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Biomedical subjects

M Bunce

Publications and source records attributed to M Bunce.

At least 19 recordsLinked to original sources

The phylogenetic position of the 'giant deer' Megaloceros giganteus.

The giant deer, or 'Irish elk', has featured extensively in debates on adaptation, sexual selection, and extinction. Its huge antlers--the largest of any deer species, living or extinct--formed a focus of much past work. Yet the phylogenetic position of the giant deer has remained an enigma. On the basis of its flattened antlers, the species was previously regarded as closely related to the living fallow deer. Recent morphological studies, however, have challenged that view and placed the giant deer closer to the living red deer or wapiti. Here we present a new phylogenetic analysis encompassing morphological and DNA sequence evidence, and find that both sets of data independently support a sister-group relationship of giant and fallow deer. Our results include the successful extraction and sequencing of DNA from this extinct species, and highlight the value of a joint molecular and morphological approach.

Animals↗

Identification of a new allele in a Sicilian individual: HLA-DPB1*0302.

We report here the identification and characterization of a novel human leucocyte antigen (HLA)-DPB1 allele that was subsequently named HLA-DPB1*0302 by the WHO Nomenclature Committee. HLA-DPB1*0302 was identified in a single Sicilian individual by a combination of sequence-specific primers, reverse line sequence-specific oligonucleotide probing and DNA sequencing-based typing. The DPB1*0302 allele is most similar to the DPB1*3101 allele, differing by a single mismatch at nucleotide position 301 (T to G).

Alleles↗

Identification of a new allele, HLA-DRB1*1360, on a DRB5 haplotype in a Brazilian individual.

The application of DNA-based methods for human leukocyte antigen (HLA) genotyping has revealed an ever-increasing degree of polymorphism within the HLA-DRB loci and has resulted in the discovery of new alleles. We have identified a new DRB1 allele that was subsequently named DRB1*1360 by the WHO Nomenclature Committee. This allele is unusual for a DRB1*13 allele, as it is present on a DRB5 haplotype rather than the normal DRB3 haplotype found in association with DRB1*13 alleles.

Base Sequence↗

HLA class-I and class-II allele frequencies and two-locus haplotypes in Melanesians of Vanuatu and New Caledonia.

HLA class-I and class-II allele frequencies and two-locus haplotypes were examined in 367 unrelated Melanesians living on the islands of Vanuatu and New Caledonia. Diversity at all HLA class-I and class-II loci was relatively limited. In class-I loci, three HLA-A allelic groups (HLA-A*24, HLA-A*34 and HLA-A*11), seven HLA-B alleles or allelic groups (HLA-B*1506, HLA-B*5602, HLA-B*13, HLA-B*5601, HLA-B*4001, HLA-B*4002 and HLA-B*2704) and four HLA-C alleles or allelic groups (HLA-Cw*04, HLA-Cw*01, HLA-Cw*0702 and HLA-Cw*15) constituted more than 90% of the alleles observed. In the class-II loci, four HLA-DRB1 alleles (HLA-DRB1*15, HLA-DRB1*11, HLA-DRB1*04 and HLA-DRB1*16), three HLA-DRB3-5 alleles (HLA-DRB3*02, HLA-DRB4*01 and HLA-DRB5*01/02) and five HLA-DQB1 alleles (HLA-DQB1*0301, HLA-DQB1*04, HLA-DQB1*05, HLA-DQB1*0601 and HLA-DQB1*0602) constituted over 93, 97 and 98% of the alleles observed, respectively. Homozygosity showed significant departures from expected levels for neutrality based on allele frequency (i.e. excess diversity) at the HLA-B, HLA-Cw, HLA-DQB1 and HLA-DRB3/5 loci on some islands. The locus with the strongest departure from neutrality was HLA-DQB1, homozygosity being significantly lower than expected on all islands except New Caledonia. No consistent pattern was demonstrated for any HLA locus in relation to malaria endemicity.

