Prospective study of insulin-like growth factor-I, insulin-like growth factor-binding protein 3, genetic variants in the IGF1 and IGFBP3 genes and risk of coronary artery disease.

Although experimental studies have suggested that insulin-like growth factor I (IGF-I) and its binding protein IGFBP-3 might have a role in the aetiology of coronary artery disease (CAD), the relevance of circulating IGFs and their binding proteins in the development of CAD in human populations is unclear. We conducted a nested case-control study, with a mean follow-up of six years, within the EPIC-Norfolk cohort to assess the association between circulating levels of IGF-I and IGFBP-3 and risk of CAD in up to 1,013 cases and 2,055 controls matched for age, sex and study enrolment date. After adjustment for cardiovascular risk factors, we found no association between circulating levels of IGF-I or IGFBP-3 and risk of CAD (odds ratio: 0.98 (95% Cl 0.90-1.06) per 1 SD increase in circulating IGF-I; odds ratio: 1.02 (95% Cl 0.94-1.12) for IGFBP-3). We examined associations between tagging single nucleotide polymorphisms (tSNPs) at the IGF1 and IGFBP3 loci and circulating IGF-I and IGFBP-3 levels in up to 1,133 cases and 2,223 controls and identified three tSNPs (rs1520220, rs3730204, rs2132571) that showed independent association with either circulating IGF-I or IGFBP-3 levels. In an assessment of 31 SNPs spanning the IGF1 or IGFBP3 loci, none were associated with risk of CAD in a meta-analysis that included EPIC-Norfolk and eight additional studies comprising up to 9,319 cases and 19,964 controls. Our results indicate that IGF-I and IGFBP-3 are unlikely to be importantly involved in the aetiology of CAD in human populations.

Mapping genetic variants associated with beta-adrenergic responses in inbred mice.

β-blockers and β-agonists are primarily used to treat cardiovascular diseases. Inter-individual variability in response to both drug classes is well recognized, yet the identity and relative contribution of the genetic players involved are poorly understood. This work is the first genome-wide association study (GWAS) addressing the values and susceptibility of cardiovascular-related traits to a selective β(1)-blocker, Atenolol (ate), and a β-agonist, Isoproterenol (iso). The phenotypic dataset consisted of 27 highly heritable traits, each measured across 22 inbred mouse strains and four pharmacological conditions. The genotypic panel comprised 79922 informative SNPs of the mouse HapMap resource. Associations were mapped by Efficient Mixed Model Association (EMMA), a method that corrects for the population structure and genetic relatedness of the various strains. A total of 205 separate genome-wide scans were analyzed. The most significant hits include three candidate loci related to cardiac and body weight, three loci for electrocardiographic (ECG) values, two loci for the susceptibility of atrial weight index to iso, four loci for the susceptibility of systolic blood pressure (SBP) to perturbations of the β-adrenergic system, and one locus for the responsiveness of QTc (p<10(-8)). An additional 60 loci were suggestive for one or the other of the 27 traits, while 46 others were suggestive for one or the other drug effects (p<10(-6)). Most hits tagged unexpected regions, yet at least two loci for the susceptibility of SBP to β-adrenergic drugs pointed at members of the hypothalamic-pituitary-thyroid axis. Loci for cardiac-related traits were preferentially enriched in genes expressed in the heart, while 23% of the testable loci were replicated with datasets of the Mouse Phenome Database (MPD). Altogether these data and validation tests indicate that the mapped loci are relevant to the traits and responses studied.

Inheritance of late flowering in natural variants of soybean cultivars under short-day conditions

The objective of this work was to determine the inheritance of the long juvenile period trait in natural variants of the Doko, BR 9 (Savana), Davis, Embrapa 1 (IAS 5RC), and BR 16 soybean cultivars. Complete diallel crosses were made between the Doko and BR 16 cultivars and their variants. A 3:1 segregation ratio was observed in the F2 populations of the 'Doko' x Doko-18T, 'Doko' x Doko-Milionária, 'Davis' x São Carlos, and 'BR 9 (Savana)' x MABR92-836 (Savanão) crosses, indicating that the long juvenile period trait is controlled by a pair of recessive genes. The difference in late flowering between the Doko cultivar and both of its variants was caused by a recessive spontaneous mutation at the same genetic locus. However, the variants Doko-18T and Doko-Milionária are identical mutants that share a pair of genes that control the long juvenile period under short-day conditions. These mutants can be used in breeding programs to develop cultivars adapted to low-latitude tropical regions.

MLL2 mutation detection in 86 patients with Kabuki syndrome: a genotype-phenotype study.

