No associations between single nucleotide polymorphisms in corticoid receptor genes and heart rate and cortisol responses to a standardized social stress test in adolescents: the TRAILS study.

Previously, sequence variation in the glucocorticoid (GR) and mineralocorticoid (MR) receptor genes (NR3C1 and NR3C2, respectively) have been found to be associated with physiological stress responses to social stress tests in small samples of adult men and oral contraceptives (OC) using women. Associations between single nucleotide polymorphisms (SNPs) in the GR (23EK-rs6190, 9beta-rs6198, BclI-rs4142324) and the MR gene (I180V-rs5522 and -2G/C (rs2070951) with cortisol and heart rate responses to a performance-related social stress task (public speaking and mental arithmetic) were examined in a large sample (n = 553) of adolescents (15-17 years). To make comparisons with previous findings, associations were tested in boys (n = 277), free-cycling (FC) girls (n = 183) and OC users (n = 93). None of the previously reported associations in adults could be replicated in this large adolescent sample. Explanations for non-replication are discussed.

No increased susceptibility to breast cancer from combined CHEK2 1100delC genotype and the HLA class III region risk factors.

CHEK2 is low-penetrance breast cancer susceptibility gene. The 1100delC mutation may interact with variants/mutations in other breast cancer susceptibility loci. We identified a risk haplotype in the HLA class III region in breast cancer patients [de Jong MM, Nolte IM, de Vries EGE, et al. The HLA class III subregion is responsible for an increased breast cancer risk. Hum Mol Genet 2003, 12, 2311-2319] and tested whether it interacted with 1100delC mutation. The CHEK2 1100delC mutation was analysed in the same series of patients and controls as in the HLA breast cancer study. In 962 unselected breast cancer patients, the 1100delC mutation was observed in 2.9% and in 367 controls in 1.4% (NS). The highest 1100delC frequency occurred in high-risk (4.4%), followed by moderate-risk (3.8%), and lowest in low genetic risk patients (2.4%, P(trend) 0.029). In HLA risk haplotype carriers no increased breast cancer risk was observed in the presence of 1100delC mutation. Patients more often had one than both genetic risk factors. The 1100delC mutation and the HLA risk haplotype confer increased breast cancer risks, but an interactive effect on breast cancer between both factors is unlikely. In contrast, the effect of 1100delC mutation on breast cancer risk was limited to individuals without HLA risk haplotype, suggesting a mutual excluding effect between these risk factors.

Association between Toll-like receptor 4 and inflammatory bowel disease.

BACKGROUND: The human Toll-like receptor 4 (TLR4) participates in the innate response. Recently, the TLR4 variant Asp299Gly has been described to affect the response of this receptor to lipopolysaccharide. As such, there is a potentially important role of TLR4 in the pathogenesis of inflammatory bowel disease (IBD). We studied the involvement of TLR4 in IBD in a large population of Dutch patients with IBD and in family-based controls. METHODS: In 781 IBD cases and 315 controls, genotyping was performed forAsp299Gly and Thr399Ile variants and for 4 microsatellite markers flanking TLR4. Association analysis and the were applied. In addition, interaction of TLR4 with the caspase recruitment domain containing protein 15 gene (CARD15) was studied in patients with Crohn's disease (CD). RESULTS: The haplotype sharing statistic showed association at microsatellite marker D9S1864 with IBD (P = 0.0019), and in particular with CD (P = 0.0025) and at TLR406 with ulcerative colitis (UC; P = 0.027). No association was found for Asp299Gly and Thr399Ile. However, the frequencies of both variant allele carriers were higher among CD cases with a disease onset > or = 40 years than among controls. No evidence for interaction between TLR4 and CARD15 was found. CONCLUSIONS: Haplotype analysis shows that TLR4 is associated with both CD and UC. The Asp299Gly and Thr399Ile variants do not show an association with CD, UC, or IBD as a group, indicating that these polymorphisms are likely not the causal ones. We propose that the 2 polymorphisms are in linkage with (the) disease susceptibility variant(s) located elsewhere on TLR4.

