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Recent studies indicate a mutual epidemiological relationship between coronary heart disease (CHD) and periodontitis. Both diseases are associated with similar risk factors and are characterized by a chronic inflammatory process. In a candidate-gene association study, we identify an association of a genetic susceptibility locus shared by both diseases. We confirm the known association of two neighboring linkage disequilibrium regions on human chromosome 9p21.3 with CHD and show the additional strong association of these loci with the risk of aggressive periodontitis. For the lead SNP of the main associated linkage disequilibrium region, rs1333048, the odds ratio of the autosomal-recessive mode of inheritance is 1.99 (95% confidence interval 1.33–2.94; P = 6.9×10−4) for generalized aggressive periodontitis, and 1.72 (1.06–2.76; P = 2.6×10−2) for localized aggressive periodontitis. The two associated linkage disequilibrium regions map to the sequence of the large antisense noncoding RNA ANRIL, which partly overlaps regulatory and coding sequences of CDKN2A/CDKN2B. A closely located diabetes-associated variant was independent of the CHD and periodontitis risk haplotypes. Our study demonstrates that CHD and periodontitis are genetically related by at least one susceptibility locus, which is possibly involved in ANRIL activity and independent of diabetes associated risk variants within this region. Elucidation of the interplay of ANRIL transcript variants and their involvement in increased susceptibility to the interactive diseases CHD and periodontitis promises new insight into the underlying shared pathogenic mechanisms of these complex common diseases., Author Summary Coronary heart disease (CHD) and periodontitis are the most widespread diseases in the Western industrialized world and pose a substantial health threat to populations worldwide. CHD is a leading cause for premature death, and periodontitis is the major cause for tooth loss in adults over 40 years. Both diseases are associated with similar risk factors such as smoking, diabetes, and gender, and both diseases are further characterized by a chronic inflammatory process. In the last year, several genome studies have identified a region of the human genome near the CDKN2A and CDKN2B genes as having an influence on CHD. We show that this genetic region, being the most important susceptibility locus for CHD to date, is also associated with a substantial risk increase of aggressive periodontitis. The associated genetic region maps to a genomic region that codes for an “antisense RNA,” which partly overlaps regulatory and coding sequences of genes CDKN2A/CDKN2B. The interplay between these common inflammatory complex diseases could be partially due to the shared genetic risk variants of this antisense RNA.

Genome-wide association studies have contributed significantly to the genetic dissection of complex diseases. In order to increase the power of existing marker sets even further, methods have been proposed to predict individual genotypes at un-typed loci from other marker sets by imputation, usually employing HapMap data as a reference. Although various imputation algorithms have been used in practice already, a comprehensive evaluation and comparison of these approaches, using genome-wide SNP data from one and the same population is still lacking. We therefore investigated four publicly available programs for genotype imputation (BEAGLE, IMPUTE, MACH, and PLINK) using data from 449 German individuals genotyped in our laboratory for three genome-wide SNP sets [Affymetrix 5.0 (500 k), Affymetrix 6.0 (1,000 k), and Illumina 550 k]. We observed that HapMap-based imputation in a northern European population is powerful and reliable, even in highly variable genomic regions such as the extended MHC on chromosome 6p21. However, while genotype predictions were found to be highly accurate with all four programs, the number of SNPs for which imputation was actually carried out ('imputation efficacy') varied substantially. BEAGLE, IMPUTE, and MACH yielded nearly identical trade-offs between imputation accuracy and efficacy whereas PLINK performed consistently poorer. We nevertheless recommend either MACH or BEAGLE for practical use because these two programs are more user-friendly and generally require less memory than IMPUTE.

Polymorphisms in the interleukin-1 (IL1) gene have been suggested to influence transcription of IL1A (interleukin-1α) and IL1B (interleukin-1β) and thereby the pathophysiology of periodontitis. This case-control association study on 415 northern European Caucasian patients with aggressive periodontitis (AgP) and 874 healthy controls was conducted to examine 10 single-nucleotide polymorphisms (SNPs) in the genes of the IL1 cluster for association with IL1A, IL1B, CKAP2L (cytoskeleton-associated protein 2-like), and IL1RN (IL-1 receptor antagonist). The results do not support an association between variants in the IL1 gene cluster and AgP. This case-control study had at least 95% power to detect genuine associations with variants carrying relative risks of at least 1.5 for heterozygous carriers and 2.25 for homozygous carriers. Previous reports of an association between IL1 promoter SNPs and periodontitis might reflect subpopulation effects and have to be interpreted with care.

Inflammatory bowel disease (IBD) typically manifests as either ulcerative colitis (UC) or Crohn's disease (CD). Systematic identification of susceptibility genes for IBD has thus far focused mainly on CD, and little is known about the genetic architecture of UC. Here we report a genome-wide association study with 440,794 SNPs genotyped in 1,167 individuals with UC and 777 healthy controls. Twenty of the most significantly associated SNPs were tested for replication in three independent European case-control panels comprising a total of 1,855 individuals with UC and 3,091 controls. Among the four consistently replicated markers, SNP rs3024505 immediately flanking the IL10 (interleukin 10) gene on chromosome 1q32.1 showed the most significant association in the combined verification samples (P = 1.35 x 10(-12); OR = 1.46 (1.31-1.62)). The other markers were located in ARPC2 and in the HLA-BTNL2 region. Association between rs3024505 and CD (1,848 cases, 1,804 controls) was weak (P = 0.013; OR = 1.17 (1.01-1.34)). IL10 is an immunosuppressive cytokine that has long been proposed to influence IBD pathophysiology. Our findings strongly suggest that defective IL10 function is central to the pathogenesis of the UC subtype of IBD.

Background & Aims: Crohn's disease (CD) and sarcoidosis (SA) are chronic inflammatory barrier diseases that share several clinical and immunological features, including the occurrence of granulomas. Methods: A 100k genome-wide association study with 83,360 single-nucleotide polymorphisms (SNPs) was performed on 382 CD patients, 398 SA patients, and 394 control individuals. The 24 SNPs that were most strongly associated in the combined CD/SA phenotype were selected for verification in an independent sample of 1,317 patients (660 CD and 657 SA) and 1,091 controls. Results: The most significant association (Bonferroni corrected P = .036) was obtained at SNP rs1398024 on chromosome 10p12.2, with an odds ratio (OR) for both diseases of 0.81 (95% confidence interval [CI], 0.69–0.96) for carriership of the rarer allele A. The P value in the overall combined sample was 4.24 × 10−6. During further follow-up, a moderate association (OR, 0.83; 95% CI, 0.72–0.96; P = .015) was observed between rs1398024 and ulcerative colitis (1,080 patients vs 1,091 controls), the second main subphenotype of inflammatory bowel disease in addition to CD. Extensive fine mapping of the 10p12.2 locus points to yet unidentified variants in the C10ORF67 gene region as the most likely underlying risk factors. Conclusion: Our study demonstrates that the combined analysis of different, albeit clinically related, phenotypes can lead to the identification of common susceptibility loci.

Stefan Schreiber and colleagues report the results of a genome-wide association study for sarcoidosis, a complex chronic inflammatory disorder. Variants near ANXA11 and PLAC9 are associated with elevated risk of the disease, with ANXA11 as the stronger candidate. Sarcoidosis is a complex chronic inflammatory disorder with predominant manifestation in the lung. In the first genome-wide association study (>440,000 SNPs) of this disease, comprising 499 German individuals with sarcoidosis and 490 controls, we detected a series of genetic associations. The strongest association signal maps to the ANXA11 (annexin A11) gene on chromosome 10q22.3. Validation in an independent sample (1,649 cases, 1,832 controls) confirmed the association (SNP rs2789679: P = 3.0 × 10−13, rs7091565: P = 1.0 × 10−5, allele-based test). Extensive fine mapping located the association signal to a region between exon 5 and exon 14 of ANXA11. A common nonsynonymous SNP (rs1049550, C > T, R230C) was found to be strongly associated with sarcoidosis. The GWAS lead SNP and additional risk variants in the region (rs1953600, rs2573346, rs2784773) were in strong linkage disequilibrium with rs1049550. Annexin A11 has complex and essential functions in several biological pathways, including apoptosis and proliferation.