Ethnicity↗

Polymorphisms in tumour necrosis factor (TNF) are associated with risk of bladder cancer and grade of tumour at presentation.

The purpose of this study is to assess the role of tumour necrosis factor (TNF) polymorphisms in the risk of developing bladder cancer and effect on tumour stage, grade and progression. In all, seven single-nucleotide polymorphisms in TNF were studied in 196 bladder cancer patients and 208 controls using a PCR-SSP genotyping technique. It was seen that there was a significant association of two polymorphisms in TNF with bladder cancer: the TNF+488A allele was found in 28.1% of patients compared with 14.9% of controls (P=0.0012). In addition, TNF-859T was found in 26.0% of patients compared with 14.4% of the controls (P=0.0036). The two loci were in tight linkage disequilibrium, that is, almost all the individuals having TNF+488A also had TNF-859T. Patients with the TNF+488A or TNF-859T were more likely to present with a moderately differentiated tumour than those patients without the uncommon allele. In all, 16.7% of patients with TNF+488A and 29.9% of patients without TNF+488A presented with a G1 tumour (P=0.015). A total of 14% of patients with TNF-859T and 30.5% of patients without TNF-859T presented with a G1 tumour (P=0.0043). There was no significant effect on time to first recurrence, stage progression or grade progression. In conclusion, a significant association between TNF polymorphisms TNF+488A and TNF-859T and risk of bladder cancer was detected in this study. Both these polymorphisms were associated with grade of tumour at presentation although there was no significant effect on subsequent tumour behaviour.

Adult↗

The contribution of human leucocyte antigen complex genes to disease phenotype in ulcerative colitis.

Linkage and association studies implicate the human leucocyte antigen (HLA) region in genetic susceptibility to ulcerative colitis (UC). However, associations with specific variants have been inconsistent, even within defined ethnic groups. A genetic basis for the disease heterogeneity of UC may account for these discrepant findings from studies in unselected populations. Here, we examine the contribution of the HLA region to the clinical phenotype of UC. We studied 321 accurately phenotyped patients recruited from a single UK centre, with a median follow-up time of 15 years. Individuals were genotyped for 340 polymorphisms constructed into 25 gene-specific allelic haplotypes between HLA-A and Tapasin. Data were analysed with respect to age of onset, disease extent and severity. Strongest association with overall susceptibility was identified with HLA-DRB1 alleles replicating previous studies (DRB1*0103, DRB1*1502 and DRB1*0401). We report a novel association with homozygosity of a tumour necrosis factor (TNF) promoter haplotype (TNF-1031T, -863C, -857C, -380G, -308G and -238G) and distal disease extent that does not extend with time (distal vs total 40.9 vs 25.7%; RR = 2.0; 95% CI 1.23-3.24). We confirm the association of DRB1*0103 with total disease and/or disease requiring colectomy and further demonstrate that DRB1*0103 is associated with shorter time to surgery. Genes in the HLA play a role in modifying disease phenotype. Further studies are required to dissect how these genes functionally interact with each other and with environmental factors to determine clinical patterns of disease

Adolescent↗

Human platelet alloantigens (HPAs): PCR-SSP genotyping of a UK population for 15 HPA alleles.

Alloimmunization to human platelet alloantigens (HPAs) is responsible for neonatal alloimmune thrombocytopenia (NAIT), post-transfusional purpura (PTP) and platelet transfusion refractoriness. HPAs may also have a role as histocompatibility antigens in transplantation as well as associations with cardiac disease. We have developed a polymerase chain reaction-sequence-specific primer (PCR-SSP) assay capable of detecting 15 HPA allelic variants. As part of the validation of the assay, 134 UK renal donors were genotyped to determine HPA allele frequencies in the UK population. The HPA allele frequencies obtained are consistent with those of the other European studies: GP1A*1 (HPA-5a) and GP1A*2 (HPA-5b), 0.914 and 0.086, respectively; GP1BA*1 (HPA-2a) and GP1BA*2 (HPA-2b), 0.925 and 0.075; GP2B*1 (HPA-3a) and GP2B*2 (HPA-3b), 0.627 and 0.373; GP3A*1 (HPA-1a) and GP3A*2 (HPA-1b), 0.840 and 0.161. The rare alleles GP2B*3 (HPA-9bw) and GP3A*3 to *8 (HPA-4b, -6b, -7bw, -8bw, -10bw and -11bw, respectively) were all absent. This comprehensive HPA genotyping assay allows rapid, accurate and reproducible results at low cost.