Influence of common variants near INSIG2, in FTO, and near MC4R genes on overweight and the metabolic profile in adolescence: the TRAILS (TRacking Adolescents' Individual Lives Survey) Study.

BACKGROUND: Overweight is a complex trait in which both environmental and genetic factors play a role. OBJECTIVE: We aimed to evaluate the influence of common genetic variants identified by genome-wide association studies on overweight and the metabolic profile in adolescence. DESIGN: In a population-based cohort of 663 girls and 612 boys aged 16 y, weight, height, skinfold thicknesses, percentage body fat, waist circumference, blood pressure, glucose, insulin, lipid profile, and DNA were obtained. We defined overweight according to international criteria. We performed multiple linear and logistic regression analyses to assess the influence of candidate single nucleotide polymorphisms near the INSIG2, in the FTO, and near the MC4R genes and repeated-measures analyses of available body mass index (BMI) and skinfold thickness data across 3 visits at ages 11, 13.5, and 16 y. RESULTS: A total of 15.1% of participants were overweight or obese at age 16 y. No associations with INSIG2 were found. Common variation in the FTO gene was associated with sex-specific z scores of BMI (B: 0.11; 95% CI: 0.03, 0.19), sum of skinfold thicknesses (B: 0.12; 95% CI: 0.04, 0.20), percentage body fat (B: 0.11; 95% CI: 0.03, 0.19), waist circumference (B: 0.11; 95% CI: 0.03, 0.19), fasting glucose (B: 0.10; 95% CI: 0.00, 0.20), and overweight (odds ratio: 1.34; 95% CI: 1.06, 1.69) at age 16 y. Repeated-measures analyses confirmed the associations for BMI and sum of skinfold thicknesses, and physical activity did not modify these associations. Common variation near the MC4R gene was associated with BMI in cross-sectional (B: 0.11; 95% CI: 0.02, 0.20) and repeated-measures (B: 0.12; 95% CI: 0.03, 0.20) analyses. CONCLUSIONS: Common variation in the FTO gene is associated with overall and abdominal adiposity. Variation near the MC4R gene is associated with BMI. These findings in adolescents strengthen and extend the results from previous research.

Multiple common variants for celiac disease influencing immune gene expression.

We performed a second-generation genome-wide association study of 4,533 individuals with celiac disease (cases) and 10,750 control subjects. We genotyped 113 selected SNPs with P(GWAS) < 10(-4) and 18 SNPs from 14 known loci in a further 4,918 cases and 5,684 controls. Variants from 13 new regions reached genome-wide significance (P(combined) < 5 x 10(-8)); most contain genes with immune functions (BACH2, CCR4, CD80, CIITA-SOCS1-CLEC16A, ICOSLG and ZMIZ1), with ETS1, RUNX3, THEMIS and TNFRSF14 having key roles in thymic T-cell selection. There was evidence to suggest associations for a further 13 regions. In an expression quantitative trait meta-analysis of 1,469 whole blood samples, 20 of 38 (52.6%) tested loci had celiac risk variants correlated (P < 0.0028, FDR 5%) with cis gene expression.

Runt-related transcription factor 3 is associated with ulcerative colitis and shows epistasis with solute carrier family 22, members 4 and 5.