Background: A genomic region on chromosome 6p21.3, including HLA-DPB1, has been linked to Wegener’s granulomatosis (WG). The basis of this association is difficult to evaluate because of the complex haplotype block architecture of this region. Objective: To identify the causative molecular genetic variation(s) using a detailed HapMap based fine-mapping approach. Methods: 282 patients with WG and 380 healthy controls were genotyped for HLA-DPB1 as well as for 35 informative single nucleotide polymorphisms (SNPs) within the respective region. 25 of these SNPs have been selected as tagging SNPs for another 219 associated SNPs. Allele and genotype frequencies were analysed separately by contingency tables and logistic regression. Finally, the coding region of RING1 was directly sequenced in subjects who carried haplotypes that were correlated with contrasting WG risks. Results: The previously reported strong association of WG with the HLA-DPB1*0401 allele was confirmed in an independent WG sample (n = 108, pc = 6.4610 28 ). When the complete cohort (n = 282) was considered, the association remained highly significant in ANCA-positive (pc = 1.26610 222 ), but not in ANCA-negative patients. An SNP 39 of HLA-DPB1 yielded the smallest p value and was associated with WG partly independently from the HLA-DPB1 alleles. Another informative SNP in the vicinity of RING1 showed significant WG association that was also partly independent of HLA-DPB1. RING1 sequencing, however, did not show any variation potentially predisposing to WG. Conclusions: The HLA-DPB1/RING1 region is strongly associated with WG in ANCA-positive subjects. Further analyses of potential cis regulatory sequences of candidate genes HLA-DPB1, RING1 and RXRB appear warranted.

Atopic dermatitis (AD) is a chronic skin disease affecting up to 15% of children in industrialized countries. AD belongs to the group of atopic disorders characterized by excessive immune reactions to ubiquitous antigens. Complex interactions between genetic and environmental factors have been suggested for atopic disorders. Dysregulation of the innate immune system appears crucial for the pathogenesis of AD. The NACHT-LRRs (NLRs) represent a group of innate immune receptors with special relevance for inflammatory processes. In order to investigate the role of variation in NLR genes for AD, we genotyped 23 single nucleotide polymorphisms (SNPs) in seven selected NLR genes (CARD4, CARD15, CARD12, NALP1, NALP3, NALP12, MHC2TA) in 392 AD patients and 297 controls by restriction enzyme digestion or TaqMan assays. Single-SNP analysis demonstrated significant associations of the CARD15_R702W variation and the NALP12_In9 T-allele with AD (P = 0.008 and P = 0.03, resp.; insignificant after Bonferroni correction). In the CARD4 gene, a rare haplotype was more frequent in AD patients than in controls. Interactions between all pairs of SNPs in the seven genes were analysed by logistic regression. Significant interactions comprised SNPs in the CARD4 gene (CARD4_In1 and CARD4_Ex6, P = 6.56 x 10(-7); CARD4_Prom und CARD4_Ex6, P = 2.45 x 10(-4)) and promoter polymorphisms in the CARD12 and NALP1 genes (P = 4.31 x 10(-4)). In conclusion, variation in individual genes from the NLR family as well as interactions within this group of innate immune receptor genes could play a role in AD pathogenesis. Investigations in other populations and functional studies are warranted to clarify contributions of NLR variation for this frequent skin disease.

Adiponectin is discussed to regulate energy balance and insulin sensitivity. Several studies indicated an association of fasting adiponectin with parameters of the metabolic syndrome. We investigated postprandial adiponectin release and its relation to traits of the metabolic syndrome. Serum adiponectin concentration after an oral glucose tolerance test and after ingestion of a standardised mixed, fat-containing meal in 110 male non-diabetic subjects was assessed. Fasting and postprandial adiponectin and the decrease of adiponectin were correlated with anthropometric and metabolic parameters. Subjects were genotyped for adiponectin − 11 388 G/A promoter single nucleotide polymorphism. Adiponectin slightly decreased after both test meals. A significant decrease was attained 5 and 6 h after the lipid load and 2 h after the glucose load. Particularly, the mixed meal postprandial adiponectin showed stronger correlations with most traits of the metabolic syndrome than fasting adiponectin: postprandial adiponectin with HDL (r 0·30) v. fasting adiponectin with HDL (r 0·23); with postprandial insulin (area under the curve): r − 0·20 v. r − 0·16; with fasting insulin: r 0·10 v. r 0·14; with BMI: r − 0·23 v. r − 0·20; with waist: r − 0·18 v. − 0·16; with systolic blood pressure: r − 0·14 v. r − 0·12; with diastolic blood pressure: r − 0·18 v. r − 0·15. In multivariate analysis, postprandial TAG were the only independent predictor of adiponectin. There was no significant association of adiponectin, NEFA and TAG with − 11 388 G/A adiponectin promoter polymorphism. Our findings favour the interpretation that postprandial adiponectin has the strongest and independent associations to postprandial TAG metabolism.

Zusammenfassung Im Gegensatz zu monogenen Erkrankungen kann bei multifaktoriellen genetisch bedingten Krankheiten aufgrund der Vielzahl genetischer und nichtgenetischer Faktoren keine klare Genotyp-Phänotyp-Beziehung erwartet werden. Mögliche Störgrößen sind u. a. das Vorliegen genetischer Heterogenität, Gen-Gen- und Gen-Umwelt-Interaktion sowie der Einfluss epigenetischer Faktoren. In der Konsequenz verringern sich die relativen Risiken einzelner genetischer Einflussfaktoren, was größere Stichprobenumfänge für ihre initiale Identifikation erforderlich macht. Aus der Prävalenz einer Erkrankung allein können meist keine Schlüsse hinsichtlich der Häufigkeit und Diversität der sie verursachenden genetischen Variation gezogen werden. Die Gewährleistung eines möglichst homogenen genetischen Hintergrunds der Studienpopulation, die Fokussierung auf möglichst einfache, klar definierte Phänotypen und die Untersuchung möglichst plausibler, genetischer und nichtgenetischer Einflussfaktoren stellen vermutlich den effizientesten Weg zur Identifikation von „Krankheitsgenen“ dar. Die Replikation von Assoziationsbefunden, insbesondere mit seltenen genetischen Varianten, hat zudem in solchen Populationen die größten Erfolgsaussichten, die der ursprünglich analysierten Gruppe genetisch möglichst ähnlich sind.

Atopic dermatitis (AD) is a chronic inflammatory skin disease affecting up to 16% of children in developed countries. A complex genetic background for AD has been suggested, with genetic as well as environmental factors influencing disease susceptibility. Among other factors, dysregulation in both the innate and the adaptive immune system has been proposed to play a role in AD pathophysiology. We present here an extended association screen for AD using microsatellite markers in 154 genes related to innate and adaptive immunity in pooled DNA samples from 150 German children with AD and 100 controls. After Bonferroni correction, no marker revealed a significant association with AD. Yet, markers representing the nuclear factor kappa B (NFKB)1 and chemokine receptor (CCR)4 genes showed differences in allelic distributions between cases and controls for both pooled DNA analysis and individual genotyping and were thus further investigated. Evaluation of additional single nucleotide polymorphisms (SNP) in the NFKB1 and CCR4 genes revealed no association of individual SNPs with AD. In contrast, haplotype analyses showed a significantly different haplotype distribution between patients and controls for CCR4 (P < 0.001). Furthermore, when SNP-SNP interaction effects were analysed for these two genes, we found significant evidence for epistatic interactions between SNPs within each of the two genes but no evidence for a gene-gene interaction, suggesting that variation in or near both the CCR4 and the NFKB1 genes might individually contribute to AD pathogenesis.

