Comparative genomics of Rothia species reveals diversity in novel biosynthetic gene clusters and ecological adaptation to different eukaryotic hosts and host niches.

Rothia species are understudied members of the phylum Actinobacteria and prevalent colonizers of the human and animal upper respiratory tract and oral cavity. The oral cavity, including the palatine tonsils, is colonized by a complex microbial community, which compete for resources, actively suppress competitors and influence host physiology. We analysed genomic data from 43 new porcine Rothia isolates, together with 112 publicly available draft genome sequences of Rothia isolates from humans, animals and the environment. In all Rothia genomes, we identified biosynthetic gene clusters predicted to produce antibiotic non-ribosomal peptides, iron scavenging siderophores and other secondary metabolites that modulate microbe-microbe and potentially microbe-host interactions. In vitro overlay inhibition assays corroborated the hypothesis that specific strains produce natural antibiotics. Rothia genomes encode a large number of carbohydrate-active enzymes (CAZy), with varying CAZy activities among the species found in different hosts, host niches and environments. These findings reveal competition mechanisms and metabolic specializations linked to ecological adaptation of Rothia species in different hosts.

Alteration of Bacterial Communities in Anterior Nares and Skin Sites of Patients Undergoing Arthroplasty Surgery: Analysis by 16S rRNA and Staphylococcal-Specific tuf Gene Sequencing.

The aim was to study alterations of bacterial communities in patients undergoing hip or knee arthroplasty to assess the impact of chlorhexidine gluconate soap decolonisation and systemic antibiotic prophylaxis. A Swedish multicentre, prospective collection of samples obtained from elective arthroplasty patients (n = 83) by swabbing anterior nares, skin sites in the groin and the site of planned surgery, before and after arthroplasty surgery, was analysed by 16S rRNA (V3-V4) gene sequencing and a complementary targeted tuf gene sequencing approach to comprehensively characterise alterations in staphylococcal communities. Significant reductions in alpha diversity was detected for both bacterial (p = 0.04) and staphylococcal (p = 0.03) groin communities after arthroplasty surgery with significant reductions in relative Corynebacterium (p = 0.001) abundance and Staphylococcus hominis (p = 0.01) relative staphylococcal abundance. In nares, significant reductions occurred for Staphylococcus hominis (p = 0.02), Staphylococcus haemolyticus (p = 0.02), and Staphylococcus pasteuri (p = 0.003) relative to other staphylococci. Staphylococcus aureus colonised 35% of anterior nares before and 26% after arthroplasty surgery. Staphylococcus epidermidis was the most abundant staphylococcal species at all sampling sites. No bacterial genus or staphylococcal species increased significantly after arthroplasty surgery. Application of a targeted tuf gene sequencing approach provided auxiliary staphylococcal community profiles and allowed species-level characterisation directly from low biomass clinical samples.

Cohort description: The Danish Blood Donor Staphylococcus aureus Carriage Study.

PURPOSE: Staphylococcus aureus carriage poses an increased risk of S. aureus infection. The aim of this study was to investigate the colonization of S. aureus among healthy individuals and to establish a prospective cohort and biobank for research in the health consequences of colonization. POPULATION AND METHODS: The Danish Blood Donor S. aureus Carriage Study (DBDSaCS) was established in 2014. So far, a total of 6082 healthy participants have been included with nasal swabs and repeated swabs are performed at subsequent donations. Samples from the first 2217 participants were cultured using a two-step method to evaluate the effect of using enrichment broth. Furthermore, 262 participants were sampled from both the nares and the throat. All participants completed a questionnaire with self-reported health, anthropometric measurements, current smoking status, and physical activity. Plasma samples, nasal swab transport media, and S. aureus isolates were stored. RESULTS: The prevalence of S. aureus nasal colonization was 41%. The prevalence of colonization was higher in men (46%) than women (34%), lower for smokers, and decreased with increasing age (<25 years: 44% vs >55 years: 35%). In participants swabbed from the nose and throat, the prevalence of S. aureus colonization after enrichment was 55% with significantly higher prevalence in the throat (45%) than in the nose (40%). The use of an enrichment broth increased the proportion of S. aureus colonization. CONCLUSION: We describe a large and growing cohort of healthy individuals established to investigate predictors for S. aureus carriage and the health consequences of carriage. Multiple projects using data from DBDSaCS linked with Danish health registers, biomarkers, and genetic markers are ongoing. Results will be published in the coming years.

