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Coenzyme Q10 (CoQ10) is a lipophilic redox molecule that is present in membranes of almost all cells in human tissues. CoQ10 is, amongst other functions, essential for the respiratory transport chain and is a modulator of inflammatory processes and gene expression. Rare monogenetic CoQ10 deficiencies show noticeable symptoms in tissues (e.g. kidney) and cell types (e.g. neurons) with a high energy demand. To identify common genetic variants influencing serum CoQ10 levels, we performed a fixed effects meta-analysis in two independent cross-sectional Northern German cohorts comprising 1300 individuals in total. We identified two genome-wide significant susceptibility loci. The best associated single nucleotide polymorphism (SNP) was rs9952641 (P value = 1.31 × 10 -8, β = 0.063, CI0.95 [0.041, 0.085]) within the COLEC12 gene on chromosome 18. The SNP rs933585 within the NRXN-1 gene on chromosome 2 also showed genome wide significance (P value = 3.64 × 10 -8, β = -0.034, CI0.95 [-0.046, -0.022]). Both genes have been previously linked to neuronal diseases like Alzheimer's disease, autism and schizophrenia. Among our 'top-10' associated variants, four additional loci with known neuronal connections showed suggestive associations with CoQ10 levels. In summary, this study demonstrates that serum CoQ10 levels are associated with common genetic loci that are linked to neuronal diseases.

Little is known about the functions of miRNAs in human longevity. Here, we present the first genome-wide miRNA study in long-lived individuals (LLI) who are considered a model for healthy aging. Using a microarray with 863 miRNAs, we compared the expression profiles obtained from blood samples of 15 centenarians and nonagenarians (mean age 96.4 years) with those of 55 younger individuals (mean age 45.9 years). Eighty miRNAs showed aging-associated expression changes, with 16 miRNAs being up-regulated and 64 down-regulated in the LLI relative to the younger probands. Seven of the eight selected aging-related biomarkers were technically validated using quantitative RT-PCR, confirming the microarray data. Three of the eight miRNAs were further investigated in independent samples of 15 LLI and 17 younger participants (mean age 101.5 and 36.9 years, respectively). Our screening confirmed previously published miRNAs of human aging, thus reflecting the utility of the applied approach. The hierarchical clustering analysis of the miRNA microarray expression data revealed a distinct separation between the LLI and the younger controls (P-value10(-5) ). The down-regulated miRNAs appeared as a cluster and were more often reported in the context of diseases than the up-regulated miRNAs. Moreover, many of the differentially regulated miRNAs are known to exhibit contrasting expression patterns in major age-related diseases. Further in silico analyses showed enrichment of potential targets of the down-regulated miRNAs in p53 and other cancer pathways. Altogether, synchronized miRNA-p53 activities could be involved in the prevention of tumorigenesis and the maintenance of genomic integrity during aging.

Micro-RNAs (miRNAs) are short, non-coding RNAs that regulate gene expression post transcriptionally. Several studies have demonstrated the relevance of miRNAs for a wide range of cellular mechanisms, however, the current knowledge on how miRNAs respond to relevant external stimuli, e.g. in disease scenarios is very limited. To generate a descriptive picture of the miRNA network associated to inflammatory responses, we quantified the levels of 330 miRNAs upon stimulation with a panel of pro-inflammatory components such as microbial pattern molecules (flagellin, diacylated lipopeptide lipopolysaccharide, muramyl dipeptide), infection with Listeria monocytogenes and TNF-α as pro-inflammatory control in primary human monocytes using real time PCR. As a result, we found distinct miRNA response clusters for each stimulus used. Additionally, we identified potential target genes of three selected miRNAs miR-129-5p, miR-146a and miR-378 which were part of PAMP-specific response clusters by transfecting THP1 monocytes with the corresponding pre- or anti-miRNAs and microfluidic PCR arrays. The miRNAs induced distinct transcriptomal signatures, e.g. overexpression of miRNA129-5p, which was selectively upregulated by the NOD2-elicitor MDP, led to an upregulation of DEFB1, IRAK1, FBXW7 and IKK γ (Nemo). Our findings on highly co-regulated clusters of miRNAs support the hypothesis that miRNAs act in functional groups. This study indicates that miRNAs play an important role in fine-tuning inflammatory mechanisms. Further investigation in the field of miRNA responses will help to understand their effects on gene expression and may close the regulatory gap between mRNA and protein expression in inflammatory diseases.

