Establishment of a PCR-SSP method for the simultaneous amplification and identification of the presence of KIR genes / 中华医学遗传学杂志

OBJECTIVE@#To develop a polymerase chain reaction-sequence specific primer (PCR-SSP) method for simultaneous amplification and identification of the KIR genes among Chinese population.@*METHODS@#Peripheral blood samples from 132 healthy donors who had given blood at Shenzhen Blood Center from January 2015 to November 2015 were selected as the study subjects. Based on the polymorphism and single nucleotide polymorphism (SNP) information of high-resolution KIR alleles in the Chinese population and the IPD-KIR database, specific primers were designed to amplify all the 16 KIR genes and the 2DS4-Normal and 2DS4-Deleted subtypes. The specificity of each pair of PCR primers was verified by using samples with known KIR genotypes. During PCR amplification of the KIR gene, co-amplification the fragment of human growth hormone (HGH) gene by multiplex PCR was used as the internal control to prevent false negative results. A total of 132 samples with known KIR genotypes were randomly selected for blind inspection to verify the reliability of the developed method.@*RESULTS@#The designed primers can specifically amplify the corresponding KIR genes, with clear and bright bands for the internal control and KIR genes. The results of detection are fully consistent with the known results.@*CONCLUSION@#The KIR PCR-SSP method established in this study can yield accurate results for the identification of the presence of KIR genes.

Association of molecular genetic polymorphism of KIR-HLA systems with acute leukemia in Southern Chinese Han / 中华医学遗传学杂志

OBJECTIVE@#To investigate the association of molecular genetic polymorphism of KIR-HLA systems with acute lymphoblastic leukemia (ALL) and acute myelocytic leukemia (AML) in southern Chinese Han.@*METHODS@#A total number of 323 cases of adult ALL patients, 350 adult AML, and 745 random healthy controls were tested by KIR PCR-SSP and HLA-A, -B, -C sequence-based typing (PCR-SBT) methods. The molecular genetic polymorphisms of KIR genes and KIR gene profiles, classⅠ HLA ligands, and KIR receptor +HLA ligand combinations were compared between patient and healthy control groups.@*RESULTS@#A total number of 32 and 33 different kinds of KIR profiles were identified in the ALL and AML patient groups. Compared with the observed frequencies of KIR profiles in healthy controls, the observed frequencies of KIR profile AA1 were significantly lower in both the ALL and AML groups (ALL group: 45.79% vs. 55.30%, Pc=0.004; AML group: 48.27% vs. 55.30%, Pc=0.030). In the ALL group, the observed frequencies of 2DL2 gene and 2DL2+HLA-C1 combination, 2DS2 gene and 2DS2+HLA-C1 combination were significantly higher than those in healthy controls (P<0.05), whereas the frequencies of 2DL3 gene, HLA-A3/A11 ligand and 3DL2+HLA-A3/A11 combination were significantly lower than those in healthy controls. However, no significant differences remained after Bonferroni correction (Pc>0.05). In AML group, the observed frequencies of both 2DS1 and 2DL5 genes were significantly higher than that in healthy controls, whereas the frequencies of HLA-C2 ligand and 2DL1+HLA-C2 combination were significantly lower than that in healthy controls(P<0.05). However, no significant difference existed after Bonferroni correction (Pc>0.05).@*CONCLUSION@#This study revealed some potential susceptibility or protective factors related to acute leukemia in southern Chinese Han, especially the protective factor KIR profile AA1, which might provide new clues and theoretical basis for the pathogenesis of acute leukemia and individualized immunotherapy.

Distribution of KIR/HLA alleles among ethnic Han Chinese patients with hepatocellular carcinoma from southern China / 中华医学遗传学杂志

OBJECTIVE@#To assess the association of KIR/HLA alleles with hepatocellular carcinoma (HCC) and hepatitis B virus (HBV) infection among ethnic Han Chinese patients from southern China.@*METHODS@#For 95 patients with HCC and 171 healthy controls, the genotype of HLA-C alleles was determined with a PCR sequence-specific oligonucleotides typing method on an Illumina GenDx NGSgo platform. Genotypes comprised of HLA-C and KIR gene alleles were also subjected to statistical analysis.@*RESULTS@#In total 16 KIR genes (2DL2, 2DS2, 2DS3, 2DS5, 3DS1, 2DS1, 2DL5, 2DS4, 3DL1, 3DP1, 2DL3, 2DP1, 3DL3, 2DL1, 3DL2 and 2DL4) were discovered in the two groups. The frequencies of KIR2DL3 alleles and combinational genotypes of KIR2DL3/HLA-C1C2 were significantly lower in the patient group compared with the controls (0.9368 vs. 0.9883, χ²>3.84; P3.84; P<0.05, RR = 0.03). The frequency of HLA-C2C2 genotype of the patient group was significantly lower than that of the controls (0.0316 vs. 0.2690, P<0.05, RR = 0.09), while the frequency of HLA-C1C2 genotype was significantly higher than that of the controls (0.2316 vs. 0.0058, P<0.05, RR = 51.23).@*CONCLUSION@#Above results suggested that the KIR2DL3 allele is associated with lower risk for HCC. There may be individual difference in patients with HCC and HBV infection but various combinations of KIR/HLA alleles.

