OBJECTIVETo detect the diversity of killer Ig-like receptor(KIR) gene content and the combination of haplotypes in Chinese Han population in Shanghai area.

METHODSDNA samples from 87 randomly unrelated healthy individuals in Shanghai Han population were genotyped with SSP/PCR method.

RESULTS(1) Frequencies of KIR genes: All of 18 known KIRs genes, such as 2DL1-5, 2DS1-5, 3DL1-3, 3DS1, KIR1D and the pseudogenes X, Xv and Z(KIR2DP1) were observed in Shanghai Hans. All individuals contain 3DL3, 2DL4, 3DL2 and 3DL1; the most common genes were 2DL3, Z, 2DL1 and X; the following were 2DS4, 1D, 2DL5, 2DS1, 3DS1 and 2DS5; the next were 2DS2, 2DL2, 2DS3 and Xv. (2) Frequencies of KIR gene haplotypes; there were 13 haplotypes detected in 87 Han individuals, among them, the most frequent one was type 2 (haplotypeA-2DS4). (3) Frequencies of KIR genotypes: 18 kinds of the combinations of the haplotypes were observed; the most frequent ones were AJ(2,2), AF (1,2). Also, In this study were identified five new genotypes FZ1 2 9 , FZ2 1 16 , FZ3 6 17 , FZ4 4 13 and FZ5 2 6 ,which had not been observed in Caucasians so far.

CONCLUSIONThese findings suggest that there are distinctive frequencies of KIR gene content, haplotype as well as genotype in Chinese Han population in Shanghai area.

China ; Gene Frequency ; Genetic Variation ; Genotype ; Haplotypes ; genetics ; Humans ; Receptors, Immunologic ; genetics ; Receptors, KIR ; Receptors, KIR2DL1 ; Receptors, KIR2DL3 ; Receptors, KIR2DL4 ; Receptors, KIR3DL1 ; Receptors, KIR3DL2 ; Receptors, KIR3DS1

OBJECTIVETo analyze the allelic polymorphism of killer cell immunoglobulin-like receptor 2DL3 (KIR2DL3) gene in Zhejiang Han population.

METHODSAllelic of KIR2DL3 was detected by PCR sequence based typing (PCR-SBT) method. Two specific fragments of KIR2DL3 were amplified using different primers and sequenced for exons 4,5,7,8,9 of KIR2DL3. All the samples would be assigned to KIR2DL3 allele according to the polymorphism site patterns.

RESULTSTwo KIR2DL3 alleles were observed, with KIR2DL3*001 having higher frequency, 0.77.

CONCLUSIONThe PCR-SBT method is reliable and there are distinctive frequencies of KIR2DL3 alleles in Zhejiang Han population.

Alleles ; Base Sequence ; China ; Gene Frequency ; Genotype ; Humans ; Polymerase Chain Reaction ; Polymorphism, Genetic ; genetics ; Receptors, KIR2DL3 ; genetics ; Sequence Analysis, DNA

The aim of this study was to analyze the promoter methylation patterns of inhibitory killer cell immunoglobulin-like receptor (KIR) which gene expression and the effect of demethylation treatment were studied, and to explore the possible regulation mechanism of inhibitory kir gene expression. The promoter methylation levels of kir2DL1 and kir2DL2/kir2DL3 in NK-92MI cell line were detected by bisulfite sequencing technique. Then NK-92MI cells were treated with 5-azacytidine to induce the demethylation of CpG islands. The levels of gene expression of kir were determined by RT-PCR. The results demonstrated that the methylation frequencies of CpG dinucleotides surrounding the promoter regions of kir2DL1 and kir2DL2/kir2DL3 genes were 25% to 88% and 5% to 80% respectively. DNA-demethylating treatment with 5-azacytidine resulted in re-expression of kir2DL1 gene and increased expressions of kir2DL1, kir2DL2 and kir2DL3 genes in NK-92MI cells. In conclusion, the promoter DNA methylation participates in the regulation of kir gene expression in NK-92MI cells.
Cell Line ; DNA Methylation ; Gene Expression ; Humans ; Killer Cells, Natural ; metabolism ; Receptors, KIR2DL1 ; genetics ; metabolism ; Receptors, KIR2DL2 ; genetics ; metabolism ; Receptors, KIR2DL3 ; genetics ; metabolism

OBJECTIVETo explore the genetic variations of HLA-Cw and 5 KIR2D loci in 2 Chinese Han populations residing at Southern and Northern mainland China, respectively, and to investigate the HLA-Cw polymorphism of a Mongolian Chinese population.

