Complement factor C4 and immunoglobulins in recurrent or chronic mucosal infections

The study assessed whether plasma concentrations of complement factors C3, C4, or immunoglobulins, serum classical pathway hemolytyic activity, or polymorphisms in the class I and II HLA genes, isotypes and gene numbers of C4, or allotypes of IgG1 and IgG3 heavy chain genes were associated with severe frequently recurring or chronic mucosal infections. According to strict clinical criteria, 188 consecutive voluntary patients without a known immunodeficiency and 198 control subjects were recruited. Frequencies of low levels in IgG1, IgG2, IgG3 and IgG4 were for the first time tested from adult general population and patients with acute rhinosinusitis. Frequently recurring intraoral herpes simplex type 1 infections, a rare form of the disease, was associated with homozygosity in HLA -A*, -B*, -C*, and -DR* genes. Frequently recurrent genital HSV-2 infections were associated with low levels of IgG1 and IgG3, present in 54% of the recruited patients. This association was partly allotype-dependent. The G3mg,G1ma/ax haplotype, together with low IgG3, was more common in patients than in control subjects who lacked antibodies against herpes simplex viruses. This is the first found immunogenetic deficiency in otherwise healthy adults that predisposes to highly frequent mucosal herpes recurrences. According to previous studies, HSV effectively evades the allotype G1ma/ax of IgG1, whereas G3mg is associated with low IgG3. Certain HLA genes were more common in patients than in control subjects. Having more than one C4A or C4B gene was associated with neuralgias caused by the virus. Low levels of IgA, IgG1, IgG2, IgG3, and IgG4 were common in the general adult population, but even more frequent in patients with chronic sinusitis. Only low IgG1 was more common chronic than in acute rhinosinusitis. Clinically, nasal polyposis and bronchial asthma were associated with complicated disease forms. The best differentiating immunologic parameters were C4A deficiency and the combination of low plasma IgG4 together with low IgG1 or IgG2, performing almost equally. The lack of C4A, IgA, and IgG4, all known to possess anti-inflammatory activity, together with a concurrently impaired immunity caused by low subclass levels, may predispose to chronic disease forms. In severe chronic adult periodontitis, any C4A or C4B deficiency combined was associated with the disease. The new quantitative analysis of C4 genes and the conventional C4 allotyping method complemented each other. Lowered levels of plasma C3 or C4 or both, and serum CH50 were found in herpes and periodontitis patients. In rhinosinusitis, there was a linear trend with the highest levels found in the order: acute > chronic rhinosinusitis > general population > blood donors with no self-reported history of rhinosinusitis. Complement is involved in the defense against the tested mucosal infections. Seemingly immunocompetent patients with chronic or recurrent mucosal infections frequently have subtle weaknesses in different arms of immunity. Their susceptibility to chronic disease forms may be caused by these. Host s subtly impaired immunity often coincides with effective immune evasion from the same arms of immunity by the disease-causing pathogens. The interpretation of low subclass levels, if no additional predisposing immunologic factors are tested, is difficult and of limited value in early diagnosis and treatment.

Complement factor H dysfunction in atypical hemolytic uremic syndrome

Alternative pathway (AP) of complement can be activated on any surface, self or non-self. In atypical hemolytic uremic syndrome (aHUS) the AP regulation on self surfaces is insufficient and leads to complement attack against self-cells resulting usually in end-stage renal disease. Factor H (FH) is one of the key regulators of AP activation on the self surfaces. The domains 19 and 20 (FH19-20) are critical for the ability of FH to discriminate between C3b-opsonized self and non-self surfaces and are a hot-spot for mutations that have been described from aHUS patients. FH19-20 contains binding sites for both the C3d part of C3b and self surface polyanions that are needed for efficient C3b inactivation. To study the dysfunction of FH19-20, crystallographic structures of FH19-20 and FH19-20 in complex with C3d (FH19-20:C3d) were solved and aHUS-associated and structurally interesting point mutations were induced to FH19-20. Functional defects caused by these mutations were studied by analyzing binding of the FH19-20 mutant proteins to C3d, C3b, heparin, and mouse glomerular endothelial cells (mGEnCs). The results revealed two independent binding interfaces between FH19-20 and C3d - the FH19 site and the FH20 site. Superimposition of the FH19-20:C3d complex on the previously published C3b and FH1-4:C3b structures showed that the FH20 site on C3d is partially occluded, but the FH19 site is fully available. Furthermore, binding of FH19-20 via the FH19 site to C3b did not block binding of the functionally important FH1-4 domains and kept the FH20 site free to bind heparin or an additional C3d. Binding assays were used to show that FH20 domain can bind to heparin while FH19-20 is bound to C3b via the FH19 site, and that both the FH19 site and FH20 are necessary for recognition of non-activator surfaces. Simultaneous binding of FH19 site to C3b and FH20 to anionic self structures are the key interactions in self-surface recognition by FH and thereby enhanced avidity of FH explains how AP discriminates between self and non-self. The aHUS-associated mutations on FH19-20 were found to disrupt binding of the FH19 or FH20 site to C3d/C3b, or to disrupt binding of FH20 to heparin or mGEnC. Any of these dysfunctions leads to loss of FH avidity to C3b bearing self surfaces explaining the molecular pathogenesis of the aHUS-cases where mutations are found within FH19-20.