A protective effect conferred by KIR3DL1 and its cognate ligand against cervical cancer among ethnic Han Chinese population and its potential mechanism / 中华医学遗传学杂志

OBJECTIVE@#To explore the role of inhibitory KIR (iKIR) and its cognate HLA ligand in the occurrence and development of cervical cancer among ethnic Han Chinese and its potential mechanism.@*METHODS@#Peripheral blood samples from 265 Han Chinese patients with cervical intraepithelial neoplasia (CIN)/cervical cancer and 200 ethnically matched healthy controls were collected. The results of KIR PCR-SSP, HLA PCR-rSSO and KIR3DL1 PCR-SBT, together with cervical cancer data from the TCGA database, were used to assess the association of iKIR genes, receptor-ligand gene combinations, iKIR transcription level in the tumor tissue and the KIR3DL1 alleles with the occurrence and development of cervical cancer.@*RESULTS@#Among the four iKIR genes (KIR2DL1, 2DL2/3, 3DL1 and 3DL2), the frequencies of KIR3DL1 and KIR3DL1-HLA-Bw4 genes among controls were significantly higher than those of the cervical cancer group (96.5% vs. 87.0%, P = 0.018; 81.5% vs. 64.8%, P=0.009). The survival rate of cervical cancer patients with a high transcription level of KIR3DL1 in tumor tissues was significantly higher than those with a low/medium transcription level (P=0.028). The frequency of strong-inhibitory and high-expression KIR3DL1*01502 allele among the healthy population was significantly higher than that of the cervical cancer group (76.0% vs. 59.3%, P =0.015).@*CONCLUSION@#Combined KIR3DL1 and KIR3DL1-HLA-Bw4 can confer a protective effect against the development of cervical cancer, which may be attributed to the strong-inhibitory and high-expression allele of KIR3DL1*01502.

Genetic polymorphisms of KIR2DS4 gene among ethnic Hans from southern China / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To study genetic polymorphisms of the KIR2DS4 gene among ethnic Hans from southern China.</p><p><b>METHODS</b>Genomic DNA was isolated from 306 unrelated individuals and amplified with KIR2DS4-specific PCR primers. KIR2DS4-positive samples were genotyped for the entire coding sequence by sequencing-based typing (SBT). Assignment of allelic genotypes was accomplished by using Assign 3.5 software. For samples with inconclusive SBT results, RT-PCR products covering the entire coding sequence of the KIR2DS4 gene were subjected to cloning and haplotype sequencing.</p><p><b>RESULTS</b>Among all tested samples, 297 were demonstrated to have carried the KIR2DS4 framework gene. For KIR2DS4-positive samples subjected to SBT for the entire coding sequences, no background was observed with the obtained sequences. Three of the seven identified alleles were of novel types, which were officially named by the KIR subcommittee of the World Health Organization Nomenclature Committee for Factors of HLA System. The observed frequencies for the 7 alleles were KIR2DS4*00101 (78.8%), *003 (10.5%), *004 (16.0%), *010 (23.2%), *017 (0.3%), *00105 (0.3%) and *018 (0.7%), respectively. Allele KIR2DS4*007 was not found. The overall frequency for normal cell-surface expression KIR2DS4 alleles including 2DS4*00101, *017 and *00105 was 79.4%, and that for non cell-surface expression alleles including 2DS4*003, *004, *010 and *018 was 50.4%. The ratio between the two was 1.6:1.</p><p><b>CONCLUSION</b>The present study has elucidated the allelic diversity of KIR2DS4 among ethnic Hans from southern China, which may provide valuable data for transplantation as well as studies on KIR-associated disease and evolution.</p>

