BACKGROUND: Human leukocyte antigen-E (HLA-E) is one of non-classical HLA class I genes. Up to now, the polymorphism analysis is mainly aimed at the variation in exon 3 of HLA-E, which determines HLA-E*01:01 or HLA-E*01:03. However, the identification of the full-length HLA-E and its novel alleles is rare reported.OBJECTIVE: To establish the method of identification of HLA-E genomic full-length sequence, and to identify its novel alleles in healthy blood donors in Shenzhen, China.METHODS: Peripheral blood DNA samples were extracted from the subjects, and the amplified primers and sequencing primers in conserved regions were designed according to the sequences of HLA-E published in the IMGT/HLA database.A high-fidelity reaction system was used to amplify the genomic full-length of HLA-E, followed by sequencing,assembling, confirming and typing.RESULTS AND CONCLUSION: Herein, we successfully established the method for amplifying genomic full-length sequence and sequence-based typing. Two novel HLA-E alleles were nominated by WHO HLA Nomenclature committee as HLA-E*01:01:01:06 and HLA-E*01:01:01:07. Compared with the most related allele HLA-E*01:01:01:01,HLA-E*01:01:01:06 had one nucleotide change at nt-26(G->T) in 5'-promoter, and HLA-E*01:01:01:07 had one nucleotide change at nt3345(T->C) in 3'-UTR. The polymorphism data of genomic full-length HLA-E in Chinese individuals need to be filled, and the method we developed here supplies the key technique for the further studies.

BACKGROUND:Many articles concerned novel al eles reported in China and outside China, but some al eles were detected lately. After that, these al eles were tested again. Because frequencies of these al eles are very low, few relevant articles are reported, so these al eles are ignored easily. OBJECTIVE:To test and identify four human leukocyte antigen rare al eles that patients and donors carries. METHODS:Genomic DNA was extracted automatical y from blood samples using quick DNA purified kit, typed by human leukocyte antigen locus commercial sequence-based typing kits and confirmed by sequence-specific primers high-resolution kits. RESULTS AND CONCLUSION:Four detected rare al eles are B*27:15, B*51:39, C*07:66 and C*08:22. The four above-mentioned human leukocyte antigen rare al eles defined by National Marrow Donor Program are not rare in China. The facts prove that C*08:22 which was defined a rare al ele by National Marrow Donor Program before is a common al ele in Chinese Han nationality.

BACKGROUND: Due to the polymorphism of HLA, a large number of ambiguities have been generated by conventional HLA typing techniques, and confirmed stereotypes of ambiguous results based on group-specific haploid full-length typing are rarely reported.OBJECTIVE: To analyze the accuracy of HLA-typing ambigulity based on group-specific haploid full-length sequencing. METHODS: The low-resolution results were used as the starting point for two ambiguous samples. Sanger sequencing (PCR-SBT) based on haploid full-length was performed after group-specific amplification. RESULTS AND CONCLUSION: One case showed a new A*02:03:01 allele, which was found a mutation in NT817 from C to T in comparison with A*11:01:01:01. The other case indicated another new C*07:02:01:01, which was found a mutation in NT879 from A to G in comparison with C*08:01:01. In conclusion, these results indicate that the group-specific haploid full-length sequencing method can be used to accurately classify HLA alleles and to discover new alleles.

Objective: To study the antitumor activity and immune modulation of extract from Chlorella pyrenoidosa Chick. (CE). Methods:Cyclophosphamide is positive control and saline is negative control. S180-bearing and HCA-bearing mice were used as animal model. The changes of the growth of S180 and HCA tumor cells in vitro, the changes of tumor weight in vivo and the effect on immune system such as phagocytosis of macrophages,proliferation of lymphocytes were investigated when CE were used. Results: The tumor cells were killed by CE in vitro. CE inhibited tumor growth markedly in S180 and HCA bearing mice and the effect was enhaced if CE and cyclophosphamide were used simultaneously. CE also enhanced the immune function. Conclusion: CE has the antitumor effect and the effect of immune modulation.

OBJECTIVETo explore the association of KIR-HLA gene polymorphism with chronic myeloid leukemia (CML) among ethnic Hans from southern China.

METHODSA 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.

RESULTSFor 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).

CONCLUSIONAbove 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.

Asian Continental Ancestry Group ; genetics ; China ; Gene Frequency ; Genetic Predisposition to Disease ; ethnology ; genetics ; Genotyping Techniques ; HLA Antigens ; genetics ; HLA-A Antigens ; genetics ; HLA-B Antigens ; genetics ; HLA-C Antigens ; genetics ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; ethnology ; genetics ; Odds Ratio ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Protein Isoforms ; genetics ; Receptors, KIR ; genetics ; Risk Factors

Objective: To investigate the expression levels and clinical significance of serum suppressor of cytokine signaling-3 (SOCS3) and associated cytokines in HIV/TB co-infected patients. Methods: The serum levels of SOCS3, IFN-γ, IL-2, IL-4, IL-6, IL-10, IL-17 and IL-22 were quantified by using enzyme-linked immunosorbent assays (ELISA) in 50 HIV-infected patients, 48 HIV/TB co-infected patients and 50 healthy donors. Pearson correlation analysis was used to analyze the correlation between SOCS3 and other seven cytokines. Results: Serum levels of SOCS3 expression in HIV/TB co-infection group were significantly higher than those in HIV-infection alone and the control group. There was also significant correlation between SOCS3 and IFN-γ, IL-4, IL-6, IL-10, IL-2 in HIV/TB co-infection group. Conclusions These findings indicated that SOCS3 may play an important role in the immune response of patients with HIV/TB co-infection and it may be helpful in the diagnosis of HIV/TB co-infection.

