OBJECTIVE: To investigate the recombinations within the human leukocyte antigen (HLA) region in two families. METHODS: Genomic DNA was extracted from the peripheral blood specimens of the different family members. HLA-A, -B, -C, -DRB1, -DQB1 and -DPB1 loci were genotyped using polymerase chain reaction-sequence specific oligonucleotide probing technique (PCR-SSO) and next-generation sequencing technique. HLA haplotype was determined by genetic analysis of the pedigree. RESULTS: The haplotypes of HLA-A*11:01~C*03:04~B*13:01~DRB1*12:02~DQB1*03:01~DPB1*05:01:01G and HLA-A*03:01~C*04:01~B*35:03~DRB1*12:01~DQB1*03:01~DPB1*04:01:01G in the family 1 were recombined between HLA-B and HLA-DRB1 loci, which formed the haplotype of HLA-A*11:01~C*03:04~B*13:01~DRB1* 12:01~DQB1*03:01~DPB1*04:01:01G. The haplotypes of HLA-A *02:06~C*03:03~B*35:01~DRB1*08:02~DQB1*04:02~ DPB1*13:01:01G and HLA-A *11:01~C*07:02~B*38:02~DRB1*15:02~DQB1*05:01~DPB1*05:01:01G in the family 2 were recombined between HLA-DQB1 and HLA-DPB1 loci, which formed the haplotype of HLA-A*02:06~C*03:03~B*35:01~ DRB1*08:02~DQB1*04:02~DPB1*05:01:01G. CONCLUSION The gene recombination events between HLA-B and -DRB1, HLA-DQB1 and -DPB1 loci were found respectively in two Chinese Han families.
Humans ; Gene Frequency ; HLA-DQ beta-Chains/genetics* ; HLA-B Antigens/genetics* ; Histocompatibility Antigens Class I/genetics* ; Haplotypes ; HLA-A Antigens/genetics* ; HLA-DRB1 Chains/genetics* ; Recombination, Genetic ; Alleles

Objective To analyze the levels of cytokines (IL-2,IFN-γ,IL-6,IL-10) associated with Th1 and Th2 cells in HLA-DQ8 transgenic mice model of ocular experimental autoimmune myasthenia gravis (oEAMG) induced by recombinant H-AChR γ subunit immunization.Methods DQ8 mice were immunized with 20 μg of AChR γ subunit,20 μg of crude E. coli extract (E. coli group),or complete Freund's adjuvant (CFA) only (CFA group). All mice were immunized on days 0,30,and 60. Mice were euthanized 28 days after the third immunization,and draining lymph node cells (LNC) and spleen lymphocytes were cultured in vitro. The supernatant was collected to observe the interleukin(IL)-2,interferon(IFN)-γ,IL-6,IL-10 production by ELISA.Results LNCs and spleen lymphocytes of H-AChR γ subunit-immunized mice exhibited significantly enhanced IFN-γ (F=76.332,P<0.001;F=34.865,P<0.001) and IL-2 (F=42.835,P<0.001;F=38.030,P<0.001),which associated with Th1 cells,as compared to E. coli group and CFA group. There were no significant differences in IL-6 (F=1.325,P=0.284;F=1.935,P=0.166) and IL-10 (F=0.908,P=0.417;F=1.189,P=0.322) levels,which secreted by Th2 cells,among these three groups.Conclusion Th1 cytokines play key roles in the pathogenesis of oEAMG,while the mechanism of Th2 cytokines for oEAMG remains unclear.
Animals ; Cytokines ; Escherichia coli ; HLA-DQ Antigens ; Interferon-gamma ; Mice ; Mice, Transgenic ; Receptors, Cholinergic ; Th1 Cells ; Th2 Cells

