Abstract: Biliary Atresia (BA) is a rare childhood disease in which bile is accumulated in the liver due to a complete or partial blockage of the biliary network. The pathogenesis of BA remains unclear. It is suggested that infection with human cytomegalovirus (HCMV) and activation of pro-inflammatory macrophages in the newborn’s liver contribute to the disease initiation and progression of BA. To test this hypothesis, we developed a human isogenic pluripotent stem cell (hiPSC)-derived cholangiocyte and macrophage co-culture to investigate how HCMV-infected macrophages interact with cholangiocytes and contribute to BA pathogenesis. hiPSC-derived macrophages were infected with HCMV (multiplicity of infection = 1.0), and the samples were divided into mock and HCMV-infected groups. Macrophages were co-cultured with hiPSC-derived cholangiocytes in a 1:1 ratio. HCMV-mediated immune responses and cholangiocyte organoid development were evaluated. HCMV infected co-culture formed smaller, multi-cystic and deformed organoids. sc-RNA-seq analysis from post-infection day 1 (PD1) showed that HCMV-infected macrophages expressed pro-inflammatory mediators (IL6 and other chemokines) and acted as pro-inflammatory macrophages. Moreover, ligand and receptor interaction revealed that macrophages derived IL-6 interacts with IL6ST receptors on cholangiocytes, and induced robust inflammatory responses in cholangiocytes, upregulated downstream target genes (SPP1, IL-8, IL-1B) and pro-inflammatory pathways. Immunohistochemistry staining revealed that IL-6, SPP1 and IL-8 expressions were significantly upregulated in HCMV+ BA liver biopsies. Our findings were further validated in the RRV (rhesus rotavirus) experimental BA mice model, in that IL6 and SPP1 expressions were significantly upregulated in experimental BA liver tissues. Our data suggest that macrophage induced inflammation of cholangiocytes via IL-6/IL6ST signaling contribute to the disease initiation and progression of BA.
Funding Source: Theme-based Research Scheme (T12-712/21-R) RGC Hong Kong SAR Government, Hong Kong SAR, China.
