C4 polymorphism and major histocompatibility complex haplotypes in IgA deficiency: association with C4A null haplotypes.

IgA-deficient individuals (n = 110) and six families comprising 9 cases of IgA deficiency were typed for HLA-A, -B, -DR, C4 and factor B. Phenotype frequencies were increased for HLA-B8 (p = 0.004), HLA-DR3 (p = 0.001) and homozygous C4AQ0 (p = 0.01) and decreased for HLA-B7 (p = 0.004), HLA-DR2 (p = 0.0001) and C4A3 (p = 0.00007) compared to controls. Homozygous C4A deficiency was found in 20% of IgA-deficient persons. As clearly suggested by investigation of families, the findings could be attributed to high prevalence of the extended major histocompatibility complex (MHC) haplotype [HLA-A1, B8, C4AQ0, C4B1, BfS, DR3] in IgA deficiency. All but 1 of the 9 IgA-deficient persons included in the family study carried this haplotype and 4 of them were homozygous. In the families, 3 persons with normal serum IgA concentrations had the same MHC haplotypes as their IgA-deficient relatives. The findings were also consistent with possible overrepresentation of other MHC haplotypes with aberrant C4 gene organization in IgA deficiency. As previously suggested, the presence of two MHC haplotypes associated with IgA deficiency appears to be a necessary but not sufficient requirement for manifestation of the condition. The putative existence of a recessive gene in the MHC with regulatory function with regard to IgA gene expression is consistent with the findings.     

Prospective analysis suggests susceptibility genes for deficiencies of IgA and several other immunoglobulins on the [HLA-B8, SC01, DR3] conserved extended haplotype

The extended major histocompatibility complex (MHC) haplotype [HLA-B8, SC01, DR3] is increased in frequency among patients with immunoglobulin (Ig)A deficiency and common variable immunodeficiency. Because the genomic region from HLA-B to HLA-DR/DQ is virtually the same on all instances of the haplotype in the general population, we reasoned that all independent instances of [HLA-B8, SC01, DR3] carry MHC susceptibility genes for these disorders. To define immunoglobulin deficiencies determined by genes on this haplotype and their mode of expression and penetrance, serum immunoglobulin class and IgG subclass concentrations were determined prospectively in homozygotes and heterozygotes of this haplotype and in Caucasian controls. Prevalence of individual immunoglobulin deficiencies in persons with [HLA-B8, SC01, DR3] ranged from 13% to 37%, significantly higher than rates in non-carriers or general controls. We found significantly increased frequencies of IgA and IgG4 deficiency only in homozygotes (13.3% and 30%, respectively) compared with heterozygotes (1.7% and 3.4%) or non-carriers (1.6% each), suggesting recessive expression. In contrast, IgD and IgG3 deficiencies were significantly more common in both homozygotes (36.7% and 30%) and heterozygotes (20.3% and 17.5%) compared with controls (4.9% and 3.4%), suggesting dominant inheritance. These results indicate multiple distinct susceptibility genes, some recessive and others dominant, for deficiency of IgA, IgD, IgG3 or IgG4 (but not for IgE, IgG1, IgG2 or IgM) on [HLA-B8, SC01, DR3]. These observations may also help to explain the observed associations of [HLA-B8, SC01, DR3] with both IgA deficiency and common variable immunodeficiency and the common occurrence of IgG subclass deficiencies in some patients with IgA deficiency.     

The [HLA-B18, F1C30, DR3] Conserved Extended Haplotype Carries a Susceptibility Gene for IgD Deficiency