Genetically determined high activities of the TNF-alpha, IL23/IL17, and NFkB pathways were associated with increased risk of ankylosing spondylitis.

BACKGROUND: Ankylosing spondylitis (AS) results from the combined effects of susceptibility genes and environmental factors. Polymorphisms in genes regulating inflammation may explain part of the heritability of AS. METHODS: Using a candidate gene approach in this case-control study, 51 mainly functional single nucleotide polymorphisms (SNPs) in genes regulating inflammation were assessed in 709 patients with AS and 795 controls. Data on the patients with AS were obtained from the DANBIO registry where patients from all of Denmark are monitored in routine care during treatment with conventional and biologic disease modifying anti-rheumatic drugs (bDMARDs). The results were analyzed using logistic regression (adjusted for age and sex). RESULTS: Nine polymorphisms were associated with risk of AS (p < 0.05). The polymorphisms were in genes regulating a: the TNF-α pathway (TNF -308 G > A (rs1800629), and - 238 G > A (rs361525); TNFRSF1A -609 G > T (rs4149570), and PTPN22 1858 G > A (rs2476601)), b: the IL23/IL17 pathway (IL23R G > A (rs11209026), and IL18-137 G > C (rs187238)), or c: the NFkB pathway (TLR1 743 T > C (rs4833095), TLR4 T > C (rs1554973), and LY96-1625 C > G (rs11465996)). After Bonferroni correction the homozygous variant genotype of TLR1 743 T > C (rs4833095) (odds ratios (OR): 2.59, 95% confidence interval (CI): 1.48-4.51, p = 0.04), and TNFRSF1A -609 G > T (rs4149570) (OR: 1.79, 95% CI: 1.31-2.41, p = 0.01) were associated with increased risk of AS and the combined homozygous and heterozygous variant genotypes of TNF -308 G > A (rs1800629) (OR: 0.56, 95% CI: 0.44-0.72, p = 0.0002) were associated with reduced risk of AS. CONCLUSION: We replicated associations between AS and the polymorphisms in TNF (rs1800629), TNFRSF1A (rs4149570), and IL23R (rs11209026). Furthermore, we identified novel risk loci in TNF (rs361525), IL18 (rs187238), TLR1 (rs4833095), TLR4 (rs1554973), and LY96 (rs11465996) that need validation in independent cohorts. The results suggest that genetically determined high activity of the TNF-α, IL23/IL17, and NFkB pathways increase risk of AS.

Genetic polymorphisms associated with psoriasis and development of psoriatic arthritis in patients with psoriasis.

BACKGROUND: Psoriasis (PsO) is a chronic inflammatory disease with predominantly cutaneous manifestations. Approximately one third of patients with PsO develop psoriatic arthritis (PsA), whereas the remaining proportion of patients has isolated cutaneous psoriasis (PsC). These two phenotypes share common immunology, but with different heredity that might in part be explained by genetic variables. METHODS: Using a candidate gene approach, we studied 53 single nucleotide polymorphisms (SNPs) in 37 genes that regulate inflammation. In total, we assessed 480 patients with PsO from DERMBIO, of whom 151 had PsC for 10 years or more (PsC10), 459 patients with PsA from DANBIO, and 795 healthy controls. Using logistic regression analysis, crude and adjusted for age and gender, we assessed associations between genetic variants and PsO, PsC10, and PsA, as well as associations between genetic variants and development of PsA in PsO. RESULTS: Eleven polymorphisms in 10 genes were nominally associated with PsO and/or PsC and/or PsA (P < 0.05). After correction for multiple testing with a false discovery rate of 5%, two SNPs remained significant: TNF (rs361525) was associated with PsO, PsC10, and PsA; and IL12B (rs6887695) was associated with PsO. CONCLUSION: Among a cohort of Danish patients with moderate-to-severe psoriasis, two SNPs in the IL12B and TNF genes were associated with susceptibility of psoriasis. None of the SNPs were specifically associated with isolated cutaneous psoriasis or psoriatic arthritis.

In vivo expression of antimicrobial peptides in atopic dermatitis.