Association of genetic polymorphisms of KIR-HLA system with chronic myeloid leukemia among ethnic Hans from southern China / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the association of KIR-HLA gene polymorphism with chronic myeloid leukemia (CML) among ethnic Hans from southern China.</p><p><b>METHODS</b>A total of 172 adult CML patients and 480 unrelated healthy controls were screened for the presence of KIR with sequence-specific primers-PCR (PCR-SSP) and sequence-based typing (SBT) of HLA-A, -B and -C loci. Polymorphisms of the KIR-HLA system were analyzed at 4 levels, and the frequencies of KIR framework genes and KIR profiles, classⅠHLA ligands, matched KIR+HLA pairs and KIR-HLA compound profile were compared between the two groups. P values were calculated using SPSS 13.0 software.</p><p><b>RESULTS</b>For the CML group, the frequencies of HLA-C2 ligand, 2DL1+HLA-C2 pair and HLA-B Bw4-80I were significantly lower than those of the control group, suggesting a protective effect against CML (HLA-C2: OR=0.386, 95%CI:0.240-0.620, P<0.01; 2DL1+HLA-C2: OR=0.316, 95%CI:0.191-0.525, P<0.01; HLA-B Bw4-80I: OR=0.576, 95%CI:0.384-0.862, P<0.01). The frequencies of KIR2DL1 ligand (HLA-C2) and KIR3DL1 ligand (HLA-B Bw4-80I) in the CML group were significantly lower than that of the control group, suggesting that the HLA-C2 and HLA-B Bw4-80I expression is probably decreased in the CML patient group, which led to reduced inhibitory signal and enhanced activating signal of KIR2DL1and/or KIR3DL1NK cells. Notably, the frequency of KIR-HLA compound profiles ID2 (KIR AA1-HLA-C1/C1-Bw6/Bw6-A3/11) in CML patients significantly increased in the CML patient group compared with the control group, suggesting that the KIR-HLA compound profiles ID2 may be a risk factor for CML (OR=2.163, 95%CI 1.198-3.906, P<0.01).</p><p><b>CONCLUSION</b>Above analysis has identified certain protective and risk factors for CML from the KIR-HLA system, which may provide a clue for the pathogenesis of leukemia and development of individualized immune therapy.</p>

Human Leukocyte Antigen-C Genotype and Killer Immunoglobulin-like Receptor-Ligand Matching in Korean Living Donor Liver Transplantation

BACKGROUND: The interaction between killer immunoglobulin-like receptors (KIRs) and HLA class I regulates natural killer (NK) cell cytotoxicity and function. The impact of NK cell alloreactivity through KIR in liver transplantation remains unelucidated. Since the frequency of HLA-C and KIR genotypes show ethnic differences, we assessed the impact of HLA-C, KIR genotype, or KIR-ligand mismatch on the allograft outcome of Korean liver allografts. METHODS: One hundred eighty-two living donor liver transplant patients were studied. Thirty-five patients (19.2%) had biopsy-confirmed acute rejection (AR), and eighteen (9.9%) had graft failure. The HLA-C compatibility, KIR genotypes, ligand-ligand, and KIR-ligand matching was retrospectively investigated for association with allograft outcomes. RESULTS: Homozygous C1 ligands were predominant in both patients and donors, and frequency of the HLA-C2 allele in Koreans was lower than that in other ethnic groups. Despite the significantly lower frequency of the HLA-C2 genotype in Koreans, donors with at least one HLA-C2 allele showed higher rates of AR than donors with no HLA-C2 alleles (29.2% vs 15.7%, P=0.0423). Although KIR genotypes also showed ethnic differences, KIR genotypes and the number of activating KIR/inhibitory KIR were not associated with the allograft outcome. KIR-ligand mismatch was expected in 31.6% of Korean liver transplants and had no impact on AR or graft survival. CONCLUSIONS: This study could not confirm the clinical impact of KIR genotypes and KIR-ligand mismatch. However, we demonstrated that the presence of HLA-C2 allele in the donor influenced AR of Korean liver allografts.