We showed previously that the conserved extended MHC haplotype [HLA-B8, SC01, DR3] carries recessive susceptibility genes for IgA and IgG4 deficiency and dominant genes for IgD and IgG3 deficiency. [HLA-B18, F1C30, DR3] has similar class II and III regions to [HLA-B8, SC01, DR3] and is common in the Basques. We therefore studied serum immunoglobulin concentrations in Basque homozygotes, heterozygotes, and noncarriers of (F1C30, DRB1*0301, DRB3*02, DQA1*0501, DQB1*0201) (F1C30, DR3). As shown by others, no subjects were deficient in IgA, IgM, or IgG subclasses. In contrast, 29% of homozygotes and three of seven double heterozygotes with (SC01, DRB1*0301, DRB3*0101, DQA1*0501, DQB1*0201) (presumed homozygotes for IgD deficiency susceptibility genes) were IgD deficient. Thus, 32% of presumed homozygotes were IgD deficient compared with 1.6% of noncarriers. Of haplotype heterozygotes, 25% were IgD deficient. The high frequency of IgD deficiency in both homozygotes and heterozygotes for (F1C30, DR3) suggests a partially penetrant dominant susceptibility gene for IgD deficiency on [HLA-B18, F1C30, DR3].     

Immunoglobulin deficiencies and susceptibility to infection among homozygotes and heterozygotes for C2 deficiency

About 25% of C2-deficient homozygotes have increased susceptibility to severe bacterial infections. C2-deficient homozygotes had significantly lower serum levels of IgG2, IgG4, IgD, and Factor B, significantly higher levels of IgA and IgG3 and levels of IgG1 and IgM similar to controls. Type I (28 bp deletion in C2 exon 6 on the [HLA-B18, S042, DR2] haplotype or its fragments) and type II (non-type I) C2-deficient patients with increased susceptibility to bacterial infection had significantly lower mean levels of IgG4 (p < 0.04) and IgA (p < 0.01) than those without infections (who had a higher than normal mean IgA level) but similar mean levels of other immunoglobulins and Factor B. Of 13 C2-deficient homozygotes with infections, 85% had IgG4 deficiency, compared with 64% of 25 without infections. IgD deficiency was equally extraordinarily common among infection-prone (50%) and noninfection-prone (70%) homozygous type I C2-deficient patients. IgD deficiency was also common (35%) among 31 type I C2-deficient heterozygotes (with normal or type II haplotypes), but was not found in 5 type II C2-deficient heterozygotes or 1 homozygote. Thus, C2 deficiency itself is associated with many abnormalities in serum immunoglobulin levels, some of which, such as in IgG4 and IgA, may contribute to increased susceptibility to infection. In contrast, IgD deficiency appears not to contribute to increased infections and appears to be a dominant trait determined by a gene or genes on the extended major histocompatibility complex (MHC) haplotype [HLA-B18, S042, DR2] (but probably not on type II C2-deficient haplotypes) similar to those previously identified on [HLA-B8, SC01, DR3] and [HLA-B18, F1C30, DR3].     

Recurrent extended HLA haplotypes in children with selective IgA deficiency

HLA supratypes as well as serum IgG, IgA and IgM levels were determined in 44 children and adolescents with severe IgA deficiency (serum IgA less than 5 mg/dl) and in first degree relatives. Frequencies of the HLA alleles B14, DR1, DQW1, C4A2 and C4B2 were significantly higher in the IgA-deficient patients than in the controls. The most recurrent haplotype among patients was B14, DR1 (p less than 10(-4) preferentially associated with A33, A28 or A blank. The supratype B14, Bfs, C4A2, C4B2, DR1, DQW1 was present in a 14-fold higher frequency than in the controls, and strongly suggests the presence of a gene in the HLA region involved in the deficiency of IgA. The fact that this supratype was not always associated with IgA deficiency in the parents and that not all IgA-deficient subjects had this supratype is discussed. Severe IgA deficiency was found in four mothers (two of the four mother-child pairs shared the B14, DR1 haplotype); three other relatives (one father and two brothers) had partial IgA deficiency and did not have the B14, DR1, DQW1 haplotype.     

