Long-term health sequelae and quality of life at least 6 months after infection with SARS-CoV-2: design and rationale of the COVIDOM-study as part of the NAPKON population-based cohort platform (POP).

PURPOSE: Over the course of COVID-19 pandemic, evidence has accumulated that SARS-CoV-2 infections may affect multiple organs and have serious clinical sequelae, but on-site clinical examinations with non-hospitalized samples are rare. We, therefore, aimed to systematically assess the long-term health status of samples of hospitalized and non-hospitalized SARS-CoV-2 infected individuals from three regions in Germany. METHODS: The present paper describes the COVIDOM-study within the population-based cohort platform (POP) which has been established under the auspices of the NAPKON infrastructure (German National Pandemic Cohort Network) of the national Network University Medicine (NUM). Comprehensive health assessments among SARS-CoV-2 infected individuals are conducted at least 6 months after the acute infection at the study sites Kiel, Würzburg and Berlin. Potential participants were identified and contacted via the local public health authorities, irrespective of the severity of the initial infection. A harmonized examination protocol has been implemented, consisting of detailed assessments of medical history, physical examinations, and the collection of multiple biosamples (e.g., serum, plasma, saliva, urine) for future analyses. In addition, patient-reported perception of the impact of local pandemic-related measures and infection on quality-of-life are obtained. RESULTS: As of July 2021, in total 6813 individuals infected in 2020 have been invited into the COVIDOM-study. Of these, about 36% wished to participate and 1295 have already been examined at least once. CONCLUSION: NAPKON-POP COVIDOM-study complements other Long COVID studies assessing the long-term consequences of an infection with SARS-CoV-2 by providing detailed health data of population-based samples, including individuals with various degrees of disease severity. TRIAL REGISTRATION: Registered at the German registry for clinical studies (DRKS00023742).

Diagnosis of monogenic liver diseases in childhood by next-generation sequencing.

Next-generation sequencing (NGS) has opened up novel diagnostic opportunities for children with unidentified, but suspected inherited diseases. We describe our single-center experience with NGS diagnostics in standard clinical scenarios in pediatric hepatology. We investigated 135 children with suspected inherited hepatopathies, where initially no causative pathogenic variant had been identified, with an amplicon-based NGS panel of 21 genes associated with acute and chronic hepatopathies. In 23 of these patients, we detected pathogenic or likely pathogenic variants in 10 different genes. We present 6 novel variants. A total of 14 of these patients presented with the characteristic phenotype of the related hepatopathy. Nine patients showed only few or atypical clinical symptoms or presented with additional signs. In another 13 out of 135 cases, we detected variants of unknown significance (VUS) in 9 different genes. Only 2 of these patients showed characteristic phenotypes conclusive with the detected variants, whereas 11 patients showed unspecific or atypical phenotypes. Our multi-gene panel is a fast and comprehensive tool to diagnose inherited pediatric hepatopathies. We also illustrate the challenge of dealing with genetic variants and highlight arising clinical questions, especially in patients with atypical phenotypes.

Rare compound heterozygous variants in PNKP identified by whole exome sequencing in a German patient with ataxia-oculomotor apraxia 4 and pilocytic astrocytoma.

Ataxia-oculomotor apraxia type 4 (AOA4) is a rare autosomal recessive neurologic disorder. The phenotype is characterized by ataxia, oculomotor apraxia, peripheral neuropathy and dystonia. AOA4 is caused by biallelic pathogenic variants in the PNKP gene encoding a polynucleotide kinase 3'-phosphatase with an important function in DNA-damage repair. By whole exome sequencing, we identified 2 variants within the PNKP gene in a 27-year-old German woman with a clinical AOA phenotype combined with a cerebellar pilocytic astrocytoma diagnosed at 23 years of age. One variant, a duplication in exon 14 resulting in the frameshift c.1253_1269dup p.(Thr424fs*49), has previously been described as pathogenic, for example, in cases of AOA4. The second variant, representing a nonsense mutation in exon 17, c.1545C>G p.(Tyr515*), has not yet been described and is predicted to cause a loss of the 7 C-terminal amino acids. This is the first description of AOA4 in a patient with central European descent. Furthermore, the occurrence of a pilocytic astrocytoma has not been described before in an AOA4 patient. Our data demonstrate compound heterozygous PNKP germline variants in a German patient with AOA4 and provide evidence for a possible link with tumor predisposition. Localization of the 2 variants in human PNKP NP_009185.2. NM_007254.3:c.1253_1269dup p.(Thr424fs*49) is predicted to cause a frameshift within the kinase domain, NM_007254.3:c.1545C>G p.(Tyr515*) is predicted to cause loss of 2 C-terminal amino acids of the kinase domain and 5 additional C-terminal amino acids.

