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.

Sequence and expression of complement factor H gene cluster variants and their roles in age-related macular degeneration risk

Purpose: To investigate how potentially functional genetic variants are coinherited on each of four common complement factor H (CFH) and CFH-related gene haplotypes and to measure expression of these genes in eye and liver tissues.

Methods: We sequenced the CFH region in four individuals (one homozygote for each of four common CFH region haplotypes) to identify all genetic variants. We studied associations between the haplotypes and AMD phenotypes in 2157 cases and 1150 controls. We examined RNA-seq profiles in macular and peripheral retina and retinal pigment epithelium/choroid/sclera (RCS) from eight eye donors and three liver samples.

Results: The haplotypic coinheritance of potentially functional variants (including missense variants, novel splice sites, and the CFHR3–CFHR1 deletion) was described for the four common haplotypes. Expression of the short and long CFH transcripts differed markedly between the retina and liver. We found no expression of any of the five CFH-related genes in the retina or RCS, in contrast to the liver, which is the main source of the circulating proteins.

Conclusions: We identified all genetic variants on common CFH region haplotypes and described their coinheritance. Understanding their functional effects will be key to developing and stratifying AMD therapies. The small scale of our expression study prevented us from investigating the relationships between CFH region haplotypes and their expression, and it will take time and collaboration to develop epidemiologic-scale studies. However, the striking difference between systemic and ocular expression of complement regulators shown in this study suggests important implications for the development of intraocular and systemic treatments.

Association of complement factor H Y402H polymorphism and age-related macular degeneration in Brazilian patients

Purpose: The aim of this study was investigate the association between complement Factor H polymorphism (Y402H) and age-related macular degeneration (AMD) in Brazilian patients. Methods: Patients with AMD aged 50 or more and age-matched healthy controls were enrolled in the study. Genomic DNA was isolated from leucocytes of patients and controls; the Y402H polymorphism of complement Factor H gene (CFH) was determined by polymerase chain reaction directed sequencing. Results: The frequency of 1277C allele of Factor H was 56.30% in patients with AMD compared with 36.51% in controls (p-value = 0.001). The genotypic distribution differed significantly between the two groups (1277CC 36.98%, 1277CT 38.65% and 1277TT 24.37% for AMD group; 1277CC 13.16%, 1277CT 46.71% and 1277TT 40.13% for controls, p-value = 0.001). The odds ratio for patients with AMD carrying only one 1277C allele was 1.36 and for those carrying two 1277C alleles was 4.63, when compared to the control group. Conclusions: These results suggest the Y402H polymorphism of CFH is a risk factor to the development of AMD in Brazilian patients. This is in accordance with findings from the majority of previous study population in Europe and North American.

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.

Molecular cloning, promoter analysis and induced expression of the complement component C9 gene in the grass carp Ctenopharyngodon idella

Complement-mediated killing of pathogens through lytic pathway is an important effector mechanism of innate immune response. C9 is the ninth member of complement components, creating the membrane attack complex (MAC). In the present study, a putative cDNA sequence encoding the 650 amino acids of C9 and its genomic organization were identified in grass carp Ctenopharyngodon idella. The deduced amino acid sequence of grass carp C9 (gcC9) showed 48% and 38.5% identity to Japanese flounder and human C9, respectively. Domain search revealed that gcC9 contains a LDL receptor domain, an EGF precursor domain, a MACPF domain and two TSP domain located in the N-terminal and C-terminal, respectively. Phylogenetic analysis demonstrated that gcC9 is clustered in a same clade with Japanese flounder, pufferfish and rainbow trout C9. The gcC9 gene consists of 11 exons with 10 introns, spacing over approximately 7 kb of genomic sequence. Analysis of gcC9 promoter region revealed the presence of a TATA box and some putative transcription factor such as C/EBP, HSF, NF-AT, CHOP-C, HNF-3B, GATA-2, IK-2, EVI- 1, AP-1, CP2 and OCT-1 binding sites. The first intron region contains C/EBPb, HFH-1 and Oct-1 binding sites. RT-PCR and Western blotting analysis demonstrated that the mRNA and protein of gcC9 gene have similar expression patterns, being constitutively expressed in all organs examined of healthy fish, with the highest level in hepatopancreas. By real-time quantitative RT-PCR analysis, gcC9 transcripts were significantly up-regulated in head kidney, spleen, hepatopancreas and down-regulated in intestine from inactivated fish bacterial pathogen Flavobacterium columnare-stimulated fish, demonstrating the role of C9 in immune response. (c) 2007 Elsevier B.V. All rights reserved.