The role of different KIR haplotypes in haplo-identical hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the role of different immunoglobulin- like receptor (KIR)haplotypes in haplo- identical hematopoietic stem cell transplantation (HSCT).</p><p><b>METHOD</b>Killer cell KIR genotyping was performed on 468 individuals from 156 unrelated families by PCR-SSP. A total of 624 KIR haplotypes from the parents were used for haplotype analysis. Ninety-two patients received haplo-identical HSCT from one of the parents.</p><p><b>RESULTS</b>The family study showed segregation of one A haplotype and at least 20 unique B haplotypes. The frequency of haplotype A was 72.92% (455/624). The most commonly observed haplotypes in group B were B1, B2, and B3, present at a frequency of 10.26%, 5.77%, and 4.48%, respectively. Compared to KIR gene matched donors (n=17), grafts from KIR gene mismatched donors (n= 14) had a positive effect on survival after haplo- identical HSCT for AML/MDS patients (OS: 88.2%vs 42.9%,P=0.015; RFS: 88.2%vs 35.7%,P=0.007). No effect was observed for ALL/NHL patients (OS: 76.0%vs 75.0%,P=0.727; RFS: 68.0%vs 65.0%,P=0.866). A significantly lower survival rate was observed for transplants from AA (n=52) and AB1/AB2 donors (n=15), compared to other group Bx donors (n=25) (OS: 53.3%vs 96.0%,P=0.017; RFS: 53.3%vs 92.0%,P=0.019). Meanwhile, the risk of relapse was much higher in AA group (n=52) compared to Bx group (n=40) (25.0%vs 5.0%,P=0.009). A higher risk of TRM was observed in AB1/AB2 group (P=0.012). In addition, transplant from donors carried Cen-B was associated with an increased survival compared with Cen-A homozygous donors (OS: 94.7%vs 68.5%,P=0.036; RFS: 89.5%vs 64.4%,P=0.045).</p><p><b>CONCLUSION</b>Overall, KIR genotyping and haplotype analyses should be useful for selection of the most optimal donors with favorable KIR gene grafts. KIR gene mismatch donors should be preferred for AML/MDS patients. Selecting donors carried Cen- B and avoiding the selection of donors of KIR genotype AA/AB1/AB2 was strongly advisable for haplo-identical HSCT.</p>

A strategy to clarify ambiguities during genotyping of functional KIR framework genes by sequencing-based typing among ethnic Hans from southern China / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To categorize ambiguous allelic combinations encountered in genotyping of functional KIR genes by sequencing-based typing of the entire coding sequence and develop an efficient approach to identify such ambiguities.</p><p><b>METHODS</b>Fourteen KIR functional framework genes from 306 ethnic Chinese individuals were genotyped for the entire encoding sequence. The number of ambiguities was directly counted. Based on the differences within each ambiguous allelic combination, group-specific PCR primers and sequencing primers for amplifying the target allelic sequence were designed. The PCR products were then subjected to sequencing in order to identify the ambiguities.</p><p><b>RESULTS</b>The 14 functional KIR genes were subjected to sequencing-based typing (SBT) for the entire encoding sequence. Six ambiguous allelic combinations were identified. The most common ambiguity 3DL2*(002, 007/010, 015) has accounted for 12.09% of the 306 tested samples. The remaining 5 ambiguities were (2DL5A*001, 2DL5B*006/2DL5A*012, 2DL5B*008), 3DL3*(001, 010/009, 048), 3DL2*(007, 008/016, 027), 3DL3*(00801, 048/01001, 026) and 3DL3*(00802, 048/01002, 026) have accounted for 5.88% (18/306), 3.59% (11/306), 2.29% (7/306), 1.31% (4/306) and 1.31% (4/306) of all samples, respectively. For two ambiguities (2DL5A*001, 2DL5B*006/2DL5A*012, 2DL5B*008) and 3DL2*(007, 008/016, 027) subjected to group-specific PCR and re-sequencing, only one demonstrable genotype was identified, While for in each of other four ambiguities subjected to group-specific PCR and re-sequencing, two different genotypes were identified.</p><p><b>CONCLUSION</b>An efficient approach by group-specific PCR and sequencing retest has been established to clarify the ambiguities during SBT testing for functional KIR framework genes, which may have a broad application in KIR sequencing-based typing.</p>

Progress in research on genetic polymorphisms and sequence-based typing of KIR genes / 中华医学遗传学杂志

Killer cell immunoglobulin-like receptors (KIRs) are members of the immunoglobulin superfamily expressed on natural killer (NK) cells and a subset of T cells. Given the receptor-ligand relationship between certain KIR and human leukocyte antigen (HLA) classⅠmolecules, the KIRs are involved in the regulation of NK cell activation through conveying activating or inhibitory signals, which plays an important role in immunities involved in transplantation, tumor, infection as well as autoimmune diseases. This paper has provided a review for the research on KIR gene polymorphisms and summarized the characteristics of the sequence-based typing method for KIR genes.