In a multicenter study, we determined the expression profiles of 863 microRNAs by array analysis of 454 blood samples from human individuals with different cancers or noncancer diseases, and validated this 'miRNome' by quantitative real-time PCR. We detected consistently deregulated profiles for all tested diseases; pathway analysis confirmed disease association of the respective microRNAs. We observed significant correlations (P = 0.004) between the genomic location of disease-associated genetic variants and deregulated microRNAs.

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.

Aims: The modulation of the intestinal expression of detoxifying proteins by relevant transcription factors, intracellular receptors and cytokines in ulcerative colitis (UC) is poorly understood. Here, we compared the intestinal expression of drug transporters, metabolizing enzymes and putative regulatory genes between inflamed and noninflamed tissue and studied their modulation by disease activity. Materials & methods: Sigmoidal biopsies of 18 UC patients and 18 healthy volunteers matched for age, gender and ABCB1 3435C>T genotype were investigated for mRNA expression levels of 43 systematically selected candidate genes by low-density array real-time PCR. Additionally, the ABCB1 gene product P-glycoprotein was visualized by immunohistochemistry and quantified by western blotting. Disease phenotype was categorized by clinical, endoscopic and histopathological examination. Disease activity was quantified by clinical activity index. Results: In inflamed sigmoidal tissue from UC patients, 11 genes (NAT1, NR2B1, CEBPB, IFG, IL8, IL10, S100A12, SPP1, DEFA5, DEFA6 and HAMP) were overexpressed. By contrast, only the major human efflux transporter ABCB1 showed significantly lower expression levels, that were inversely correlated with those of certain antimicrobial peptides (DEFA5/6) and cytokines (IL1β and IL8). Cell culture experiments revealed a time-dependent decrease of ABCB1 expression upon IL8 exposure. Disease activity profoundly modified ABCB1 expression, indicated by an inverse correlation of clinical activity index values with ABCB1 mRNA expression (r = -0.603; p = 0.017) and markedly reduced protein expression in UC patients with moderate and severe symptomology (p = 0.011). Conclusion: Cytokine-mediated downregulation of the major human efflux transporter ABCB1 in inflamed intestine of UC patients is presumably dependent on disease activity, with a possible contribution from IL8.

Human longevity is heritable with a genetic component of 25-32%. Variation in genes regulating the levels of somatic maintenance and DNA repair functions is thought to modulate the aging process and to contribute to survival at advanced age. We tested 92 non-synonymous SNPs in 49 DNA repair genes for a possible association with longevity in a sample of 395 German centenarians and 411 controls. The obtained association signal in exonuclease 1 (EXO1) was further investigated by fine mapping and mutation detection, leading to the identification of the functionally relevant SNP rs1776180. Our detailed analyses revealed that the C allele of this promoter SNP is significantly enriched in female centenarians. This finding replicated in 455 female French centenarians and 109 controls. The C allele leads to the loss of a binding site for the basic helix-loop-helix transcription factor E47, resulting in higher EXO1 expression. Thus, we have detected a hitherto undescribed role for E47 as a negative regulator of EXO1 transcription and a genetic variant in the EXO1 promoter that counteracts the E47-mediated repression of the gene. Given the survival advantage that is associated with the C allele of rs1776180, EXO1 can be considered a candidate for a novel longevity-enabling gene. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Although many properties of the innate immune system are shared among multicellular animals, the evolutionary origin remains poorly understood. Here we characterize the innate immune system in Hydra, one of the simplest multicellular animals known. In the complete absence of both protective mechanical barriers and mobile phagocytes, Hydra's epithelium is remarkably well equipped with potent antimicrobial peptides to prevent pathogen infection. Induction of antimicrobial peptide production is mediated by the interaction of a leucine-rich repeats (LRRs) domain containing protein with a TIR-domain containing protein lacking LRRs. Conventional Toll-like receptors (TLRs) are absent in the Hydra genome. Our findings support the hypothesis that the epithelium represents the ancient system of host defence. (C) 2008 Elsevier Ltd. All rights reserved.