OBJECTIVETo study the genetic polymorphisms of human leukocyte antigen (HLA)- A, B, C, DRB1, DQA1, DQB1, DPA1and DPB1among ethnic Hans from southern China.

METHODS481 randomly selected individuals were genotyped using a polymerase chain reaction (PCR) sequence-based typing (SBT) method for the above genes. Their allele frequencies were determined by direct counting.

RESULTSIn total, 28 HLA-A, 57 HLA-B, 28 HLA-C, 40 HLA-DRB1, 18 HLA-DQA1, 17 HLA-DQB1, 6 HLA-DPA1and 21 HLA-DPB1alleles were identified. Among these, common alleles (with allelic frequencies > 0.05) included A*1101, A*2402, A*0207, A*3303, A*0201, B*40:01, B*46:01, B*58:01, B*13:01, B*15:02, C*01:02, C*07:02, C*03:04, C*03:02, C*08:01, C*03:03, C*04:01, DRB1*09:01, DRB1*15:01, DRB1*12:02, DRB1*08:03, DRB1*03:01, DRB1*04:05, DRB1*11:01, DQA1*01:02, DQA1*03:02, DQA1*03:03, DQA1*06:01, DQA1*01:03, DQA1*05:05, DQA1*01:04, DQA1*03:01, DQA1*05:01, DQB1*03:01, DQB1*03:03, DQB1*06:01, DQB1*05:02, DQB1*03:02, DQB1*02:01, DQB1*03:02, DQB1*06:02, DPA1*02:02, DPA1*01:03, DPA1*02:01, DPB1*05:01, DPB1*02:01, DPB1*13:01, DPB1*04:01and DPB1*02:02.For each of the locus, the overall frequencies of common alleles were 75.57%, 52.81%, 78.28%, 62.16%, 86.70%, 77.23%, 95.32% and 81.59%, respectively.

CONCLUSIONThe allelic frequencies of the 8 selected HLA loci among ethnic Hans from southern China may served as a reference for anthropology, legal medicine, transplantation and disease association studies.

Alleles ; Asian Continental Ancestry Group ; genetics ; China ; Gene Frequency ; Genotype ; Genotyping Techniques ; methods ; HLA-A Antigens ; genetics ; HLA-B Antigens ; genetics ; HLA-C Antigens ; genetics ; HLA-DP Antigens ; genetics ; HLA-DQ alpha-Chains ; genetics ; HLA-DQ beta-Chains ; genetics ; HLA-DRB1 Chains ; genetics ; Histocompatibility Antigens Class I ; genetics ; Histocompatibility Antigens Class II ; genetics ; Humans ; Linkage Disequilibrium ; Polymerase Chain Reaction ; Polymorphism, Genetic

OBJECTIVETo study genetic polymorphisms of the KIR2DS4 gene among ethnic Hans from southern China.

METHODSGenomic 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.

RESULTSAmong 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.

CONCLUSIONThe 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.

Alleles ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; China ; Gene Frequency ; Genotype ; Genotyping Techniques ; methods ; Haplotypes ; Humans ; Polymorphism, Genetic ; Receptors, KIR ; genetics ; Sequence Analysis, DNA ; methods

OBJECTIVETo study the distribution of MICA alleles among ethnic Han Chinese blood donors from Shenzhen and their linkage disequilibrium with HLA-B gene.

METHODSFor 143 randomly selected blood donors, the MICA and HLA-B alleles were determined with a PCR-sequence based typing (SBT) method. Allelic frequency, haplotypic diversity and linkage disequilibrium were analyzed with a Pypop software.

RESULTSThirteen MICA and 35 HLA-B alleles were identified among the 143 blood donors, among which MICA*008:01 had the highest frequency (76/286), whilst MICA*008:01-HLA-B*40:01 and MICA*010-HLA-B*46:01 were the most common haplotypes. No novel allele was identified.

CONCLUSIONThe allele frequencies, haplotype diversities and linkage disequilibrium parameters under a high resolution can facilitate further studies and applications of the MICA and HLA-B genes.

OBJECTIVETo investigate the genetic basis for a novel allele HLA-C*01:78.

METHODSGenomic DNA was extracted from peripheral blood using a QIAGEN quick DNA extraction kit. The regions encompassing HLA-C from exon 1 to intron 3 and intron 3 to 3'UTR were amplified and cloned using a cloning sequencing kit in order to split the two alleles apart. Selected clones were sequenced to include exons 2 to 4.

RESULTSSequencing results have indicated the HLA-C alleles of the proband to be a novel C*03:04 allele. The sequence has been submitted to GenBank (KF049216). BLAST analysis has confirmed the novel allele to have one nucleotide difference as C*01:03 at genomic nt316C>A (codon 82CGC>AGC) in exon 2, which has resulted in replacement of one amino acid (82R>S).

CONCLUSIONThe novel allele has been officially named as C*01:78 by the WHO Nomenclature Committee. The HLA allele type of the proband was therefore A*02:07, 24:02; B*40:01, 46:01; C*01:78, 03:04; DQB1*05:02, 05:02; DRB1*16:02, 16:02.

Alleles ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; Exons ; Female ; HLA-C Antigens ; genetics ; Humans ; Introns ; Leukemia ; genetics ; Male ; Molecular Sequence Data ; Polymorphism, Single Nucleotide ; Sequence Analysis, DNA