Alleles↗

Identification of two new alleles in a single Korean individual, HLA-B*1568 and HLA-DRB1*1208.

We have identified a new HLA-B*15 allele and a new HLA-DRB1*12 allele, named B*1568 and DRB1*1208, respectively. The alleles were identified using a combination of sequence specific primers, reverse line sequence specific oligonucleotide probing and sequence-based typing. Both alleles were identified in a single individual of Korean origin. HLA-B*1568 appears to be an HLA-B*4801/B*1507 hybrid combining the exon 2 sequence of B*4801 and the exon 3 and 4 sequences of B*1507. Exon 2 of DRB1*1208 was most similar to DRB1*1201 or 1206, with a single mismatch at nucleotide position 165 (A to C). At the protein level, this substitution results in a phenylalanine substitution at position 26 that creates an identical amino acid sequence to DRB3*0202 between amino acid positions 17 and 36.

Amino Acid Sequence↗

High resolution MIC genotyping: design and application to the investigation of inflammatory bowel disease susceptibility.

The highly polymorphic nonclassical MHC class I chain-related (MIC) genes MICA and MICB encode stress inducible glycoproteins expressed on a variety of epithelial cells including intestinal cells. Interaction with the receptor NKG2D is likely to provide an important costimulatory signal for activation and proliferation of NK cells, activated macrophages and CD8 alphabeta and gammadelta T cells. Fifty-four MICA and 17 MICB alleles have been described to date. Although the functional significance of this polymorphism is not known, the high degree of nonconservative substitution, concentration to the putative ligand-binding site and recent observation that different MICA alleles bind to NKG2D with varying affinity has generated much interest. The MIC genes are attractive functional and positional candidate genes for inflammatory bowel disease susceptibility as a consequence of their position in the HLA region and expression on the gastrointestinal epithelium. We developed a robust, high-resolution PCR-SSP genotyping method that can be incorporated into the standard 'Phototyping' system and which effectively identifies 46 of 54 MICA alleles, and all 17 MICB alleles. We applied this system in combination with microsatellite genotyping of the exon 5 variable number of tandem repeats (VNTR) to the investigation of genetic susceptibility to the inflammatory bowel diseases, ulcerative colitis and Crohn's disease. We studied 248 patients with Crohn's disease, 329 with ulcerative colitis and 354 ethnically matched controls. Linkage disequilibrium patterns between HLA-B, MICA and MICB are presented. Analysis by individual allele or by multilocus haplotype failed to identify any significant disease associations.

Alleles↗

Comprehensive hereditary hemochromatosis genotyping.

Hereditary hemochromatosis (HH) is an iron-overload disease common in populations of Northern European origin. Patients display increased iron absorption leading to excessive iron deposition and potential multiorgan failure. Using polymerase chain reaction sequence-specific primer (PCR-SSP) technology, we have developed an HH diagnosis assay capable of detecting 19 non-synonymous HFE mutations (including a previously unreported mutation, V295A) and several TFR2, SLC11A3 and H ferritin alleles implicated in HH. As part of the validation process, 159 UK renal donors were genotyped to determine HH allele frequencies in the UK population. The alleles nominally identified as HFE*01 (C282Y), HFE*02 (H63D) and HFE*03 (S65C) were found at frequencies of 0.085, 0.173 and 0.009, respectively. All other potential HH-associated alleles were absent, confirming their low prevalence in this population. This assay enables comprehensive routine HH genotyping, producing rapid, accurate and reproducible results at low cost.

