All memory T cells mount an accelerated response on antigen reencounter, but significant functional heterogeneity is present within the respective memory T-cell subsets as defined by CCR7 and CD45RA expression, thereby warranting further stratification. Here we show that several surface markers, including KLRB1, KLRG1, GPR56, and KLRF1, help define low, high, or exhausted cytokine producers within human peripheral and intrahepatic CD4 + memory T-cell populations. Highest simultaneous production of TNF and IFN-γ is observed in KLRB1 + KLRG1 + GPR56 + CD4 T cells. By contrast, KLRF1 expression is associated with T-cell exhaustion and reduced TNF/IFN-γ production. Lastly, TCRβ repertoire analysis and in vitro differentiation support a regulated, progressive expression for these markers during CD4 + memory T-cell differentiation. Our results thus help refine the classification of human memory T cells to provide insights on inflammatory disease progression and immunotherapy development.

Memory T cells mount an accelerated response upon re-challenge but are heterogeneous in phenotype and function. Traditionally memory T cells were classified into central memory, effector memory and terminally differentiated effector memory (T EMRA ) cells based on expression of CCR7 and CD45RA. Functional heterogeneity even within these subsets demonstrated the need for more suitable markers. We applied bulk and single gene expression profiling of human CD4 + memory T cells and identified surface markers, KLRB1, KLRG1, GPR56 and KLRF1, allowing classification into "low", "high" or "exhausted" cytokine producers. In contrast to common understanding KLRG1 expression was not associated with exhaustion and highest production of multiple cytokines was observed in KLRB1 + KLRG1 + GPR56 + T cells. Only additional KLRF1expression was associated with a decline in cytokine production. The superiority of KLRF1 to define exhausted cytokine producers compared to classical T EMRA identification was best exemplified for intrahepatic T cells in patients with inflammatory liver diseases.

The intestine mediates a delicate balance between tolerogenic and inflammatory immune responses. The continuous pathogen encounter might also augment immune cell responses contributing to complications observed upon intestinal transplantation (ITx). We thus hypothesized that ITx patients show persistent signs of immune cell activation affecting both the adaptive and innate immune cell compartment. Information on the impact of intestinal grafts on immune cell composition, however, especially in the long-term is sparse. We here assessed activated and differentiated adaptive and innate immune subsets according to time, previous experience of cellular or antibody-mediated rejections or type of transplant after ITx applying multi-parametric flow cytometry, gene expression, serum cytokine and chemokine profiling. ITx patients showed an increase in CD16 expressing monocytes and myeloid dendritic cells (DCs) compared to healthy controls. This was even detectable in patients who were transplanted more than 10 years ago. Also, conventional CD4 + and CD8 + T cells showed persistent signs of activation counterbalanced by increased activated CCR4 + regulatory T cells. Patients with previous cellular rejections had even higher proportions of CD16 + monocytes and DCs, whereas transplanting higher donor mass with multi-visceral grafts was associated with increased T cell activation. The persistent inflammation and innate immune cell activation might contribute to unsatisfactory results after ITx.