Synergistic Expression of Histone Deacetylase 9 and Matrix Metalloproteinase 12 in M4 Macrophages in Advanced Carotid Plaques.

OBJECTIVE/BACKGROUND: Expression patterns and association with cell specific gene expression signatures of the epigenetic regulator histone deacetylase 9 (HDAC9) and matrix metalloproteinase 12 (MMP12) in human plaque are not known. METHODS: This was a prospective cohort study. Genome wide expression analysis was performed in carotid, femoral, aortic plaques (n = 68) and left internal thoracic (LITA) controls (n = 28) and plaque histological severity assessed. Correlation and hierarchical cluster analysis was utilised. RESULTS: HDAC9 was associated with MMP12 expression in carotid plaques (r = .46, p = .012) and controls (r = -.44, p = .034). HDAC9 and MMP12 clustered with inflammatory macrophage markers but not with smooth muscle cell (SMC) rich markers. In plaques from all arterial sites, MMP12 but not HDAC9 showed positive correlation (p < .05) with M2 and M4 polarized macrophage markers, and negative correlation with SMC rich signatures. In the carotid plaques, all M4 macrophage markers associated with MMP12 and HDAC9. The negative association of MMP12 with SMC rich signatures was pronounced in the carotid plaques. Neither HDAC9 nor MMP12 associated consistently with plaque stabilisation or thrombosis related genes. Immunohistochemistry further supported the association between HDAC9 and MMP12 in atherosclerotic plaques. CONCLUSION: M4 macrophages are a possible source for HDAC9 and MMP12 expression in advanced human plaques.

Childhood asthma is associated with mutations and gene expression differences of ORMDL genes that can interact.

BACKGROUND: Genomewide association studies identified ORMDL3 as a plausible asthma candidate gene. ORMDL proteins regulate sphingolipid metabolism and ceramide homeostasis and participate in lymphocyte activation and eosinophil recruitment. Strong sequence homology between the three ORMDL genes and ORMDL protein conservation among different species suggest that they may have shared functions. We hypothesized that if single nucleotide polymorphisms (SNPs) in ORMDL3 alter its gene expression and play a role in asthma, variants in ORMDL1 and ORMDL2 might also be associated with asthma. METHODS: Asthma associations of 44 genotyped SNPs were determined in at least 1303 subjects (651 asthmatics). ORMDL expression was evaluated in peripheral blood mononuclear cells (PBMC) from 55 subjects (eight asthmatics) before and after allergen stimulation, and in blood (n = 60, 5 asthmatics). Allele-specific cis-effects on ORMDL expression were assessed. Interactions between human ORMDL proteins were determined in living cells. RESULTS: Sixteen SNPs in all three ORMDLs were associated with asthma (14 in ORMDL3). Baseline expression of ORMDL1 (P = 1.7 × 10(-6) ) and ORMDL2 (P = 4.9 × 10(-5) ) was significantly higher in PBMC from asthmatics, while induction of ORMDLs upon stimulation was stronger in nonasthmatics. Disease-associated alleles (rs8079416, rs4795405, rs3902920) alter ORMDL3 expression. ORMDL proteins formed homo- and heterooligomers and displayed similar patterns of interaction with SERCA2 and SPT1. CONCLUSIONS: Polymorphisms in ORMDL genes are associated with asthma. Asthmatics exhibit increased ORMDL levels, suggesting that ORMDLs contribute to asthma. Formation of heterooligomers and similar interaction patterns with proteins involved in calcium homeostasis and sphingolipid metabolism could indicate shared biological roles of ORMDLs, influencing airway remodeling and hyperresponsiveness.

Increased efficacy of omalizumab in atopic dermatitis patients with wild-type filaggrin status and higher serum levels of phosphatidylcholines.

Omalizumab, a monoclonal antibody targeting IgE, is an established therapy for severe allergic asthma and has shown efficacy in chronic spontaneous urticaria. Small-scale studies indicated some beneficial effect also in atopic dermatitis (AD). To evaluate the efficacy of omalizumab in AD and to identify markers associated with treatment response, we conducted a prospective 28-week open-label trial on 20 adults with moderate-to-severe AD. Our results confirm previous observations of a positive response in a subgroup of patients and suggest that responders are characterized by the absence of filaggrin mutations and altered lipid metabolite profiles with high levels of various glycerophospholipids.