Enhanced surface plasmon resonance immunoassay for human complement factor 4

Using an enhanced surface plasmon resonance (SPR) immunosensor, we have determined the concentration of human complement factor 4 (C4). Antibody protein was concentrated into a carboxymethyldextran-modified gold surface by electrostatic attraction force and a simultaneous covalent immobilization of antibody based on amine coupling reaction took place. The sandwich method was applied to enhance the response signal and the specificity of antigen binding assay. The antibody immobilized surface had good response to C4 in the range of 0.02-20 mug/ml by this enhanced immunoassay. The regeneration effect by pH 2 glycine-HC1 buffer was also investigated. The same antibody immobilized surface could be used more than 80 cycles of C4 binding and regeneration. In addition, the ability to determinate C4 directly from serum sample without any purification was investigated. The sensitivity, specificity and reproducibility of the enhanced immunoassay are satisfactory. The results clearly demonstrate the advantages of the enhanced SPR technique for C4 immunoassay.

Deletion of complement factor H-related genes CFHR1 and CFHR3 is associated with atypical hemolytic uremic syndrome.

Atypical hemolytic uremic syndrome (aHUS) is associated with defective complement regulation. Disease-associated mutations have been described in the genes encoding the complement regulators complement factor H, membrane cofactor protein, factor B, and factor I. In this study, we show in two independent cohorts of aHUS patients that deletion of two closely related genes, complement factor H-related 1 (CFHR1) and complement factor H-related 3 (CFHR3), increases the risk of aHUS. Amplification analysis and sequencing of genomic DNA of three affected individuals revealed a chromosomal deletion of approximately 84 kb in the RCA gene cluster, resulting in loss of the genes coding for CFHR1 and CFHR3, but leaving the genomic structure of factor H intact. The CFHR1 and CFHR3 genes are flanked by long homologous repeats with long interspersed nuclear elements (retrotransposons) and we suggest that nonallelic homologous recombination between these repeats results in the loss of the two genes. Impaired protection of erythrocytes from complement activation is observed in the serum of aHUS patients deficient in CFHR1 and CFHR3, thus suggesting a regulatory role for CFHR1 and CFHR3 in complement activation. The identification of CFHR1/CFHR3 deficiency in aHUS patients may lead to the design of new diagnostic approaches, such as enhanced testing for these genes.

Genetic variants of complement factor H gene are not associated with premature coronary heart disease:A family-based study in the Irish population

Background: The complement factor H (CFH) gene has been recently confirmed to play an essential role in the development of age-related macular degeneration (AMD). There are conflicting reports of its role in coronary heart disease. This study was designed to investigate if, using a family-based approach, there was an association between genetic variants of the CFH gene and risk of early-onset coronary heart disease. Methods: We evaluated 6 SNPs and 5 common haplotypes in the CFH gene amongst 1494 individuals in 580 Irish families with at least one member prematurely affected with coronary heart disease. Genotypes were determined by multiplex SNaPshot technology. Results: Using the TDT/S-TDT test, we did not find an association between any of the individual SNPs or any of the 5 haplotypes and early-onset coronary heart disease. Conclusion: In this family-based study, we found no association between the CFH gene and early-onset coronary heart disease. © 2007 Meng et al; licensee BioMed Central Ltd.

An Ecological Correlation Study of Late Age-Related Macular Degeneration and the Complement Factor H Y402H Polymorphism

PURPOSE:
To investigate whether variation in the distribution of the risk allele frequency of the Y402H single-nucleotide polymorphism (SNP) across various ethnicities and geographic regions reflects differences in the prevalence of late age-related macular degeneration (AMD) in those ethnicities.

