C4 Null Alleles Research Articles

Epidemiology, Evaluation and Management of Anti-Phospholipid Syndrome

Antiphospholipid syndrome is defined as the presence of antiphospholipid antibodies in the setting of thrombosis and/or pregnancy loss (APLS).Antiphospholipid syndrome (APS) is an autoimmune thromboinflammatory condition that affects individuals and their families in a negative and often fatal way. There are the two types of APS: Primary APS, which develops on its own, and secondary APS, which is linked to another autoimmune disorder, most often systemic lupus erythematosus (SLE). The HLA-DR7, DR4, DRw53, DQw7, and C4 null alleles have all been linked to APLS. APS is frequently misdiagnosed due to clinical diversity and a lack of diagnostic test consistency. The classification criteria were created to categorise APS patients for research reasons, but they can also be used by professionals to establish diagnoses. Unprovoked thrombosis is currently treated with long-term warfarin or another vitamin K antagonist drug. To avoid obstetric complications, low-dose aspirin and prophylactic heparin, primarily low-molecular-weight heparin, are utilised.In this article we’ll be looking at Anti-phospholipid Syndrome, it’s etiology, epidemiology, evaluation and management.

C4A deficiency in children and adolescents with recurrent respiratory infections

Increased susceptibility to recurrent viral and bacterial respiratory infections in children and young adults is not well understood. To evaluate the role of complement factor C4 in the defense against respiratory infections, we studied complement factor C4 allotypes C4A and C4B and copy numbers of C4A and C4B genes in 84 children and young adults with recurrent acute otitis media, sinusitis, or pneumonia and in 74 healthy controls. The occurrence of C4A gene deficiency was significantly higher in patients compared with controls (26% vs 14%, p = 0.048). Girls predominated in the group of patients with C4A deficiency (73% girls and 27% boys, p = 0.004). The lectin pathway of complement was more often functionally impaired in patients with C4A deficiency than in patients with no C4A deficiency (41% vs 13%, p = 0.033). Classical and alternative pathways were normal in individuals with C4 null alleles. C4A deficiency is 1 of the minor defects of the innate immunity that may predispose children and young adults to recurrent respiratory infections. C4 gene testing should be added to the list of investigations when the cause for recurrent acute otitis media, maxillary sinusitis, or pneumonia in children and young adults is sought.

Assessment of complement C4 gene copy number using the paralog ratio test.

The complement C4 locus is in the class III region of the MHC, and exhibits copy number variation. Complement C4 null alleles have shown association with a number of diseases including systemic lupus erythematosus (SLE). However, most studies to date have used protein immunophenotyping and not direct interrogation of the genome to determine C4 null allele status. Moreover, a lack of accurate C4 gene copy number (GCN) estimation and tight linkage disequilibrium across the disease-associated MHC haplotypes has confounded attempts to establish whether or not these associations are causal. We have therefore developed a high throughput paralog ratio test (PRT) in association with two restriction enzyme digest variant ratio tests (REDVRs) to determine total C4 GCN, C4A GCN, and C4B GCN. In the densely genotyped CEU cohort we show that this method is accurate and reproducible when compared to gold standard Southern blot copy number estimation with a discrepancy rate of 9%. We find a broad range of C4 GCNs in the CEU and the 1958 British Birth Cohort populations under study. In addition, SNP-C4 CNV analyses show only moderate levels of correlation and therefore do not support the use of SNP genotypes as proxies for complement C4 GCN.

Prolonged clinically asymptomatic evolution after HIV-1 infection is marked by the absence of complement C4 null alleles at the MHC.

The length of time after which persons infected with HIV-1 progress to AIDS is variable. Certain alleles at the MHC have been shown to influence negatively the clinical outcome of HIV-1-infected persons and to be associated with special clinical manifestations. We investigated the MHC class I, class II and class III antigens in 54 Caucasian HIV-1-infected persons. The MHC profile of individuals with a prolonged period before AIDS is marked by a lower frequency of C4 null alleles.

Effects of C4 null alleles and homoduplications on quantitative expression of C4A and C4B.