Gm allotypes in IgA deficiency

Gm phenotypes were examined in 90 Swedish IgA-deficient (less than 0.05 g/litre of serum IgA) donors and 40 normal first and second degree relatives of six of these donors. The G1m1,2, G3m5 and Km1 frequency in the group of IgA-deficient donors did not differ from that found in the normal population. Among the relatives, HLA and/or Gm identical normal sibs were observed. Anti-IgA antibodies were present in 29 of the IgA-deficient donors and anti-IgG in seven. No association between the two was found. A statistically significant association between the G1m-2 phenotype and the presence of anti-IgA antibodies was observed. When subdivided according to HLA type, a non-random distribution of Gm phenotypes was seen in HLA-B8/DR3 positive individuals with anti-IgA antibodies (HLA-B8/DR3 being the haplotype associated with IgA deficiency). These data suggest an association between IgA deficiency, anti-IgA and the studied Gm allotypes.     

Central MHC genes affect IgA levels in the human: reciprocal effects in IgA deficiency and IgA nephropathy

This study investigates the hypothesis that alternative alleles of one or more genes in the central major histocompatibility complex (MHC) predispose carriers to IgA deficiency (IgAD) or IgA Nephropathy (IgAN). Australian caucasian IgAD, IgAN patients, and controls were typed at HLA loci, single nucleotide polymorphisms, and microsatellites in the MHC. Alleles of the D6S273 microsatellite exhibited strong associations with IgAD and IgAN. D6S273*129 and *139 were more frequent in IgAD and less frequent in IgAN patients than controls. The reverse was true for D6S273*133 and *131. Alleles of other microsatellites exhibited weak associations with IgAD or IgAN. D6S273*129 is found on the 65.1 ancestral haplotype [HLA-B14(65),DR1], which has been reported to be increased in IgAD, but the majority of IgAD patients with D6S273*129 did not have other alleles of the haplotype. D6S273*139 is characteristic of the 8.1 ancestral haplotype (HLA-A1,B8,DR3), which was common in IgAD and rare in IgAN patients. Further studies of the 8.1 haplotype in Australian, German and Spanish caucasian subjects revealed that HLA-DR3, in the absence of -B8, is not associated with IgAD. However -B8 is associated with IgAD in the absence of -DR3, consistent with a susceptibility locus in the central MHC. Provisional mapping within this region is discussed.     

Two variants of DRw52, DR3, and DQw2 specificities distinguish two different DR3-bearing extended haplotypes

HLA-DR beta and DQ beta MHC antigens present on B-lymphoblastoid cell lines homozygous for either [HLA-B8,DR3,SC01] or [HLA-B18,DR3,F1C30] were studied by two-dimensional gel electrophoresis. Comparison of neuraminidase-treated DR proteins from these cells showed that DR3-bearing cells express two DR beta genes. However, one DR beta chain (beta a) is nonpolymorphic, whereas the other beta chain (beta b) is polymorphic. Two variants of DR beta (DR3 or DRw52) and two of DQ beta (DQw2) were found, with all the examples of each extended haplotype carrying only one of these sets of variants. The variants thus absolutely distinguished the two DR3-bearing extended haplotypes. These results support the hypothesis of extended haplotypes at the protein level, and demonstrate the fixity of alleles on them in the HLA-B-D/DR region.     

Molecular analysis of the IGHA and MHC class III region genes in one family with IgA and C4 deficiencies

We report molecular studies in 2 IgA-deficient persons. One of them had an unusual association with an acute lymphoblastic leukaemia; his sister was also IgA deficient and shared an HLA haplotype and a complotype known to be associated to IgA deficiencies. The 2 IgA-deficient siblings also had low C4 serum levels due to C4A*Q0 allele. We showed that both defects were transmitted independently in the family. Molecular analysis revealed no major structural defects of the IGHA coding and switch regions, whereas a broad C4A-21-OHA deletion was responsible for the C4A*Q0 phenotype. These results confirm previous data showing that IgA deficiencies seem to be, in most cases, a regulatory defect rather than a structural defect of the coding IGHA region itself. These data were further supported by another molecular study in a patient with a recurrent Landry-Guillain-Barre syndrome who showed total absence of serum IgA and sIgA+B cells with no major structural defect of the IGHA region.     