The aim of this review is to present findings on expression of antimicrobial peptides (AMPs) in atopic dermatitis (AD) skin, focusing only on in vivo studies, and to discuss differences in results obtained using various skin sampling techniques and different methodology for analysis of AMPs. The review also includes a discussion of the effect of frequently used treatments on AMP expression. Many studies have shown a reduced level of AMPs in lesional AD skin when compared to psoriatic skin, explaining the high frequency of AD-related infections. Interestingly, however, non-lesional AD skin has shown the same upregulation of AMPs after barrier disruption as non-lesional psoriatic skin. Various methods have been used to analyse AMP expression in the skin, and when comparing these methods, differences are revealed in AMP expression depending on the method used for sampling and analysis. Comparisons indicate that analyses of mRNA levels of AMPs may find greater differences in expression than analyses of protein levels. Few studies evaluate the effect of topical treatments on the expression of AMPs, and these indicate an inhibition of AMP expression, particularly after use of corticosteroids. AMPs are important components of the skin as a defense against infections, and despite much research, the clinical importance of the effect of common treatments, including systemic treatments for AD and the interplay between AMPs and the skin microbiome, is still largely unknown.

Polymorphisms in the Toll-Like Receptor and the IL-23/IL-17 Pathways Were Associated with Susceptibility to Inflammatory Bowel Disease in a Danish Cohort.

BACKGROUND: The inflammatory bowel diseases (IBD), Crohn's disease (CD) and ulcerative colitis (UC), result from the combined effects of susceptibility genes and environmental factors. Previous studies have shown that polymorphisms in the Toll-like receptor (TLR), the apoptosis, the IL-23/IL-17 and the interferon gamma (IFNG) pathways are associated with risk of both CD and UC. METHODS: Using a candidate gene approach, 21 functional single nucleotide polymorphisms (SNPs) in 15 genes were assessed in a clinical homogeneous group of severely diseased ethnic Danish patients consisting of 624 patients with CD, 411 patients with UC and 795 controls. The results were analysed using logistic regression. RESULTS: The polymorphisms TLR5 (rs5744174) and IL12B (rs6887695) were associated with risk of CD, and TLR1 (rs4833095) and IL18 (rs187238) were associated with risk of both CD and UC (p<0.05). After Bonferroni correction for multiple testing, the homozygous variant genotype of TLR1 743 T>C (rs4833095) was associated with increased risk CD (OR: 3.15, 95% CI: 1.59-6.26, p = 0.02) and CD and UC combined (OR: 2.96, 95% CI: 1.64-5.32, p = 0.005). CONCLUSION: Our results suggest that genetically determined high activity of TLR1 and TLR5 was associated with increased risk of both CD and UC and CD, respectively. This supports that the host microbial composition or environmental factors in the gut are involved in risk of IBD. Furthermore, genetically determined high activity of the IL-23/IL-17 pathway was associated with increased risk of CD and UC. Overall, our results support that genetically determined high inflammatory response was associated with increased risk of both CD and UC.

Genetic Variations in Pattern Recognition Receptor Loci Are Associated with Anti-TNF Response in Patients with Rheumatoid Arthritis.

OBJECTIVES: To determine whether genetic variation within genes related to the Toll-like receptor, inflammasome and interferon-γ pathways contributes to the differences in treatment response to tumour necrosis factor inhibitors (anti-TNF) in patients with rheumatoid arthritis (RA). METHODS: In a retrospective case-case study, we assessed 23 functional single nucleotide polymorphisms (SNPs) in 15 genes. We included 538 anti-TNF naïve Danish RA patients from the nationwide DANBIO database. Multivariable logistic regression analyses were performed to detect associations (p-value<0.05) between genotypes and European League Against Rheumatism (EULAR) treatment responses. False Discovery Rate corrections for multiple testing (q-value) and stratified analyses were performed to investigate association with individual therapies and IgM-rheumatoid factor (RF) status. RESULTS: Six of twenty successfully genotyped polymorphisms were nominally associated with EULAR treatment response. Three of these were in weak to moderate linkage disequilibrium with polymorphisms previously reported associated with anti-TNF treatment response. TLR5(rs5744174) variant allele carriers (odds ratio(OR) = 1.7(1.1-2.5),p = 0.010,q = 0.46) and TLR1(rs4833095) homozygous variant carriers (OR = 2.8(1.1-7.4),p = 0.037,q = 0.46) had higher odds for a positive treatment response. NLRP3(rs10754558) variant allele carriers (odds ratio(OR) = 0.6(0.4-1.0),p = 0.045,q = 0.46) were more likely to have a negative treatment response. The association in TLR5(rs5744174) remained significant after correction for multiple comparisons among patients negative for RF (OR = 6.2(2.4-16.3),p = 0.0002,q = 0.024). No other association withstood correction for multiple testing. Post hoc analyses showed that change in Patient Global score on a visual analogue scale (VAS) and change in pain VAS were the main factors responsible for the association. CONCLUSIONS: We reproduced previously reported associations between genetic variation in the TLR10/1/6 gene cluster, TLR5, and NLRP3 loci and response to anti-TNF treatment in RA. Changes in VAS pain and patient global scores were the main contributors to the association found for TLR5. Furthermore, we identified other candidate genes that require replication in independent cohorts.