The role of KIR2DL3/HLA-C*0802 in Brazilian patients with rheumatoid vasculitis

OBJECTIVES: Rheumatoid arthritis is a polygenically controlled systemic autoimmune disease. Rheumatoid vasculitis is an important extra-articular phenotype of rheumatoid arthritis that can result in deep cutaneous ulcers. The objective of this study was to establish a correlation between the frequency of major histocompatibility complex class I/II alleles and killer immunoglobulin-like receptor genotypes in patients with cutaneous rheumatoid vasculitis. METHODS: Using the Scott & Bacon 1984 criteria to diagnose rheumatoid vasculitis and after excluding any other causes such as diabetes, atherosclerosis, adverse drug reactions, infection, and smoking, patients who met the criteria were selected. All of the selected rheumatoid vasculitis patients presented deep cutaneous ulcers. Identification of the major histocompatibility complex class I/II and killer immunoglobulin-like receptor genotypes was performed by polymerase chain reaction assays of samples collected from the 23 rheumatoid vasculitis patients as well as from 80 controls (40 non-rheumatoid vasculitis RA control patients and 40 healthy volunteers). RESULTS: An association between the presence of the HLA-DRB1*1402 and HLA-DRB1*0101 alleles and cutaneous lesions in rheumatoid vasculitis patients and a correlation between the inhibitor KIR2DL3 and the HLA-C*0802 ligand in rheumatoid vasculitis patients were found. CONCLUSION: An association was found between the presence of the HLA-DRB1*1402 and HLA-DRB1*0101 alleles and the development of cutaneous lesions in rheumatoid vasculitis patients. Additionally, the HLA-C*0802 ligand protects these individuals from developing cutaneous lesions. .

Novel Association of Killer Cell Immunoglobulin-like Receptor Genes with EBV-infectious Diseases in Children / 生物医学与环境科学（英文）

Killer cell immunoglobulin-like receptors (KIRs) which are mainly expressed on natural killer (NK) cells are implicated in many virus infections. However, it is unclear whether or not KIRs are associated with susceptibility to Epstein-Barr virus (EBV) infection related diseases. Therefore, the purpose of our study was to investigate possible correlation between polymorphisms of KIR genes and infectious mononucleosis (IM)/EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH). The polymorphisms of KIR genes were detected by polymerase chain reaction with sequence-specific primers (PCR-SSP). The results would contribute to clarify the association of KIRs with EBV induced diseases, and provide new insights into the role of NK cells and innate immune response against viral infections and/or subsequent progression.

Frequency of HLA-Cw and Its Corresponding Killer Cell Immunoglobulin-like Receptors (KIR) among Blood Donors in Chinese Nanjing Han Ethnic Group / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the frequency of HLA-Cw and its KIR2D genotypes in Han blood donor population in Chinese Nanjing area and to analyze the match and distinguish modes of them so as to provide the basis for further studying their roles in incidence and development of disease.</p><p><b>METHODS</b>The PCR-SSP was used to genotyping of HLA-Cw and KIR2D for 241 Han blood donors in Jiangsu Provincial blood center; according to distingush modes of HLA-Cw and KIR2D genes, the distingushed results of HLA-Cw and corresponding activating or inhibitory KIR2D receptors of individuals were analyzed.</p><p><b>RESULTS</b>The frequency of HLA-C1 expression in donor population of Nanjing area was 76.35% which was much higher than that of HLA-C2 expression (23.65%); the expression C1/C2 alleles was accorded with Handy-weinberg balance. The expression frequencies of 5 KIR2Ds (L1, L2, L3, S1 and S2) matched to HLA-Cw were 97.93%, 29.05%, 98.34%, 29.05% and 21.16%, respectively. The match of HLA-C1/C2 to 2DL1⁺/2DL2⁻/2DL3⁺/2DS1⁻/2DS2⁻ was predominated (75/241). Couclusion: The polymorphism data of HLA-Cw and 5 KIR2Ds from blood donors in Chinese Nanjing area has been provided in this study. The match analysis found that the expression of inhibitory HLA-Cw-KIR is higher than that of activated HLA-Cw-KIR, suggesting that the HLA-Cw/KIR2D combination is characterized by preponderance of inhibitory signal pathway.</p>

Polymorphisms of killer cell immunoglobulin-like receptor and its ligand HLA-I gene among northern Chinese Han population / 中华医学遗传学杂志

OBJECTIVE To investigate the distribution of killer cell immunoglobulin-like receptors (KIR) and their specific ligands human leukocyte antigen-I (HLA-I) gene in northern China. METHODS One hundred and eighty-four unrelated northern Chinese Han individuals were recruited. Genotypes of the KIR and HLA-ABC genes were studied by sequence-specific primer polymerase chain reaction (SSP-PCR). RESULTS Sixteen KIR genes were detected among the 184 unrelated individuals. In all individuals, the four framework genes were present. The frequencies for those carrying the remaining 12 KIR genes have ranged from 16.3% to 99.5%. Twenty-four KIR genotypes were identified, for which half were detected in a single individual. A new genotype comprised of KIR2DL3, 3DL1, 2DP1 and the framework genes was detected in one subject. Respectively, 12, 27 and 11 specificities of HLA alleles were identified on the HLA-A, B, C loci. CONCLUSION The distribution of polymorphisms of KIR and its ligand HLA-ABC genes among northern Chinese Han population have been ascertained. The frequencies of 9 KIR/HLA combinations in the above population have been determined for the first time.