Fine-mapping of IgE-associated loci 1q23, 5q31, and 12q13 using 1000 Genomes Project data.

BACKGROUND: Genome-wide association studies (GWAS) repeatedly identified 1q23 (FCER1A), 5q31 (RAD50-IL13 and IL4), and 12q13 (STAT6) as major susceptibility loci influencing the regulation of total serum IgE levels. As GWAS may be insufficient to capture causal variants, we performed fine-mapping and re-genotyping of the three loci using 1000 Genomes Project datasets. METHODS: Linkage disequilibrium tagging polymorphisms and polymorphisms of putative functional relevance were genotyped by chip technology (24 polymorphisms) or MALDI-TOF-MS (40 polymorphisms) in at least 1303 German children (651 asthmatics). The effect of polymorphisms on total serum IgE, IgE percentiles, and atopic diseases was assessed, and a risk score model was applied for gene-by-gene interaction analyses. Functional effects of putative causal variants from these three loci were studied in silico. RESULTS: Associations from GWAS were confirmed and extended. For 1q23 and 5q31, the majority of associations were found with mild to moderately elevated IgE levels, while in the 12q13 locus, single-nucleotide polymorphisms (SNPs) were associated with strongly elevated IgE levels. Gene-by-gene interaction analyses suggested that the presence of mutations in all three loci increases the risk for elevated IgE up to fourfold. CONCLUSION: This fine-mapping study confirmed previous associations and identified novel associations of SNPs in 1q23, 5q31, and 12q13 with different levels of serum IgE and their concomitant contribution to IgE regulation.

A common atopy-associated variant in the Th2 cytokine locus control region impacts transcriptional regulation and alters SMAD3 and SP1 binding.

BACKGROUND: Type 2 immune responses directed by Th2 cells and characterized by the signature cytokines IL4, IL5, and IL13 play major pathogenic roles in atopic diseases. Single nucleotide polymorphisms in the human Th2 cytokine locus in particular in a locus control region within the DNA repair gene RAD50, containing several RAD50 DNase1-hypersensitive sites (RHS), have been robustly associated with atopic traits in genome-wide association studies (GWAS). Functional variants in IL13 have been intensely studied, whereas no causative variants for the IL13-independent RAD50 signal have been identified yet. This study aimed to characterize the functional impact of the atopy-associated polymorphism rs2240032 located in the human RHS7 on cis-regulatory activity and differential binding of transcription factors. METHODS: Differential transcription factor binding was analyzed by electrophoretic mobility shift assays (EMSAs) with Jurkat T-cell nuclear extracts. Identification of differentially binding factors was performed using mass spectrometry (LC-MS/MS). Reporter vector constructs carrying either the major or minor allele of rs2240032 were tested for regulating transcriptional activity in Jurkat and HeLa cells. RESULTS: The variant rs2240032 impacts transcriptional activity and allele-specific binding of SMAD3, SP1, and additional putative protein complex partners. We further demonstrate that rs2240032 is located in an RHS7 subunit which itself encompasses repressor activity and might be important for the fine-tuning of transcription regulation within this region. CONCLUSION: The human RHS7 critically contributes to the regulation of gene transcription, and the common atopy-associated polymorphism rs2240032 impacts transcriptional activity and transcription factor binding.

Increased amino acids levels and the risk of developing of hypertriglyceridemia in a 7-year follow-up.

BACKGROUND: Recently, five branched-chain and aromatic amino acids were shown to be associated with the risk of developing type 2 diabetes (T2D). AIM: We set out to examine whether amino acids are also associated with the development of hypertriglyceridemia. MATERIALS AND METHODS: We determined the serum amino acids concentrations of 1,125 individuals of the KORA S4 baseline study, for which follow-up data were available also at the KORA F4 7 years later. After exclusion for hypertriglyceridemia (defined as having a fasting triglyceride level above 1.70 mmol/L) and diabetes at baseline, 755 subjects remained for analyses. RESULTS: Increased levels of leucine, arginine, valine, proline, phenylalanine, isoleucine and lysine were significantly associated with an increased risk of hypertriglyceridemia. These associations remained significant when restricting to those individuals who did not develop T2D in the 7-year follow-up. The increase per standard deviation of amino acid level was between 26 and 40 %. CONCLUSIONS: Seven amino acids were associated with an increased risk of developing hypertriglyceridemia after 7 years. Further studies are necessary to elucidate the complex role of these amino acids in the pathogenesis of metabolic disorders.