The availability of MoAbs now allows the accurate quantification of the individual C4 isotypes, C4A and C4B. Using a sensitive two-site immunoradiometric technique to measure serum levels of C4A and C4B, we studied the relationship between genotype and phenotype and physiological factors affecting C4 expression in 129 fully genotyped healthy subjects. Our results confirm that there is extensive phenotypic overlap between genotypic groups and it was not possible to determine the presence of single null alleles from total serum C4. Of the factors which may influence C4 expression, we found that age contributes a very small influence but that gender has no effect and there was no evidence for the presence of feedback of null alleles on the expression of remaining genes. Potential problems in quantifying C4 arising from the complex relationship between isotypic identity and serotypic recognition were highlighted by the finding of reversed antigenic expression of a C4B*5 molecule which was recognized as C4A by the anti-Rg:1 monoclonal used in these studies. We also confirmed that the extended MHC haplotype associated with Felty's syndrome, HLA-B44, C4A*3, C4BQ*O, HLA-DR4, encodes an expressed, duplicated, C4A gene.

Identification of two independent risk factors for lupus within the MHC in United Kingdom families.

The association of the major histocompatibility complex (MHC) with SLE is well established yet the causal variants arising from this region remain to be identified, largely due to inadequate study design and the strong linkage disequilibrium demonstrated by genes across this locus. The majority of studies thus far have identified strong association with classical class II alleles, in particular HLA-DRB1*0301 and HLA-DRB1*1501. Additional associations have been reported with class III alleles; specifically, complement C4 null alleles and a tumor necrosis factor promoter SNP (TNF-308G/A). However, the relative effects of these class II and class III variants have not been determined. We have thus used a family-based approach to map association signals across the MHC class II and class III regions in a cohort of 314 complete United Kingdom Caucasian SLE trios by typing tagging SNPs together with classical typing of the HLA-DRB1 locus. Using TDT and conditional regression analyses, we have demonstrated the presence of two distinct and independent association signals in SLE: HLA-DRB1*0301 (nominal p = 4.9 × 10−8, permuted p < 0.0001, OR = 2.3) and the T allele of SNP rs419788 (nominal p = 4.3 × 10−8, permuted p < 0.0001, OR = 2.0) in intron 6 of the class III region gene SKIV2L. Assessment of genotypic risk demonstrates a likely dominant model of inheritance for HLA-DRB1*0301, while rs419788-T confers susceptibility in an additive manner. Furthermore, by comparing transmitted and untransmitted parental chromosomes, we have delimited our class II signal to a 180 kb region encompassing the alleles HLA-DRB1*0301-HLA-DQA1*0501-HLA-DQB1*0201 alone. Our class III signal importantly excludes independent association at the TNF promoter polymorphism, TNF-308G/A, in our SLE cohort and provides a potentially novel locus for future genetic and functional studies.

Defective prevention of immune precipitation in autoimmune diseases is independent of C4A*Q0

Increased prevalence of C4 null alleles is a common feature of autoimmune diseases. We have shown previously that complement-dependent prevention of immune precipitation (PIP) is defective in patients with systemic lupus erythematosus (SLE), and correlated this defect with C4A*Q0 and low levels of the C4A isotype. To further clarify the role of C4A in the aetiology of SLE, we now extend our studies to other diseases which have been associated with C4A*Q0. The frequency of C4A*Q0 was increased in Icelandic patients with coeliac disease (0.50; P < 0.001), Grave's disease (0.30; P = 0.002) and insulin-dependent diabetes mellitus (0.23; P = 0.04) and in British patients with dermatitis herpetiformis (0.42; P = 0.002) and this was reflected in low levels of C4A. In spite of this, PIP was normal in these patients, and in marked contrast to our previous observations on connective tissue diseases, PIP measurements in these patient groups correlated more strongly with levels of C4B (r = 0.51, P = 0.0000004) than C4A. Patients with increased levels of anti-C1q antibodies had significantly lower PIP than patients without such antibodies (P < 0.01) and a negative association of PIP with anti-C1q antibodies was also reflected in an increased prevalence (P = 0.006) and levels (P = 0.006) of anti-C1q antibodies in patients with subnormal PIP, as well as a negative correlation between PIP and anti-C1q antibodies (r = - 0.25, P = 0.02). These results show that the PIP defect cannot be explained by low levels of C4A alone and suggest that measurements of anti-C1q antibodies may be useful in future studies on the molecular cause of the PIP defect in autoimmune connective tissue disease.