METHODSHLA-Cw genotyping was performed in a total of 293 healthy individuals including 1 Southern Han population living in Hunan Province (n=112), 1 Northern Han population (n=98) and 1 Mongolian Chinese population(n=83) in the Inner Mongolia Autonomous Region, using polymerase chain reaction-sequence specific primer(PCR-SSP) technique. Dimorphism at residue 80 of domain in the HLA-Cw molecule was examined by an additional set of PCR-SSP reactions. PCR-SSP was also used to detect the presence or absence of inhibitory KIR2DL1/2DL2/2DL3 loci and activating KIR2DS1/2DS2 loci for the 2 Han populations.

RESULTSThe main findings were: (1) Very significant frequency difference in the HLA-Cw alleles and dimorphism at codon 80 was detected between Hunan Han and Northern Han population, and between Hunan Han and Mongolian population (P < 0.001),while there was no such difference between the 2 Northern Chinese populations (P> 0.05); (2) There was no significant difference in frequencies of either the 5 individual KIR2D genes or the genotype distributions between the 2 Han populations (P> 0.05); (3) Asn(80)ls/Asn(80), 2DL1+/2DL2-/2DL3+/2DS1-/2DS2- predominated in both Han populations (45/112, 29/98), followed by Asn(80)/Asn(80), 2DL1+/2DL2-/2DL3+/2DS1+/2DS2- (18/112,16/98) and Asn(80)/Lys(80), 2DL1+/2DL2-/2DL3+/2DS1-/2DS2-(11/112,17/98). Among the 12 types of HLA-Cw codon 80 and KIR2D combinations, only Lys(80)/Lys(80), 2DL1+/2DL2-/2DL3+/2DS1-/2DS2- showed marginally significant frequency difference between the 2 Han populations(1/112 vs 8/98; Fisheros P was 0.0134).

CONCLUSIONOur study provided the polymorphism data of HLA-Cw gene for 3 Chinese populations with different geographic and/or ethnic background, we further analyzed the distribution of 5 KIR2D receptor genes in 2 Han populations. Our data suggest that in spite of HLA-Cw heterogeneity, remarkable similarities may exist between the Southern and Northern Chinese Han populations at the combinational level of HLA-Cw and KIR2D, which are characterized by preponderant inhibitory signal pathways.

Asian Continental Ancestry Group ; Genetic Variation ; genetics ; HLA-C Antigens ; genetics ; Humans ; Polymerase Chain Reaction ; Polymorphism, Single Nucleotide ; genetics ; Receptors, KIR ; genetics ; Receptors, KIR2DL1 ; genetics ; Receptors, KIR2DL2 ; genetics ; Receptors, KIR2DL3 ; genetics

In order to investigate the effect of demethylating treatment on the expression of inhibitory KIR and the cytolytic activity of NK-92MI cells, and to study the possible relationship between the demethylation of inhibitory kir gene and the function of NK cells. NK-92MI cells were treated with 5-azacytidine to induce DNA demethylation. The expression of KIR3DL1, KIR2DL2/KIR2DL3, KIR2DL1 and the viability of NK-92MI cells were detected by flow cytometry. The KIR3DL1 positive and the KIR3DL1 negative NK-92MI cells were also sorted by flow cytometry. The cytotoxicity of NK-92MI against K562 cells was detected by the LDH release assay. The results demonstrated that the expressions of KIR3DL1, KIR2DL2/KIR2DL3 and KIR2DL1 in NK-92MI cells all increased after treating with 1.0, 2.5 and 5 micromol/L of 5-azacytidine for 72 hours. And the cytotoxicity of NK-92MI against K562 cells decreased. In these dose range, 5-azacytidine did not influence the viability of NK-92MI cells. Additionally, the cytotoxicity of KIR3DL1 positive NK-92MI cells was lower than that of the KIR3DL1 negative cells. It is concluded that the demethylating treatment suppresses the cytotoxicity of NK-92MI cells through increasing the expression of inhibitory KIR.
Cytotoxicity, Immunologic ; immunology ; DNA Methylation ; Flow Cytometry ; Gene Expression Regulation ; Humans ; K562 Cells ; Killer Cells, Natural ; immunology ; metabolism ; Receptors, KIR2DL1 ; metabolism ; Receptors, KIR2DL3 ; metabolism ; Receptors, KIR3DL1 ; metabolism ; Receptors, KIR3DL2 ; metabolism