Objective: To analyze family-based haplotype frequencies of HLA-A, -B, -C, -DRB1 and -DQB1 genes and their clinical significance. Methods: The data of HLA genotyping in 3568 families undergoing related haploidentical transplantation between 2012 and 2017 at the First Affiliated Hospital of Soochow University were retrospectively evaluated. The HLA genotyping was performed by PCR amplification with sequence-based typing (PCR-SBT) and sequence-specific oligonucleotide probe (PCR-SSOP) methods. The family genetic analysis and haplotype frequencies were also investigated. Results: All the families were divided into 3 groups, including group1 of 1 422 entire families; group2 of 1 310 patients and either of their parents or one of their children; group3 of 836 patients and their HLA≥5/10 matched sibling donors. In the haplotypes with frequencies greater than 0.1% in group1+ group2, the frequency of A*11∶01-B*40∶01-C*03∶04-DRB1*11∶01-DQB1*03∶01, A*02∶07-B*51∶01-C*14∶02-DRB1*09:01-DQB1*03∶03 were significantly different between group1 and group2 (P=0.029, 0.033) . The frequency of A*11∶01-B*46∶01-C*01∶02∶01G-DRB1*09∶01-DQB1*03∶03 was significantly different between group1 and group3 (P=0.035) . The frequency of A*02∶01-B*40∶01-C*07∶02-DRB1*09∶01-DQB1*03∶03 was significantly different between group1 and group2 (P=0.034) , or group1 and group3 (P=0.034) . The frequency of A*24∶02-B*13∶01-C*03∶04-DRB1*12∶02-DQB1*03:01 was significantly different between group2 and group3 (P=0.046) . Conclusion: In this study, we summarize the prevalence of haplotype frequencies in terms of HLA-A, -B, -C, -DRB1 and-DQB1. Based on the database of family haplotype analysis, patients and donor candidates are sorted with matched HLA genotype while unmatched HLA haplotype. Even in patients without entire family information, HLA haplotype analysis assists in choosing the optimal related or unrelated donors.
Alleles ; Child ; Gene Frequency ; HLA-A Antigens ; HLA-B Antigens ; HLA-C Antigens ; HLA-DQ beta-Chains ; HLA-DRB1 Chains ; Haplotypes ; Humans ; Retrospective Studies

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 observe the correlation between constitution of yin deficiency syndrome (YDS) and polymorphism of HLA-DQA1/treatment response of Peg-lFNalpha therapy in HBeAg positive chronic hepatitis B (CHB) patients, and to explore constitution of Chinese medicine (CM) in response of interferon therapy.

METHODSTotally 120 HBeAg positive CHB patients who were treated with Peg-IFNalpha were enrolled, and assigned to YDS group (59 cases) and non-YDS group (61 cases) according to classification of CM constitutions. All patients were subcutaneously injected with Peg-IFNalpha-2b (1.0 microg/kg body weight) or Peg-IFNalpha-2a (180 microg), once per week. Effective efficacy was primarily judged when complete response (CR) or partial response (PR) was obtained at month 6. Those with CR or PR completed 1 year therapeutic course. HLA-DQA1 gene types were detected by polymerase chain reaction sequence specific primers (PCR-SSP). The distribution difference of CM constitutions in patients with CR or PR and their inter-group HLA-DQA1 allele frequency were compared.

RESULTSDifferent treatment responses of Peg-IFNalpha were observed in CHB patients of two different CM constitutions. The ratio of CR + PR was 61.0% (36/59) in YDS group, obviously lower than that in NYDS group [78.7% (48/61), P < 0. 05]. Patients with CR had a lower allele frequency of HLA-DQA1 * 0501 than those with no-response [14.8% (8/54) vs. 30.6% (22/72)] with statistical difference (P < 0.05). Patients with CR had a higher allele frequency of HLA-DQA1 * 0601 than those with no-response [18.5% (10/54) vs. 5.6% (4/72)] with statistical difference (P < 0.05). The allele frequency of HLA-DQA1 * 0301 was lower in YDS group than in non-YDS group [2. 5% (3/118) vs. 9.8% (12/122)] with statistical difference (P < 0.05). The allele frequency of HLA-DQA1 * 0501 was higher in YDS group than in non-YDS group [33.9% (40/118) vs. 18.9% (23/122)] with statistical difference (P < 0.05). Yet statistical significance was lost after adjustment (Pc > 0.05 for both).

CONCLUSIONSBoth constitutions of CM and HLA-DQA1 gene polymorphism af- fect HBeAg positive CHB patients' response to Peg-INFalpha. Constitutions of YDS and HLA-DQA1 * 0501 was not favorable to response, their association needed to be further studied.

Antiviral Agents ; therapeutic use ; Gene Frequency ; HLA-DQ alpha-Chains ; genetics ; Hepatitis B e Antigens ; blood ; Hepatitis B, Chronic ; drug therapy ; genetics ; Humans ; Interferon-alpha ; therapeutic use ; Medicine, Chinese Traditional ; Polyethylene Glycols ; therapeutic use ; Polymorphism, Genetic ; Recombinant Proteins ; therapeutic use ; Remission Induction ; Yin Deficiency ; genetics

OBJECTIVETo establish a novel HLA genotyping method for preimplantation genetic diagnonis (PGD) using multiple displacement amplification-polymerase chain reaction-sequencing based technique (MDA-PCR-SBT).

METHODSPeripheral blood samples and 76 1PN, 2PN, 3PN discarded embryos from 9 couples were collected. The alleles of HLA-A, B, DR loci were detected from the MDA product with the PCR-SBT method. The HLA genotypes of the parental peripheral blood samples were analyzed with the same protocol. The genotypes of specific HLA region were evaluated for distinguishing the segregation of haplotypes among the family members, and primary HLA matching was performed between the embryos.