The Impact of HLA and KIR Ligand Mismatching on Unrelated Allogeneic Hematopoietic Stem Cell Transplantation in Korean Adult Patients

BACKGROUND: The impact of HLA and KIR ligand mismatching on the outcome of hematopoietic stem cell transplantation (HSCT) remains unclear. Previous reports have identified considerable ethnic differences in the impact of HLA and KIR ligand mismatches, as well as KIR ligand status, on HSCT; however, to date, no data has been acquired in Korean adult patients. METHODS: We investigated the association of high-resolution HLA matching on five loci (HLA-A, -B, -C, -DRB1, and -DQB1), KIR ligand mismatching, and KIR ligand status on the outcome of allogeneic HSCT from unrelated donors in 154 Korean adult patients treated at Seoul National University Hospital. RESULTS: In a multivariate analysis, less than 9/10 allelic matches in five HLA loci was an independent risk factor for acute graft-versus-host disease (GVHD) (grade II to IV) (P=0.019, odds ratio [OR]=2.7). In addition, HLA-A allele mismatching was increasingly prevalent in patients with acute GVHD compared to patients without (61.9% vs. 34.5%, P=0.06). For KIR ligand status, the patient and donor combination of both C1/C1 ligands showed better event-free and overall survival than combinations with C2 ligand patients or donors (P=0.048, P=0.034, respectively) by log-rank test. CONCLUSIONS: Korean adult transplant patients with less than 9 of 10 HLA allele matches in the HLA-A, -B, -C, -DRB1, and DQB1 loci have a higher likelihood of developing acute GVHD (grade II to IV). Impact of KIR ligand status on clinical outcome should be further studied in a larger patient population.

Impact of recipient/donor killer cell immunoglobulin-like receptor gene mismatch on outcomes of renal transplant / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the impact of killer cell immunoglobulin-like receptor (KIR) gene mismatch on the outcomes of renal transplantation.</p><p><b>METHODS</b>We collected the data from 111 donor-recipient pairs of kidney transplant and analyzed the status of KIR gene matching, acute rejection (AR), and 1-year and 3-year survival of the recipients who were followed continuously for over 37 months.</p><p><b>RESULTS</b>Seventeen KIR genes were expressed in both recipient and donor groups, and the frequency of KIR3DS1 was significantly higher in the recipients than in the donors (38.75% vs 24.66%, OR=2.17, χ² = 3.94, P<0.05). The average rate of donor-recipient KIR matching was 82.53%. The donor-recipient KIR2DS1 matching rate was significantly higher in AR group than in no-AR group (85.00% vs 54.95%, χ² = 6.19, P<0.05). The rate of donor-recipient KIR AB-AB genotype was significantly higher in AR group than in no-AR group (33.33% vs 8.00%, P<0.05). The 1- and 3-year survival rates was 94.59% and 82.88% in these recipients, respectively. The frequency of donor KIR-AB genotpye was significantly higher in recipients with poor outcomes (57.89% vs 29.63%, χ² = 8.19, P<0.05); the frequency of both donor and recipient KIR-AB genotype was also significantly higher in recipients with poor prognoses (36.84% vs 9.78%, χ² = 14.87, P<0.05).</p><p><b>CONCLUSIONS</b>KIR3DS1 may be the susceptible gene associated with uremia. A KIR-AB genotype of either the donor or the recipient can increase the risk of AR and reduce the 1- and 3-year survival rate. This finding can be of ethically importance in choosing a living related donor.</p>

Importância das disparidades genéticas nos genes HLA e KIR na resposta de pacientes submetidos ao transplante alogênico de células progenitoras hematopoiéticas para o tratamento de doenças onco-hematologicas / Importance of genetic differences in HLA and KIR genes in the response of patients undergoing allogeneic hematopoietic stem cell transplantation for treatment of onco-hematological diseases