To test whether mutations in γ-aminobutyric acid type A receptor (GABAA -R) subunit genes contribute to the etiology of rolandic epilepsy (RE) or its atypical variants (ARE).We performed exome sequencing to compare the frequency of variants in 18 GABAA -R genes in 204 European patients with RE/ARE versus 728 platform-matched controls. Identified GABRG2 variants were functionally assessed for protein stability, trafficking, postsynaptic clustering, and receptor function.Of 18 screened GABAA -R genes, we detected an enrichment of rare variants in the GABRG2 gene in RE/ARE patients (5 of 204, 2.45%) in comparison to controls (1 of 723, 0.14%; odds ratio = 18.07, 95% confidence interval = 2.01-855.07, p = 0.0024, pcorr = 0.043). We identified a GABRG2 splice variant (c.549-3TG) in 2 unrelated patients as well as 3 nonsynonymous variations in this gene (p.G257R, p.R323Q, p.I389V). Functional assessment showed reduced surface expression of p.G257R and decreased GABA-evoked currents for p.R323Q. The p.G257R mutation displayed diminished levels of palmitoylation, a post-translational modification crucial for trafficking of proteins to the cell membrane. Enzymatically raised palmitoylation levels restored the surface expression of the p.G257R variant γ2 subunit.The statistical association and the functional evidence suggest that mutations of the GABRG2 gene may increase the risk of RE/ARE. Restoring the impaired membrane trafficking of some GABRG2 mutations by enhancing palmitoylation might be an interesting therapeutic approach to reverse the pathogenic effect of such mutants.

Short tandem repeat (STR) markers are widely and continuously used in forensic applications. However, past research has demonstrated substantial allelic association between STR markers on both autosomes and the X chromosome, leading to partially redundant information that these markers can provide. Here, we quantify the allelic association between Y-chromosomal STR markers that are part of established forensic panels, separately for three different continental groups. We further propose a sequential marker selection procedure that is based on Shannon's equivocation and that accounts for allelic association between STR markers, leading to a maximal gain in independent information. In application to three real-world data sets, we demonstrate the procedure's superior performance when compared to single-locus diversity selection strategies, resulting in the optimal marker set for a given data set in the majority of marker subsets. Noting the inferior performance of the established Y-STR marker panels in a retrospective investigation, we suggest that future forensic marker selection should be guided, besides by other technical selection criteria, by an equivocation-based approach to obtain maximally discriminatory marker sets at minimal cost.

Gene mapping refers to the localization of disease genes on the human gene map. The currently used mathematical methods for gene mapping are outlined and compared. The example of Gilles de la Tourette syndrome is discussed in detail.

The purpose of this work is to quantify the effects that errors in genotyping have on power and the sample size necessary to maintain constant asymptotic Type I and Type II error rates (SSN) for case-control genetic association studies between a disease phenotype and a di-allelic marker locus, for example a single nucleotide polymorphism (SNP) locus. We consider the effects of three published models of genotyping errors on the chi-square test for independence in the 2 × 3 table. After specifying genotype frequencies for the marker locus conditional on disease status and error model in both a genetic model-based and a genetic model-free framework, we compute the asymptotic power to detect association through specification of the test’s non-centrality parameter. This parameter determines the functional dependence of SSN on the genotyping error rates. Additionally, we study the dependence of SSN on linkage disequilibrium (LD), marker allele frequencies, and genotyping error rates for a dominant disease model. Increased genotyping error rate requires a larger SSN. Every 1% increase in sum of genotyping error rates requires that both case and control SSN be increased by 2–8%, with the extent of increase dependent upon the error model. For the dominant disease model, SSN is a nonlinear function of LD and genotyping error rate, with greater SSN for lower LD and higher genotyping error rate. The combination of lower LD and higher genotyping error rates requires a larger SSN than the sum of the SSN for the lower LD and for the higher genotyping error rate.

Background: Clinical features and animal models of essential tremor (ET) suggest gamma-aminobutyric acid A receptor (GABA(A)R) subunits and GABA transporters as putative candidate genes. Methods: A total of 503 ET cases and 818 controls were investigated for an association between polymorphisms in 15 GABA(A)R and four GABA transporter genes and ET. Results: Nine nominally significant tagging SNPs (P values from 4.9 x 10(-2) to 5.2 x 10(-4)) were found in the hypothesis generation stage. Five SNPs were followed up in a second verification stage but failed to reach significance. (P values from 0.30 to 0.77). Discussion: In our samples, no evidence of association between GABAAR and GABA transporter genes with ET was detected. Further studies are necessary to clarify the role of these genes in ET.

We identified 15q13.3 microdeletions encompassing the CHRNA7 gene in 12 of 1,223 individuals with idiopathic generalized epilepsy (IGE), which were not detected in 3,699 controls (joint P = 5.32 x 10(-8)). Most deletion carriers showed common IGE syndromes without other features previously associated with 15q13.3 microdeletions, such as intellectual disability, autism or schizophrenia. Our results indicate that 15q13.3 microdeletions constitute the most prevalent risk factor for common epilepsies identified to date.

In a previous study on Sicilian men, an association of the functional polymorphism Asp299Gly in the TLR4 gene was reported with longevity and a reduced risk of myocardial infarction (MI). Here, we sought to confirm the findings in our extensive collection of 273 long-lived men (95-107 years), 606 male early-onset MI patients and 594 appropriate controls from Germany. Despite sufficient statistical power to replicate the findings observed in the South Italians, our results rule out a noteworthy influence of the TLR4 polymorphism upon human longevity or MI in German men. Therefore, the previously described associations in Sicilians might reflect either population-specific or potentially false-positive results.

Material and Methods: Patients were randomized 1:1:1 to receive either SOF+RBV for 16 or 24 weeks or SOF+PEG/RBV for 12 weeks and stratified by HCV genotype and cirrhosis status. All patients received SOF 400mg daily and RBV 1000–1200mg in a divided daily dose. PEG was administered as 180mg weekly injection. The primary end point was sustained virologic response 12 weeks after treatment (SVR12). Results: Of 592 patients randomized and treated, 92% had GT3 HCV, 67% were male, 84% white, 53% treatment experienced, 62% had non-CC IL28B genotypes, and 37% had cirrhosis. GT2 treatmentexperienced patients with cirrhosis had high SVR12 rates in all treatment groups: 87% of those receiving SOF+RBV for 16 weeks, 100% of those receiving SOF+RBV for 24 weeks, and 94% of those receiving SOF+PEG/RBV for 12 weeks. Among GT3 patients, SVR12 rates were highest in those receiving SOF+PEG/RBV for 12 weeks (93%) as compared to SOF+RBV for 24 (84%, p 0.008) or 16 weeks (71%, p < 0.001) (Table). The most common adverse events in all arms were fatigue, headache, insomnia, and nausea. Overall, 6 (1%) patients discontinued treatment due to adverse events; one of them was treated with SOF+PEG/RBV.