Complotypes, extended haplotypes, male segregation distortion, and disease markers

From our studies in Caucasian families of HLA, complement, and glyoxalase alleles have developed the concepts of the complotype and the extended haplotype. complotypes are clusters of the four genes for complement proteins encoded within the MHC designated (in arbitrary order) by their BF, C2, C4A, and C4B alleles. They are inherited in families and occur in populations as functionally single genetic units and exhibit linkage disequilibrium with HLA-B and HLA-DR alleles which are complotype, rather than complement gene allele, specific. In Caucasians, there are 10-12 common sets of HLA-B, DR, complotype sets that show significant linkage disequilibrium. These haplotypes constitute 25-30% of all MHC haplotypes in Caucasians. Because there is evidence for relative fixity of alleles on these chromosomes to an unknown extent beyond the HLA-B-DR interval, they have been called extended MHC haplotypes. It appears likely that it is these extended haplotypes that provide most of the known linkage disequilibrium pairs previously reported for MHC alleles as well as many of the known MHC allele-disease associations. The most common extended haplotype [HLA-B8, DR3, SC01], when it carries GLO2, is increased in type I diabetes mellitus and probably a number of other diseases, including gluten-sensitive enteropathy and membranoproliferative glomerulonephritis. In the families with these disorders studied by us, this haplotype exhibits male segregation distortion, a feature displayed by t-mutants found in wild mouse populations. This feature constitutes an important selective advantage for the chromosome and may contribute to the accumulation of susceptibility mutations for a variety of diseases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Genetic prediction of nonresponse to hepatitis B vaccine

In previous studies of the antibody response to hepatitis B vaccine in 598 subjects who received a full course of vaccination, we observed a bimodal response, with about 14 percent producing less than approximately 1000 radioimmunoassay (RIA) units. An analysis of the major histocompatibility complex (MHC) HLA and complement types of 20 of the subjects with the lowest responses indicated a greater-than-expected number of homozygotes for the extended or fixed MHC haplotype [HLA-B8, SC01, DR3]. This finding suggested that the lack of a normal response was a recessive MHC-linked trait. In this study, we prospectively vaccinated five homozygotes and nine heterozygotes for this haplotype in the expectation that the homozygotes would produce much lower levels of antibody than the heterozygotes. When the antibody response was assessed two months after the third injection, four of the five homozygotes had produced very low levels (approximately 1000 units or less) of antibody (mean, 467 RIA units; range, less than 8 to 1266), whereas all nine heterozygotes produced more than 2500 RIA units (mean, 15,608; range, 2655 to 28,900) (P less than 0.01). We conclude that the usual response to hepatitis B surface antigen is due to the presence of a dominant immune-response gene in the MHC and that a low response is due to the absence of such a gene and the presence on both chromosomes of MHC haplotypes (such as [HLA-B8, SC01, DR3]) that indicate such a response.     

The genetic basis for the association of the 8.1 ancestral haplotype (A1, B8, DR3) with multiple immunopathological diseases

Summary: An individual's major histocompatibility complex (MHC) ancestral haplotype (AH) is the dearest single determinant of susceptibility to MHC associated immunopathological disease, as it defines the alleles carried at all loci in the MHC. However, the direct effects of any of the 150–200 genes that constitute the MHC are difficult to determine since recombination only occurs at defined hotspots. This review concerns the 8.1 AH (HLA-A1, C7, B8, C4AQ0, C4B1, DR3, DQ2), which is carried by most Caucasians with HLA-B8. It is associated with accelerated human immunodeficiency virus (HIV) disease, and susceptibility to insulin-dependent diabetes mellitns (IDDM), systemic lupus erythematosus, dermatitis herpetiformis, common variable immunodeficiency and IgA deficiency, myasthenia gravis and several other conditions. We have mapped susceptibility genes for HIV, IDDM and myasthenia gravis co the central MHC between HLA-B and the tumour necrosis factor or complement genes. Here we consider which of the remaining 8.1-associated diseases are more closely associated with HLA-DR3 and/or DQ2. Several candidate genes in the central MHC have the potential to modulate immune or inflammatory responses in an antigen-independent manner, as is seen in studies of cultured cells from healthy carriers of the 8.1 AH. Hence these genes may act as a common co-factor in the diverse immunopathological conditions associated with the 8.1 AH.     