Increased Frequency of C4B*Q0 Alleles in Patients with Henoch-Schonlein Purpura

Henoch-Schonlein purpura (HSP) is a vasculitis of unknown aetiology, possibly involving immune complexes. The complement system is essential for the clearance of immune complexes. Our aim was to explore the hypothesis that patients with HSP have abnormal complements, contributing to the development of the disease. The study included 56 patients diagnosed with HSP at the Children's Hospital, Iceland between 1984 and 2000, and 98 blood donors as controls. Serum levels of immunoglobulin A, C4A, C4B and mannan-binding lectin were measured and compared between the two groups. C4 null alleles were significantly more common in HSP patients than in controls (P = 0.018) and were carried by 66.1% of the patients compared with 41.2% of the controls. This difference was due to an increased frequency of C4B*Q0 allele in the HSP group (0.25 versus 0.11 in the control group; P = 0.002). The fact that the majority of our patients carried a C4 null allele indicates that children with C4 deficiencies may have an increased risk of developing HSP. This may reflect inadequate complement activity and possibly present an opportunity to identify patients at risk of developing serious morbidity associated with HSP.

HLA and hypocomplementemia: the disadvantage of carrying the HLA-B35 and the silent alleles of the C4 complement component.

Hypocomplementemia is an extremely complex phenomenon: we devoted our attention to its immunogenetic basis, particularly to the HLA haplotypes involved and to the study of C4 polymorphic genes. With this in mind we analyzed a group of unrelated patients with hypocomplementemia and 15 families suffering from specific C4 deficiency. Firstly, we performed a population analysis in order to identify a statistically significant association: HLA-B35 and C4BQ0 alleles, in the total group of hypocomplementemic individuals, seem to be associated with the primary disease. Secondly, we defined HLA haplotypes clear-cut segregation in the hypocomplementemic families and we identified the most common HLA haplotypes carrying B35 and C4 null allele associated with this condition. With the aid of correspondence analysis and the Transmission Disequilibrium Test (TDT), we measured the strength of this association. In this work, mainly through family analysis, we envisaged a potentially interesting genomic trait, within HLA, close to B locus, that seems to be involved in hypocomplementemia itself and perhaps in hypocomplementemia-related disorders.

C3 and C4 allotypes in anti-neutrophil cytoplasmic autoantibody (ANCA)-positive vasculitis.

In ANCA-associated small vessel vasculitis few genetic factors have proven to be of importance for disease susceptibility, an exception being deficiency of alpha1-anti-trypsin, the main inhibitor of proteinase 3 (PR3). Alerted by our finding that myeloperoxidase has affinity for C3, and the finding of an increased frequency of the C3F allele in systemic vasculitis in a British cohort, we examined polymorphism of C3 and C4 in patients with ANCA+ small vessel vasculitis. After identification of all patients at our department with a positive ANCA test during the period 1991-95 and a diagnosis of small vessel vasculitis, blood samples were collected after informed consent. The 67 included patients were grouped according to ANCA serology and disease phenotype using the Chapel Hill nomenclature. The gene frequency of C3F was found to be increased (0. 32) compared with controls (0.20; P < 0.05) in the PR3-ANCA+ subgroup. The frequency of C4A3 was increased in the group as a whole, but no increase of C4 null alleles was seen. The findings imply a role for the complement system in the pathogenesis of ANCA-associated small vessel vasculitis.

Circulating immune complexes and complement C4 null alleles in patients in patients operated on for premature atherosclerotic peripheral vascular disease.

