RRV-induced biliary atresia in neonatal mice involves CD8 + T lymphocyte killer cells and the Notch signaling pathway.

BACKGROUND

Persistent inflammation induced by viral infection may contribute to the pathogenesis of biliary atresia (BA). Moreover, CD4+ helper cells and CD8+ killer cells are the main effector cells involved in BA and intrahepatic bile duct injuries.

OBJECTIVE

Thus, we aimed to explore the dynamics of inflammatory cell infiltration and inflammation-regulated pathways in liver-specific inflammatory responses.

METHODS

Neonatal Balb/C mice were intraperitoneally infected with 1 × 106 PFU rhesus rotavirus (RRV; BA + group), 1 × 105 PFU RRV (BA- group), or DMEM (control group). Mice were sacrificed 7 or 14 days post-infection and their bile ducts, livers, and spleen-derived tissues were examined via H & E staining. The number of CD4+T lymphocytes helper cells (CD4+Th), CD8+T lymphocytes killer cells (CD8+Tc), natural killer (NK) cells, and macrophages (Mac) in the liver and spleen were quantified by flow cytometry. The expression of inflammatory genes was analyzed via a PCR-array. Western blotting was conducted to quantify the protein expression of Notch receptor active fragments (NICD). Finally, some mice were injected with DAPT (a γ-secretase inhibitor) 12 h post-infection followed by analysis of liver and bile duct tissues after 14 days.

RESULTS

The numbers of CD4+Th cells were increased in the livers of BA- mice after 14 days (P < 0.05). After RRV infection, the number of CD8+Tc, CD4+Th, NK, and Mac were increased in the livers of BA + mice after 7 and 14 days. Notably, NK cell numbers remained elevated in the BA + group, but the number of Mac first increased and then decreased in both the treatment groups. PCR-array analyses indicated that the expression of many genes related to T cell proliferation and differentiation significantly increased in the livers of BA. The most upregulated gene was Jagged2 (20.34-fold). Increased NICD (Notch receptor active fragments) protein expression was found in the BA + group. Finally, DAPT injection could reduce inflammation, CD8+Tc infiltration, NICD expression, and bile duct damage after RRV infection. We found that CD8+Tc played the most important role in damaging bile ducts and promoting BA.

CONCLUSION

The DAPT-based intervention could reduce expression of CD8+Tc and bile duct damage in BA mouse livers post-RRV infection. We believe that the Notch signaling pathway regulates CD8+Tc functions and inflammatory dynamics in BA mouse livers.