Pierre Mercier

Influence of shared epitope–negative HLA–DRB1 alleles on genetic susceptibility to rheumatoid arthritis

OBJECTIVE Most patients with rheumatoid arthritis (RA) express the shared epitope (SE). It is not known whether SE-negative HLA-DRB1 alleles influence the development of RA. This study examined the influence of SE-negative HLA-DR alleles (DRB1*X) on the development of RA in 3 different French populations. METHODS HLA-DRB1 alleles were defined by polymerase chain reaction with sequence-specific oligonucleotide hybridization or sequence-specific primers. SE-negative alleles were classified according to the electric charge of their P4 pocket. HLA-DRB1 alleles *0103, *0402, *07, *08, *11 (except *1107), *12, and *13 have a neutral or negative P4 charge and are called DRB1*XP4n. HLA-DRB1*03, *0403, *0406, *0407, *0901, *1107, *14, *15, and *16 have a positive P4 charge and are called DRB1*XP4p. RESULTS Among the SE-negative subjects, DRB1 genotypes with 1 or 2 DRB1*XP4n alleles were significantly overrepresented in the control subjects compared with the RA patients, whereas DRB1*XP4p/XP4p genotypes were equally represented in the patients and controls. In single-dose SE-positive subjects, SE/XP4n genotypes were equally represented in the patients and controls. However, SE/XP4p genotypes were significantly overrepresented in the RA patients. CONCLUSION The DRB1*X allele polymorphism influences susceptibility to RA. Alleles that have a neutral or negative electric charge in their P4 pocket (DRB1*XP4n), such as DRB1*0103, *0402, *07, *08, *11 (except *1107), *12, and *13, protect against RA. Alleles that have a positive electric charge in their P4 pocket (DRB1*XP4p), such as DRB1*03, *0403, *0406, *0407, *0901, *1107, *14, *15, and *16, have no influence on the predisposition to RA.

HLA-DRB1 alleles and Jka immunization

BACKGROUND: In transfusion medicine, anti‐Jka has been implicated in hemolytic transfusion reactions. Development of anti‐Jka after transfusion does not always occur after Jk(a–) patients receive at least 1 unit of Jk(a+) blood unit. This study was designed to identify HLA‐DRB1 alleles associated with predisposition to Jka immunization after blood transfusion or pregnancy.

HLA-DRB1 polymorphism is associated with Kell immunisation

K immunisation is observed in some polytransfused patients and pregnant women but does not occur in all cases of K incompatibility. This study analysed the role of genetic background in this selective response to K antigen by investigating HLA‐DRB1 alleles associated with K immunisation in a southern European population. HLA‐DRB1 genotyping was performed by polymerase chain reaction sequence‐specific oligonucleotide/sequence‐specific primer procedures in 54 K immunised patients and 200 healthy controls. The frequency of HLA‐DRB1*11 was significantly higher in K immunised patients than healthy controls: 31 of 54 (57%) vs. 56 of 200 (28%) (Pc < 0·001). In the remaining K immunised HLA‐DRB1*11‐negative patients, the frequency of HLA‐DRB1*13 was increased: 14 of 23 (61%) vs. 49 of 144 in healthy controls (34%) (P < 0·02). The combined frequency of the two HLA‐DRB1 alleles (HLA‐DRB1*11 and HLA‐DRB1*13) was 83% in K immunised patients when compared with 52% in healthy controls (Pc < 0·001). K and k differ by a single amino acid T193 (M). The DRB1*11 and DRB1*13 alleles share a HLA‐DRB1 gene sequence containing S in position 13, D in 70 and A in 74, and coding for the P4 pocket within the HLA‐DR binding groove. This feature of the HLA‐DRB1 gene could be involved in the K peptide presentation through a polymorphism ligand specific for the T193 (M) of K. In conclusion, this study demonstrated a high frequency of HLA‐DRB1*11 or HLA‐DRB1*13 alleles in K immunised patients, which could be due to specific K peptide presentation by HLA‐DR molecules.

The HLA-D system: At least two loci and four distinct phenotypic traits per haplotype. Introduction to component typing in families and population by primed lymphocyte typing

Using a number of intrafamilial PLTs raised against identical HLA haplotypes it has been possible to construct a model in an informative family defining the HLA-D region as a genetic system. This system consists of at least two regions separated by a recombination between HLA-D and GLO. In relation to the site of recombination, a minimum of one centromeric and three telomeric components can be identified per haplotype.—Fourteen PLTs raised and defined within the family were subsequently tested in a Caucasian population (n=84) and in 13 unrelated, complete families.—It is concluded that the hypothetical model proposed for the HLA-D region as a genetic system of linked loci, coding at the cell surface for associated but distinct components (at least four per haplotype), allows for typing of the components of the HLA-D system of any given haplotype. Serological typing of HLA-D components should, in the near future, provide a more convenient way of establishing component phenotypes than the present use of primed lymphocyte typing reagents. Among the components isolated, some have a high association with the classic alleles defined either by homozygous typing cells or DR serology. Others form the basis of cross-reactivity but their presence does not interfere with standard typing. Others, however, seem by their mere presence to be responsible for false assignments.—The concept of HLA-D as a genetic system clarifies many of the inconsistencies observed with a one-locus system.