Circulating immune complexes can lead to vascular inflammation and premature atherosclerosis and the fourth component of complement, C4, plays an important role in the removal of immune complexes. The objective of this study was to analyze the relation between circulating immune complexes and C4 null alleles in patients operated on for peripheral vascular disease before the age of 50. The prevalence of circulating immune complexes and null alleles of C4 (C4Q0) was determined in 62 patients with peripheral atherosclerosis requiring surgery before 50 years of age and in a matched control group. C4A and C4B null alleles (C4A*Q0, C4B*Q0) were determined by electrophoresis of plasma, followed by immunofixation. C4A and C4B concentrations were measured by ELISA. Circulating immune complexes were determined by sucrose density gradient centrifugation and gel filtration. There was no difference in the distribution of C4Q0 between patients and controls. The patients had higher prevalences and levels of circulating immune complexes. This was correlated with the presence of C4Q0, especially C4A*Q0. There was an inverse correlation of concentration of circulating immune complexes with C4A levels and with ratio of C4A/B levels. Thus, a significant proportion of patients with premature peripheral atherosclerosis had circulating immune complexes and C4A*Q0 enhanced the propensity to immune complex formation. This might represent one mechanism for vascular damage in this patient group.

C4 null alleles seem to be a genetic marker for HCV-related mixed cryoglobulinemia

C4 null alleles seem to be a genetic marker for HCV-related mixed cryoglobulinemia

Polymorphism of Complement C4 and Susceptibility to IDDM and Microvascular Complications

The aim of this study was to investigate whether or not the inherited polymorphism of complement C4 is associated with genetic susceptibility to microvascular complications in IDDM as previously reported. We determined C4 phenotypes in 241 patients with IDDM and 140 healthy control subjects by agarose gel electrophoresis and immunoprecipitation. C4 allotype frequencies were compared between patients and healthy control subjects. In addition, we compared allotype frequencies of 83 patients with nephropathy with those of 80 patients without nephropathy and compared those of 50 patients with proliferative retinopathy with those of 68 patients without retinopathy or background retinopathy. Duration of IDDM in control patients was at least 21 years. Patients and healthy control subjects differed at both the C4A (P < 0.00001) and C4B (P < 0.0005) loci. The C4 null allele C4AQ0 was significantly increased in IDDM patients (26.8 vs. 11.8%, P < 0.005). C4B2 was more frequently observed in patients (14.5 vs. 6.8%, P < 0.05) compared with healthy control subjects. No differences were observed in C4 allotype distribution between patients with and without nephropathy or retinopathy. These data confirm previous reports of an association between the C4 null allele C4AQ0 and IDDM. Our results do not support an association of the inherited polymorphism of complement C4 with genetic susceptibility to microvascular complications in patients with IDDM.

C4 NULL ALLELES IN A SWEDISH POPULATION

The distribution of C4 null alleles (C4Q0) was studied in a Swedish population comprising 410 male individuals from Stockholm County and Uppsala City. The prevalances of homozygous C4A*Q0 and C4B*Q0 determined by analysing the gene products in serum were 5.0 and 5.6%, respectively, in the Stockholm population and 4.0% each in the Uppsala population. These values are higher than those previously reported from other Caucasian populations. The finding reflects a different genetic composition of the Swedish population and may have relevance to the disease spectrum in this population as compared to other Caucasian populations.

Association Between Circulating Immune Complexes, Complement C4 Null Alleles, and Myocardial Infarction Before Age 45 Years

One hundred patients who had survived a myocardial infarction before the age of 45 years and 90 age- and sex-matched healthy individuals were investigated for circulating immune complexes (CICs) and the presence of complement C4 null alleles (C4Q0). CICs were found in increased concentrations in 20% of patients and 6.7% of control subjects. Patients and control subjects had the same prevalence of C4Q0. CICs were present in all patients and in 36% of the control subjects homozygous for C4Q0. Patients and control subjects heterozygous for C4Q0 had CICs in 71% and 0%, respectively. The high prevalence and a high concentration of CICs were particularly associated with C4A*Q0. Patients homozygous for C4A*Q0 had concentrations of LDL that were lower than found in other patients. The increased concentration of CICs associated with genetic deficiencies of the complement factor C4 might thus be an additional etiological factor for the development of chronic vascular damage and premature myocardial infarction.