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.

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.

Cross-reactivity of antipneumococcal surface protein C (PspC) antibodies with different strains and evaluation of inhibition of human complement factor H and secretory IgA binding via PspC

Pneumococcal surface protein C (PspC) is an important candidate for a cost-effective vaccine with broad coverage against pneumococcal diseases. Previous studies have shown that Streptococcus pneumoniae is able to bind to both human factor H (FH), an inhibitor of complement alternative pathway, and human secretory IgA (sIgA) via PspC. PspC was classified into 11 groups based on variations of the gene. In this work, we used three PspC fragments from different groups (PspC3, PspC5, and PspC8) to immunize mice for the production of antibodies. Immunization with PspC3 induced antibodies that recognized the majority of the clinical isolates as analyzed by Western blotting of whole-cell extracts and flow cytometry of intact bacteria, while anti-PspC5 antibodies showed cross-reactivity with the paralogue pneumococcal surface protein A (PspA), and anti-PspC8 antibodies reacted only with the PspC8-expressing strain. Most of the isolates tested showed strong binding to FH and weaker interaction with sIgA. Preincubation with anti-PspC3 and anti-PspC5 IgG led to some inhibition of binding of FH, and preincubation with anti-PspC3 partially inhibited sIgA binding in Western blotting. The analysis of intact bacteria through flow cytometry showed only a small decrease in FH binding after incubation of strain D39 with anti-PspC3 IgG, and one clinical isolate showed inhibition of sIgA binding by anti-PspC3 IgG. We conclude that although anti-PspC3 antibodies were able to recognize PspC variants from the majority of the strains tested, partial inhibition of FH and sIgA binding through anti-PspC3 antibodies in vitro could be observed for only a restricted number of isolates.

Multifactor effects and evidence of potential interaction between complement factor H Y402H and LOC387715 A69S in age-related macular degeneration.

BACKGROUND: Variants in the complement cascade genes and the LOC387715/HTRA1, have been widely reported to associate with age-related macular degeneration (AMD), the most common cause of visual impairment in industrialized countries. METHODS/PRINCIPAL FINDINGS: We investigated the association between the LOC387715 A69S and complement component C3 R102G risk alleles in the Finnish case-control material and found a significant association with both variants (OR 2.98, p = 3.75 x 10(-9); non-AMD controls and OR 2.79, p = 2.78 x 10(-19), blood donor controls and OR 1.83, p = 0.008; non-AMD controls and OR 1.39, p = 0.039; blood donor controls), respectively. Previously, we have shown a strong association between complement factor H (CFH) Y402H and AMD in the Finnish population. A carrier of at least one risk allele in each of the three susceptibility loci (LOC387715, C3, CFH) had an 18-fold risk of AMD when compared to a non-carrier homozygote in all three loci. A tentative gene-gene interaction between the two major AMD-associated loci, LOC387715 and CFH, was found in this study using a multiplicative (logistic regression) model, a synergy index (departure-from-additivity model) and the mutual information method (MI), suggesting that a common causative pathway may exist for these genes. Smoking (ever vs. never) exerted an extra risk for AMD, but somewhat surprisingly, only in connection with other factors such as sex and the C3 genotype. Population attributable risks (PAR) for the CFH, LOC387715 and C3 variants were 58.2%, 51.4% and 5.8%, respectively, the summary PAR for the three variants being 65.4%. CONCLUSIONS/SIGNIFICANCE: Evidence for gene-gene interaction between two major AMD associated loci CFH and LOC387715 was obtained using three methods, logistic regression, a synergy index and the mutual information (MI) index.