Genetic variation in the vaspin gene affects circulating serum vaspin concentrations.

OBJECTIVE: Visceral adipose tissue-derived serine protease inhibitor (vaspin) is an adipokine potentially linking obesity, insulin resistance and type 2 diabetes. Here, we searched for genetic determinants that could explain the variability in serum vaspin concentrations. RESEARCH DESIGN AND METHODS: First, we conducted a genome-wide association study (GWAS) for serum vaspin in the Sorbs cohort (N=826). Subsequently, 26 single-nucleotide polymorphisms (SNPs) covering genetic variation in the vaspin locus were genotyped in the Sorbs. In addition, we measured serum vaspin concentrations in 1806 samples from Augsburg/the Cooperative Health Research in the Region of Augsburg (KORA) for replication of the association signals. Finally, we conducted association analyses of vaspin SNPs with metabolic traits in the Sorbs (N=1013), KORA (N=1813) and a further cohort from Germany (Leipzig: N=1857). RESULTS: Six SNPs mapping between serpinA1 and serpinA4, including the vaspin locus, on chromosome 14 reached P-values < or = 10(-8) in the GWAS in the Sorbs. The fine mapping of variants within the vaspin locus in the Sorbs and subsequent replication in the KORA sample revealed several SNPs significantly associated with serum vaspin concentrations reaching P-values of up to 10(-35). However, no significant association with type 2 diabetes or related traits was found in either cohort after the Bonferroni correction for multiple comparisons. CONCLUSION: Our data show that the variability in serum vaspin concentrations might be explained by its genetic variants.

Polymorphisms in the IRF-4 gene, asthma and recurrent bronchitis in children.

BACKGROUND: Interferon-regulatory factors (IRFs) play a crucial role in immunity, not only influencing interferon expression but also T cell differentiation. IRF-4 was only recently recognized as a further major player in T cell differentiation. OBJECTIVE: As IRF-1 polymorphisms were shown to be associated with atopy and allergy, we comprehensively investigated effects of IRF-4 variants on allergy, asthma and related phenotypes in German children. METHODS: Fifteen tagging single nucleotide polymorphisms (SNPs) in the IRF-4 gene were genotyped by MALDI-TOF MS in the cross-sectional ISAAC phase II study population from Munich and Dresden (age 9-11; N = 3099). Replication was performed in our previously established genome-wide association study (GWAS) data set (N = 1303) consisting of asthma cases from the Multicenter Asthma Genetic in Childhood (MAGIC) study and reference children from the ISAAC II study. RESULTS: SNPs were not significantly associated with asthma but with bronchial hyperresponsiveness, atopy and, most interestingly, with recurrent bronchitis in the first data set. The IRF-4 variant rs9378805 was associated with recurrent bronchitis in the ISAAC population and replicated in the GWAS data set where further SNPs showed associations with recurrent bronchitis and asthma. CONCLUSIONS: We found genetic associations in IRF-4 to be associated with recurrent bronchitis in our two study populations. Associated polymorphisms are localized in a putative regulatory element in the 3'UTR region of IRF-4. These findings suggest a putative role of IRF-4 in the development of bronchitis.

Integrative genetic and metabolite profiling analysis suggests altered phosphatidylcholine metabolism in asthma.

BACKGROUND: Genome-wide association studies (GWAS) have identified many risk loci for asthma, but effect sizes are small, and in most cases, the biological mechanisms are unclear. Targeted metabolite quantification that provides information about a whole range of pathways of intermediary metabolism can help to identify biomarkers and investigate disease mechanisms. Combining genetic and metabolic information can aid in characterizing genetic association signals with high resolution. This work aimed to investigate the interrelation of current asthma, candidate asthma risk alleles and a panel of metabolites. METHODS: We investigated 151 metabolites, quantified by targeted mass spectrometry, in fasting serum of asthmatic and nonasthmatic individuals from the population-based KORA F4 study (N = 2925). In addition, we analysed effects of single-nucleotide polymorphisms (SNPs) at 24 asthma risk loci on these metabolites. RESULTS: Increased levels of various phosphatidylcholines and decreased levels of various lyso-phosphatidylcholines were associated with asthma. Likewise, asthma risk alleles from the PDED3 and MED24 genes at the asthma susceptibility locus 17q21 were associated with increased concentrations of various phosphatidylcholines with consistent effect directions. CONCLUSIONS: Our study demonstrated the potential of metabolomics to infer asthma-related biomarkers by the identification of potentially deregulated phospholipids that associate with asthma and asthma risk alleles.