No organismo humano, as moléculas HLA (Human Leukocyte Antigens) são proteínas expressas na superfície da maioria das células nucleadas e são codificadas por genes localizados no braço curto do cromossomo 6 na região do Complexo Principal de Histocompatibilidade (CPH). Essas proteínas são caracterizadas pelo alto grau de polimorfismo, e também faz a ligação com receptores KIR (Immunoglobulin-like Receptors), expressos nas células Natural Killer. Os receptores KIR, que reconhecem moléculas do complexo HLA de classe I, estão entre os principais receptores inibidores dos linfócitos NK. Células infectadas por vírus e células tumorais perdem ou têm diminuída a expressão de moléculas HLA de classe I e, por isso, são eliminadas pela ausência de ligação entre moléculas HLA e receptores KIR inibitórios. Atualmente, muitos estudos têm destacado a importância dos genes KIR e HLA no Transplante de Células Progenitoras Hematopoiéticas (TCPH). O TCPH é o tratamento de escolha para muitas doenças hematológicas e dependem de vários fatores incluindo o estágio da doença, o regime de condicionamento, a fonte de células, o grau de identidade HLA entre doador e receptor e o desenvolvimento da doença do enxerto contra o hospedeiro (DECH). Estudos recentes indicam que a presença de células NK alorreativas no enxerto representa um fator favorável à recuperação de pacientes, uma vez que essas células têm a capacidade de eliminar células tumorais residuais pela ausência ou diminuição da expressão de moléculas HLA e sem a indução da DECH. Também outros fatores podem estar envolvidos na resposta pós-transplante, como a presença e ausência de determinados alelos HLA e genes KIR, os quais podem estar ligados à melhor ou pior resposta pós-transplante.

In the human organism, the HLA (human leukocyte antigens) are proteins expressed on the surface of most nucleated cells and are encoded by genes located on the short arm of chromosome 6 in the region of the Major Histocompatibility Complex (MHC). These proteins are characterized by a high degree of polymorphism, and also make the connection with KIR (Immunoglobulin-like Receptors), expressed in Natural Killer cells. KIR receptors that recognize HLA molecules of class I are among the major inhibitory receptors of NK-cells. Virus infected cells and tumor cells have lost or diminished expression of HLA class I molecules and therefore are eliminated by the absence of binding between HLA molecules and inhibitory KIR receptors. Currently, many studies have highlighted the importance of KIR and HLA genes in Hematopoietic Stem Cell Transplantation (HSCT). HPCT is the treatment of choice for many hematological malignancies and depends on various factors including stage of disease, the conditioning regimen, the source of cells, the degree of identity between donor and recipient HLA and development of chronic graft-versus-host (GVHD). Recent studies indicate that the presence of alloreactive NK cells in the graft is a factor aiding the recovery of patients, since these cells have the ability to eliminate residual tumor cells by the absence or diminution of expression of HLA molecules and without inducing GVHD.

Targeted resequencing of candidate genes reveals novel variants associated with severe Behcet's uveitis

Behcet's disease (BD) is a chronic systemic inflammatory disorder characterized by four major manifestations: recurrent uveitis, oral and genital ulcers and skin lesions. To identify some pathogenic variants associated with severe Behcet's uveitis, we used targeted and massively parallel sequencing methods to explore the genetic diversity of target regions. A solution-based target enrichment kit was designed to capture whole-exonic regions of 132 candidate genes. Using a multiplexing strategy, 32 samples from patients with a severe type of Behcet's uveitis were sequenced with a Genome Analyzer IIx. We compared the frequency of each variant with that of 59 normal Korean controls, and selected five rare and eight common single-nucleotide variants as the candidates for a replication study. The selected variants were genotyped in 61 cases and 320 controls and, as a result, two rare and seven common variants showed significant associations with severe Behcet's uveitis (P<0.05). Some of these, including rs199955684 in KIR3DL3, rs1801133 in MTHFR, rs1051790 in MICA and rs1051456 in KIR2DL4, were predicted to be damaging by either the PolyPhen-2 or SIFT prediction program. Variants on FCGR3A (rs396991) and ICAM1 (rs5498) have been previously reported as susceptibility loci of this disease, and those on IFNAR1, MTFHR and MICA also replicated the previous reports at the gene level. The KIR3DL3 and KIR2DL4 genes are novel susceptibility genes that have not been reported in association with BD. In conclusion, this study showed that target enrichment and next-generation sequencing technologies can provide valuable information on the genetic predisposition for Behcet's uveitis.