Approaches to the genetics of cardiovascular disease through genetic field work. Successful molecular genetic studies of complex disease require exact, careful phenotypization, which is more difficult than that performed for monogenic diseases. We have developed a family-oriented field working approach, which relies on index patients, their primary care physicians, and a minimum number of field working staff. The patients are responsible for recruiting their family members. Packets containing an explanatory pamphlet, an informed consent statement, a questionnaire, and blood cuvettes are provided. Data are transferred from questionnaire and from the laboratory into a computer program that facilitates construction of the family tree. We have applied this genetic field working approach primarily to patients with lipid disorders. Coupling results from genetic field working with modern DNA diagnostic tests such as the oligonucleotide ligation assay, has enabled us to effectively identify patients with familial hypercholesterolemia in the German population. We are now extending genetic field working to hypertension. Hypertension is much more difficult to study, because the phenotype is more difficult to discern and document. Both complex diseases have the disadvantage that the parents of the index patients are likely to already be dead. Nevertheless, we concentrate on the recruitment of large pedigrees, sibling pairs with parents whenever possible, and trios consisting of index patient and both parents or index patient, parent and sibling. With these constellations we can conduct association studies, linkage analysis, and novel combinations of both approaches.

AimMany studies investigated the role of genetic variants in periodontitis, but few were established as risk factors. We aimed to validate the associations of recent candidate genes in aggressive periodontitis (AgP).Material and MethodsWe analysed 23 genes in 600 German AgP patients and 1441 controls on the Illumina custom genotyping array Immunochip. We tested a suggestive association in a Dutch and German/Austrian AgP case-control sample, and a German chronic periodontitis (CP) case-control sample using Sequenom iPlex assays. We additionally tested the common known risk variant rs1333048 of the gene ANRIL for its association in a Turkish and Italian population.ResultsNone of the analysed genes gave statistical evidence for association. Upon covariate adjustment for smoking and gender, in the pooled German-Austrian AgP sample, IL10 SNP rs6667202 was associated with p = 0.016, OR = 0.77 (95% CI = 0.6-0.95), and in the Dutch AgP sample, adjacent IL10 SNP rs61815643 was associated with p = 0.0009, OR = 2.31 (95% CI = 1.4-3.8). At rs61815643, binding of the transcription factor PPARG was predicted. ANRIL rs1333048 was associated in the Turkish sample (pallelic = 0.026, OR = 1.67 [95% CI = 1.11-2.60]).ConclusionsPrevious candidate genes carry no susceptibility factors for AgP. Association of IL-10 rs61815643 with AgP is suggested. ANRIL is associated with periodontitis across different populations.

The sterolin locus (ABCG5/ABCG8) confers susceptibility for cholesterol gallstone disease in humans. Both the responsible variant and the molecular mechanism causing an increased incidence of gallstones in these patients have as yet not been identified. Genetic mapping utilized patient samples from Germany (2,808 cases, 2,089 controls), Chile (680 cases, 442 controls), Denmark (366 cases, 766 controls), India (247 cases, 224 controls), and China (280 cases, 244 controls). Analysis of allelic imbalance in complementary DNA (cDNA) samples from human liver (n = 22) was performed using pyrosequencing. Transiently transfected HEK293 cells were used for [(3) H]-cholesterol export assays, analysis of protein expression, and localization of allelic constructs. Through fine mapping in German and Chilean samples, an ∼250 kB disease-associated interval could be defined for this locus. Lack of allelic imbalance or allelic splicing of the ABCG5 and ABCG8 transcripts in human liver limited the search to coding single nucleotide polymorphisms. Subsequent mutation detection and genotyping yielded two disease-associated variants: ABCG5-R50C (P = 4.94 × 10(-9) ) and ABCG8-D19H (P = 1.74 × 10(-10) ) in high pairwise linkage disequilibrium (r(2) = 0.95). [(3) H]-cholesterol export assays of allelic constructs harboring these genetic candidate variants demonstrated increased transport activity (3.2-fold, P = 0.003) only for the ABCG8-19H variant, which was also superior in nested logistic regression models in German (P = 0.018), Chilean (P = 0.030), and Chinese (P = 0.040) patient samples.This variant thus provides a molecular basis for biliary cholesterol hypersecretion as the mechanism for cholesterol gallstone formation, thereby drawing a link between "postgenomic" and "pregenomic" pathophysiological knowledge about this common complex disorder. (HEPATOLOGY 2012).

Idiopathic focal epilepsy (IFE) with rolandic spikes is the most common childhood epilepsy, comprising a phenotypic spectrum from rolandic epilepsy (also benign epilepsy with centrotemporal spikes, BECTS) to atypical benign partial epilepsy (ABPE), Landau-Kleffner syndrome (LKS) and epileptic encephalopathy with continuous spike and waves during slow-wave sleep (CSWS)(1,2). The genetic basis is largely unknown. We detected new heterozygous mutations in GRIN2A in 27 of 359 affected individuals from 2 independent cohorts with IFE (7.5%; P = 4.83 x 10(-18), Fisher's exact test). Mutations occurred significantly more frequently in the more severe phenotypes, with mutation detection rates ranging from 12/245 (4.9%) in individuals with BECTS to 9/51 (17.6%) in individuals with CSWS (P = 0.009, Cochran-Armitage test for trend). In addition, exon-disrupting microdeletions were found in 3 of 286 individuals (1.0%; P = 0.004, Fisher's exact test). These results establish alterations of the gene encoding the NMDA receptor NR2A subunit as a major genetic risk factor for IFE.

BACKGROUND AND AIMS: High frequency electrosurgery has a key role in the broadening application of liver surgery. Its molecular signature, i.e. the metabolites evolving from electrocauterization which may inhibit hepatic wound healing, have not been systematically studied. METHODS: Human liver samples were thus obtained during surgery before and after electrosurgical dissection and subjected to a two-stage metabolomic screening experiment (discovery sample: N = 18, replication sample: N = 20) using gas chromatography/mass spectrometry. RESULTS: In a set of 208 chemically defined metabolites, electrosurgical dissection lead to a distinct metabolic signature resulting in a separation in the first two dimensions of a principal components analysis. Six metabolites including glycolic acid, azelaic acid, 2-n-pentylfuran, dihydroactinidiolide, 2-butenal and n-pentanal were consistently increased after electrosurgery meeting the discovery (p

Background To date, liver biopsy is the only means of reliable diagnosis for fatty liver disease (FLD). Owing to the inevitable biopsy-associated health risks, however, the development of valid noninvasive diagnostic tools for FLD is well warranted. Aim We evaluated a particular metabolic profile with regard to its ability to diagnose FLD and compared its performance to that of established phenotypes, conventional biomarkers and disease-associated genotypes. Methods The study population comprised 115 patients with ultrasound-diagnosed FLD and 115 sex- and age-matched controls for whom the serum concentration was measured of 138 different metabolites, including acylcarnitines, amino acids, biogenic amines, hexose, phosphatidylcholines (PCs), lyso-PCs and sphingomyelins. Established phenotypes, biomarkers, disease-associated genotypes and metabolite data were included in diagnostic models for FLD using logistic regression and partial least-squares discriminant analysis. The discriminative power of the ensuing models was compared with respect to area under curve (AUC), integrated discrimination improvement (IDI) and by way of cross-validation (CV). Results Use of metabolic markers for predicting FLD showed the best performance among all considered types of markers, yielding an AUC of 0.8993. Additional information on phenotypes, conventional biomarkers or genotypes did not significantly improve this performance. Phospholipids and branched-chain amino acids were most informative for predicting FLD. Conclusion We show that the inclusion of metabolite data may substantially increase the power to diagnose FLD over that of models based solely upon phenotypes and conventional biomarkers.

