Adaptive features of natural killer (NK) cells have been reported in various species with different underlying mechanisms. It is unclear, however, which NK cell populations are capable of mounting antigen-specific recall responses and how such functions are regulated at the molecular level. Here, we identify and characterize a discrete population of CD49a+CD16− NK cells in the human liver that displays increased epigenetic potential to elicit memory responses and has the functional properties to exert antigen-specific immunity in the skin as an effector site. Integrated chromatin-based epigenetic and transcriptomic profiling revealed unique characteristics of hepatic CD49a+CD16− NK cells when compared with conventional CD49a−CD16+ NK cells, thereby defining active genomic regions and molecules underpinning distinct NK cell reactivity. In contrast to conventional NK cells, our results suggest that adaptive CD49a+CD16− NK cells are able to bypass the KIR receptor-ligand system upon antigen-specific stimulation. Furthermore, these cells were highly migratory toward chemokine gradients expressed in epicutaneous patch test lesions as an effector site of adaptive immune responses in the skin. These results define pathways operative in human antigen-specific adaptive NK cells and provide a roadmap for harnessing this NK cell subset for specific therapeutic or prophylactic vaccine strategies.

BackgroundTumor-associated macrophages (TAM) constitute the most abundant immune cells in the tumor stroma initiating pro-inflammatory (M1) or immunosuppressive (M2) responses depending on their polarization status. Advances in tumor immunotherapy call for a detailed understanding of potential immunogenic mechanisms of irradiation routinely applied in rectal cancer patients.MethodsTo test the effects of radiotherapy on TAM, we ex vivo irradiated tissue samples of human rectal cancer and assessed the phenotype by flow cytometry. We furthermore evaluated the distribution of leucocyte subsets in tissue sections of patients after short-course radiotherapy and compared findings to non-pretreated rectal cancer using an immunostaining approach. Organotypic assays (OTA) consisting of macrophages, cancer-associated fibroblast and cancer cell lines were used to dissect the immunological consequences of irradiation in macrophages.ResultsWe demonstrate that short-course neoadjuvant radiotherapy in rectal cancer patients is associated with a shift in the polarization of TAM towards an M1-like pro-inflammatory phenotype. In addition, ex vivo irradiation caused an increase in the phagocytic activity and enhanced expression of markers associated with stimulatory signals necessary for T-cell activation. In OTA we observed that this alteration in macrophage polarization could be mediated by extracellular vesicles (EV) derived from irradiated tumor cells. We identified high mobility group box 1 in EV from irradiated tumor cells as a potential effector signal in that crosstalk.ConclusionsOur findings highlight macrophages as potential effector cells upon irradiation in rectal cancer by diminishing their immunosuppressive phenotype and activate pro-inflammation. Our data indicate that clinically applied short-term radiotherapy for rectal cancer may be exploited to stimulate immunogenic macrophages and suggest to target the polarization status of macrophages to enhance future immunotherapeutic strategies.

Mounting experimental evidence hints to an import role for natural killer (NK) cells in adaptive immune responses to pathogens. NK cells with adaptive features are heterogeneous and belong to different subsets according to their phenotype as well as the nature of their adaptive recall reactions. Three types of adaptive NK cell responses have been described: (i) NK cells with long-lived memory of multiple different haptens and viral antigens were described in murine liver tissue with a possible human counterpart; (ii) infection of human and mouse cytomegalovirus is associated with an expansion of NKG2C + and Ly49H + NK cells, respectively, that selectively recognize CMV-encoded peptides thereby facilitating recall responses; (iii) cytokine-stimulated NK cells respond to different stimuli with enhanced production of IFN-γ after re-stimulation. These exciting findings not only support the idea of NK cells with adaptive features, but define a novel field of harnessing memory NK cell subsets for therapeutic strategies.