RESULTSThe 76 embryos were subjected to MDA and 74 (97.4%) were successfully amplified. For the 34 embryos from the single blastomere group, the amplification rate was 94.1%, and for the 40 embryos in the two blastomeres group, the rate was 100%. The dropout rates for DQ allele and DR allele were 1.3% and 0, respectively. The positive rate for MDA in the single blastomere group was 100%, with the dropout rates for DQ allele and DR allele being 1.5% and 0, respectively. The positive rate of MDA for the two blastomere group was 100%, with the dropout rates for both DQ and DR alleles being 0. The recombination rate of fetal HLA was 20.2% (30/148). Due to the improper classification and abnormal fertilized embryos, the proportion of matched embryos HLA was 20.3% (15/74),which was lower than the theoretical value of 25%.

CONCLUSIONPGD with HLA matching can facilitate creation of a HLA-identical donor (saviour child) for umbilical cord blood or bone marrow stem cells for its affected sibling with a genetic disease. Therefore, preimplantation HLA matching may provide a tool for couples desiring to conceive a potential donor progeny for transplantation for its sibling with a life-threatening disorder.

Blastocyst ; cytology ; metabolism ; Female ; Genotype ; Genotyping Techniques ; methods ; HLA Antigens ; genetics ; HLA-DQ beta-Chains ; genetics ; HLA-DRB1 Chains ; genetics ; Humans ; Polymerase Chain Reaction ; methods ; Pregnancy ; Preimplantation Diagnosis ; methods ; Reproducibility of Results ; Sequence Analysis, DNA ; methods

BACKGROUNDKeshan disease (KD) is an endemic cardiomyopathy in China. The etiology of KD is still under debate and there is no effective approach to preventing and curing this disease. Young women of child-bearing age are the most frequent victims in rural areas. The aim of this study was to determine the differences between molecular pathogenic mechanisms in male and female KD sufferers.

METHODSWe extracted RNA from the peripheral blood mononuclear cells of KD patients (12 women and 4 men) and controls (12 women and 4 men). Then the isolated RNA was amplified, labeled and hybridized to Agilent human 4×44k whole genome microarrays. Gene expression was examined using oligonucleotide microarray analysis. A quantitative polymerase chain reaction assay was also performed to validate our microarray results.

RESULTSAmong the genes differentially expressed in female KD patients we identified: HLA-DOA, HLA-DRA, and HLA-DQA1 associated with spontaneous autoimmunity; BMP5 and BMP7, involved in cardiomyocyte differentiation defect; and ADAMTS 8, CCL23, and TNFSF15, implicated in anti-angiogenic activities. These genes are involved in the canonical pathways and networks recognized for the female KD sufferers and might be related to the pathogenic mechanism of KD.

CONCLUSIONOur results might help to explain the higher susceptibility of women to this disease.

ADAM Proteins ; genetics ; ADAMTS Proteins ; Adult ; Autoimmunity ; genetics ; physiology ; Bone Morphogenetic Protein 5 ; genetics ; Bone Morphogenetic Protein 7 ; genetics ; Cardiomyopathies ; genetics ; pathology ; Cell Differentiation ; genetics ; physiology ; Chemokines, CC ; genetics ; Enterovirus Infections ; genetics ; pathology ; Female ; Gene Expression Profiling ; HLA-D Antigens ; genetics ; HLA-DQ alpha-Chains ; genetics ; HLA-DR alpha-Chains ; genetics ; Humans ; Male ; Middle Aged ; Myocytes, Cardiac ; cytology ; metabolism ; Oligonucleotide Array Sequence Analysis ; Sex Factors ; Tumor Necrosis Factor Ligand Superfamily Member 15 ; genetics

OBJECTIVETo investigate the association of single nucleotide polymorphisms in the HLA-DP and DQ genes with the outcome of chronic hepatitis B virus infection.

METHODSTwo hundred and four healthy subjects, 255 clearance subjects, 204 asymptomatic HBV carriers (AsC), 136 chronic hepatitis B (CHB), 68 liver cirrhosis (LC) and hepatocellular carcinoma (HCC) were enrolled. Genotypes of rs3077, rs9277535 and rs2647050 were determined by sequence specific primers-PCR (PCR-SSP).

RESULTSBy using healthy subjects and clearance subjects as the control groups, rs3077 and rs9277535 were significantly associated with chronic HBV infection under additive and dominant models (P< 0.05). Meanwhile, haplotypes GGA, AGA, AAA appeared to be protective factors against chronic HBV infection (P < 0.05). By using AsC as the control group, comparison with the CHB, LC and HCC groups showed no association of the 3 SNPs or haplotypes with the clinical outcome (P > 0.05).

CONCLUSIONHLA-DP gene polymorphisms are strongly associated with chronic HBV infection. The presence of A allele at rs3077 and rs9277535 of the HLA-DP gene may decreased the risk for chronic HBV infection.