C4 Chido 3 and 6 distinguish two diabetogenic haplotypes: HLA-B49, SC01,DR4,DQw8 and B8,SC01,DR3,DQw2.

The combination of the HLA complement allotypes BFS, C2C, C4AQ0 (deleted gene) and C4B1, termed SC01 complotype, usually present in the HLA-B8,DR3,DQw2 diabetogenic haplotype, has also been found in a novel "low frequency" HLA-B49,DR4,DQw8 haplotype associated with Spanish insulin-dependent diabetes mellitus (IDDM). Family studies of C4 antigenic determinants Rodgers/Chido and their specific C4d nucleotide sequences confirm that this novel haplotype bearing Chido -3, -6 is not due to a recent recombination from the common HLA-B8,DR3 haplotype bearing Chido 3,6; moreover, Chido analysis at the serological or DNA level is presently the only way to distinguish both SC01 complotypes, since BF, C2, steroid 21-hydroxylase and C4 genes do not reveal other differences by restriction fragment analysis. On the other hand, HLA-B49,SC01,DR4 is the first DR4-bearing IDDM-susceptible haplotype with a deleted C4 gene described so far and the only DR4-bearing haplotype found in the Spanish population. This report further supports the fact that extended haplotypes with deleted (or "not duplicated") genes in the class III region contain IDDM-susceptibility more often than non-deleted (or "duplicated") haplotypes in the Spanish and other Mediterranean populations.     

The MHC type 1 diabetes susceptibility gene is centromeric to HLA-DQB1

HLA-DQB1 is widely considered to be the major histocompatibility complex (MHC) susceptibility gene for type 1 diabetes (T1D). However, since inheritance of the gene in T1D is recessive, the presence of the protective HLA-DQB1 0602 allele with normal nucleotide sequence in some patients raises the question of whether HLA-DQB1 is not the susceptibility locus itself but merely a good marker. HLA-DQB1 0602 is part of a conserved extended haplotype (CEH) [HLA-B7, SC31, DR2] (B7, DR2) with fixed DNA over more than 1Mb of genomic DNA that normally carries a protective allele at the true susceptibility locus. We postulated that, in patients with HLA-DQB1 0602, the protective allele at the susceptibility locus has been replaced by a susceptibility allele through an ancient crossover at meiosis centromeric to HLA-DQB1. We analyzed single nucleotide polymorphisms (SNPs) distinguishing the HLA-DQA2 (the first expressed gene centromeric to HLA-DQB1) allele on the normal HLA-B7, DR2 CEH from those on susceptibility CEHs in T1D patients and controls with HLA-DQB1 0602. All but 1 of 20 healthy control HLA-DQB1 0602 haplotypes had identical (consensus) first intron HLA-DQA2 5-SNP haplotypes. Fifteen of 19 patients with HLA-DQB1 0602 were homozygous for 1 or more HLA-DQA2 SNPs differing from consensus HLA-DQA2 SNPs, providing evidence of crossover involving the HLA-DQA2 locus. The remaining 4 patients were heterozygous at all positions and therefore uninformative. The loss of dominant protection usually associated with HLA-DQB1 0602 haplotypes is consistent with a locus centromeric to HLA-DQB1 being a major determinant of MHC-associated susceptibility, and perhaps the true T1D susceptibility locus.     

