytes induced upregulation of<br>proinflammatory factors, proliferation and downregulation of cholangiocytes markers.<br>Together these findings indicated that HCMV infection induced abnormal development and<br>proliferation of bile duct cells, and promoted inflammation in liver. Our co-culture allows us<br>to model BA and to investigate the virus-macrophages-cholangiocytes interactions in the<br>pathogenesis of BA.</p>