Numerous studies of human populations in Europe and Asia have revealed a concordance between their extant genetic structure and the prevailing regional pattern of geography and language. For native South Americans, however, such evidence has been lacking so far. Therefore, we examined the relationship between Y-chromosomal genotype on the one hand, and male geographic origin and linguistic affiliation on the other, in the largest study of South American natives to date in terms of sampled individuals and populations. A total of 1,011 individuals, representing 50 tribal populations from 81 settlements, were genotyped for up to 17 short tandem repeat (STR) markers and 16 single nucleotide polymorphisms (Y-SNPs), the latter resolving phylogenetic lineages Q and C. Virtually no structure became apparent for the extant Y-chromosomal genetic variation of South American males that could sensibly be related to their inter-tribal geographic and linguistic relationships. This continent-wide decoupling is consistent with a rapid peopling of the continent followed by long periods of isolation in small groups. Furthermore, for the first time, we identified a distinct geographical cluster of Y-SNP lineages C-M217 (C3*) in South America. Such haplotypes are virtually absent from North and Central America, but occur at high frequency in Asia. Together with the locally confined Y-STR autocorrelation observed in our study as a whole, the available data therefore suggest a late introduction of C3* into South America no more than 6,000 years ago, perhaps via coastal or trans-Pacific routes. Extensive simulations revealed that the observed lack of haplogroup C3* among extant North and Central American natives is only compatible with low levels of migration between the ancestor populations of C3* carriers and non-carriers. In summary, our data highlight the fact that a pronounced correlation between genetic and geographic/cultural structure can only be expected under very specific conditions, most of which are likely not to have been met by the ancestors of native South Americans., Author Summary In the largest population genetic study of South Americans to date, we analyzed the Y-chromosomal makeup of more than 1,000 male natives. We found that the male-specific genetic variation of Native Americans lacks any clear structure that could sensibly be related to their geographic and/or linguistic relationships. This finding is consistent with a rapid initial peopling of South America, followed by long periods of isolation in small tribal groups. The observed continent-wide decoupling of geography, spoken language, and genetics contrasts strikingly with previous reports of such correlation from many parts of Europe and Asia. Moreover, we identified a cluster of Native American founding lineages of Y chromosomes, called C-M217 (C3*), within a restricted area of Ecuador in North-Western South America. The same haplogroup occurs at high frequency in Central, East, and North East Asia, but is virtually absent from North (except Alaska) and Central America. Possible scenarios for the introduction of C-M217 (C3*) into Ecuador may thus include a coastal or trans-Pacific route, an idea also supported by occasional archeological evidence and the recent coalescence of the C3* haplotypes, estimated from our data to have occurred some 6,000 years ago.

Background Human ß-defensins are a family of antimicrobial peptides located at the mucosal surface. Both sequence multi-site variations (MSV) and copy-number variants (CNV) of the defensin-encoding genes are associated with increased risk for various diseases, including cancer and inflammatory conditions such as psoriasis and acute pancreatitis. In a case–control study, we investigated the association between MSV in DEFB104 as well as defensin gene (DEF) cluster copy number (CN), and pancreatic ductal adenocarcinoma (PDAC) and chronic pancreatitis (CP). Results Two groups of PDAC (N=70) and CP (N=60) patients were compared to matched healthy control groups CARLA1 (N=232) and CARLA2 (N=160), respectively. Four DEFB104 MSV were haplotyped by PCR, cloning and sequencing. DEF cluster CN was determined by multiplex ligation-dependent probe amplification. Neither the PDAC nor the CP cohorts show significant differences in the DEFB104 haplotype distribution compared to the respective control groups CARLA1 and CARLA2, respectively. The diploid DEF cluster CN exhibit a significantly different distribution between PDAC and CARLA1 (Fisher’s exact test P=0.027), but not between CP and CARLA2 (P=0.867). Conclusion Different DEF cluster b CN distribution between PDAC patients and healthy controls indicate a potential protective effect of higher CNs against the disease.

Massively parallel sequencing of whole genomes and exomes has facilitated a direct assessment of causative genetic variation, now enabling the identification of genetic factors involved in rare diseases (RD) with Mendelian inheritance patterns on an almost routine basis. Here, we describe the illustrative case of a single consanguineous family where this strategy suffered from the difficulty to distinguish between two etiologically distinct disorders, namely the co-occurrence of hereditary hypophosphatemic rickets (HRR) and congenital myopathies (CM), by their phenotypic manifestation alone. We used parametric linkage analysis, homozygosity mapping and whole exome-sequencing to identify mutations underlying HRR and CM. We also present an approximate approach for assessing the probability of co-occurrence of two unlinked recessive RD in a single family as a function of the degree of consanguinity and the frequency of the disease-causing alleles. Linkage analysis and homozygosity mapping yielded elusive results when assuming a single RD, but whole-exome sequencing helped to identify two mutations in two genes, namely SLC34A3 and SEPN1, that segregated independently in this family and that have previously been linked to two etiologically different diseases. We assess the increase in chance co-occurrence of rare diseases due to consanguinity, i.e. under circumstances that generally favor linkage mapping of recessive disease, and show that this probability can increase by several orders of magnitudes. We conclude that such potential co-occurrence represents an underestimated risk when analyzing rare or undefined diseases in consanguineous families and should be given more consideration in the clinical and genetic evaluation.

Colorectal cancer (CRC), one of the most frequent neoplasias worldwide, has both genetic and environmental causes. As yet, however, gene–environment (G × E) interactions in CRC have been studied mostly for a small number of candidate genes only. Therefore, we investigated the possible interaction, in CRC etiology, between single-nucleotide polymorphisms (SNPs) on the one hand, and overweight, smoking and alcohol consumption on the other, at a genome-wide level. To this end, we adopted a two-tiered approach comprising a case-only screening stage I (314 cases) and a case–control validation stage II (259 cases, 1,002 controls). Interactions with the smallest p value in stage I were verified in stage II using multiple logistic regression analysis adjusted for sex and age. In addition, we specifically studied known CRC-associated SNPs for possible G × E interactions. Upon adjustment for sex and age, and after allowing for multiple testing, however, only a single SNP (rs1944511) was found to be involved in a statistically significant interaction, namely with overweight (multiplicity-corrected p = 0.042 in stage II). Several other G × E interactions were nominally significant but failed correction for multiple testing, including a previously reported interaction between rs9929218 and alcohol consumption that also emerged in our candidate SNP study (nominal p = 0.008). Notably, none of the interactions identified in our genome-wide analysis was with a previously reported CRC-associated SNP. Our study therefore highlights the potential of an “agnostic” genome-wide approach to G × E analysis.

Recent trials and guidelines have established the use of neoadjuvant chemotherapy for resectable UICC stage II to IV gastric cancers. In this setting, preoperative staging is pivotal for correct patient selection. This cohort study was designed to assess the accuracy of endoscopic ultrasound (EUS) and the ability to select correctly patients for neoadjuvant chemotherapy on the basis of survival outcome.Eighty-two consecutive Caucasian patients (46 male; median age 72 years) with gastric cancer underwent EUS staging and subsequent surgery without perioperative chemotherapy or radiotherapy. Patients were followed for a median of 800 days postoperatively. Pathology and EUS UICC and T stages were compared and evaluated as predictors of survival using Kaplan-Meier and Cox regression analysis.The overall accuracy of EUS for UICC classification compared with pathology was 62 %, and the accuracy for delineation of UICC I was 89 %. For the therapeutically relevant differentiation of early gastric cancer (UICC stage I), EUS (mean survival, 2,298 days, R2 = 0.23) and pathology (2,461 days, R2 = 0.24) predicted survival equally well. Similar results were obtained for T staging by EUS (mean survival, 2,065 days for uT1/2, R2 = 0.24) or pathology (2,185 days, R2 = 0.22).EUS identifies the low risk subgroup (uUICC stage I or uT1/2) with similar performance as pUICC stage I or stage pT1/2 in gastric cancer and very similar survival characteristics. EUS thus may be the noninvasive method of choice for preoperative selection of patients for immediate resection versus neoadjuvant chemotherapy.