The objective of this study was to elucidate the correlation of killer immunoglobulin-like receptor (KIR) gene diversity with nasopharyngeal carcinoma (NPC) in the Chinese southern Han population. KIR genotyping of peripheral blood samples from 67 patients with NPC and 77 randomly-selected healthy controls was performed by PCR-SSP, the relative risk (RR) value was calculated by means of Wolf method. The results showed that the KIR2DL3 gene frequency in NPC patient group was significantly lower than that in healthy controls (χ²>3.84, p < 0.05, RR = 0.08), whereas the KIR2DS5 and KIR2DL5B gene frequencies in patient group were significantly higher than those in healthy controls (χ²>3.84, p < 0.05, RR > 1), the other KIR gene frequencies were no statistically different between two groups. It is concluded that the KIR2DL3, KIR2DS5 and KIR2DL5B genes may be correlated with pathogenesis of NPC in the Chinese southern Han population.
Adult ; Aged ; Asian Continental Ancestry Group ; genetics ; Case-Control Studies ; Female ; Gene Frequency ; Genotype ; Humans ; Male ; Middle Aged ; Nasopharyngeal Neoplasms ; genetics ; Receptors, KIR ; genetics ; Receptors, KIR2DL3 ; genetics ; Receptors, KIR2DL5 ; genetics

The study was purposed to investigate the polymorphism of killer cell immunoglobulin-like receptor (KIR) gene of the patients with leukemia and to explore the correlation between the KIR gene and susceptibility of leukemia. The KIR genotype of 50 patients with leukemia and 60 healthy controls in northern. Hans were analyzed by PCR-SSP. The results indicated that the present known 18 KIR genes were detected and identified. The frequencies of KIR 3DL3, 3DL2 and 2DL4 were 100% in all subjects, with the most frequent genotype KIR 3DP1 (0.86) followed by 2DP1, 2DL3, 3DL1, 2DL1, 3DS1, 2DL5, 2DS4, 2DS2, 1D, 2DS5, 2DL2, 2DS1, 2DS3 and 3DP1v in leukemia successively. Compared with the control, the KIR 3DL1 (0.60) and 2DL1 (0.57) were significantly lower in the leukemia patient group than that in the control group (1.00) (P < 0.01). It is concluded that the polymorphism of KIR gene is associated with susceptibility of leukemia in Hans. There may be a negative correlation between pathogenesis of leukemia and KIR 3DL1, KIR 3DS1, KIR 2DL1, KIR 2DL5 genes.
Adolescent ; Adult ; Child, Preschool ; Female ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; Male ; Middle Aged ; Polymorphism, Genetic ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; Receptors, Immunologic ; genetics ; Receptors, KIR ; Receptors, KIR2DL1 ; Receptors, KIR2DL3 ; Receptors, KIR2DL4 ; Receptors, KIR3DL1 ; Receptors, KIR3DL2 ; Receptors, KIR3DS1

The study was aimed at the exploration of relationship between T cells expressing killer cell inhibitor receptors (KIR, CD158 and CD94) and graft-versus-host disease (GVHD) after hematopoietic stem cell transplantation. The expression rates of CD158a, CD158b and CD94 on T cells and NK cell were detected by flow cytometry and donor/recipient HLA-Cw was analyzed using PCR after peripheral blood stem cell transplantation (PBSCT) and umbilical cord blood transplantation (UCBT). After both PBSCT and UCBT, the rates of CD3(+)CD158a(+) and CD3(+)CD158b(+) T cells increased, especially the rate of CD8(+)CD158b(+) T cells. In both acute and chronic GVHD groups, the rate of CD3(+)CD158b(+) T cells increased, especially in acute GVHD. The CD94 mainly expressed on CD3(+)CD8(+) T cells. The percentage of the expression of CD94 on CD4(+) and CD8(+) cells after UCBT and PBSCT increased significantly. The expression of KIR in GVHD (early stage of transplantation) increased but the expression of KIR in chronic GVHD (advanced stage of transplantation) decreased. Five patients who HLA-Cw matched had no severe GVHD. In four patients who underwent allo-PBSCT and UCBT from related HLA-matched donors, only 2 patients had no aGVHD. Four patients underwent transplantation from unrelated HLA-matched donors had GVHD. These observations suggested that there is some relationship between GVHD and KIR expression on T cells. CD158b might be an inhibitory molecule of T cell activated at early stage after transplantation. Understanding the mechanism of GVHD with the expression of KIR on T cells, especially those binding the HLA-Cw might shed light on the establishment of the specific immunotolerance for the prevention of GVHD. To pay attention to HLA-Cw typing is very important to reduce GVHD and increase GVL effect in related or unrelated HLA-matched transplantation.
Antigens, CD ; analysis ; Antigens, Differentiation, T-Lymphocyte ; analysis ; Genotype ; Graft vs Host Disease ; etiology ; HLA-C Antigens ; genetics ; Hematopoietic Stem Cell Transplantation ; Humans ; Lectins, C-Type ; Receptors, Immunologic ; analysis ; Receptors, Interleukin-2 ; analysis ; Receptors, KIR ; Receptors, KIR2DL1 ; Receptors, KIR2DL3 ; T-Lymphocytes ; immunology

OBJECTIVETo analyze the changes of inhibitory killer cell immunoglobulin-like receptors (KIRs), NKG2D receptor and the cytotoxicity of natural killer (NK) cells induced by persistent exposure to CNE2 cells.