BACKGROUND: Inflammatory bowel disease (IBD), comprising Crohn's disease (CD) and ulcerative colitis (UC), are intestinal inflammatory disorders with a complex genetic background. Mice deficient for the runt-domain-transcription-factor3 (Runx3) develop spontaneous colitis. Human RUNX3 resides in an IBD-susceptibility locus. We studied the association of RUNX3 in a cohort of IBD patients and analyzed the interaction with SLC22A4/5. RUNX3 and OCTN1 mRNA expression was assessed in inflamed and noninflamed mucosa from patients and controls. METHODS: 543 IBD patients (309 CD / 234 UC) and 296 controls were included. Four single nucleotide polymorphisms (SNPs) and 4 microsatellite markers were studied for RUNX3. Five SNPs (including SNP-207G-->C and SNP1672C-->T) were analyzed for SLC22A4/5. RUNX3, and OCTN1 expression in mucosal tissue from 30 patients (14 UC / 16 CD) and 6 controls were determined by quantitative polymerase chain reaction. RESULTS: A significant association between RUNX3-SNP rs2236851 and UC (OR 1.61; 95% confidence interval [CI] 1.11-2.32, P = 0.020) was found. Carriership is associated with pancolitis (odds ratio [OR] 1.86; 95% CI 1.08-3.21). SLC22A4/5-SNPs rs272893 and rs273900 are associated with CD (OR 2.16; 95% CI 1.21-3.59 and OR 2.40; 95% CI 1.43-4.05). We found epistasis for carriership of a risk-associated allele in RUNX3 and SLC22A4/5 for UC patients versus CD patients (OR 3.83; 95% CI 1.26-11.67). RUNX3 mRNA expression is increased (P = 0.01) in inflamed colonic mucosa of UC patients compared to noninflamed mucosa and controls. CONCLUSIONS: We provide evidence for the genetic association of RUNX3 with UC and for CD with the IBD5 locus including SLC22A4/5. An epistatic effect of RUNX3 and SLC22A4 was associated with an increased risk for UC. Our data suggest a role for RUNX3 in UC susceptibility.

Association of interleukin-1 receptor-associated kinase M (IRAK-M) and inflammatory bowel diseases.

OBJECTIVE: Inflammatory bowel diseases (IBD) have a complex genetic background. The interleukin receptor associated kinase-M (IRAK-M) is a NF-kappaB-mediated, negative regulator of Toll-like receptor (TLR) signaling. A functional mutation in a negative regulator might induce impaired endotoxin tolerance and increased inflammatory responses. IRAK-M is situated on chromosome 12q14, a susceptibility locus for IBD, which makes it a good candidate gene. The objective of the study was to analyze a large cohort of IBD patients for the association between IBD and IRAK-M. MATERIAL AND METHODS: A total of 542 patients with IBD (309 Crohn's disease (CD), 233 ulcerative colitis (UC)) and 305 controls were studied. Two single nucleotide polymorphisms (V147I and V270I) and six microsatellite markers were evaluated using association analysis and the haplotype sharing statistic. Results were stratified for CARD15 mutations R702W, G908R and 1007fsinsC. RESULTS: No significant differences in IRAK-M allele frequencies were observed between IBD, UC, CD or subgroups of CD or UC and controls. Five out of 36 UC patients (13.9%) with an IBD-associated CARD15 mutation were carriers versus 2/167 (1.2%) in non-carriers (OR 13.1, 95% CI 1.0-164.5). No interaction was observed for CD. CONCLUSIONS: No evidence was found to suggest an association between IBD, CD, UC or subsets of CD and UC and IRAK-M. However, an interaction was found between IRAK-M and CARD15 in UC patients. In CARD15 mutant patients, the production of IRAK-M upon stimulation might be impaired. Further studies are needed to determine whether an impaired negative regulation of the TLR-signaling pathway might be partly responsible for the development of IBD.

Absence of association between the multidrug resistance (MDR1) gene and inflammatory bowel disease.

OBJECTIVE: The multidrug resistance (MDR1) gene encodes for P-glycoprotein, a drug efflux pump. Mice deficient for the MDR1a gene spontaneously develop colitis. In humans, a polymorphism in exon 26 (C3435T) is associated with reduced expression levels and function of MDR1. Currently there are controversial data on the association between MDR1 and inflammatory bowel disease (IBD). The purpose of this study was to examine the involvement of this gene in IBD in a large population of Dutch patients with IBD and family-based controls. MATERIAL AND METHODS: A total of 781 IBD cases and 315 controls were investigated. CD phenotypes were determined according to the Vienna Classification. Individuals were genotyped for six single nucleotide polymorphisms (SNPs) close to and in the MDR1 locus. This included the C3435T variant and six microsatellite markers close to and in the MDR1 locus. Single locus association analysis, haplotype association analysis and haplotype sharing statistic (HSS) were used to search for differences between patients and controls. RESULTS: No association was observed for any of the SNPs with IBD as a group, or for ulcerative colitis, Crohn's disease and Crohn's disease phenotypes, either by single locus or haplotype association analysis or by HSS. CONCLUSIONS: No association was observed between the MDR1 gene and IBD. This suggests that it is unlikely that MDR1 plays a role in IBD susceptibility.