HLA-Cw alleles associated with HLA extended haplotypes and C2 deficiency

Abstract: There are four MHC-linked complement genes, BF, C2, C4A and C4B, that are inherited as single DNA units, known as complotypes. Extended haplotypes were initially defined by studying the distribution of complotypes in relation to HLA-B and HLA-DR loci in Caucasian families. In order to analyze the distribution of HLA-Cw alleles in relation to extended haplotypes, we studied a large panel of MHC homozygous and heterozygous cell lines representing previously described Caucasian-derived extended haplotypes and 14 patients with complete C2 deficiency. HLA alleles were assigned using sequence-specific oligonucleotide probe hybridization (SSOP). Family analysis served to assign haplotypes for heterozygous samples. We found distinctive HLA-Cw alleles for each independent extended haplotype. Their association in each instance was statistically significant All patients with C2 deficiency carrying the haplotype [HLA-B18, S042, DR2] were associated with HLA-Cw*1203. These conserved allelic combinations may become an important tool for the study of human evolution and may contribute to the expeditious selection of prospective donors in clinical transplantation.     

Genetic and immunologic analysis of a family containing five patients with common-variable immune deficiency or selective IgA deficiency

A family with 13 members included 2 subjects with selective IgA deficiency (IgA-D) and 3 subjects with common-variable immune deficiency (CVID), diseases which usually occur sporadically. Reciprocal combinations of B and T cellsin vitro between one normal and two immune-deficient family members and normal subjects revealed that defective Ig synthesis was determined by the B cells, while the patient T cells functioned normally. Normal T helper and suppressor function was demonstrated even in one patient with CVID who developed a T-cell lymphoproliferative disorder associated with elevated IgM; this patient's B cells made only IgMin vitro. Immune deficiencies were inherited in this family in a pattern consistent with an autosomal dominant trait with incomplete penetrance. All the immune-deficient patients in this family possessed at least one copy of an MHC haplotype previously shown to be abnormally frequent in IgA-D and CVID: HLA-DQB1*0201, HLA-DR3, C4B-Sf, C4A-deleted, G11-15, Bf-0.4, C2-a, HSP70-7.5, TNF-5, HLA-B8, and HLA-A1. The patient who developed the lymphoproliferative disorder was homozygous for this haplotype. Four immunologically normal members, one of whom was 80 years old, also possessed this MHC haplotype, indicating that its presence is not sufficient for disease expression. A small segment of another MHC haplotype associated with Ig deficiency in the population also occurred in this family, but it was not associated with immune deficiency. The presence of neutral amino acids at position 57 of DQ, previously correlated with IgA-D, was associated with disease in this family approximately to the same degree reported previously in unrelated patients. Thus the expression of immunodeficiency in individuals bearing a disease-associated MHC haplotype appears to require either additional genes or an environmental trigger.     

Increased Apoptosis of CD20+ IgA+ B Cells is the Basis for IgA Deficiency: The Molecular Mechanism for Correction In Vitro by IL-10 and CD40L

IgA deficiency is the most common primary immunodeficiency in humans. Comparative analysis of gene expression in PBMC from IgA-deficient (IgAd) and normal donors using functional multiplex panels showed overexpression of the Caspase-1 (CASP-1) gene. Cells from all the IgAd donors (n=7) expressed 4–10-fold caspase-1 mRNA over normal controls (n=5). CD19⁺ B cells from all IgAd donors produced IgA in cultures following IL-10 and CD40L with Staphylococcus aureus (Cowan) (SAC) or tetanus toxoid (TT) treatments. In CD19⁺ B cells from IgAd donors, reconstitution of IgA secretion was associated with protection of the CD20⁺ B cell population that underwent apoptosis in the absence of IL-10, CD40L, and TT (triple treatment). Caspase-1 gene expression was decreased in the reconstituted cells. Furthermore, treatment with a caspase-1 inhibitor also independently protected against B cell apoptosis in vitro. An apoptosis-specific cDNA array showed differential expression of 4 out of 96 genes and a shift towards survival-related gene expression from the apoptotic to the protected B cells after triple treatment. There was an increase in the expression of the IAP-2 (inhibitor of apoptosis) gene in the reconstituted cells. Upregulation of the IAP-2 gene protects B cells from deletion and allows for IgA secretion in this system. The inability to detect secreted IgA in IgAd patients could result from the loss of IgA-committed B cells that express high levels of caspase-1.