NOD-like receptors (NLRs) comprise a family of cytosolic proteins that have been implicated as ancient cellular sentinels mediating protective immune responses elicited by intracellular pathogens or endogenous danger signals. Genetic variants in NLR genes have been associated with complex chronic inflammatory barrier diseases (e.g. Crohn disease, bronchial asthma). In this review, we focus on the molecular pathophysiology of NLRs in the context of chronic inflammatory diseases and pinpoint recent advances in the evolutionary understanding of NLR biology. We propose that the field of NLRs may serve as a prototype for how a comprehensive understanding of an element of the immunological barrier will eventually lead to the development of targeted diagnostic, therapeutic and/or preventive strategies.

Genome-wide association (GWA) studies offer a powerful unbiased method for the identification of multiple susceptibility genes for complex diseases. Here we report the results of a GWA study for Crohn's disease (CD) using family trios from the Quebec Founder Population (QFP). Haplotype-based association analyses identified multiple regions associated with the disease that met the criteria for genome-wide significance, with many containing a gene whose function appears relevant to CD. A proportion of these were replicated in two independent German Caucasian samples, including the established CD lociNOD2andIBD5. The recently describedIL23Rlocus was also identified and replicated. For this region, multiple individuals with all major haplotypes in the QFP were sequenced and extensive fine mapping performed to identify risk and protective alleles. Several additional loci, including a region on 3p21 containing several plausible candidate genes, a region nearJAKMIP1on 4p16.1, and two larger regions on chromosome 17 were replicated. Together with previously published loci, the spectrum of CD genes identified to date involves biochemical networks that affect epithelial defense mechanisms, innate and adaptive immune response, and the repair or remodeling of tissue.

Background: The association between diet and fatty liver disease (FLD) has predominantly been analyzed for single nutrients or foods, and findings have been inconsistent. Objective: We aimed to compare associations of hypothesis-driven and exploratory dietary pattern scores with liver fat content. Design: Liver fat was measured by using magnetic resonance imaging as liver signal intensity (LSI) in a population-based, crosssectional study that included 354 individuals. We applied partial least-squares regression to derive an exploratory dietary pattern score that explained variation in both, the intake of 38 food groups, which were assessed by using a food-frequency questionnaire, and LSI. The hypothesis-driven score was calculated on the basis of published studies. Multivariable linear or logistic regression was used to investigate associations between dietary pattern scores and LSI or FLD. Results: A higher percentage of LSI variation was explained by the exploratory (12.6%) compared with the hypothesis-driven (2.2%) dietary pattern. Of the 13 most important food groups of the exploratory dietary pattern, intakes of green and black tea, soups, and beer were also individually associated with LSI values. A 1-unit increase in the exploratory dietary pattern score was positively associated with FLD (OR: 1.56; 95% CI: 1.29, 1.88). Furthermore, a 1-unit increase in the hypothesis-driven dietary pattern score, which consisted of alcohol, soft drinks, meat, coffee, and tea, was positively associated with FLD (OR: 1.25; 95% CI: 1.10, 1.43). Conclusion: We defined a hypothesis-driven dietary pattern and derived an exploratory dietary pattern, both of which included alcohol, meat (poultry), and tea, associated with liver fat content independent from confounders, which should be explored in prospective studies. Am J Clin Nutr doi: 10.3945/ajcn.113. 070219.

Sugar beet ( Beta vulgaris L.) is a biennial species. Shoot elongation (bolting) starts after a period of low temperature. The dominant allele of locus B causes early bolting without cold treatment. This allele is abundant in wild beets whereas cultivated beets carry the recessive allele. Fifteen AFLP markers, tightly linked to the bolting locus, have been identified using bulked segregant analysis. The F(2)-population consisted of 2,134 individuals derived after selfing a single F(1)-plant ( Bb). In a first step, a linkage map was established with 249 markers based on 775 F(2)-individuals with a coverage of 822.3 cM. The loci are dispersed over nine linkage groups corresponding to the haploid chromosome number of Beta species. Seventeen marker loci were placed at a distance less than 3.2 cM around the bolting gene. In a second step, four of those markers most closely linked to B were mapped with the entire F(2)-population. Two of the markers were mapped flanking the B gene at distances of 0.14 and 0.23 cM. The other two markers were mapped at a distance of 0.5 cM from the gene. The tight linkage could be verified by testing 88 unrelated plants from a breeding program. The closely linked markers will enable breeders to select for the non-bolting character without laborious test crossings. Moreover, these markers are being used for map-based cloning of the bolting gene.