A large number of short tandem repeat (STR) markers spanning the entire human X chromosome have been described and established for use in forensic genetic testing. Due to their particular mode of inheritance, X-STRs often allow easy and informative haplotyping in kinship analyses. Moreover, some X-STRs are known to be tightly linked so that, in combination, they constitute even more complex genetic markers than each STR taken individually. As a consequence, X-STRs have proven particularly powerful in solving complex cases of disputed blood relatedness. However, valid quantification of the evidence provided by X-STR genotypes in the form of likelihood ratios requires that the recombination rates between markers are exactly known. In a collaborative family study, we used X-STR genotype data from 401 two- and three-generation families to derive valid estimates of the recombination rates between 12 forensic markers widely used in forensic testing, namely DXS10148, DXS10135, DXS8378 (together constituting linkage group I), DXS7132, DXS10079, DXS10074 (linkage group II), DXS10103, HPRTB, DXS10101 (linkage group III), DXS10146, DXS10134 and DXS7423 (linkage group IV). Our study is the first to simultaneously allow for mutation and recombination in the underlying likelihood calculations, thereby obviating the bias-prone practice of excluding ambiguous transmission events from further consideration. The statistical analysis confirms that linkage groups I and II are transmitted independently from one another whereas linkage groups II, III and IV are characterised by inter-group recombination fractions that are notably smaller than 50%. Evidence was also found for recombination within all four linkage groups, with recombination fraction estimates ranging as high as 2% in the case of DXS10146 and DXS10134.

Somatic copy number (CN) alterations are major drivers of tumorigenesis and growth. Although next-generation sequencing (NGS) technologies enable a deep genomic analysis of cancers, the analysis of the data remains subject to biases and multiple sources of error, including varying local read coverage. The currently existing algorithms for NGS-based detection of CN abberations do not incorporate information on the local coverage quality. We have developed a new algorithm, copy number estimation with controlled support (CoNCoS) that increases the accuracy of CN estimation in paired tumor/normal exome sequencing data sets by assessing and optimizing the support for a site-specific CN estimate. We show by simulations and in a benchmarking study against single nucleotide polymorphism (SNP) microarray data that our approach outperforms the commonly used methods CNAnorm and VarScan2. Our algorithm is suitable to increase the accuracy of somatic CN analysis by a support-optimized estimation approach.

BACKGROUNDDupuytren's disease is a benign fibromatosis of the hands and fingers that leads to flexion contractures. We hypothesized that multiple genetic and environmental factors influence susceptibility to this disease and sought to identify susceptibility genes to better understand its pathogenesis.METHODSWe conducted a genomewide association study of 960 Dutch persons with Dupuytren's disease and 3117 controls (the discovery set) to test for association between the disease and genetic markers. We tested the 35 single-nucleotide polymorphisms (SNPs) most strongly associated with Dupuytren's disease (PRESULTSInitially, we observed a significant genomewide association between Dupuytren's disease and 8 SNPs at three loci. Tests of replication and joint analysis of all data from 2325 patients with Dupuytren's disease and 11,562 controls yielded an association with 11 SNPs from nine different loci (PCONCLUSIONSThis study implicates nine different loci involved in genetic susceptibility to Dupuytren's disease. The fact that six of these nine loci harbor genes encoding proteins in the Wnt-signaling pathway suggests that aberrations in this pathway are key to the process of fibromatosis in Dupuytren's disease.

Cytosine methylation provides an epigenetic level of cellular plasticity that is important for development, differentiation and cancerogenesis. We adopted microdroplet PCR to bisulfite treated target DNA in combination with second generation sequencing to simultaneously assess DNA sequence and methylation. We show measurement of methylation status in a wide range of target sequences (total 34 kb) with an average coverage of 95% (median 100%) and good correlation to the opposite strand (rho = 0.96) and to pyrosequencing (rho = 0.87). Data from lymphoma and colorectal cancer samples for SNRPN (imprinted gene), FGF6 (demethylated in the cancer samples) and HS3ST2 (methylated in the cancer samples) serve as a proof of principle showing the integration of SNP data and phased DNA-methylation information into “hepitypes” and thus the analysis of DNA methylation phylogeny in the somatic evolution of cancer.

Next-generation sequencing and the availability of high-density genotyping arrays have facilitated an analysis of somatic and meiotic mutations at unprecedented level, but drawing sensible conclusions about the functional relevance of the detected variants still remains a formidable challenge. In this context, the study of allelic imbalance in intermediate RNA phenotypes may prove a useful means to elucidate the likely effects of DNA variants of unknown significance. We developed a statistical framework for the assessment of allelic imbalance in next-generation transcriptome sequencing (RNA-seq) data that requires neither an expression reference nor the underlying nuclear genotype(s), and that allows for allele miscalls. Using extensive simulation as well as publicly available whole-transcriptome data from European-descent individuals in HapMap, we explored the power of our approach in terms of both genotype inference and allelic imbalance assessment under a wide range of practically relevant scenarios. In so doing, we verified a superior performance of our methodology, particularly at low sequencing coverage, compared to the more simplistic approach of completely ignoring allele miscalls. Because the proposed framework can be used to assess somatic mutations and allelic imbalance in one and the same set of RNA-seq data, it will be particularly useful for the analysis of somatic genetic variation in cancer studies. Hum Mutat 32:98-106, 2011. (C) 2010 Wiley-Liss, Inc.

Next-generation sequencing (NGS) will likely facilitate a better understanding of the causes and consequences of human genetic variability. In this context, the validity of NGS-inferred single-nucleotide variants (SNVs) is of paramount importance. We therefore developed a statistical framework to assess the fidelity of three common NGS platforms. Using aligned DNA sequence data from two completely sequenced HapMap samples as included in the 1000 Genomes Project, we unraveled remarkably different error profiles for the three platforms. Compared to confirmed HapMap variants, newly identified SNVs included a substantial proportion of false positives (3–17%). Consensus calling by more than one platform yielded significantly lower error rates (1–4%). This implies that the use of multiple NGS platforms may be more cost-efficient than relying upon a single technology alone, particularly in physically localized sequencing experiments that rely upon small error rates. Our study thus highlights that different NGS platforms suit different practical applications differently well, and that NGS-based studies require stringent data quality control for their results to be valid.

Essential tremor (ET) and Parkinson's disease (PD) are the most common movement disorders and show clinical, genetic, and pathophysiological overlap. Single-nucleotide polymorphisms (SNPs) in the leucine-rich repeat (LRR) and immunoglobulin (Ig) domain-containing, Nogo receptor-interacting protein gene (LINGO1) are associated with ET. LINGO1 is overexpressed in the substantia nigra (SN) of PD patients and inhibition of LINGO1 confers neuroprotection in a rodent model of PD. In this study we test the hypothesis whether SNPs in the LINGO1 gene that are associated with ET are also associated with PD. Three large German case–control samples from Kiel, Lubeck, and Tubingen (total: 1,798 cases and 1,482 controls) were genotyped for the three LINGO1 SNPs associated with ET. Association was assessed using allele- and genotype-based tests in each of the three samples separately, in the combined sample, and in subsets of patients with early-onset PD (

Background Sarcoidosis (SA) is a systemic granulomatous inflammatory disorder with complex etiology and strong clustering in families. Genome-wide association studies have been successful in the identification of common risk variants for the disease. To reveal susceptibility variants with low frequencies but strong effects, we performed a genome-wide linkage scan in a large sample of SA families. Methods We genotyped 528 members of 181 German SA families for 3,882 single nucleotide polymorphism assays from the SNPlex System Human Linkage Mapping Set 4K. Results Nonparametric linkage analysis revealed one region of suggestive linkage on chromosome 12p13.31 at 20 cM (logarithm of odds [LOD] = 2.53; local P value=.0003) and another linkage peak of nearly suggestive linkage on 9q33.1 at 134 cM (LOD = 2.12; local P value=.0009). The latter has been reported to show suggestive evidence for linkage in a sample of 229 African American SA families previously. Analysis of acute and chronically affected families revealed a subphenotype-specific linkage pattern and an additional, nearly suggestive linkage peak on chromosome 16p13.11 at 38 cM (LOD = 2.09; local P value=.001), which was confined to acute SA. Conclusion Our results propose that the respective regions might harbor yet-unidentified, possibly subphenotype-specific risk factors for the disease (eg, with immune-related functions like the tumor necrosis factor receptor 1). They should be proved to be important for SA pathogenesis and investigated in detail with an emphasis on rare variants. Subphenotype-specific risk factors might serve for prognosis of the clinical course of the disease.