Effectiveness of anti-tumour necrosis factor-α therapy in Danish patients with inflammatory bowel diseases.

INTRODUCTION: The objective of this study was to evaluate the outcome of anti-tumour necrosis factor-α (anti-TNF) treatment in a large cohort of patients with inflammatory bowel diseases (IBD), including Crohn's disease (CD) and ulcerative colitis (UC) in clinical practice and to establish a cohort for future studies of genetic markers associated with treatment response. METHODS: A national, clinically based cohort of previously naïve anti-TNF treated patients from 18 medical departments was established. The patients were screened for tuberculosis prior to treatment initiation. By combining the unique personal identification number of Danish citizens (the CPR number) from blood samples with data from the National Patient Registry, patients with International Classification of Diseases, Version 10 (ICD-10) codes K50-K63 were identified. Treatment efficacy reflected the maximum response within 22 weeks. RESULTS: Among 492 patients with CD and 267 patients with UC, 74%/13%/14% and 65%/12%/24% were responders, partial responders and non-responders to anti-TNF therapy, respectively. More patients with UC than with CD were non-responders (odds ratio (OR) = 1.96, 95% confidence interval (CI): 1.34-2.87, p = 0.001). Young age was associated with a beneficial response (p = 0.03), whereas smoking ≥ 10 cigarettes/day was associated with non-response among patients with CD (OR = 2.33, 95% CI: 1.13-4.81, p = 0.03). CONCLUSION: In this clinically based cohort of Danish patients with IBD treated with anti-TNF, high response rates were found. Heavy smoking was associated with non-response, whereas young age at treatment initiation was associated with a beneficial response among patients with CD. Thus, the results obtained in this cohort recruited from clinical practice were similar to those previously obtained in clinical trials. FUNDING: The work was funded by Health Research Fund of Central Denmark Region, Colitis-Crohn Foreningen and the University of Aarhus (PhD grant). TRIAL REGISTRATION: Clinicaltrials NCT02322008.

Anti-TNF treatment response in rheumatoid arthritis patients is associated with genetic variation in the NLRP3-inflammasome.

OBJECTIVE: Many patients with rheumatoid arthritis (RA) benefit from tumor necrosis factor-α blocking treatment (anti-TNF), but about one third do not respond. The objective of this study was to replicate and extend previously found associations between anti-TNF treatment response and genetic variation in the TNF-, NF-κB- and pattern recognition receptor signalling pathways. METHODS: Forty-one single nucleotide polymorphisms (SNPs), including 34 functional, in 28 genes involved in inflammatory pathways were assessed in 538 anti-TNF naive Danish RA patients with clinical data. Multivariable logistic regression analyses were performed to test associations between genotypes and treatment response at 3-6 months using the European League Against Rheumatism (EULAR) response criterion. American College of Rheumatology treatment response (ACR50) and relative change in 28-joint disease activity score (relDAS28) were used as secondary outcomes. Subgroup analyses were stratified according to smoking status, type of anti-TNF drug and IgM-Rheumatoid Factor (IgM-RF) status. False discovery rate (FDR) controlling was used to adjust for multiple testing. RESULTS: Statistically significant associations with EULAR response were found for two SNPs in NLRP3(rs4612666) (OR (odds ratio) for good/moderate response = 0.64 (95% confidence interval: 0.44-0.95), p = 0.025, q = 0.95) and INFG(rs2430561) (OR = 0.40 (0.21-0.76), p = 0.005, q = 0.18) and among IgM-RF positive patients for TNFRS1A(rs4149570) (0.59 (0.36-0.98), p = 0.040, q = 0.76). Current smokers who carried the NLRP3(rs4612666) variant allele were less likely to benefit from anti-TNF treatment (OR = 0.24 (0.10-0.56), p = 0.001, q = 0.04). CONCLUSIONS: In a population of Danish RA patients, we confirm the NLRP3 gene as associated with EULAR anti-TNF response as previously reported. The NLRP3 variant (T) allele is associated with lower treatment response, in particular among current smokers. Furthermore, we find that a functional polymorphism in the interferon-γ gene is associated with anti-TNF response. All findings should be tested by replication in independent validation cohorts and augmented by assessing cytokine levels and activities of the relevant gene products.