METHODSThe HLA-class I genotypes of CNE2 cells and KIR genotypes were determined by PCR with sequence-specific primers (PCR-SSP). The expressions of KIR2DL1, KIR2DL3, KIR3DL1, and NKG2D by the NK cells (freshly isolated NK cells, NK cells cocultured with 100 U/ml IL2 or with 100 U/ml IL2 and CNE2 cells as the control, IL2 and CNE2 groups, respectively) were analyzed by flow cytometry. Cytotoxicity of NK cells against CNE2 cells were detected by LDH releasing assay.

RESULTSThe HLA genotypes of CNE2 cells were A2, 24, B18, 35, Cw4, 7. NK cells isolated from 3 healthy donors expressed KIR2DL1, KIR2DL3, and KIR3DL1. After 4, 24 and 48 h of culture, NK cells in CNE2 group displayed higher KIR2DL1, KIR2DL3 but lower NKG2D expression than those in the control and IL2 groups (P<0.01), whereas the latter two groups showed no significant difference in KIR2DL1, KIR2DL3, and NKG2D expressions (P>0.05), and no difference in KIR3DL1 expression was found between the 3 groups (P>0.05). After 24 h of culture, the cytotoxicity against CNE2 cells mediated by the NK cells in IL2 and CNE2 groups were (26.96-/+1.47) % and (2.74-/+1.64) % at E:T ratios of 10:1, and (35.74-/+3.59)% and (4.57-/+2.41) % at E:T ratio of 20:1, respectively. NK cells in CNE2 group displayed lower cytotoxicity than those in IL2 group (P<0.01).

CONCLUSIONSPersistent exposure to tumor cells expressing NKG2D ligands can lead to downregulated expression of NKG2D receptor, increased expression of KIRs and reduction of NK-mediated cytolysis. These results elucidate the molecular mechanism of reduced cytotoxicity mediated by the edited NK cells and indicate that blocking HLA-class I-bound KIRs or enhancing the expression of NKG2D may promote NK cell-mediated cytolysis.

Cell Line, Tumor ; Cell Survival ; immunology ; Cytotoxicity, Immunologic ; immunology ; Flow Cytometry ; Humans ; Killer Cells, Natural ; cytology ; immunology ; metabolism ; NK Cell Lectin-Like Receptor Subfamily K ; Nasopharyngeal Neoplasms ; immunology ; metabolism ; pathology ; Receptors, Immunologic ; metabolism ; Receptors, KIR ; metabolism ; Receptors, KIR2DL1 ; metabolism ; Receptors, KIR2DL3 ; metabolism ; Receptors, Natural Killer Cell

UNLABELLEDThe purpose of this study was to evaluate the effects of cellular immunity activation on P58(+) cells expressing killer cell inhibitory receptor (KIR) and their regulatory function on cellular immunity, and provid theoretical data for preventing graft-vers-host disease (GVHD) in stem cell transplantation therapy. The mononuclear cells from human peripheral blood were incubated with IL-2, Con A and Lipostin (LP) for 72 hours. The KIR expressing cells, P58.1(+) and P58.2(+) cells, were analyzed by flow cytometry. The results showed that the percentages of CD3(+), CD4(+), CD8(+), CD16(+)CD56(+), P58.1(+) and P58.2(+) cells were greatly increased after treated with IL-2, Con A and LP, separately or in combination, and the percentages of above cells in combined treatment groups were higher than those of single stimulated groups, especially the percentage of cells in the IL-2 + LP group was significantly higher than those in IL-2 and LP singly treated groups.

IN CONCLUSIONIL-2, Con A and LP possess the ability to induce the expression of KIR and stimulate proliferation of P58.1(+) and P58.2(+) cells while to activate the celluar immunity response, the expression of P58 gene may be regulated by the activation of cellular immunity.

Adult ; CD3 Complex ; analysis ; CD4 Antigens ; analysis ; CD56 Antigen ; analysis ; CD8 Antigens ; analysis ; Cell Count ; Cell Division ; drug effects ; Concanavalin A ; pharmacology ; Flow Cytometry ; Humans ; Interleukin-2 ; pharmacology ; Leukocytes, Mononuclear ; cytology ; drug effects ; immunology ; Receptors, IgG ; analysis ; Receptors, Immunologic ; analysis ; Receptors, KIR ; Receptors, KIR2DL3