Association of single nucleotide polymorphisms in the diamine oxidase gene with diamine oxidase serum activities.

BACKGROUND: Histamine intolerance (HIT) is associated with an excess of histamine because of an impaired function of the histamine-degrading enzyme diamine oxidase (DAO). The genetic background of HIT is unknown yet. METHODS: Case-control association study of all haplotype tagging and four previously reported DAO SNPs and one HNMT Single nucleotide polymorphism with symptoms of HIT and DAO serum activity in 484 German individuals including 285 patients with clinical symptoms of HIT and 199 controls. RESULTS: Diamine oxidase serum activity was significantly associated with seven SNPs within the DAO gene. The minor allele at rs2052129, rs2268999, rs10156191 and rs1049742 increased the risk for a reduced DAO activity whereas showing a moderate protective effect at rs2071514, rs1049748 and rs2071517 in the genotypic (P = 2.1 × 10(-8) , 7.6 × 10(-10) , 8.3 × 10(-10) , 0.009, 0.005, 0.00001, 0.006, respectively) and allelic genetic model (P = 2.5 × 10(-11) , 5.4 × 10(-13) , 8.9 × 10(-13) , 0.00002, 0.006, 0.0003, 0.005, respectively). Reporter gene assays at rs2052129 revealed a lower promoter activity (P = 0.016) of the minor allele. DAO mRNA expression in peripheral blood mononuclear cells of homozygous carriers of the minor allele at rs2052129, rs2268999, rs10156191 was lower (P = 0.002) than homozygous carriers of the major allele. Diamine oxidase variants were not associated with the HIT phenotype per se, only with DAO activity alone and the subgroup of HIT patients displaying a reduced DAO activity. CONCLUSIONS: DAO gene variants strongly influence DAO expression and activity but alone are not sufficient to fully effectuate the potentially associated disease state of HIT, suggesting an interplay of genetic and environmental factors.

A common FADS2 promoter polymorphism increases promoter activity and facilitates binding of transcription factor ELK1.

Fatty acid desaturases (FADS) play an important role in the formation of omega-6 and omega-3 highly unsaturated fatty acids (HUFAs). The composition of HUFAs in the human metabolome is important for membrane fluidity and for the modulation of essential physiological functions such as inflammation processes and brain development. Several recent studies reported significant associations of single nucleotide polymorphisms (SNPs) in the human FADS gene cluster with HUFA levels and composition. The presence of the minor allele correlated with a decrease of desaturase reaction products and an accumulation of substrates. We performed functional studies with two of the associated polymorphisms (rs3834458 and rs968567) and showed an influence of polymorphism rs968567 on FADS2 promoter activity by luciferase reporter gene assays. Electrophoretic mobility shift assays proved allele-dependent DNA-binding ability of at least two protein complexes to the region containing SNP rs968567. One of the proteins binding to this region in an allele-specific manner was shown to be the transcription factor ELK1 (a member of ETS domain transcription factor family). These results indicate that rs968567 influences FADS2 transcription and offer first insights into the modulation of complex regulation mechanisms of FADS2 gene transcription by SNPs.

A unique genetic defect on chromosome 3 is responsible for juvenile obesity in the Berlin Fat Mouse.