Alpha-defensin DEFA1A3 gene copy number elevation in Danish Crohn's disease patients.

BACKGROUND AND PURPOSE OF STUDY: Extensive copy number variation is observed for the DEFA1A3 gene encoding alpha-defensins 1-3. The objective of this study was to determine the involvement of alpha-defensins in colonic tissue from Crohn's disease (CD) patients and the possible genetic association of DEFA1A3 with CD. METHODS: Two-hundred and forty ethnic Danish CD patients were included in the study. Reverse transcriptase PCR assays determined DEFA1A3 expression in colonic tissue from a subset of patients. Immunohistochemical analysis identified alpha-defensin peptides in colonic tissue. Copy number of DEFA1A3 and individual alleles, DEFA1 and DEFA3, were compared with those for controls, by use of combined real-time quantitative PCR and pyrosequencing, and correlated with disease location. RESULTS: Inflammatory-dependent mRNA expression of DEFA1A3 (P < 0.001), and the presence of alpha-defensin peptides, were observed in colonic tissue samples. Higher DEFA1A3 gene copy number (CD: mean copy number, 7.2 vs. controls 6.7; P < 0.001) and individual DEFA1 alleles (CD mean copy number 5.6 vs. controls 5.1; P < 0.01) were associated with CD, with strong association with colonic location (P < 0.001). CONCLUSIONS: Alpha-defensins are involved in the inflammation of CD, with local mRNA and peptide expression. In combination with the findings that a high DEFA1A3 copy number is significantly linked to CD, these results suggest that a high DEFA1A3 copy number might be important in hindering the normal inflammatory response in CD, particularly colonic CD.

Infliximab dependency is related to decreased surgical rates in adult Crohn's disease patients.

BACKGROUND: Infliximab dependency in children with Crohn's disease (CD) has recently been described and found to be associated with a decreased surgery rate. AIM: To assess infliximab dependency of adult CD patients, evaluate the impact on surgery, and search for possible clinical and genetic predictors. METHODS: Two hundred and forty-five CD patients treated with infliximab were included from Danish and Czech Crohn Colitis Database (1999-2006). Infliximab response was assessed as immediate outcome, 1 month after infliximab start: complete, partial, and no response. Three months outcome, after last intended infusion: prolonged response (maintenance of complete/partial response), infliximab dependency (relapse requiring repeated infusions to regain complete/partial response or need of infliximab >12 months to sustain response). RESULTS: Forty-seven percent obtained prolonged response, 29% were infliximab dependent and 24% nonresponders. The cumulative probability of surgery 40 months after infliximab start was 20% in prolonged responders, 23% in infliximab-dependent patients and 76% in nonresponders (P<0.001). The cumulative probability of surgery at 40 months in patients on maintenance versus on demand regime was 33 and 31%, respectively (P=0.63). No relevant clinical or genetic predictors were identified. CONCLUSION: The infliximab dependency response seems to be equivalent to the prolonged response in adult CD patients when comparing surgery rates.

Fabry disease mimicking hypertrophic cardiomyopathy: genetic screening needed for establishing the diagnosis in women.