Methylenetetrahydrofolate reductase (MTHFR) and susceptibility for (pre)neoplastic cervical disease.

Methylenetetrahydrofolate reductase (MTHFR) is a critical enzyme regulating the metabolism of folate and methionine. The potential influence of MTHFR activity on DNA methylation and on the availability of uridylates and thymidylates for DNA synthesis and repair presents MTHFR as a candidate for being a cancer-predisposing gene. In the present study, we have examined a large study population to determine whether the C677T polymorphism at the MTHFR locus affects susceptibility for cervical cancer or its precursor, cervical intraepithelial neoplasia (CIN). In addition, we have investigated whether this polymorphism is causal, and not merely associated, by typing microsatellite markers in the region surrounding the MTHFR gene. A total of 311 CIN and 695 cervical cancer patients and 115 family-based and 586 unrelated controls was analysed. Association analysis showed a decreased cervical cancer risk for individuals heterozygous or homozygous for the T-allele, both for squamous cell carcinoma (heterozygous odds ration [OR] 0.66 [0.51-0.86]; homozygous OR 0.76 [0.49-1.16]) and adenocarcinoma (heterozygous OR 0.71 [0.49-1.03]; homozygous OR 0.34 [0.14-0.81]). No difference was found for high grade CIN (heterozygous OR 1.03 [0.76-1.40]; homozygous OR 0.91 [0.54-1.55]). A microsatellite haplotype containing the C allele was associated with an increased risk for cervical cancer and CIN (both among squamous cell carcinomas, adenocarcinomas and CIN II-III; OR = 2.61 [1.59-4.27]). Our study thus lends further support to the hypothesis that the MTHFR C677T polymorphism is involved in susceptibility cervical cancer but also illustrates that, despite the large sample size analysed, still larger studies are needed to establish fully the nature of this association.

The HLA class III subregion is responsible for an increased breast cancer risk.

BRCA1 and BRCA2 germline mutations account for <5% of breast cancer cases. Less penetrant breast cancer susceptibility genes are likely to exist. Earlier studies have suggested involvement of the HLA region. The HLA region was genotyped with 24 microsatellite markers and markers for two single nucleotide polymorphisms (SNPs) in TNFalpha and TNFbeta, in germline DNA from 956 breast cancer patients and 1271 family-based controls. Association analyses and the haplotype sharing statistic (HSS) were used to search for differences in haplotype sharing between patients and controls. Based on criteria known to influence genetic breast cancer risk, patients were divided into groups of high, moderate and low risk. The HSS revealed a significant difference in mean haplotype sharing between patients and controls for four consecutive markers (D6S2671, TNFa, D6S2672 and MICA), the highest being at D6S2671 (P=0.017). Subgroup analyses showed that moderate-risk patients were responsible for this difference, with the strongest association for D6S2672 (P=0.0009). A single haplotype was more frequent and longer in moderate-risk patients than in controls. The results were confirmed with association analyses. Individuals homozygous for haplotype 110-184 (D6S2672-MICA) were observed in 9.0% of moderate-risk patients and 1.5% of controls [odds ratio (OR)=7.14], while heterozygotes were at a lower risk (OR=1.41), suggesting a recessive effect. No association was observed between the two SNPs in TNFalpha (-308) and TNFbeta (intron 1) and breast cancer risk. The results reveal a potential role of the HLA class III subregion in susceptibility to breast cancer in patients at moderate familial risk.