Major histocompatibility complex class II and C4 alleles in Mexican Americans with systemic lupus erythematosus.

Few data exist on associations of class II and class III alleles of the major histocompatibility complex (MHC) and susceptibility to systemic lupus erythematosus (SLE) in Mexican Americans, a group of predominantly mixed Spanish and Native American ancestry. Therefore, MHC class II alleles (HLA-DRB1, DQA1, DQB1, DPA1 and DPB1 alleles) and C4 allotypes were determined in 52 Mexican American SLE patients and 105 ethnic-matched controls. HLA-DRB1*0301 and C4A*Q0 were each increased in the SLE patients, especially HLA-DRB1*0301 in those with anti-Ro/SSA autoantibodies. C4A*Q0 was associated with HLA-DRB1*0301 only in a minority of patients and controls. Anti-U1RNP antibodies were significantly associated with the presence of HLA-DQB1*0302, and the risk for the production of anti-Ro antibodies was heightened by the presence of at least three (out of four possible) DQA1 chains possessing a glutamine at position 34 and/or DQB1 chains a leucine at position 26 of their outermost domains. Thus the HLA class II and C4 null allele associations that have been noted in other ethnic groups are also found in Mexican Americans, suggesting shared susceptibility factors across ethnic lines in predisposition to SLE.

Increased frequency of the C4A*6 rare allele in rheumatic heart disease.

The aim of the present investigation was to determine whether the alleles of the MHC class III complement proteins BF, C2 and C4 (C4A and C4B) could be markers for RHD in the Brazilian population. Forty-nine patients with chronic RHD were studied. The controls included 65 healthy unrelated individuals, matched with the patients according to sex, age and ethnical background. BF, C2, C4A and C4B allotypes were determined by standard technologies including Western blots for C2 and C4 variants with monoclonal and policlonal antibodies. The results showed a significantly elevated presence of the C4A*6 rare allele (p = 0.003 RR = 11.85) and a decrease of C4A*3 in the patients. In addition, C4 null and BF and C4 rare alleles were more frequent in patients than in the controls. Considering that in this investigation only RHD patients were included, further studies are necessary in order to clarify whether C4A6 is a marker for the cardiac form or for the disease itself.

Complement polymorphism in herpes gestationis: Association with C4 null allele

Herpes gestationis (HG) is a rare, pregnancy-related skin disease characterized by the production of an autoantibody to a component of the hemidesmosome. It is associated with the class II antigens HLA-DR3 and HLA-DR4, but its potential association with the "class III antigens" C2, C4, and factor B has not previously been studied. Our purpose was to study complement polymorphism in HG. Using electrophoresis and immunofixation techniques, we determined the allele frequencies of C4A, C4B, C3, and factor B in 42 patients with a history of HG. Ninety percent of patients carried a C4 null allele (C4*QO). No statistically significant association with C3 or factor B alleles was seen. HG is associated with the presence of a C4*QO. Whether the C4*QO is the primary genetic association, or whether the C4*QO is related to its linkage disequilibrium with DR3 and DR4 has yet to be determined.

Early-onset autoimmune hepatitis is associated with a C4A gene deletion

Background: Autoimmune hepatitis is an immunologically mediated disorder with some similarities to systemic lupus erythematosus, including an association with HLA-A1, B8, DR3. This haplotype includes a C4A, 21-OHA gene deletion. Low serum levels of complement and C4 null alleles have been reported in autoimmune hepatitis, but studies have been at the protein level only.Methods: Twenty-four white patients with autoimmune hepatitis were studied by Southern blots using a C4A gene complementary DNA probe. HLA A, B, and C typing was determined using standard microcytotoxicity assays, and DR and DQ specificities were determined by restriction fragment length polymorphism analysis. Results: Thirteen of 24 patients had the C4A gene deletion compared with 12 of 90 controls. HLA-A1 and B8 were increased in patients with autoimmune hepatitis, as were HLA-DR3 (DR17), Dw24, DQ2. Patients with a C4A gene deletion presented at a younger age than those without the deletion and had significantly lower serum C3 and C4 levels. The C4A gene deletion was associated with HLA-A1, B8, DR3 in all but 1 patient who was HLA-DR3 negative. Conclusions: A C4A gene deletion is found in patients with autoimmune hepatitis, especially those presenting at a young age. This complement gene deletion may be an important factor in the development of this disease.

