The human liver microenvironment shapes the homing and function of CD4(+) T-cell populations

OBJECTIVE: Tissue-resident memory T cells (T(RM)) are vital immune sentinels that provide protective immunity. While hepatic CD8(+) T(RM) have been well described, little is known about the location, phenotype and function of CD4(+) T(RM). DESIGN: We used multiparametric flow cytometry, histological assessment and novel human tissue coculture systems to interrogate the ex vivo phenotype, function and generation of the intrahepatic CD4(+) T-cell compartment. We also used leukocytes isolated from human leukocyte antigen (HLA)-disparate liver allografts to assess long-term retention. RESULTS: Hepatic CD4(+) T cells were delineated into three distinct populations based on CD69 expression: CD69(−), CD69(INT) and CD69(HI). CD69(HI)CD4(+) cells were identified as tissue-resident CD4(+) T cells on the basis of their exclusion from the circulation, phenotypical profile (CXCR6(+)CD49a(+)S1PR1(−)PD-1(+)) and long-term persistence within the pool of donor-derived leukcoocytes in HLA-disparate liver allografts. CD69(HI)CD4(+) T cells produced robust type 1 polyfunctional cytokine responses on stimulation. Conversely, CD69(INT)CD4(+) T cells represented a more heterogenous population containing cells with a more activated phenotype, a distinct chemokine receptor profile (CX(3)CR1(+)CXCR3(+)CXCR1(+)) and a bias towards interleukin-4 production. While CD69(INT)CD4(+) T cells could be found in the circulation and lymph nodes, these cells also formed part of the long-term resident pool, persisting in HLA-mismatched allografts. Notably, frequencies of CD69(INT)CD4(+) T cells correlated with necroinflammatory scores in chronic hepatitis B infection. Finally, we demonstrated that interaction with hepatic epithelia was sufficient to generate CD69(INT)CD4(+) T cells, while additional signals from the liver microenvironment were required to generate liver-resident CD69(HI)CD4(+) T cells. CONCLUSIONS: High and intermediate CD69 expressions mark human hepatic CD4(+) T(RM) and a novel functionally distinct recirculating population, respectively, both shaped by the liver microenvironment to achieve diverse immunosurveillance.