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

Abstract

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. 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. Hepatic CD4⁺ T cells were delineated into three distinct populations based on CD69 expression: CD69^-, CD69^INT and CD69^HI. CD69^HICD4⁺ 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^HICD4⁺ T cells produced robust type 1 polyfunctional cytokine responses on stimulation. Conversely, CD69^INTCD4⁺ T cells represented a more heterogenous population containing cells with a more activated phenotype, a distinct chemokine receptor profile (CX₃CR1⁺CXCR3⁺CXCR1⁺) and a bias towards interleukin-4 production. While CD69^INTCD4⁺ 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^INTCD4⁺ T cells correlated with necroinflammatory scores in chronic hepatitis B infection. Finally, we demonstrated that interaction with hepatic epithelia was sufficient to generate CD69^INTCD4⁺ T cells, while additional signals from the liver microenvironment were required to generate liver-resident CD69^HICD4⁺ T cells. 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.