Genetic Characterization of the Population of Grande Comore Island (Njazidja) According to Major Blood Groups

The Comorian population is historically considered a blend of influences from African Bantus, Arabs, and possibly Austronesians. In this study we present the first genetic data on the current Comorian population. Serologic analysis of the six major blood group systems (ABO, RH, KEL, FY, JK, and MNS) was performed on 164 individuals from Grande Comore Island (Njazidja). In addition, Duffy genotypes were determined by polymerase chain reaction using allele-specific primers. Our findings establish a high frequency of the Fy(a– b–) phenotype (86%), presenting the same genetic background as in sub-Saharan Africa. Analysis of genetic frequencies, distances, and admixture with other populations indicates that African Bantus made the main contribution to the gene pool (73.2% ± 15.5%). The Arab contribution from the Arabian peninsula was smaller (24.2% ± 7%) and the Indonesian contribution was minor (2.6% ± 9%). The major Bantu contribution was commensurate with the Bantu cultural influence. The contribution from the Arabian peninsula seemed in relation to its permeating religious and linguistic influence. As with the language, the Indonesian contribution to the Comorian gene pool was small. These results are in agreement with historical, sociological, and linguistic data.

HLA-DRB1 and DQB1 polymorphisms in Southern France and genetic relationships with other Mediterranean populations

This study presents the results of HLA-DRB1 and DQB1 sequence-specific oligonucleotide probe (SSOP) typing for a population sample of 181 individuals originating from southern France. On the basis of allele and haplotype frequencies, we compared our population with others from the Mediterranean area. Allele frequencies are comparable to those found in other western European populations (France, Portugal, Spain) and indicate neighboring exchanges. The haplotype frequencies showed relationships with North Africans and Jewish populations, as well as the common origin of Moroccan and Lebanese Jews. Therefore, allele frequencies seem to be more able to show recent exchanges while haplotype frequencies might show ancestral relationships. These results may serve as references for future studies of HLA and disease in southern France.

Murine H-2Dd-reactive monoclonal antibodies recognize shared antigenic determinant(s) on human HLA-B7 or HLA-B27 molecules or both

We have evaluated the serological relationships between the murine H-2Dd and human HLA molecules using four H-2Dd-reactive monoclonal antibodies (mAbs) produced in the A.BY (KbIbDb) anti-A.TL (KsIkDd) combination. In the mouse, these reagents exhibited three distinct reactivity patterns: Dd, Ks, and H-2u (mAb 81.L); Dd, H-2p, and H-2u (mAb 81.R); and Dd, Kd, H-2p, H-2u, and H-2v (mAbs 97.G and 97.H). Sequential immunoprecipitation and cross-competitive mAb binding experiments revealed that these mAbs recognized determinants in two spatially distinct polymorphic domains on the H-2Dd molecule of B10.A(5R) cells (defined by mAbs 81.L and 81.R, 97.H, and 97.G, respectively). MAbs 81.R, 97.G, and 97.H, but not 81.L, also defined an HLA-linked polymorphism in the human, the main characteristics of which can be summarized as follows: (i) on B lymphoblastoid cell lines, mAbs 81.R and 97.H bound to cells expressing the HLA-B7, HL-B27 or -Bw40 cross-reacting specificities, (ii) on peripheral blood lymphocyte (PBL) panel mAb 81.R exerted C dependent cytotoxicity to 118 of 400 cells tested, including almost all HLA-B7 or HLA-B27 cells or both (r: 0.952), (iii) the expression of the 81.R. cross-reacting determinant segregated in an informative family with the parental haplotype carrying the HLA-B7 allele, and (iv) mAbs 81.R, 97.G, and 97.H recognized topologically related determinants on the same class I molecule(s) of the human B lymphoblastoid cells JY (HLA-A2,2, -B7,7). These data support the view that some, but not all H-2Dd allotopes have been conserved throughout evolution and are associated in the human with the HLA-B7, -B27 cross-reacting specificities.

HLA DRB1, DMA, and DMB gene polymorphisms in Rheumatoid Arthritis

OBJECTIVE To study the influence of DMA and DMB genes on susceptibility to Rheumatoid Arthritis (RA). METHODS HLA-DRB1, DMA and DMB polymorphisms were defined by PCR SSOP in 203 European Mediterranean RA patients and 181 unrelated healthy controls. RESULTS No significant difference in the phenotype frequencies of DMA and DMB alleles was observed between patients and controls. We found decreased frequencies of DMA*0102 and DMB*0104 in patients but this did not reach significance. These decreased frequencies could be due to a positive linkage disequilibrium with DRB1*0701, an allele which is underrepresented in RA patients. In stratified analysis with RA susceptibility Epitope positive (SE) DRB1 alleles, there was no significant difference in DMA and DMB phenotype frequencies between SE/SE, SE/X, and X/X patients versus controls. Among SE/X subjects, no significant difference in DM distribution frequencies was observed in DRB1*0101/X, 0102/X, 0401/X, 0404/X and 0405/X groups. CONCLUSION DMA and DMB polymorphism does not seem to influence susceptibility to develop RA. Differences in DMA phenotype frequencies between patients and controls are secondary to linkage disequilibrium with DRB1 alleles.