COMPLEMENT DEFICIENCY AND DISEASE

Summary of complement deficiencies Component Number of cases Chromosome location of gene Disease associations Classical pathway proteins Clq Clr/Cls C4 C2 I>40 10 17 t>100 A, B chains 1,p; C chain ? Closely linked on 12,p13 Two genes C4A and C4B in MHC on chromosome 6 MHC locus on chromosome 6, adjacent to factor B Alternative pathway components Factor D 1 ? ?X Properdin I>50 X C3, factors H and I C3 16 19 Factor I 15 4 Factor H 12 1 in RCA cluster Terminal components C5 19 9 C6 >150 5 closely linked to C7 C7 26 5 C8 32 c~, 13 closely linked on 1, y on 9 C9 5 (Caucasoid) 5 Japanese (many) SLE in majority; pyogenic infections including meningitis As above As above Pyogenic infections; SLE; many healthy Neisserial infections Neisserial infections; rarely other pyogenic infection Pyogenic infections; glomerulonephritis; SLE As above As above; haemolytic uraemic syndrome Neisserial infections; rarely SLE As above As above As above Neisserial infection; weak association with neisserial infection Data derived in part from Ref. 24 and updated to the end of 1990 by newly described cases. and preliminary data suggest that deficiency is often the result of a defect in synthesis of the B chain 7. The genes encoding the A and B chains are closely linked on chromosome lp, whereas that encoding the C chain has yet to be assigned. Deficiencies of Clr or Cls are rare (total of ten cases) and result from a failure to synthesize these subcompo- nents. Most of the individuals so far described have had a combined deficiency of the two subcomponents, a prob- able consequence of their close genetic linkage (Table 1). Typically, these individuals have no Clr and reduced levels of Cls (20-40O/o) 7. C4 is encoded by two closely linked, highly poly- morphic genes, located within the MHC on chromosome 6, which give rise to the isotypic forms, C4A and C4B. Complete C4 deficiency thus requires the inheritance of null alelles at both C4 loci and only about 16 individuals with homozygous deficiency have been identified, most of whom have presented with early onset, severe, SLE 8. Single null alleles are common at both loci, resulting in the frequent detection of individuals with heterozygous deficiencies of one or both isotypes or, more rarely, homozygous deficiency of one isotype 8. It has been esti- mated that only about 60% of the population have four functioning C4 genes, making partial deficiency of C4 the commonest immune deficiency in man. Most null alleles do not contribute to C4 protein production and the serum concentration is thus related, approximately, to the number of functional genes. However, as there is considerable overlap of C4 concentrations between the different genotypes, it is impossible to ascertain C4 null alleles simply by measuring C4 concentration. The strong link between homozygous complement deficiency and SLE led to the formulation of the hypoth- esis that partial inherited complement deficiency might also cause increased disease susceptibility. Initial studies showed that there is indeed an increased prevalence of null alleles, mainly of C4A, among Caucasoid patients with SLE but the analysis was confounded by strong linkage disequilibrium between C4A null alleles and HLA-DR3 (Ref. 9), which effectively prevented separ- ation of the relative contribution of these two putative disease susceptibility genes. One approach to at- tempt to disentangle the relative contributions of HLA- DR3 and C4A Q*0 to disease susceptibility is examin- ation of the MHC associations with SLE in different racial groups, in which haplotypes containing different combinations of the allotypic variants of the different MHC gene products are found. With this type of ap- proach, it has been found that the association of SLE with