Adult ; Asian Continental Ancestry Group ; ethnology ; genetics ; Case-Control Studies ; China ; ethnology ; Female ; Genotype ; HLA-DP Antigens ; genetics ; HLA-DQ Antigens ; genetics ; Hepatitis B virus ; physiology ; Hepatitis B, Chronic ; ethnology ; genetics ; virology ; Humans ; Male ; Middle Aged ; Polymorphism, Single Nucleotide

OBJECTIVETo analyze the status of HLA-DPA1 and DPB1 matching for unrelated donor-recipient pairs matched at high-resolution allele level for HLA-A, B, C, DRB1 and DQB1 loci.

METHODSA total of 76 unrelated donor-recipient pairs matching at allele level for HLA-A, B, C, DRB1 and DQB1 loci were subjected to HLA-DPA1 and DPB1 sequence-based typing (SBT). HLA-DPA1and DPB1 matching status at high-resolution allelic level was also analyzed.

RESULTSThe allelic identity ratio for single HLA-DPA1 and DPB1 were 17.1% and 9.2%, respectively. HLA-DPA1 and DPB1 allelic identity ratio were both very low. The majority of unrelated donor-recipient pairs (73.7%) had an incompatibility at 1 HLA-DPA1 allele, 9.2% of pairs had an incompatibility at 2 DPA1 alleles. As for the high-polymorphic HLA-DPB1 gene, 57.9% of studied donor-recipient pairs had an incompatibility at 1 HLA-DPB1 allele, almost 1/3 (32.9%) of them were completely incompatible. When HLA-DPA1 and DPB1 genes were analyzed together, the donor-recipient pairs matched at 2/4 was the most common (51.4%), 4/4 allelic complete matched pairs accounted for 5.6%, and 0/4 matched pairs accounted for 8.3%.

CONCLUSIONOur results indicated that the ratio of HLA-DPA1 and DPB1 complete match in the unrelated donor-recipient pairs matching at allelic level for HLA-A, B, C, DRB1 and DQB1 loci were very low. The effect of HLA-DPA1 and DPB1 matching status on clinical unrelated stem cell transplantation still needs to be elucidated.

Alleles ; HLA-DP alpha-Chains ; genetics ; HLA-DP beta-Chains ; genetics ; HLA-DQ beta-Chains ; genetics ; Histocompatibility Antigens Class I ; genetics ; Histocompatibility Testing ; methods ; Humans ; Transplantation ; methods ; Unrelated Donors

BACKGROUND: Narcolepsy is a neurologic disorder characterized by excessive daytime sleepiness, symptoms of abnormal rapid eye movement (REM) sleep, and a strong association with HLA-DRB1*1501, -DQA1*0102, and -DQB1*0602. Here, we investigated the clinico-physical characteristics of Korean patients with narcolepsy, their HLA types, and the clinical utility of high-resolution PCR with sequence-specific primers (PCR-SSP) as a simple typing method for identifying DRB1*15/16, DQA1, and DQB1 alleles. METHODS: The study population consisted of 67 consecutively enrolled patients having unexplained daytime sleepiness and diagnosed narcolepsy based on clinical and neurological findings. Clinical data and the results of the multiple sleep latency test and polysomnography were reviewed, and HLA typing was performed using both high-resolution PCR-SSP and sequence-based typing (SBT). RESULTS: The 44 narcolepsy patients with cataplexy displayed significantly higher frequencies of DRB1*1501 (Pc= 0.003), DQA1*0102 (Pc=0.001), and DQB1*0602 (Pc=0.014) than the patients without cataplexy. Among patients carrying DRB1*1501-DQB1*0602 or DQA1*0102, the frequencies of a mean REM sleep latency of less than 20 min in nocturnal polysomnography and clinical findings, including sleep paralysis and hypnagogic hallucination were significantly higher. SBT and PCR-SSP showed 100% concordance for high-resolution typing of DRB1*15/16 alleles and DQA1 and DQB1 loci. CONCLUSIONS: The clinical characteristics and somnographic findings of narcolepsy patients were associated with specific HLA alleles, including DRB1*1501, DQA1*0102, and DQB1*0602. Application of high-resolution PCR-SSP, a reliable and simple method, for both allele- and locus-specific HLA typing of DRB1*15/16, DQA1, and DQB1 would be useful for characterizing clinical status among subjects with narcolepsy.
Adolescent ; Adult ; Aged ; Alleles ; Cataplexy/genetics ; *DNA Probes, HLA ; Female ; Gene Frequency ; Genetic Predisposition to Disease ; Genotype ; HLA-DQ Antigens/genetics ; HLA-DRB1 Chains/genetics ; *Histocompatibility Testing ; Humans ; Male ; Middle Aged ; Narcolepsy/*diagnosis/genetics ; Phenotype ; Polymerase Chain Reaction