ObjectivePostprandial (pp) lipid metabolism is associated with insulin resistance and type 2 diabetes. In young men, pp triglycerides (TGs) are more strongly associated with traits of metabolic syndrome (MS) than fasting TGs. We established a cohort of middle-aged men selected for traits of MS and pp lipid metabolism to determine if fasting TGs or pp TGs are more closely related to MS.Research design and methodsA total of 1558 men were characterized for MS. A total of 755 men underwent an oral metabolic tolerance test consisting of a standardized high-fat meal and an oral glucose tolerance test. Blood samples were drawn in the fasting state and hourly until 9 h to determine pp TGs and free fatty acids. Glucose and insulin were analyzed until 5 h pp.ResultsIn the overall cohort, 329 subjects (21.1%) had a complete MS based on the Adult Treatment Panel III criteria, and 650 subjects (41.7%) had a complete MS based on the International Diabetes Federation criteria. The association of pp TGs with MS parameters was not stronger than the association of fasting TGs with them. Pp TGs were independently associated with β-cell function.ConclusionsPp TGs did not show a higher correlation with MS traits than fasting TGs. This finding is probably due to the high incidence of overweight subjects in this middle-aged cohort.

COX-2 plays an important role in periodontitis by mediating inflammatory reactions in periodontal tissues, and the COX-2 polymorphisms rs20417 and rs689466 have been reported to be associated with periodontitis in populations of Taiwanese and Chinese ethnicity. To test whether these variants were associated with periodontitis in populations of European ethnicity, we genotyped the single-nucleotide polymorphisms (SNPs) rs689466 and rs6681231, the latter a haplotype tagging SNP (htSNP) for rs20417 (r2>0.95), in our large-analysis population of individuals with aggressive (n = 532) and chronic periodontitis (n = 1052), and 2873 healthy control individuals. The rare G allele of htSNP rs6681231 was associated with aggressive periodontitis prior to and after adjustment for the covariates smoking, diabetes, and gender, with an odds ratio of 1.57 (95% confidence interval 1.18–2.08; p = 0.002). The validation of the association of rs20417 by the htSNP rs6681231 provides evidence for a general genetic risk of COX-2 variants in the pathogenesis of periodontitis.

BACKGROUND & AIMS: Genome-wide association studies have mapped loci that are associated with serum levels of bilirubin. Bilirubin is a major component of gallstones so we investigated whether these variants predict gallstone bilirubin content and overall risk for gallstones. METHODS: Loci that were identified in a meta-analysis to attain a genome-wide significance level of a P value less than 1.0 107 (UGT1A1, SLCO1B1, LST-3TM12, SLCO1A2) were analyzed in 1018 individuals with known gallstone composition. Gallstone risk was analyzed in 2606 German choleystecomized individuals and 1121 controls and was replicated in 210 cases and 496 controls from South America. RESULTS: By using the presence of bilirubin as a phenotype, variants rs6742078 (UGT1A1; P .003), rs4149056 (SLCO1B1; P .003), and rs4149000 (SLCO1A2; P .015) were associated with gallstone composition. In regression analyses, only UGT1A1 and SLCO1B1 were independently retained in the model. UGT1A1 (rs6742078; P .018) was associated with overall gallstone risk. In a sex-stratified analysis, only male carriers of rs6742078 had an increased risk for gallstone disease (P 2.1 107; odds ratiorecessive, 2.34; Pwomen .47). The sex-specific association of rs6742078 was confirmed in samples from South America (Pmen .046; odds ratiorecessive, 2.19; Pwomen .96). CONCLUSIONS: The UGT1A1 Gilbert syndrome variant rs6742078 is associated with gallstone disease in men; further studies are required regarding the sex-specific physiology of bilirubin and bile acid metabolism. Variants of ABCG8 and UGT1A1 are the 2 major risk factors for overall gallstone disease, they contribute a population attributable risk of 21.2% among men. Fil: Buch, Stephan. University Hospital Schleswig-Holstein; Alemania Fil: Schafmayer, Clemens. University Hospital Schleswig-Holstein; Alemania Fil: Völzke, Henry. University of Greifswald; Alemania Fil: Seeger, Marcus. University Hospital Schleswig-Holstein; Alemania Fil: Miquel, Juan F.. Pontificia Universidad Católica de Chile; Chile Fil: Sookoian, Silvia Cristina. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Houssay. Instituto de Investigaciones Medicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; Argentina Fil: Egberts, Jan H.. University Hospital Schleswig-Holstein; Alemania Fil: Arlt, Alexander. University Hospital Schleswig-Holstein; Alemania Fil: Pirola, Carlos Jose. University Hospital Schleswig-Holstein; Alemania. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Lerch, Markus M.. University of Greifswald; Alemania Fil: John, Ulrich. University of Greifswald; Alemania Fil: Franke, Andre. University Hospital Schleswig-Holstein; Alemania Fil: von Kampen, Oliver. University Hospital Schleswig-Holstein; Alemania Fil: Brosch, Mario. University Hospital Schleswig-Holstein; Alemania Fil: Nothnagel, Michael. University Hospital Schleswig-Holstein; Alemania Fil: Kratzer, Wolfgang. Universitat Ulm; Alemania Fil: Boehm, Bernhard O.. Universitat Ulm; Alemania Fil: Bröring, Dieter C.. University Hospital Schleswig-Holstein; Alemania Fil: Schreiber, Stefan. University Hospital Schleswig-Holstein; Alemania Fil: Krawczak, Michael. University Hospital Schleswig-Holstein; Alemania Fil: Hampe, Jochen. University Hospital Schleswig-Holstein; Alemania

Periodontitis is a widespread, complex inflammatory disease of the mouth, which results in a loss of gingival tissue and alveolar bone, with aggressive periodontitis (AgP) as its most severe form. To identify genetic risk factors for periodontitis, we conducted a genome-wide association study in German AgP patients. We found AgP to be strongly associated with the intronic SNP rs1537415, which is located in the glycosyltransferase gene GLT6D1. We replicated the association in a panel of Dutch generalized and localized AgP patients. In the combined analysis including 1758 subjects, rs1537415 reached a genome-wide significance level of P= 5.51 x 10(-9), OR = 1.59 (95% CI 1.36-1.86). The associated rare G allele of rs1537415 showed an enrichment of 10% in periodontitis cases (48.4% in comparison with 38.8% in controls). Fine-mapping and a haplotype analysis indicated that rs1537415 showed the strongest association signal. Sequencing identified no further associated variant. Tissue-specific expression analysis of GLT6D1 indicated high transcript levels in the leukocytes, the gingiva and testis. Analysis of potential transcription factor binding sites at this locus predicted a significant reduction of GATA-3 binding affinity, and an electrophoretic mobility assay indicated a T cell specific reduction of protein binding for the G allele. Overexpression of GATA-3 in HEK293 cells resulted in allele-specific binding of GATA-3, indicating the identity of GATA-3 as the binding protein. The identified association of GLT6D1 with AgP implicates this locus as an important susceptibility factor, and GATA-3 as a potential signaling component in the pathophysiology of periodontitis.