DNA Mutational Analysis↗

Immune responses to native beta(2)-glycoprotein I in patients with systemic lupus erythematosus and the antiphospholipid syndrome.

OBJECTIVE: To identify HLA class II associations with anti beta(2)-glycoprotein I (beta2GPI) antibodies in a cohort of Caucasian patients with systemic lupus erythematosus (SLE) and to determine whether these HLA genotypes act as restriction elements for lymphocyte proliferation to native human beta2GPI in vitro. METHODS: Anti-beta2GPI antibodies were detected in patient sera using enzyme-linked immunosorbent assays (ELISAs). HLA class II alleles (DRB1, DQB1) were determined by polymerase chain reaction-based DNA genotyping. In vitro peripheral blood mononuclear cell (PBMC) responses to native human beta2GPI were measured in a 7-day proliferation assay. RESULTS: We identified three groups of Caucasian SLE patients using these ELISAs. In group 1, 16 out of 18 SLE patients (89%) with anti-beta2GPI antibodies were positive for HLA-DRB1*0401/4/8, DR11 or DRB1*1302 (P=0.001 vs controls) compared with 23 out of 53 patients (43%) in group 2 with anti-cardiolipin antibodies only, 57 out of 151 patients (38%) in group 3 (SLE patients without anticardiolipin antibodies) and 109 out of 225 controls (48%). Fourteen patients with anti-beta2GPI antibodies had greater median stimulation indices to beta2GPI in vitro compared with the 15 controls studied (P=0.04). CONCLUSION: The HLA class II and PBMC proliferation data suggest that beta2GPI may be both a T- and B-cell autoantigen in SLE.

Adolescent↗

Mother-to-child transmission of HIV infection and CTL escape through HLA-A2-SLYNTVATL epitope sequence variation.

Cytotoxic T lymphocytes (CTL) play a central role in containment of HIV infection. Evasion of the immune response by CTL escape is associated with progression to disease. It is therefore hypothesised that transmitted viruses encode escape mutations within epitopes that are required for successful control of viraemia. In order to test this hypothesis, escape through the dominant HLA-A2-restricted CTL epitope SLYNTVATL (p17 Gag residues 77-85 SL9) in the setting of mother-to-child-transmission (MTCT) was investigated. Initial data from two families in which the HIV-infected mother expressed HLA-A*0201 and had transmitted the virus to other family members were consistent with this hypothesis. In addition, analysis of the gag sequence phylogeny in one family demonstrated that CTL escape variants can be successfully transmitted both horizontally and vertically. To test the hypothesis further, a larger cohort of transmitting mothers (n=8) and non-transmitters (n=14) were studied. Variation within the SL9 epitope was associated with expression of HLA-A2 (P=0.04) but overall no clear link between variation from the SL9 consensus sequence and MTCT was established. However, the high level of background diversity within p17 Gag served to obscure any possible association between escape and MTCT. In conclusion, these studies highlighted the obstacles to demonstrating CTL escape arising at this particular epitope. Alternative strategies likely to be more definitive are discussed.

Amino Acid Sequence↗

Evolution and transmission of stable CTL escape mutations in HIV infection.

Increasing evidence indicates that potent anti-HIV-1 activity is mediated by cytotoxic T lymphocytes (CTLs); however, the effects of this immune pressure on viral transmission and evolution have not been determined. Here we investigate mother-child transmission in the setting of human leukocyte antigen (HLA)-B27 expression, selected for analysis because it is associated with prolonged immune containment in adult infection. In adults, mutations in a dominant and highly conserved B27-restricted Gag CTL epitope lead to loss of recognition and disease progression. In mothers expressing HLA-B27 who transmit HIV-1 perinatally, we document transmission of viruses encoding CTL escape variants in this dominant Gag epitope that no longer bind to B27. Their infected infants target an otherwise subdominant B27-restricted epitope and fail to contain HIV replication. These CTL escape variants remain stable without reversion in the absence of the evolutionary pressure that originally selected the mutation. These data suggest that CTL escape mutations in epitopes associated with suppression of viraemia will accumulate as the epidemic progresses, and therefore have important implications for vaccine design.

