Subsets of thymic resident NKT cells show differential requirements for CD28 co-stimulation during antigen activation

Abstract

Abstract Natural killer T (NKT) cells rapidly respond to antigenic stimulation with cytokine production and direct cytotoxicity. These innate-like characteristics arise from their differentiation into mature effector cells during thymic development. A subset of mature NKT cells remain thymic resident, but their activation and function remain poorly understood. We examined the roles of CD28 and CTLA-4 in driving activation of these thymic resident NKT cells. Unlike previous studies with peripheral NKT cells, the proliferation of thymic NKT cells was significantly impaired when CD28 engagement was blocked - but unaffected with CTLA-4 activation or blockade. Within NKT cell subsets, however, stage 3 NKT cells marked by higher NK1.1 expression were significantly more sensitive to the loss of CD28 signals compared to NK1.1 negative stage 2 NKT cells. In concordance with this, CD28 blockade suppressed all NKT cell cytokines, lowering the ratio of IFN-γ:IL-4 production by NK1.1 positive NKT cells. Intriguingly, the activation-dependent upregulation of the master transcription factor PLZF did not require CD28 co-stimulation in either NK1.1 positive or negative thymic NKT subsets – underlining a dichotomy between requirements for early activation and the subsequent proliferation and effector function of these cells. Collectively, our studies demonstrate the ability of CD28 co-stimulation to fine tune subset-specific responses by thymic resident NKT cells and contextually shape the milieu in this primary lymphoid organ.