KIR Gene And Its Match With HLA-Cw Impact On NK Cell Cytotoxicity

Objective:The aim was to study how the killer immuneglobulin-like inhibition receptor(KIR) and its match HLA-Cw impact on NK cell activity;To investigate how the KIR2DS1(the surface of killer cell immunoglobulin-like inhibitory receptors,KIR) gene expression effected on NK(natural killer) cells killing activities against AML(acute myeloid leukemia) target cells;To further exploring the KIR / HLA-Cw(human leucocyte antigen-Cw,HLA-Cw loci) mismatches destruction mechanism。Method:The KIR and HLA gene typing of 27 healthy donors and 30 AML patients were performed by PCR -SSP;High-purified NK cells separated by DNYAL bead negative selection were taken as effector cells,and the freshly isolated bone marrow mononuclear cells from newly diagnosed AML patients as target ceils.The anti-CD158a, CD158b monoclonal antibody was used to block KIR2DL1,KIR2DL2/3 receptors of NK cells;The NK cells cytotoxicities against target cells with different KIR/HLA-Cw combinations before and after KIR receptors blockades were detected by methyl thiazolyl tetrazolium(MTT) reduction assay in vitro。Results:The purity of NK cell CD3-/CD56+16+ is 90.8%±6.08%,anaylsed by Flow cytometer(FCM);The less number of KIR /HLA-Cw matches the the more activity of NK cells,0 match of NK cell/target cell KIR/HLA-Cw,cytotoxicity was 50.66%±8.40%,1 match and 2 matches were 38.28%±6.71%,19.74%±4.15%,F=20.226,P＜0.001;NK cells expressed KIRs also had relationship with cytotoxicity,inhibitory KIRs expressed＞50%,the cytotoxicity is 10%,inhibitory KIRs expressed 20%-50%,the cytotoxicity is 20%, inhibitory KIRs＜25%,the cytotoxicity is 55%,F=16.276,p＜0.001;The killing effect of NK ceils significantly enhanced after inhibitory receptor KIRs blockades(t =- 3.00,P = 0.005);The cell lysis of C1 group,KIR2DS1+ NK cells against C2 group target ceils (57.370%±1.400%) was significantly higher than that against C1 group(44.190%±4.666%)and C1/C2 group(36.770%±6.56%)target cells(F = 11.87,P = 0.021), furthermore the differences of cell lysis among the above three groups become more evident after the KIR receptors blockades(F = 18.72,P = 0.009)。Concision:NK cell kill AML leukemic cells mechanism follow KIR/HLA-Cw mismatch theory;the level of inhibitory KIRs expressed on NK cell surface showed the percentage of NK cells that could be inhibited by the HLA-Cw ligands;The interaction between inhibitory KIR and HLA ligands,and also KIRs expression on NK cells both impact significantly on NK cell function;That less KIR/HLA-Cw match and less expression of KIRs on NK ceils result in more NK cell activated with stronger killing effect;The activity of NK cells increased after the inhibitory KIR receptors blockades;And the killing activities of KIR2DS1 + NK cells was higher than KIR2DS1- NK cells;C1 group, KIR2DS1 + NK cells lysed more C2 group target cells than the C1 and C1/C2 group target ceils in vitro。...