Adult↗

Donor cytokine genotype influences the development of acute rejection after renal transplantation.

BACKGROUND: Acute allograft rejection remains an important cause of morbidity after kidney transplantation, and has been shown to be a crucial determinant of long-term graft function. Although rejection is mediated by recipient lymphocytes, both donor and recipient factors contribute to the local environment that influences the nature, severity, and duration of the rejection response. Cytokines are a major determinant of this milieu, and this study sought to explore the impact of donor cytokine and cytokine receptor gene polymorphisms on acute rejection after renal transplantation. METHODS: A total of 145 cadaveric renal allograft donors were selected for analysis according to the presence or absence of graft rejection in the first 30 days after transplantation. DNA was genotyped for 20 polymorphisms in 11 cytokine and cytokine receptor genes using the polymerase chain reaction with sequence specific primers. Associations were assessed using contingency table analysis and the chi2 test, using a two-set design. RESULTS: A polymorphism at position -174 of the donor IL-6 gene was associated with the incidence (P=0.0002) and severity (P=0.000007) of recipient acute rejection. This finding was independent of HLA-DR matching. Acute rejection was not influenced by recipient IL-6 genotype, or by donor-recipient matching of IL-6 genotype. CONCLUSION: This study identifies donor IL-6 genotype as a major genetic risk factor for the development of acute rejection after renal transplantation. This provides evidence that donor-derived cytokines play a major role in determining outcome after transplantation, and will contribute to the development of therapeutic algorithms to predict individuals at particularly high risk of acute rejection.

Acute Disease↗

Substantial differences in specificity of HIV-specific cytotoxic T cells in acute and chronic HIV infection.

Cytotoxic T lymphocytes (CTLs) play a vital part in controlling viral replication during human viral infections. Most studies in human infections have focused on CTL specificities in chronic infection and few data exist regarding the specificity of the initial CTL response induced in acute infection. In this study, HIV-1 infection in persons expressing human histocompatibility leukocyte antigen (HLA)-A*0201 was used as a means of addressing this issue. In chronic infection, the dominant HLA-A*0201-restricted CTL response is directed towards the epitope SLYNTVATL ("SL9") in p17 Gag (residues 77-85). This epitope is targeted by 75% of HLA-A*0201-positive adults, and the magnitude of this A*0201-SL9 response shows a strong negative association with viral load in progressive infection. Despite using the highly sensitive peptide-major histocompatibility complex tetramer and intracellular cytokine assays, responses to the SL9 epitope were not detectable in any of 11 HLA-A*0201-positive subjects with acute HIV-1 infection (P = 2 x 10(-6)), even when assays were repeated using the SL9 peptide variant that was encoded by their autologous virus. In contrast, multiple responses (median 3) to other epitopes were evident in 7 of the 11 A*0201-positive subjects. Longitudinal study of two subjects confirmed that the A*0201-SL9 response emerged later than other CTL responses, and after viral set point had been reached. Together, these data show that the CTL responses that are present and that even may dominate in chronic infection may differ substantially from those that constitute the initial antiviral CTL response. This finding is an important consideration in vaccine design and in the evaluation of vaccine candidates.

Acute Disease↗

Low frequency of HLA DRB1*03 - DQB1*02 and DQB1*0302 haplotypes in Romania is consistent with the country's low incidence of Type I diabetes.

