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.