METHODS:
Published data were obtained via a systematic search. Study samples were grouped into clusters by ethnicity and geographic location and the Spearman correlation coefficient of the prevalence of late AMD and risk allele frequencies was calculated across clusters.

RESULTS:
Across all ethnicities, AMD prevalence was seen to increase with age. Populations of European descent had both higher risk allele frequencies and prevalence of late AMD than did Japanese, Chinese, and Hispanic descendants. Results for African descendants were anomalous: although allele frequency was similar to that in European populations, the age-specific prevalence of late AMD was considerably lower. The correlation coefficient for the association between allele frequency and AMD prevalence was 0.40 (95% confidence interval [CI] = -0.36 to 0.84, P = 0.28) in all populations combined and 0.71 (95% CI = 0.02-0.94, P = 0.04) when people of African descent were excluded.

CONCLUSIONS:
Evidence was found at the population level to support a positive association between the Y204H risk allele and the prevalence of AMD after exclusion of studies undertaken on persons of African ancestry. Data in African, Middle Eastern, and South American populations are needed to provide a better understanding of the association of late AMD genetic risk across ethnicities.

The association between complement component 2/complement factor B polymorphisms and age-related macular degeneration: A HuGE review and meta-analysis

The authors performed a systematic review of the association of complement component 2(C2)/complement factor B (CFB) gene polymorphisms with age-related macular degeneration (AMD). In total, data from 19 studies published between 2006 and 2011 were pooled for 4 polymorphisms: rs9332739 and rs547154 in the C2 gene and rs4151667 and rs641153 in the CFB gene. Data extraction and assessments for risk of bias were independently performed by 2 reviewers. Allele frequencies and allele and genotypic effects were pooled. Heterogeneity and publication bias were explored. Pooled minor allele frequencies for all 4 SNPs were between 4.7% and 9.6% for all polymorphisms, except for an Indian population in which the C allele at rs9332739 was the major allele. For the C2 polymorphisms, the minor C allele at rs9332739 and the minor T allele at rs547154 carried estimated relative risks (odds ratios) of 0.55 (95% confidence interval (CI): 0.46, 0.65) and 0.47 (95% CI: 0.39, 0.57), respectively. For the CFB polymorphisms, the minor A alleles at rs4151667 and rs614153 carried estimated risks of 0.54 (95% CI: 0.45, 0.64) and 0.41 (95% CI: 0.34, 0.51), respectively. These allele effects contributed to an absolute lowering of the risk of all AMD in Caucasian populations by 2.0%-6.0%. This meta-analysis provides a robust estimate of the protective association of C2/CFB with AMD.

Complement factor H is a serum-binding protein for adrenomedullin, and the resulting complex modulates the bioactivities of both partners

Adrenomedullin (AM) is an important regulatory peptide involved in both physiological and pathological states. We have previously demonstrated the existence of a specific AM-binding protein (AMBP-1) in human plasma. In the present study, we developed a nonradioactive ligand blotting assay, which, together with high pressure liquid chromatography/SDS-polyacrylamide gel electrophoresis purification techniques, allowed us to isolate AMBP-1 to homogeneity. The purified protein was identified as human complement factor H. We show that AM/factor H interaction interferes with the established methodology for quantification of circulating AM. Our data suggest that this routine procedure does not take into account the AM bound to its binding protein. In addition, we show that factor H affects AM in vitro functions. It enhances AM-mediated induction of cAMP in fibroblasts, augments the AM-mediated growth of a cancer cell line, and suppresses the bactericidal capability of AM on Escherichia coli. Reciprocally, AM influences the complement regulatory function of factor H by enhancing the cleavage of C3b via factor I. In summary, we report on a potentially new regulatory mechanism of AM biology, the influence of factor H on radioimmunoassay quantification of AM, and the possible involvement of AM as a regulator of the complement cascade.

Plasma Complement factor H in Alzheimer's Disease.

Biomarkers for Alzheimer's disease (AD) should meet several criteria, including simplicity of testing. Inappropriate activation of the complement cascade has been implicated in the pathogenesis of AD. Complement factor H (CFH) is a regulator of the cascade, but studies on plasma CFH levels in AD have provided mixed results. This study compared plasma CFH levels in 317 AD cases with 254 controls using an immunodiffusion assay. The sample had an 80% power to detect a difference of 23 mg/L between cases and controls, but no difference was evident. Plasma CFH may not be a suitable biomarker for AD.