AIMS: Fabry disease, an X-linked storage disorder caused by defective lysosomal enzyme alpha-galactosidase A activity, may resemble sarcomere-gene-associated hypertrophic cardiomyopathy (HCM). The 'cardiac variant' of Fabry disease which only affects the heart may be missed unless specifically tested for. METHODS AND RESULTS: We evaluated 90 consecutively recruited HCM probands and their relatives. Probands without sarcomere-gene mutations were tested for alpha-galactosidase A gene (GLA) mutations. Of the 90 families, 31 (34%) had sarcomere gene mutations and were therefore excluded. In the remaining 59 probands, 3 (5%) had GLA mutations as follows. The first proband, a female with asymmetric septal hypertrophy (ASH), a significant left ventricular outflow tract gradient, and chronic obstructive pulmonary disease, was heterozygous for a novel missense mutation (p.N139S). The second proband, a male with ASH and multiple episodes of ventricular tachycardia, was hemizygous for a missense mutation (p.A156T). His daughter was heterozygous, but had normal enzyme activity. The third proband was a female with ASH, and no other indices of Fabry disease. She was heterozygous for a GLA missense mutation (p.G271S). She had one affected daughter but her two other children were unaffected. The affected daughter had three children, of whom two were also affected--a boy aged 8 and a daughter aged 10 years. CONCLUSION: This is the first report of systematic mutation screening of GLA in HCM patients without sarcomere gene mutations. GLA mutations were found in 3/90 (3%) of HCM families and in 2/20 (10%) of females without sarcomere-gene mutations. None of the probands presented other indices of Fabry disease. This, in combination with putative reversibility of cardiac changes by enzyme replacement therapy, supports systematic testing for Fabry disease. Enzyme measurements are sufficient in men, but genetic testing is needed in women.

HLA-A alleles and infectious mononucleosis suggest a critical role for cytotoxic T-cell response in EBV-related Hodgkin lymphoma.

A proportion of classical Hodgkin lymphoma (HL) is believed to be causally related to infection with the ubiquitous lymphotropic EBV. The determining factors for development of EBV-related HL remain poorly understood, but likely involve immunological control of the viral infection. Accordingly, markers of the HLA class I region have been associated with risk of EBV-related HL. To study the host genetic component of EBV-related HL further, we investigated the lymphoma's association with HLA-A*01 and HLA-A*02 simultaneously in the setting of infectious mononucleosis (IM), a risk factor for EBV-related HL, in a case-series analysis including 278 EBV-related and 656 EBV-unrelated cases of HL. By logistic regression, HLA-A*01 alleles [odds ratio (OR) per allele, 2.15; 95% CI, 1.60-2.88] were associated with increased and HLA-A*02 alleles (OR per allele, 0.70; 95% CI, 0.51-0.97) with decreased risk of EBV-related HL. These allele-specific associations corresponded to nearly 10-fold variation in risk of EBV-related HL between HLA-A*01 and HLA-A*02 homozygotes. History of IM was also associated with risk of EBV-related HL (OR, 3.40; 95% CI, 1.74-6.66). The association between history of IM and EBV-related HL was not seen in the presence of HLA-A*02 because this allele appeared to neutralize the effect of IM on EBV-related HL risk. Our findings suggest that HLA class I-restricted EBV-specific cytotoxic T-cell responses and events in the early immune response to EBV infection in IM play critical roles in the pathogenesis of EBV-related HL.

Associations between COX-2 polymorphisms, blood cholesterol and risk of acute coronary syndrome.

BACKGROUND: The use of specific COX-2 inhibitors in cancer prevention has been associated with higher risk of acute coronary syndrome (ACS) and myocardial infarction. The aim of this study was to investigate if the polymorphisms COX2 T8473C (rs5275), and COX2 A-1195G (rs689466), which modify the enzyme levels of COX-2, were associated with risk of ACS and if alcohol intake, smoking, and use of NSAID would modify the associations. We also wanted to investigate associations with blood lipid levels. METHODS: A case-cohort study including 1031 ACS cases and a sub-cohort of 1703 persons was nested within the population-based prospective study Diet, Cancer and Health of 57,053 individuals aged 55-64 at recruitment 1993-1997. RESULTS: Male variant allele carriers of COX-2 T8473C were at lower risk of ACS (IRR=0.75, CI=0.61-0.93, p=0.008) than homozygous wildtype carriers. There were no statistically significant interactions between genotypes and alcohol intake, smoking and NSAID use in relation to risk of ACS. Among males, there was interaction between COX-2 T8473C and alcohol in relation to total cholesterol, non-HDL cholesterol and LDL levels (p for interaction: 0.003, 0.007 and 0.01, respectively), such that variant allele carriers with low alcohol intake had the lowest lipid levels. No statistically significant associations were observed in females. CONCLUSION: This study suggests that genetically determined COX-2 levels are associated with risk of ACS and blood lipid levels among males. No consistent associations were found for females.