AIMS/HYPOTHESIS: Our study aimed to determine the association of HLA class II HLA-DQB1 alleles with Type I (insulin-dependent) diabetes mellitus and the frequencies of these alleles in the Romanian population, which has one of the lowest incidences of Type I diabetes in children aged 0-14 years in Europe at 3-4 cases per 100,000 person-years. METHODS: We used the sequence specific primer-polymerase chain reaction (PCR-SSP) technique to type HLA-DQB1 alleles, the HLA-DRB1alleles DRB1*03 and one single nucleotide polymorphism (SNP) in the insulin gene (INS). We studied 204 Type I diabetic Romanian families, 196 of which were simplex with 70.3 % of subjects diagnosed under 14 years of age. Data was analysed using a modified version of the Transmission Disequilibrium Test, the Transmission Disequilibrium Test itself, and the affected family-based control method. RESULTS: We found, as expected, the strong positive DQB1*02-DRB1*03 and DQB1*0302, and negative DQB1*0602, HLA class II allele associations with Type I diabetes in these Romanian families. However, using the affected family-based control method, we found relatively low population frequencies of DQB1*02-DRB1*03 and DQB1*0302 alleles in Romania (15.8%) compared with Sardinia (31.3%), a high incidence European region (35 cases per 100,000 person-years in children aged 0-14years). The INS locus had a strong effect in this data set with 80.5 % transmission of the susceptible INS allele from parents to affected siblings (relative risk = 4.1). CONCLUSION/INTERPRETATION: Part of the explanation for the low incidence of Type I diabetes in Romania could be the lower frequency of the DRB1*03 DQB1*02 and DQBI*0302 susceptibility haplotypes in this country.

Child↗

CD1 genotyping of patients with Mycobacterium malmoense pulmonary disease.

Mycobacterium malmoense is an opportunistic mycobacterium that occasionally causes disease in non-immunosuppressed individuals. As only a few individuals exposed to these organisms actually develop clinical disease, it is possible there is a genetic component to susceptibility. CD1 molecules are capable of presenting antigens from more virulent mycobacteria to T cells; therefore, we were interested in discovering whether recently described polymorphisms in CD1 molecules modulated susceptibility to M. malmoense pulmonary disease. The CD1 system comprises five genes (CD1A, -B, -C, -D, and -E) located on chromosome 1 (1q22-23). CD1 molecules are structurally and functionally related to major histocompatibility complex (MHC) class I molecules and are expressed on dedicated antigen-presenting cells. The primary function of CD1 molecules is to present lipid and glycolipid antigens to T cells. We have developed an allele-specific polymerase chain reaction-sequence-specific primer (PCR-SSP) method of CD1 genotyping. Using this method, we compared the allele and haplotype frequencies of CD1 in 49 HIV-negative patients with M. malmoense pulmonary disease with those in 342 normal controls. The CD1A and CD1E alleles were nominally identified as CD1A*01, CD1A*02, CD1E*01 and CD1E*02, and the control gene frequencies were found to be 5%, 95%, 67% and 33%, respectively. No significant difference was observed between the patient and control cohorts. Positive linkage disequilibrium values of 0.73 were observed between CD1A*02 and CD1E*01 (P<0.0001; chi2 test), and 0.94 between CD1A*01 and CD1E*02 (P<0.0001; chi2 test). Typing was also performed for two previously described CD1D alleles (CD1D*01 and CD1D*02), although only CD1D*01 was detected.

Antigens, CD1↗

Review article: the genetics of inflammatory bowel disease.

Recent epidemiological, clinical and molecular studies have provided strong evidence that inherited predisposition is important in the pathogenesis of chronic inflammatory bowel diseases. The model most consistent with the epidemiological data suggests that Crohn's disease and ulcerative colitis are related polygenic diseases, sharing some but not all susceptibility genes. Investigators throughout the world have applied the complementary techniques of genome-wide scanning and candidate gene analysis. Four areas of linkage have been widely replicated on chromosomes 16 (IBD1), 12 (IBD2), 6 (IBD3-the HLA region), and most recently on chromosome 14. Fine mapping of these regions is underway. Of the 'positional' candidate genes, most attention has centred on the genes of the major histocompatibility complex. Genes within this region may determine disease susceptibility, behaviour, complications and response to therapy. Hope continues that studies of inflammatory bowel disease genetics will provide fresh insight into disease pathogenesis and soon deliver clinical applications.

Chromosomes, Human, Pair 12↗