The Regulation of AMD Pathobiology by Complement Factor H

Complement factor H (CFH) is a major susceptibility gene for age-related macular degeneration (AMD); however, its impact on AMD pathobiology is unresolved. Here, the role of CFH in the development of AMD pathology in vivo was interrogated by analyzing aged Cfh+/- and Cfh-/- mice fed a high fat, cholesterol-enriched diet. Strikingly, decreased levels of CFH led to increased sub-retinal pigmented epithelium (RPE) deposit formation, specifically basal laminar deposits, following high fat diet. Mechanistically, our data show that deposits are due to CFH competition for lipoprotein binding sites in Bruch’s membrane. Interestingly and despite sub-RPE deposit formation occurring in both Cfh+/- and Cfh-/- mice, RPE damage accompanied by loss of vision occurred only in old Cfh+/- mice. We demonstrate that such pathology is a function of excess complement activation and C5a production, associated with monocyte recruitment, in Cfh+/- mice versus complement deficiency in Cfh-/- animals. Due to the CFH dependent increase in sub-RPE deposit height we interrogated the potential of CFH as a novel regulator of Bruch’s membrane lipoprotein binding and show, using human Bruch’s membrane explants, that CFH removes endogenous human lipoproteins in aged donors. Interestingly, although the CFH H402 variant shows altered binding to BrM, this does not affect its ability to remove endogenous lipoproteins. This new understanding of the complicated interactions of CFH in AMD-like pathology provides an improved foundation for the development of targeted therapies for AMD.

Brain-derived neurotrophic factor (BDNF) gene : no major impact on antidepressant treatment response

The brain-derived neurotrophic factor (BDNF) has been suggested to play a pivotal role in the aetiology of affective disorders. In order to further clarify the impact of BDNF gene variation on major depression as well as antidepressant treatment response, association of three BDNF polymorphisms [rs7103411, Val66Met (rs6265) and rs7124442] with major depression and antidepressant treatment response was investigated in an overall sample of 268 German patients with major depression and 424 healthy controls. False discovery rate (FDR) was applied to control for multiple testing. Additionally, ten markers in BDNF were tested for association with citalopram outcome in the STAR*D sample. While BDNF was not associated with major depression as a categorical diagnosis, the BDNF rs7124442 TT genotype was significantly related to worse treatment outcome over 6 wk in major depression (p=0.01) particularly in anxious depression (p=0.003) in the German sample. However, BDNF rs7103411 and rs6265 similarly predicted worse treatment response over 6 wk in clinical subtypes of depression such as melancholic depression only (rs7103411: TThad main effects on antidepressant treatment response in ANOVA models when the remaining SNPs were considered as covariates. The STAR*D analyses did not yield significant results at any of the ten BDNF markers. Our results do not support an association between genetic variation in BDNF and antidepressant treatment response or remission. Post-hoc analyses provide some preliminary support for a potential minor role of genetic variation in BDNF and antidepressant treatment outcome in the context of melancholic depression.

Investigation of brain derived neurotrophic factor (BDNF) gene variants in migraine

