SAT-069 FUNCTIONAL SPECIALISATION OF NATURAL KILLER CELL SUBSETS IN HUMAN KIDNEY TRANSPLANT REJECTION

Abstract

Human natural killer (NK) cells are innate lymphocytes implicated in kidney transplant rejection. However, the respective contributions of functionally distinct NK cell subsets (CD56bright cytokine-producing versus CD56dim cytotoxic effector) in episodes of allograft rejection remain uncertain, with current immunohistochemical methods unable to differentiate these discrete subsets. This study uses an innovative multi-colour flow cytometric-based approach to unequivocally define and evaluate NK cell subsets in human allograft rejection. We extracted renal lymphocytes from human transplant biopsies stratified based on the histopathological diagnosis of (a) non-rejection, (b) borderline, (c) T cell-mediated rejection (TCMR), (d) antibody-mediated rejection (ABMR) and (e) mixed (TCMR+ABMR) rejection. NK cell subsets were identified, enumerated and phenotyped (activation state, chemokine receptors,) by fourteen-colour flow cytometry. Dissociation supernatants were harvested and levels of soluble proteins were determined using the LEGENDplex™ Human CD8/NK Panel multiplex bead-based assay. Absolute numbers of only CD56bright NK cells were significantly elevated in TCMR biopsies. In contrast, both CD56bright and CD56dim NK cell numbers were significantly increased in biopsies with ABMR (ABMR and mixed). The CD56dim NK cells in ABMR biopsies also expressed significantly higher levels of activation marker CD69 compared to CD56dim NK cells in non-rejection biopsies. The dissociation supernatants from biopsies of patients diagnosed with ABMR also had significantly elevated levels of cytotoxic effector molecules: perforin, granzyme A and granulysin. Chemokine receptor profiling of the NK cell compartment showed fractalkine ligand CX3CR1 to be exclusively expressed on CD56dim NK cells. Our results indicate that the NK cell subsets are differentially recruited and activated during distinct types of rejection, suggestive of specialised functional roles. Further functional dissection of these subsets may provide useful diagnostic and prognostic information for patients experiencing allograft rejection.