Periodontal diseases are complex inflammatory diseases and affect up to 20% of the worldwide population. An unbalanced reaction of the immune system toward microbial pathogens is considered as the key factor in the development of periodontitis. Defensins have a strong antimicrobial function and are important contributors of the immune system toward maintaining health. Here, we present the first systematic association study of DEFB1. Using a haplotype-tagging single nucleotide polymorphism (SNP) approach, including described promoter SNPs of DEFB1, we investigated the associations of the selected variants in a large population (N=1337 cases and 2887 ethnically matched controls). The 3′ untranslated region SNP, rs1047031, showed the most significant association signal for homozygous carriers of the rare A allele (P=0.002) with an increased genetic risk of 1.3 (95% confidence interval: 1.11–1.57). The association was consistent with the specific periodontitis forms: chronic periodontitis (odds ratio=2.2 (95% confidence interval: 1.16–4.35), P=0.02), and aggressive periodontitis (odds ratio=1.3 (95% confidence interval 1.04–1.68), P=0.02). Sequencing of regulatory and exonic regions of DEFB1 identified no other associated variant, pointing toward rs1047031 as likely being the causative variant. Prediction of microRNA targets identified a potential microRNA-binding site at the position of rs1047031.

Genotype imputation for single nucleotide polymorphisms (SNPs) has been shown to be a powerful means to include genetic markers in exploratory genetic association studies without having to genotype them, and is becoming a standard procedure. A number of different software programs are available. In our experience, user-friendliness is often the deciding factor in the choice of software to solve a particular task. We therefore evaluated the usability of three publicly available imputation programs: BEAGLE, IMPUTE and MACH. We found all three programs to perform well with HapMap reference data, with little effort needed for data preparation and subsequent association analysis. Each of them has different strengths and weaknesses, however, and none is optimal for all situations.

At least in principle, most instances of complex kinship testing can be reduced to pairwise kinship cases involving two critical family members that either link or separate presumed sub-branches of a family. In the European population, the 34 short tandem repeats (STRs) currently used in forensic genetics are sufficiently powerful to allow assessment of disputed first and second but not lower degrees of pairwise blood relatedness. We provide estimates of the means and variances of marker-specific log-likelihood ratios, using large-sample approximation and assuming different scenarios of pairwise kinship analysis. These estimates allow power calculations to be performed for any combination of the available STRs. Since some of the markers considered are physically linked, chromosome-wide likelihood calculations in kinship cases other than parent-child duos (and trios) have to take the reduced rates of meiotic inter-marker recombination into account. We show by simulation that this requirement may be ignored when discriminating distant hypotheses about kinship, but that linkage may play an important role in the biostatistical analysis of more intricate cases.

Risk alleles for age-related diseases are expected to decrease in frequency in the population strata of increasing age. Consistent with this hypothesis, earlier studies showed a depletion of the Alzheimer's disease risk factor APOE*epsilon4 in long-lived individuals (LLIs). To evaluate whether this observation also holds for a previously suggested Alzheimer's disease risk haplotype in the A2M gene, we analyzed this particular haplotype in 1042 German LLIs (aged 95-100 years) and 1040 younger individuals (aged 60-75 years). Our results show a significant depletion of this haplotype in LLIs, thus confirming it as a mortality factor in the elderly. Consequently, our data support an involvement of the suggested A2M risk haplotype in the pathogenesis of Alzheimer's disease and adds new evidence to the risk-allele depletion hypothesis.

The filaggrin (FLG) gene is one of the most widely replicated susceptibility genes for atopic dermatitis (AD) so far. Yet, FLG mutations cannot fully account for the original linkage peak on chromosome 1q21, a region comprising the so-called epidermal differentiation complex (EDC). Since the EDC contains numerous genes relevant for epidermal differentiation, we sought to evaluate variation in other genes located in this region in a German AD case-control cohort. Thirty-two single nucleotide polymorphisms (SNPs) in 21 genes across the EDC were genotyped in 402 unrelated AD patients and 325 non-atopic controls by means of restriction enzyme digestion or TaqMan assays. Allele and genotype frequencies were tested for differences between patients and controls by logistic regression. Haplotype frequencies were evaluated using the famhap software. Except for the already known association with FLG, we did not identify any additional significant associations of EDC genes with AD. Thus, in this German cohort, there is no evidence that additional genes in the EDC region apart from FLG contribute substantially to AD pathogenesis.

Background: NOD proteins are part of innate immunity mechanisms. They play a role in epithelial barrier functions and inflammatory responses to bacteria. Single nucleotide polymorphisms (SNPs) in the NOD1 gene have proven to be associated with inflammatory bowel disease (IBD) and asthma. Objective: To investigate SNPs in the NOD1 gene in relation to aggressive periodontitis (AgP), a multifactorial, inflammatory disease of the supporting tissues of the teeth. Materials and Methods: Five SNPs in the NOD1 gene (4 intronic and 1 exonic) were tested for association in a total of 415 AgP patients and 874 controls both of Northern European ancestry. Results: The frequencies of the rare SNP alleles ranged between 21% and 26% among cases, and 20—27% among controls, and were not statistically different between cases and controls. Two SNPs were in strong linkage disequilibrium (r2 = 0.97 in cases and 0.94 in controls). The overall haplotype distributions did not differ between cases and controls. We observed 8 haplotypes with a frequency of !1% among cases and/or controls, but none of these haplotype frequencies differed significantly among cases and controls. Logistic regression analyses with adjustment for gender and smoking status did not reveal significant associations with AgP for any of the 5 SNPs. This study had a power of !95% to detect associations with variants carrying relative risks of !1.5 for heterozygote carriers and !2.25 for homozygote carriers. Conclusions: Although SNPs in the NOD1 gene have been strongly associated with cases of IBD, the current study failed to show an association of NOD1 SNPs with AgP.

9 páginas, 3 figuras, 2 tablas.-- et al., Genetic matching potentially provides a means to alleviate the effects of incomplete Mendelian randomization in population-based gene–disease association studies. We therefore evaluated the genetic-matched pair study design on the basis of genome-wide SNP data (309 790 markers; Affymetrix GeneChip Human Mapping 500K Array) from 2457 individuals, sampled at 23 different recruitment sites across Europe. Using pair-wise identity-by-state (IBS) as a matching criterion, we tried to derive a subset of markers that would allow identification of the best overall matching (BOM) partner for a given individual, based on the IBS status for the subset alone. However, our results suggest that, by following this approach, the prediction accuracy is only notably improved by the first 20 markers selected, and increases proportionally to the marker number thereafter. Furthermore, in a considerable proportion of cases (76.0%), the BOM of a given individual, based on the complete marker set, came from a different recruitment site than the individual itself. A second marker set, specifically selected for ancestry sensitivity using singular value decomposition, performed even more poorly and was no more capable of predicting the BOM than randomly chosen subsets. This leads us to conclude that, at least in Europe, the utility of the genetic-matched pair study design depends critically on the availability of comprehensive genotype information for both cases and controls., This work was supported by the Netherlands Forensic Institute (M Ka), Affymetrix (M Ka and M Kr), the German National Genome Research Network and the German Federal Ministry of Education and Research (H-EW, SS, M Kr and PN); the Helmholtz Zentrum München – German Research Center for Environmental Health, Neuherberg and the Munich Center of Health Sciences as part of LMUinnovativ (H-EW), the Netherlands Organization for Scientific Research (AGU: NWO 175.010.2005.011), the European Commission (AGU: GEFOS; 201865, AS: LD Europe; QLG2-CT-2001-00916); the Czech Ministry of Health (MM: VZFNM 00064203 and IGA NS/9488-3), Helse-Vest, Regional Health Authority Norway (LAB), the Swedish National Board of Forensic Medicine (GH: RMVFoU 99:22, 02:20) and the Academy of Finland (AS: 80578, OMLL, JP: 109265 and 111713).