Modeling the HLA component in rheumatoid arthritis: sensitivity to DRB1 allele frequencies.

Rheumatoid arthritis is an inflammatory disease for which positive associations have been described with some HLA‐DRB1 alleles. The associated alleles share a similar amino acid sequence in the third hypervariable region, the shared epitope, but differ at position 71 and 86. It has been suggested that HLA susceptibility to rheumatoid arthritis could be due not only to the shared epitope but could also be influenced by specific amino acids at positions 71 and 86. In this study, we investigated the role of these amino acids in rheumatoid arthritis on 203 unrelated patients. An involvement of amino acid 71 was detected but no conclusion was possible regarding amino acid 86. A study of the sensitivity of the conclusions to marker allele frequencies was performed. We showed that the results obtained for amino acid 71 are not very sensitive to allele frequencies but those obtained at position 86 are highly sensitive. This emphasizes the importance of studying the robustness of results to variations in allele frequencies before conclusions are drawn. Genet. Epidemiol. 19:422–428, 2000.

Birth of a new allele in a sibling: cis or trans gene conversion during meiosis?

The immune response in higher vertebrates is a complex phenomenon mediated by a network of cellular interactions. Major histocompatibility complex (MHC) class II DR, DQ, and DP are highly polymorphic cell surface glycoproteins which play a central role in immunity by binding and presenting peptides to helper T lymphocytes (Brown et al. 1993). These molecules are composed of non-covalently associated a and b chains encoded respectively by A and B genes. The first domains of these chains form a groove where foreign or self peptides are bound and presented to T-cell receptors (TCR). HLA-DR, DQ, and DP molecules are encoded within the human MHC by A and B genes located on multiple loci of the short arm of chromosome 6 (Hardy et al. 1986) which might have been generated by duplication events occurring at least 5 million years ago (Gyllensten et al. 1991 a; Rollini et al. 1985). The number of coding (DRB1, B3, B4, and B5) or non-coding (DRB2, B6, B7, and B9) DRB genes differs from one haplotype to another and is strongly linked to the allelic variant at the DRB1 locus (Andersson et al. 1987; BoÈhme et al. 1985; Spies et al. 1985). Thus, for example, DRB1*04 alleles are found on a haplotype which also carries the DRB4 gene coding for the DR53 serological determinant. The polymorphism of these genes, mainly restricted to the first domain, arises in the 39 part from a patchwork of short sequences, which are shared by several alleles belonging to different homologous class II genes and exchanged between them probably by gene conversion or reciprocal exchanges (Abastado 1993; Andersson et al. 1991; Coppin et al. 1990; Erlich and Gyllensten 1991; Gorski 1989; Gorski and Mach 1986; Gustafsson et al. 1984; Wu et al. 1986). Whereas the DRB1 gene is highly polymorphic, with 184 alleles described in the WHO Nomenclature Report (1997), the polymorphism of the other loci is more restricted with, for instance, only nine alleles described for the DRB4 gene (Bodmer et al. 1997). Nevertheless, this low additional polymorphism increases the global class II polymorphism on the cell surface and increases the DRB1 diversity by providing a source of new sequences which can be transferred to DRB1 alleles by unidirectional or reciprocal intra or inter-chromosomal exchanges. The first domain of the DRB molecule is composed of a b sheet (amino acids 9 to 53) followed by an a-helix ranging from amino acids 54 to 78 (Brown et al. 1993), and the sequences of the DRB second exon which code for these two structures appear to reflect different evolutionary histories. Thus the b sheet in some alleles may be more than 20 million years old (Gyllensten et al. 1991a,b), may have arisen before speciation, and appears to have followed evolutionary pathways identical to those of the third exon. Selection seems to support the conservation of a limited number of epitopes in these regions. The a-helix is composed of sequences shared by different DR generic groups and probably exchanged between homologous loci by various recombinational events. These variations are mediated in part by intra exon segment transfers and appear to be favored by a Chi(c)-like sequence overlapping the two parts of the second exon. One such c-like sequence identified at codons 51 to 55 (Gyllensten et al. 1991a,b; Wu et al. 1990) very similar to the consensus c-like recombination sequence observed in the minisatellite 33.15 (Jeffreys et al. 1985), could represent a general breakpoint for the exchange of sequence elements. The c-site in Escherechia coli is indeed well known as a sequence element which stimulates RecBCD enzyme dependent recombinations in its vicinity (Dixon and Kowalczykowski 1993), while the hypervariable minisatellite consensus core sequence which The nucleotide sequence data reported in this paper have been submitted to the EMBL/GenBank nucleotide sequence database and have been assigned the accession number Z71541. The name DRB1*0424 was officially assigned by the WHO Nomenclature Committee in July 1996 (Bodmer et al. 1997)