Recently, pathogenic variants in the MLL2 gene were identified as the most common cause of Kabuki (Niikawa-Kuroki) syndrome (MIM#147920). To further elucidate the genotype-phenotype correlation, we studied a large cohort of 86 clinically defined patients with Kabuki syndrome (KS) for mutations in MLL2. All patients were assessed using a standardized phenotype list and all were scored using a newly developed clinical score list for KS (MLL2-Kabuki score 0-10). Sequencing of the full coding region and intron-exon boundaries of MLL2 identified a total of 45 likely pathogenic mutations (52%): 31 nonsense, 10 missense and four splice-site mutations, 34 of which were novel. In five additional patients, novel, i.e. non-dbSNP132 variants of clinically unknown relevance, were identified. Patients with likely pathogenic nonsense or missense MLL2 mutations were usually more severely affected (median 'MLL2-Kabuki score' of 6) as compared to the patients without MLL2 mutations (median 'MLL2-Kabuki score' of 5), a significant difference (p < 0.0014). Several typical facial features such as large dysplastic ears, arched eyebrows with sparse lateral third, blue sclerae, a flat nasal tip with a broad nasal root, and a thin upper and a full lower lip were observed more often in mutation positive patients.

Modification of the monomer composition of polyhydroxyalkanoate synthesized in Saccharomyces cerevisiae expressing variants of the beta-oxidation-associated multifunctional enzyme.

Expression by Saccharomyces cerevisiae of a polyhydroxyalkanoate (PHA) synthase modified at the carboxy end by the addition of a peroxisome targeting signal derived from the last 34 amino acids of the Brassica napus isocitrate lyase (ICL) and containing the terminal tripeptide Ser-Arg-Met resulted in the synthesis of PHA. The ability of the terminal peptide Ser-Arg-Met and of the 34-amino-acid peptide from the B. napus ICL to target foreign proteins to the peroxisome of S. cerevisiae was demonstrated with green fluorescent protein fusions. PHA synthesis was found to be dependent on the presence of both the enzymes generating the beta-oxidation intermediate 3-hydroxyacyl-coenzyme A (3-hydroxyacyl-[CoA]) and the peroxin-encoding PEX5 gene, demonstrating the requirement for a functional peroxisome and a beta-oxidation cycle for PHA synthesis. Using a variant of the S. cerevisiae beta-oxidation multifunctional enzyme with a mutation inactivating the B domain of the R-3-hydroxyacyl-CoA dehydrogenase, it was possible to modify the PHA monomer composition through an increase in the proportion of the short-chain monomers of five and six carbons.

Coronary artery anomalies and variants: technical feasibility of assessment with coronary MR angiography at 3 T.

The purpose of this study was to prospectively use a whole-heart three-dimensional (3D) coronary magnetic resonance (MR) angiography technique specifically adapted for use at 3 T and a parallel imaging technique (sensitivity encoding) to evaluate coronary arterial anomalies and variants (CAAV). This HIPAA-compliant study was approved by the local institutional review board, and informed consent was obtained from all participants. Twenty-two participants (11 men, 11 women; age range, 18-62 years) were included. Ten participants were healthy volunteers, whereas 12 participants were patients suspected of having CAAV. Coronary MR angiography was performed with a 3-T MR imager. A 3D free-breathing navigator-gated and vector electrocardiographically-gated segmented k-space gradient-echo sequence with adiabatic T2 preparation pulse and parallel imaging (sensitivity encoding) was used. Whole-heart acquisitions (repetition time msec/echo time msec, 4/1.35; 20 degrees flip angle; 1 x 1 x 2-mm acquired voxel size) lasted 10-12 minutes. Mean examination time was 41 minutes +/- 14 (standard deviation). Findings included aneurysms, ectasia, arteriovenous fistulas, and anomalous origins. The 3D whole-heart acquisitions developed for use with 3 T are feasible for use in the assessment of CAAV.

FTO genetic variants, dietary intake and body mass index: insights from 177,330 individuals.