Polymorphisms in inflammation genes, tobacco smoke and furred pets and wheeze in children.

Persistent wheeze in childhood is associated with airway inflammation. The present study investigated relationships between polymorphisms in inflammatory genes, exposure to tobacco smoke and furred pets and risk of recurrent wheeze in children. Within a birth cohort of 101,042 children we identified 1111 eighteen month old cases with recurrent wheeze and 735 wheeze-free controls among 11942 children recruited in the Copenhagen area. Polymorphisms in IL-4R, IL-8, IL-13, SPINK5, and CD14 were genotyped. Interviews at gestational wks 12 and 30, and at age 6 and 18 months included questions on number of episodes with wheeze (18 months), exposure to tobacco smoke and pet-keeping. Recurrent wheeze was defined as at least four episodes of wheeze before the child was 18 months old. There was a statistically significant association between the IL-13 Arg144Gln polymorphism and risk of recurrent wheeze (p = 0.01). Furthermore, there was a statistically significant interaction between this polymorphism and exposure to tobacco smoke during pregnancy, though this was probably a chance finding. There were no other statistically significant effects of the polymorphisms or interactions with exposure to tobacco smoke in relation to the risk of recurrent wheeze. Polymorphisms in IL-8 affected the association between pet-keeping and risk of wheeze. Polymorphisms in inflammation genes might affect the association between environmental exposures and risk of recurrent wheeze in early childhood.

PPARgamma Pro12Ala polymorphism and risk of acute coronary syndrome in a prospective study of Danes.

BACKGROUND: Acute coronary syndrome (ACS) is a major cause of morbidity and mortality in the western world. Peroxisome proliferator-activated receptor gamma (PPARgamma) plays a key role in the regulation of the energy balance, adipocyte differentiation and lipid biosynthesis. The aim was to investigate if the polymorphism PPARgamma2 Pro12Ala, which encodes a less efficient transcription factor, was associated with risk of acute coronary disease and if there were interactions between this polymorphism and factors that modify PPARgamma activity, such as alcohol intake, smoking, and use of non-steroidal anti-inflammatory medicine. METHODS: A case-cohort study including 1031 ACS cases and a sub-cohort of 1703 persons was nested within the population-based prospective study Diet, Cancer and Health of 57,053 individuals. RESULTS: Homozygous male variant allele carriers of PPARgamma2 Pro12Ala were at higher risk of ACS (HR = 2.12, 95% CI: 1.00-4.48) than homozygous carriers of the Pro-allele. Among men, there was a statistically significant interaction between genotypes and alcohol intake such that homozygous variant allele carriers with a low alcohol intake were at higher risk of ACS (HR = 25.3, CI: 16.5-38.7) compared to homozygous common allele carriers (p for interaction < 0.0001). Overall, the association was only observed among homozygous variant allele carriers. Thus, all the observed associations were obtained in subgroups including small numbers of cases. It is therefore possible that the observed associations were due to chance. CONCLUSION: In the present study, there were no consistent associations between PPARgamma Pro12Ala and risk of ACS, and no consistent interaction with alcohol, BMI, NSAID or smoking in relation to ACS.

Diagnostic yield, interpretation, and clinical utility of mutation screening of sarcomere encoding genes in Danish hypertrophic cardiomyopathy patients and relatives.

