T-bet+ tissue-resident regulatory T cells are enriched in the neonatal mouse liver and promoted by IL-27 and IFN-γ.

Neonatal regulatory T (Treg) cells in secondary lymphoid organs have greater proliferative capacity and more potent suppressive functions than adult Treg cells. However, the phenotypic and functional features of Tregs in neonatal nonlymphoid organs are not well understood. Our prior work demonstrated that thymus-derived Treg cells entering the neonatal mouse liver enhance immune tolerance and periportal liver maturation. Compared to splenic Treg cells, these hepatic Tregs have faster turnover and superior suppression of naïve T-cell proliferation. To further define this population, we conducted single-cell transcriptomic and immunophenotypic analyses of liver- and spleen-derived Tregs from neonatal and adult mice. Our analysis revealed a distinct T-box transcription factor Tbx21 (T-bet)+ effector Treg subset uniquely enriched in the neonatal liver. These cells possess liver-homing properties, clonal expansion of unique T-cell receptor (TCR) repertoires, and heightened suppressive activity. Interleukin-27 (IL-27) and interferon gamma (IFN-γ), cytokines highly expressed in the neonatal liver, play a critical role in promoting T-bet expression and the acquisition of an ectonucleoside triphosphate diphosphohydrolase 1 (CD39)+ C-X-C chemokine receptor type 3 (CXCR3)+ double-positive phenotype in Treg cells. Collectively, our findings characterize a liver-specific neonatal T-bet+ Treg population shaped by the hepatic microenvironment, highlighting its unique tissue-resident signature and immunoregulatory role.