FTO is the strongest known genetic susceptibility locus for obesity. Experimental studies in animals suggest the potential roles of FTO in regulating food intake. The interactive relation among FTO variants, dietary intake and body mass index (BMI) is complex and results from previous often small-scale studies in humans are highly inconsistent. We performed large-scale analyses based on data from 177,330 adults (154 439 Whites, 5776 African Americans and 17 115 Asians) from 40 studies to examine: (i) the association between the FTO-rs9939609 variant (or a proxy single-nucleotide polymorphism) and total energy and macronutrient intake and (ii) the interaction between the FTO variant and dietary intake on BMI. The minor allele (A-allele) of the FTO-rs9939609 variant was associated with higher BMI in Whites (effect per allele = 0.34 [0.31, 0.37] kg/m(2), P = 1.9 × 10(-105)), and all participants (0.30 [0.30, 0.35] kg/m(2), P = 3.6 × 10(-107)). The BMI-increasing allele of the FTO variant showed a significant association with higher dietary protein intake (effect per allele = 0.08 [0.06, 0.10] %, P = 2.4 × 10(-16)), and relative weak associations with lower total energy intake (-6.4 [-10.1, -2.6] kcal/day, P = 0.001) and lower dietary carbohydrate intake (-0.07 [-0.11, -0.02] %, P = 0.004). The associations with protein (P = 7.5 × 10(-9)) and total energy (P = 0.002) were attenuated but remained significant after adjustment for BMI. We did not find significant interactions between the FTO variant and dietary intake of total energy, protein, carbohydrate or fat on BMI. Our findings suggest a positive association between the BMI-increasing allele of FTO variant and higher dietary protein intake and offer insight into potential link between FTO, dietary protein intake and adiposity.

The TREAT-NMD DMD Global Database: Analysis of More than 7,000 Duchenne Muscular Dystrophy Mutations.

Analyzing the type and frequency of patient-specific mutations that give rise to Duchenne muscular dystrophy (DMD) is an invaluable tool for diagnostics, basic scientific research, trial planning, and improved clinical care. Locus-specific databases allow for the collection, organization, storage, and analysis of genetic variants of disease. Here, we describe the development and analysis of the TREAT-NMD DMD Global database (http://umd.be/TREAT_DMD/). We analyzed genetic data for 7,149 DMD mutations held within the database. A total of 5,682 large mutations were observed (80% of total mutations), of which 4,894 (86%) were deletions (1 exon or larger) and 784 (14%) were duplications (1 exon or larger). There were 1,445 small mutations (smaller than 1 exon, 20% of all mutations), of which 358 (25%) were small deletions and 132 (9%) small insertions and 199 (14%) affected the splice sites. Point mutations totalled 756 (52% of small mutations) with 726 (50%) nonsense mutations and 30 (2%) missense mutations. Finally, 22 (0.3%) mid-intronic mutations were observed. In addition, mutations were identified within the database that would potentially benefit from novel genetic therapies for DMD including stop codon read-through therapies (10% of total mutations) and exon skipping therapy (80% of deletions and 55% of total mutations).

Hepatitis C virus variants resistant to macrocyclic NS3-4A inhibitors subvert IFN-β induction by efficient MAVS cleavage.

BACKGROUND & AIMS: The hepatitis C virus (HCV) NS3-4A protease is essential for the HCV life cycle and a prime target of antiviral treatment strategies. Protease inhibitors, however, are limited by emergence of resistance-associated amino acid variants (RAVs). The capacity to cleave and inactivate mitochondrial antiviral-signaling protein (MAVS) in the RIG-I-signaling pathway is a cardinal feature of NS3-4A, by which HCV blocks induction of interferon-(IFN)-β, thereby promoting viral persistence. Here, we aimed to investigate the impact of NS3-4A RAVs on MAVS cleavage. METHODS: The impact of NS3-4A RAVs on MAVS cleavage was assessed using immunoblot analyses, luciferase reporter assays and molecular dynamics simulations to study the underlying molecular principles. IFN-β was quantified in serum from patients with different NS3-4A RAVs. RESULTS: We show that macrocyclic NS3-4A RAVS with substitutions at residue D168 of the protease result in an increased capacity of NS3-4A to cleave MAVS and suppress IFN-β induction compared with a comprehensive panel of RAVs and wild type HCV. Mechanistically, we show the reconstitution of a tight network of electrostatic interactions between protease and the peptide substrate that allows much stronger binding of MAVS to D168 RAVs than to the wild-type protease. Accordingly, we could show IFN-β serum levels to be lower in patients with treatment failure due to the selection of D168 variants compared to R155 RAVs. CONCLUSIONS: Our data constitutes a proof of concept that the selection of RAVs against specific classes of direct antivirals can lead to the predominance of viral variants with possibly adverse pathogenic characteristics.

Expanding the Mutation Spectrum Affecting αIIbβ3 Integrin in Glanzmann Thrombasthenia: Screening of the ITGA2B and ITGB3 Genes in a Large International Cohort.