Background & Aims: Micro-RNAs (miRNAs) are short, non-coding RNAs that regulate gene expression post transcriptionally. Several studies have demonstrated distinct miRNA profiles in a wide range of human disorders including inflammatory bowel disease and colon cancer. Surprisingly little is known about miRNA responses in immune cells that are challenged with relevant proinflammatory stimuli, i.e. cytokines or pathogen-associated microbial patterns. With this study we thus aim to generate a prototypic picture of inflammatory miRNA networks in monocytes as a relevant immune cell population and to understand functional consequences of selected regulated miRNAs on inflammatory signalling pathways. Methods: Levels of 330 miRNAs upon stimulation with a panel of pro-inflammatory components such as microbial pattern molecules (flagellin, diacylated lipopeptide lipopolysaccharide, muramyl dipeptide), infection with Listeria monocytogenes and TNF-alpha as pro-inflammatory control in primary human monocytes were qunatified using real time PCR. Selected miRNAs were verified in parallel in LPMNCs and biopsies from IBD patients. Transfection of premiRs and antago-miRs in THP1 monocytic cells were employed in order to understand functional consequences of miRNA regulation. Results: We identified distinct miRNA response clusters pointing to unique and shared mechanisms for different proinflammatory stimuli even in a single cell type. Additionally, we identified potential target genes of three selected miRNAs miR-129-5p, miR-146a and miR-378 which were part of PAMP-specific response clusters by transfecting THP1 monocytes with the corresponding preor antimiRNAs . The miRNAs induced distinct transcriptomal signatures, e.g. overexpression of miRNA129-5p, which was selectively upregulated by the NOD2-elicitor MDP, led to an upregulation of DEFB1, IRAK1, FBXW7 and IKK-gamma (Nemo). Interestingly, regulation of several "overlap" miRNAs shared between stimuli could also be detected in samples from IBD patients. Conclusions: Our findings on co-regulated clusters of miRNAs support the hypothesis that miRNAs act in functional groups. This study indicates that miRNAs play an important role in fine-tuning inflammatory mechanisms. Investigation in the field of miRNA responses in intestinal inflammation will help to understand their effects on gene expression and may close the regulatory gap between mRNA and protein expression in inflammatory diseases.

Nat. Genet, 40, 1103–1106 (2008), published online 10 August 2008; corrected after print 24 March 2009 In the version of this article initially published, the SNP rs1049550 listed in the abstract was indicated incorrectly as a T>C change. It is a C>T change. On the third page of the article, the haplotype containing rs1049550 was incorrectly listed as TATACC.

The reduced form of Coenzyme Q10 (CoQ10), ubiquinol (Q10H2), serves as a potent antioxidant in mitochondria and lipid membranes. There is evidence that Q10H2 protects against oxidative events in lipids, proteins and DNA. Serum gamma-glutamyltransferase (GGT) activity is associated with cardiovascular diseases. In a physiological range, activity of GGT is a potential early and sensitive marker of inflammation and oxidative stress. In this study, we first examined the relationship between CoQ10 status and serum GGT activity in 416 healthy participants between 19 and 62 years of age in a cross-sectional study (cohort I). In the second step, 53 healthy males (21–48 years of age; cohort II) underwent a 14-day Q10H2 supplementation (150 mg/d) to evaluate the effect of Q10H2 supplementation on serum GGT activity and GGT1 gene expression. There was a strong positive association between CoQ10 status and serum GGT activity in cohort I. However, a gender-specific examination revealed differences between male and female volunteers regarding the association between CoQ10 status and serum GGT activity. Q10H2 supplementation (cohort II) caused a significant decrease in serum GGT activity from T0 to T14 (p