The American Heart Association (AHA) recommends family screening for hypertrophic cardiomyopathy (HCM). We assessed the outcome of family screening combining clinical evaluation and screening for sarcomere gene mutations in a cohort of 90 Danish HCM patients and their close relatives, in all 451 persons. Index patients were screened for mutations in all coding regions of 10 sarcomere genes (MYH7, MYL3, MYBPC3, TNNI3, TNNT2, TPM1, ACTC, CSRP3, TCAP, and TNNC1) and five exons of TTN. Relatives were screened for presence of minor or major diagnostic criteria for HCM and tracking of DNA variants was performed. In total, 297 adult relatives (>18 years) (51.2%) fulfilled one or more criteria for HCM. A total of 38 HCM-causing mutations were detected in 32 index patients. Six patients carried two disease-associated mutations. Twenty-two mutations have only been identified in the present cohort. The genetic diagnostic yield was almost twice as high in familial HCM (53%) vs. HCM of sporadic or unclear inheritance (19%). The yield was highest in families with an additional history of HCM-related clinical events. In relatives, 29.9% of mutation carriers did not fulfil any clinical diagnostic criterion, and in 37.5% of relatives without a mutation, one or more criteria was fulfilled. A total of 60% of family members had no mutation and could be reassured and further follow-up ceased. Genetic diagnosis may be established in approximately 40% of families with the highest yield in familial HCM with clinical events. Mutation-screening was superior to clinical investigation in identification of individuals not at increased risk, where follow-up is redundant, but should be offered in all families with relatives at risk for developing HCM.

Rapid tumour-like growth of giant filiform polyposis in a patient without a history of chronic bowel inflammation.

Long-term inflammation of the colonic mucosa during chronic inflammatory bowel disease with alternating periods of ulceration and healing may lead to the formation of finger-like projections, so-called filiform polyps. In rare cases, several filiform polyps form large tumour masses, termed giant filiform polyposis. We present a case of giant obstructing filiform polyposis in a patient without previous evidence of chronic bowel inflammation. The resected ascending colon from a 37-year-old woman was evaluated macroscopically and microscopically, and the presence of gene polymorphisms was studied by means of multiplex capillary electrophoresis single-strand conformation polymorphism assay, DNA sequencing, TaqMan analysis, and restriction enzyme cleavage. The giant filiform polyposis was restricted to a 15 cm segment of the ascending colon, and the remaining colonic mucosa was entirely without inflammatory changes. During the post-operative follow-up period, the patient developed symptoms and signs of distal bowel inflammation. Gene polymorphism studies were inconclusive as to Crohn's disease. In conclusion, we present an unusual pathological entity of giant filiform polyposis, which developed relatively rapidly in a colon without any history or macroscopic changes suggestive of chronic inflammatory bowel disease. Although the patient subsequently developed symptoms in keeping with Crohn's disease, studies of genetic polymorphism were unable to confirm this notion, and colorectal tissue has not been sampled postoperatively for histological evaluation.

Disease concordance, zygosity, and NOD2/CARD15 status: follow-up of a population-based cohort of Danish twins with inflammatory bowel disease.

OBJECTIVES: A Danish cohort of twins with inflammatory bowel disease (IBD), Crohn's disease (CD) and ulcerative colitis (UC), has previously been collected. The aim of the present study was to reassess this cohort in order to compare clinical characteristics in concordant versus discordant twin pairs, test twin zygosity genetically, follow-up on disease concordance, and examine NOD2/CARD15 genetic status. METHODS: The Danish cohort is one of two population-based cohorts worldwide and consists of 103 twin pairs. After median 13 yr of follow-up, all twins were contacted and hospital files were scrutinized to reassess disease concordance and obtain phenotype data. DNA was obtained from 123 twins for analysis of zygosity and prevalence of the three common NOD2/CARD15 mutations. RESULTS: Zygosity tested genetically was consistent with the former assessment based on questionnaires. The proband concordance for CD remained fairly stable: 63.6% among monozygotic (MZ) twins and 3.6% among dizygotic (DZ) twins. Clinical characteristics were similar in twins from concordant versus discordant pairs. Forty-four percent of patients with CD were positive for >or=1 mutant allele of NOD2/CARD15 compared to 2% of UC patients (p < 0.001) and 19% of healthy twins (p= 0.02). The allele mutation frequency was 43% among the healthy twins to patients with CD versus 9% among twins to UC patients (p= 0.01). CONCLUSIONS: Previous questionnaire assessment of twin zygosity was confirmed by genetic test. Concordance for CD remained quite stable and was significantly higher among MZ than DZ twins. A high NOD2/CARD15 mutation frequency was observed both among CD twins and their healthy siblings.