We report the largest international study on Glanzmann thrombasthenia (GT), an inherited bleeding disorder where defects of the ITGA2B and ITGB3 genes cause quantitative or qualitative defects of the αIIbβ3 integrin, a key mediator of platelet aggregation. Sequencing of the coding regions and splice sites of both genes in members of 76 affected families identified 78 genetic variants (55 novel) suspected to cause GT. Four large deletions or duplications were found by quantitative real-time PCR. Families with mutations in either gene were indistinguishable in terms of bleeding severity that varied even among siblings. Families were grouped into type I and the rarer type II or variant forms with residual αIIbβ3 expression. Variant forms helped identify genes encoding proteins mediating integrin activation. Splicing defects and stop codons were common for both ITGA2B and ITGB3 and essentially led to a reduced or absent αIIbβ3 expression; included was a heterozygous c.1440-13_c.1440-1del in intron 14 of ITGA2B causing exon skipping in seven unrelated families. Molecular modeling revealed how many missense mutations induced subtle changes in αIIb and β3 domain structure across both subunits, thereby interfering with integrin maturation and/or function. Our study extends knowledge of GT and the pathophysiology of an integrin.

Common Variants of TLR1 Associate with Organ Dysfunction and Sustained Pro- Inflammatory Responses during Sepsis

Background: Toll-like receptors (TLRs) are critical components for host pathogen recognition and variants in genes participating in this response influence susceptibility to infections. Recently, TLR1 gene polymorphisms have been found correlated with whole blood hyper-inflammatory responses to pathogen-associated molecules and associated with sepsis-associated multiorgan dysfunction and acute lung injury (ALI). We examined the association of common variants of TLR1 gene with sepsis-derived complications in an independent study and with serum levels for four inflammatory biomarker among septic patients. Methodology/Principal Findings: Seven tagging single nucleotide polymorphisms of the TLR1 gene were genotyped in samples from a prospective multicenter case-only study of patients with severe sepsis admitted into a network of intensive care units followed for disease severity. Interleukin (IL)-1 b, IL-6, IL-10, and C-reactive protein (CRP) serum levels were measured at study entry, at 48 h and at 7th day. Alleles -7202G and 248Ser, and the 248Ser-602Ile haplotype were associated with circulatory dysfunction among severe septic patients (0.001<=p <= 0.022), and with reduced IL-10 (0.012<= p <=0.047) and elevated CRP (0.011<= p <=0.036) serum levels during the first week of sepsis development. Additionally, the -7202GG genotype was found to be associated with hospital mortality (p =0.017) and ALI (p =0.050) in a combined analysis with European Americans, suggesting common risk effects among studies Conclusions/Significance: These results partially replicate and extend previous findings, supporting that variants of TLR1 gene are determinants of severe complications during sepsis.

Clear Cell Papillary Renal Cell Carcinoma and Renal Angiomyoadenomatous Tumor: Two Variants of a Morphologic, Immunohistochemical, and Genetic Distinct Entity of Renal Cell Carcinoma.

Clear cell papillary renal cell carcinoma (ccpRCC) and renal angiomyoadenomatous tumor (RAT) share morphologic similarities with clear cell (ccRCC) and papillary RCC (pRCC). It is a matter of controversy whether their morphologic, immunophenotypic, and molecular features allow the definition of a separate renal carcinoma entity. The aim of our project was to investigate specific renal immunohistochemical biomarkers involved in the hypoxia-inducible factor pathway and mutations in the VHL gene to clarify the relationship between ccpRCC and RAT. We investigated 28 ccpRCC and 9 RAT samples by immunohistochemistry using 25 markers. VHL gene mutations and allele losses were investigated by Sanger sequencing and fluorescence in situ hybridization. Clinical follow-up data were obtained for a subset of the patients. No tumor recurrence or tumor-related death was observed in any of the patients. Immunohistochemistry and molecular analyses led to the reclassification of 3 tumors as ccRCC and TFE3 translocation carcinomas. The immunohistochemical profile of ccpRCC and RAT samples was very similar but not identical, differing from both ccRCC and pRCC. Especially, the parafibromin and hKIM-1 expression exhibited differences in ccpRCC/RAT compared with ccRCC and pRCC. Genetic analysis revealed VHL mutations in 2/27 (7%) and 1/7 (14%) ccpRCC and RAT samples, respectively. Fluorescence in situ hybridization analysis disclosed a 3p loss in 2/20 (10%) ccpRCC samples. ccpRCC and RAT have a specific morphologic and immunohistochemical profile, but they share similarities with the more aggressive renal tumors. On the basis of our results, we regard ccpRCC/RAT as a distinct entity of RCCs.

Eosinophilic fasciitis: Typical abnormalities, variants and differential diagnosis of fasciae abnormalities using MR imaging.

Eosinophilic fasciitis is a rare condition. It is generally limited to the distal parts of the arms and legs. MRI is the ideal imaging modality for diagnosing and monitoring this condition. MRI findings typically evidence only fascial involvement but on a less regular basis signal abnormalities may be observed in neighboring muscle tissue and hypodermic fat. Differential diagnosis of eosinophilic fasciitis by MRI requires the exclusion of several other superficial and deep soft tissue disorders.