OBJECTIVE: This study aimed at the mapping and estimation of genetic and sex effects contributing to the obese phenotype of the Berlin Fat Mouse Inbred line 860 (BFMI860). This mouse line is predisposed for juvenile obesity. BFMI860 mice accumulate 24% total fat mass at 10 weeks of age under a standard maintenance diet. DESIGN: A total of 471 mice of a (BFMI860 x C57BL/6NCrl) F2 intercross population were fed a standard maintenance diet and were analysed for body composition at 10 weeks when they finished their rapid growth phase. RESULTS: The most striking result was the identification of a novel obesity locus on chromosome 3 (Chr 3) at 40 Mb, explaining 39% of the variance of total fat mass in the F2 population under a standard diet. This locus was named jObes1 (juvenile obesity 1). The BFMI860 allele effect was recessive. Males and females homozygous at jObes1 had on average 3.0 and 3.3 g more total fat mass at 10 weeks than the other two genotype classes, respectively. The effect was evident in all white adipose tissues, brown adipose tissue and also in liver. The position of the Chr 3 effect is syntenic to an obesity locus in humans. Additional loci for total fat mass and different white adipose tissue weights with minor effects were detected on mouse Chr 5 and 6. Another locus on Chr 4 had influence especially on liver weight. Many loci including jObes1 affected males and females to a different extent. CONCLUSION: The major locus on Chr 3 for juvenile obesity and its interaction with sex is unique and makes the BFMI860 mice an interesting resource for the discovery of novel genetic factors predisposing obesity, which might also contribute to obesity in humans. The results suggested that metabolic and regulatory pathways differed between the sexes.

Genes and lifestyle factors in obesity: results from 12,462 subjects from MONICA/KORA.

BACKGROUND: Data from meta-analyses of genome-wide association studies provided evidence for an association of polymorphisms with body mass index (BMI), and gene expression results indicated a role of these variants in the hypothalamus. It was consecutively hypothesized that these associations might be evoked by a modulation of nutritional intake or energy expenditure. OBJECTIVE: It was our aim to investigate the association of these genetic factors with BMI in a large homogenous population-based sample to explore the association of these polymorphisms with lifestyle factors related to nutritional intake or energy expenditure, and whether such lifestyle factors could be mediators of the detected single-nucleotide polymorphism (SNP)-association with BMI. It was a further aim to compare the proportion of BMI explained by genetic factors with the one explained by lifestyle factors. DESIGN: The association of seven polymorphisms in or near the genes NEGR1, TMEM18, MTCH2, FTO, MC4R, SH2B1 and KCTD15 was analyzed in 12,462 subjects from the population-based MONICA/KORA Augsburg study. Information on lifestyle factors was based on standardized questionnaires. For statistical analysis, regression-based models were used. RESULTS: The minor allele of polymorphism rs6548238 C>T (TMEM18) was associated with lower BMI (-0.418 kg m(-2), P=1.22 × 10(-8)), and of polymorphisms rs9935401 G>A (FTO) and rs7498665 A>G (SH2B1) with increased BMI (0.290 kg m(-2), P=2.85 × 10(-7) and 0.145 kg m(-2), P=9.83 × 10(-3)). The other polymorphisms were not significantly associated. Lifestyle factors were correlated with BMI and explained 0.037% of the BMI variance as compared with 0.006% of explained variance by the associated genetic factors. The genetic variants associated with BMI were not significantly associated with lifestyle factors and there was no evidence of lifestyle factors mediating the SNP-BMI association. CONCLUSIONS: Our data first confirm the findings for TMEM18 with BMI in a single study on adults and also confirm the findings for FTO and SH2B1. There was no evidence for a direct SNP-lifestyle association.

Analysis of the high affinity IgE receptor genes reveals epistatic effects of FCER1A variants on eczema risk.

BACKGROUND: High levels of total and allergen-specific IgE levels are a key feature in allergic diseases. The high-affinity receptor for IgE, which is composed of one alpha (FCER1A), one beta (FCER1B), and two gamma (FCER1G) subunits, represents the central receptor of IgE-induced reactions. In a genome-wide association scan, we recently identified associations between functional FCER1A variants and total serum IgE levels. Previous studies had reported linkage and association of FCER1B variants with IgE and atopic traits. The FCER1G gene has not yet been investigated with regard to atopy. Filaggrin (FLG) is the strongest known risk gene for eczema, in particular the allergic subtype of eczema. METHODS: We investigated the association of FCER1A, FCER1B, and FCER1G variants with IgE in a large population-based cohort (n = 4261) and tested for epistatic effects using the model-based multifactor dimensionality reduction (MB-MDR) method. In addition, we investigated a potential interaction between FLG and FCER1A variants in a large collection of eczema cases (n = 1018) and population controls. RESULTS: Three strongly correlated FCER1A polymorphisms were significantly associated with total and specific IgE levels as well as allergic sensitization. No associations were seen for FCER1B and FCER1G. After adjustment for FLG effects, a significant epistatic effect of the FCER1A variants rs10489854 and rs2511211 on eczema risk was detected. CONCLUSIONS: These results suggest that FCER1A variants by themselves and in combination influence IgE levels and act synergistically to influence eczema risk.