Influence of donor activating or inhibitory KIR on prognosis of unmanipulated allogeneic hematopoietic stem cell transplantation / 中国实验血液学杂志

This study was purposed to investigate the role of NK-alloreactivity and donor-inhibiting or activating KIR gene in predicting prognosis under unmanipulated allogeneic blood and marrow transplantation. A modified polymerase chain reaction sequence specific primers (PCR-SSP) method was used to typing KIR and HLA genotype of donors and recipients. The relationship between donor activating or inhibitory KIR and recipient HLA genotypes on event free survival (EFS), cumulative incidence of malignant relapse and transplant-related mortality (TRM) were investigated retrospectively in 67 patients undergoing hematopoietic stem cell transplantation. The results showed that no effect of 'KIR/HLA mismatched' was detected on acute graft-versus-host disease (aGVHD) and relapse. The EFS of KIR/HLA mismatched group was lower, especially KIR2DL1/HLA-C2 mismatched group (44.8% vs 69.2%, P = 0.043). However, EFS was better for the presence of donor-activating KIR2DS2 (81.3% vs 52.6%, P = 0.052), and the relapse rate was significantly lower for the presence of this genotype (7.7% vs 34.2%, P = 0.05). EFS was worse in patients homozygous for group 1 HLA-C (C1) when donor carries the activating KIR2DS1 (KIR2DS1 positive/HLA-C2-negative group, P = 0.028), and the incidence of aGVHD in this group was significantly higher than that in any other groups (P = 0.028). In multivariate analysis, advanced disease stage, more than two donor-activating KIR, donor KIR2DS2-negative genotype were associated with an reduced disease-free survival (HR = 3.34, 2.19, 3.18;and P = 0.005, 0.053, 0.066). Donor KIR2DS2-negative genotype were also associated with an increased risk of relapse (HR = 6.72, 9.43; and P = 0.019, 0.047). And donor KIR2DS1 positive/recipient HLA-C2 negative group was the only risk factor of TRM (HR = 3.27, 95% CI 1.78 - 9.06, P = 0.023). It is concluded that missing ligand for the donor inhibitory KIR has weak effect on the outcome of unmanipulated HSCT. The activating KIR play an important role in the EFS, relapse and TRM after HSCT. Donor KIR2DS1-positive/recipient HLA-C2-negative group and donor KIR2DS1 gene negative predict poor prognosis. Analysis of KIR genotype and its ligand is important for the selection of best donor and prognostic evaluation in unmanipulated allogeneic HSCT.

CD16 expression is an independent prognostic factor for extranodal NK/T cell lymphoma of nasal type / 中华病理学杂志

<p><b>OBJECTIVE</b>To analyze the clinicopathologic features of extranodal NK/T cell lymphoma, nasal type (ENKTCL-N), to explore the expression of NK cell-associated receptors in ENKTCL-N and the relationship with prognosis, and to establish a prognostic model.</p><p><b>METHODS</b>One hundred and twenty-six cases of ENKTCL-N were selected from the files of the Department of Pathology, West China Hospital of Sichuan University. The relevant clinical and follow-up data were collected, and the histopathology was reviewed. All specimens were stained immunohistochemically for CD16, ICAM-1 and LFA-1. RT-PCR was used to detect the expression of CD94, NKG2 and KIR. The relationship between the prognosis of ENKTCL-N, clinical features, histopathological characteristics and expression of these markers were also analyzed.</p><p><b>RESULTS</b>ENKTCL-N mainly occurred in middle-age and young patients (median age, 41 years). The male to female ratio was 3.2:1. Sites more commonly involved were the nose and upper aerodigestive tract whereas those for the non-nasal type were the skin and gut. Only six cases involved two or more extranodal sites. Most (86.5%, 109/126) of the patients were in clinical stages I/II. The tumors showed predominately medium-sized tumor cells and large-sized tumor cells accounted for only 9.5% (12/126). Coagulative necrosis was present in all cases. The expression rates of CD56, CD16, CD94, LFA-1 and ICAM-1 were 82.6% (95/115), 15.1% (19/126), 55.4% (41/74), 40.5% (51/126) and 0, respectively. The expression rate of NKG2 receptor was 90.5% (67/74) overall. NKG2 receptor expression was independent of CD94. The overall expression rate of KIR receptor was 33.8% (25/74) and KIR receptor restriction was not detected in 20.8% (5/24) of the cases. Follow-up data was available in all patients, with median and average survival time being 15 months and 20.2 months, respectively. Survival analysis showed that prognostic factors included the gender, age, disease type, extranodal involvement, stage, the expression of CD16, LFA-1 and CD94. Cox's proportional hazard regression analysis revealed four factors, age, involved site, stage and CD16 expression, were independent prognostic factors.</p><p><b>CONCLUSIONS</b>The age, disease type, stage and CD16 expression are independent prognostic factors. Establishment of a prognostic model based on the above four factors can be more accurate in the prognostication of ENKTCL-N. The differences in the clinical features, prognosis, and expression of NK cell-associated receptors are obvious between nasal NK-cell lymphoma and non-nasal NK-cell lymphoma.</p>