A number of observations have suggested that brain derived neurotrophic factor (BDNF) plays a role in migraine pathophysiology. This study investigates whether variants in the BDNF gene are associated with migraine in an Australian case-control population. Background. Brain derived neurotrophic factor (BDNF) has an important role in neural growth, development and survival in the central nervous system and is an important modulator of central and peripheral pain responses. Variants in BDNF, in particular the functional Val66Met polymorphism (rs6265), have been found to be associated with a number of psychiatric disorders, cognitive function and obesity. As BDNF has been found to be differentially expressed in a number of aspects related to migraine, we tested for association between single nucleotide polymorphisms (SNPs) in BDNF and migraine. Methods. Five SNPs in the BDNF locus (rs1519480, rs6265, rs712507, rs2049046 and rs12273363) were genotyped initially in a cohort of 277 migraine cases, including 172 diagnosed with migraine with aura (MA) and 105 with migraine without aura (MO), and 277 age- and sex-matched controls. Three of these SNPs (rs6265, rs2049046 and rs12273363) were subsequently genotyped in a second cohort of 580 migraineurs, including 473 diagnosed with MA and 105 with O, and 580 matched controls. Results. – BDNF SNPs rs1519480, rs6265, rs712507 and rs12273363 were not significantly associated with migraine. However, rs2049046 showed a significant association with migraine, and in particular, MA in the first cohort. In the second cohort, although an increase in the rs2049046 T-allele frequency was observed in migraine cases, and in both MA and MO subgroups, it was not significantly different from controls. Analysis of data combined from both cohorts for rs2049046 showed significant differences in the genotypic and allelic distributions for this marker in both migraine and the MA sub-group. Conclusion. This study confirmed previous studies that the functional BDNF SNP rs6265 (Val66Met) is not associated with migraine. However, we found that rs2049046, which resides at the 5’ end of 3 one the BDNF transcripts, may be associated with migraine, suggesting that further investigations of this SNP may be warranted.

The effect of transforming growth factor β1 gene polymorphisms in ankylosing spondylitis

Objectives. To determine whether genetic polymorphisms in or near the transforming growth factor β1 (TGFB1) locus were associated d with susceptibility to or severity of ankylosing spondylitis (AS). Methods. Five intragenic single-nucleotide polymorphisms (SNP) and three microsatellite markers flanking the TGFB1 locus were genotyped. Seven hundred and sixty-two individuals from 184 multiplex families were genotyped for the microsatellite markers and two of the promoter SNPs. One thousand and two individuals from 212 English and 170 Finnish families with AS were genotyped for all five intragenic SNPs. A structured questionnaire was used to assess the age of symptom onset, disease duration and disease severity scores, including the BASDAI (Bath Ankylosing Spondylitis Disease Activity Index) and BASFI (Bath Ankylosing Spondylitis Functional Index). Results. A weak association was noted between the rare TGFB1 + 1632 T allele and AS in the Finnish population (P = 0.04) and in the combined data set (P = 0.03). No association was noted between any other SNPs or SNP haplotype and AS, even among those families with positive non-parametric linkage scores. The TGFB1 +1632 polymorphism was also associated with a younger age of symptom onset (English population, allele 2 associated with age of onset greater by 4.2 yr, P = 0.05; combined data set, allele 2 associated with age of onset greater by 3.2 yr, P = 0.02). A haplotype of coding region SNPs (TGFB1 +869/ +915+1632 alleles 2/1/2) was associated with age of symptom onset in both the English parent-case trios and the combined data set (English data set, haplotype 2/1/2 associated with age of onset greater by 4.9 yr, P = 0.03; combined data set, haplotype 2/1/2 associated with greater age of onset by 4.2 yr, P = 0.006). Weak linkage with AS susceptibility was noted and the peak LOD score was 1.3 at distance 2 cM centromeric to the TGFB1 gene. No other linkage or association was found between quantitative traits and the markers. Conclusion. This study suggests that the polymorphisms within the TGFB1 gene play at most a small role in AS and that other genes encoded on chromosome 19 are involved in susceptibility to the disease.

DNA structural variability as a factor in gene expression and evolution

Redundant DNA can buffer sequence dependent structural deviations from an ideal double helix. Buffering serves a mechanistic function by reducing extraneous conformational effects which could interfere with readout or which would impose energetic constraints on evolution. It also serves an evolutionary function by allowing for gradual variations in conformation-dependent regulation of gene expression. Such gradualism is critical for the rate of evolution. The buffer structure concept provides a new interpretation for repetitive DNA and for exons and introns.