Association of a variant in the muscarinic acetylcholine receptor 2 gene (CHRM2) with nicotine addiction.

Genetic factors contribute to the overall risk of developing nicotine addiction, which is the major cause of preventable deaths in western countries. However, knowledge regarding specific polymorphisms influencing smoking phenotypes remains scarce. In the present study we provide evidence that a common single nucleotide polymorphism (SNP) in the 5' untranslated region of CHRM2, the gene coding for the muscarinic acetylcholine receptor 2 is associated with nicotine addiction. CHRM2 was defined as a candidate gene for nicotine addiction based on previous evidence that linked variations in CHRM2 to alcohol and drug dependence. A total of more than 5,500 subjects representative of the German population were genotyped and assessed regarding their smoking habits. The impact of three SNPs in CHRM2 on smoking behavior/nicotine addiction was investigated using logistic regression models or a quasi-Poisson regression model, respectively. We found the T allele of SNP rs324650 to be associated with an increased risk of smoking/nicotine dependence according to three different models, the recessive models of regular or heavy smokers vs. never-smokers (odds ratio 1.17 in both analyses) and according to the Fagerström index of nicotine addiction. In the analysis stratified by gender this association was only found in females. Our data provide further evidence that variations in CHRM2 may be associated with the genetic risk of addiction in general or with certain personality traits that predispose to the development of addiction. Alternatively, variations in CHRM2 could modulate presynaptic auto-regulation in cholinergic systems and may thereby affect an individual's response to nicotine more specifically.

Autoantibodies to zinc transporter 8 and SLC30A8 genotype stratify type 1 diabetes risk.

AIMS/HYPOTHESIS: Our aim was to determine the relationships between autoantibodies to zinc transporter 8 (ZnT8), genotypes of the ZnT8-encoding gene SLC30A8 and type 1 diabetes risk. METHODS: ZnT8 autoantibodies (ZnT8A) were measured in sera of 1,633 children with a first-degree family history of type 1 diabetes and who were prospectively followed from birth. Antibodies were measured by Protein A-based radiobinding assays and COOH-terminal (R325, W325 or Q325 variants) or NH(2)-terminal constructs of human ZnT8. SLC30A8 genotyping at single-nucleotide polymorphism (SNP) rs13266634 was performed on 1,170 children. RESULTS: Antibodies against COOH-terminal ZnT8 constructs (ZnT8A-COOH) developed in 58 children as early as 9 months of age (median 3 years). They were detected in 55 of 128 (43%) children with autoantibodies to insulin, GAD and/or insulinoma-associated protein 2 and 34 of 42 (81%) who progressed to diabetes. The additional presence of ZnT8A-COOH stratified diabetes risk in islet autoantibody-positive children (p < 0.0001). SLC30A8 genotype strongly influenced ZnT8A type and diabetes risk in ZnT8A-COOH-positive children. Antibody binding against the ZnT8 R325 variant was strictly correlated with the number of the corresponding SLC30A8 R325-encoding alleles, whereas binding against the W325 variant was highest in children who had SLC30A8 W325-encoding alleles (p = 0.001). Moreover, ZnT8A-COOH-positive children who carried homozygous SLC30A8 SNP rs13266634 genotypes progressed faster to diabetes than those who were heterozygous (59% [95% CI 42.3-75.7%] vs 22% [95% CI 0-44.3%] within 5 years; p = 0.01). CONCLUSIONS/INTERPRETATION: Autoimmunity against the COOH-terminal region of ZnT8 is a highly relevant prognostic feature in childhood type 1 diabetes. Risk stratification in ZnT8A-COOH-positive children is further improved by SLC30A8 genotyping.

Association of nicotinic acetylcholine receptor subunit alpha 4 polymorphisms with nicotine dependence in 5500 Germans.

Polymorphisms in the CHRNA4 gene coding the nicotinic acetylcholine receptor subunit alpha 4 have recently been suggested to play a role in the determination of smoking-related phenotypes. To examine this hypothesis, we conducted a genetic association study in three large samples from the German general population (N(1)=1412; N(2)=1855; N(3)=2294). Five single-nucleotide polymorphisms in CHRNA4 were genotyped in 5561 participants, including 2707 heavily smoking cases (regularly smoking at least 20 cigarettes per day) and 2399 never-smoking controls (