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NATURAL KILLER (NK) CELL CROSSMATCH: FUNCTIONAL ANALYSIS OF INHIBITORY KIR RECEPTORS AND THEIR HLA LIGANDS.
M. Fallena, MD, M. Han, PhD and P. Stastny, MD. Dallas TX, UT Southwestern Med Cntr, Internal Medicine.

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NK cells may influence outcome after hematopoietic cell transplantation. Their function depends on a complex interaction between inhibitory and stimulating receptors and HLA ligands. The goal of this study was to establish a functional assay to correlate with the genetics of these systems. NK cells were tested for cytotoxicity against Con A blasts in a 4-hour ⁵¹Cr release assay. HLA-A, B, C and KIR typing was performed by PCR-SSP in healthy volunteers. Surface expression of KIR receptors was determined by flow cytometry using monoclonal antibodies CD158a, CD158b, NKB1, and CD94. The KIR genotype and the HLA ligands were compared in order to establish the number of inhibitory KIR-HLA matches. Flow cytometry was used to estimate the percentage of NK cells expressing each KIR type that could be inhibited by the HLA ligands in target cells. The K562 cell line was used as positive control and showed cytotoxicity greater than 50% in all cases at E:T ratio 12.5:1. Autologous killing was on average less than 10%. Cytotoxicity was greater than 50% in cases when one KIR was matched, ranged between 7 and 32% when two matches are present and was less than 10% when 3 matches were observed. The number of matches could not be determined only by genotype because of variable expression of KIR receptors. We observed an NK cell donor genotyping for KIR3DL1 but with no surface expression by flow cytometry suggesting this person may carry a null allele. In some cases, cytotoxicity could not be predicted by typing or flow cytometry. Further study is necessary to investigate if the percentage of cytotoxicity in the NK cell crossmatch is clinically correlated to the graft versus leukemia effect or decreased relapse risk after bone marrow transplantation.