Interethnic Differences in the Distribution of Clinically Relevant Vascular Endothelial Growth Factor Genetic Polymorphisms

Vascular endothelial growth factor (VEGF) is a homodimeric glycoprotein produced mostly in endothelial cells and its transcription is regulated by a variety of growth factors and cytokines. VEGF plays many relevant roles, and three functional polymorphisms in the promoter region of the VEGF gene (C-2578A, G-1154A, and G-634C) have been associated with disease conditions. Although some studies suggest that interethnic differences exist in the distribution of these variants, no previous study has examined this hypothesis in admixed populations. We examined the distribution of these three clinically relevant VEGF single-nucleotide polymorphisms in 175 white and 185 black subjects. We have also estimated the haplotype distribution and assessed associations between these variants. Although the A-2578 and A-1154 variants were more common in whites (39% and 29%, respectively) than in blacks (29% and 16%, respectively; both p < 0.05), no significant interethnic differences were found with regards to the G-634C polymorphism. While the haplotype including the C-2578, G-1154, and G-634 variants was the most common in both ethnic groups, it was more common in blacks than in whites (p < 0.05). The haplotype including the C-2578, A-1154, and G-634 alleles and the haplotype including the C-2578, A-1154, and C-634 alleles were more common in whites than in blacks (both p < 0.05). These results show marked interethnic differences in the distribution of genetic variants of VEGF that may explain, at least in part, interethnic disparities in the susceptibility to cardiovascular diseases.

Endothelial Nitric Oxide Synthase Haplotypes Associated with Aura in Patients with Migraine

There is strong evidence implicating nitric oxide (NO) in the pathophysiology of migraine and aura. Therefore, genetic polymorphisms in the endothelial NO synthase (eNOS) gene have been studied as candidate markers for migraine susceptibility. We compared for the first time the distribution of eNOS haplotypes including the three clinically relevant eNOS polymorphisms (T(-786)C in the promoter, rs2070744; Glu298Asp in exon 7, rs1799983; and a 27 bp variable number of tandem repeats in intron 4) and two additional tagging single-nucleotide polymorphisms (rs3918226 and rs743506) in 178 women with migraine (134 without aura and 44 with aura) and 117 healthy controls (control group). Genotypes were determined by TaqMan allele discrimination assay, real-time polymerase chain reaction, and polymerase chain reaction followed by fragment separation by electrophoresis. The GA (rs743506) genotype was more common in the control group than in women with migraine (odds ratio = 0.47, 95% confidence interval [CI] 0.29-0.78, p<0.01). No significant differences were found in allele distributions for the five eNOS polymorphisms. However, the haplotypes including the variants ""C C a Glu G"" and the variants ""C C b Glu G"" were more common in women with migraine with aura than in women with migraine without aura (odds ratio = 30.71, 95% CI = 1.61-586.4 and odds ratio = 17.26, 95% CI = 1.94-153.4, respectively; both p<0.0015625). These findings suggest that these two eNOS haplotypes affect the susceptibility to the presence of aura in patients with migraine.

Molecular and geographic analyses of vampire bat-transmitted cattle rabies in central Brazil

Background: Vampire bats are important rabies virus vectors, causing critical problems in both the livestock industry and public health sector in Latin America. In order to assess the epidemiological characteristics of vampire bat-transmitted rabies, the authors conducted phylogenetic and geographical analyses using sequence data of a large number of cattle rabies isolates collected from a wide geographical area in Brazil. Methods: Partial nucleoprotein genes of rabies viruses isolated from 666 cattle and 18 vampire bats between 1987 and 2006 were sequenced and used for phylogenetic analysis. The genetic variants were plotted on topographical maps of Brazil. Results: In this study, 593 samples consisting of 24 genetic variants were analyzed. Regional localization of variants was observed, with the distribution of several variants found to be delimited by mountain ranges which served as geographic boundaries. The geographical distributions of vampire-bat and cattle isolates that were classified as the identical phylogenetic group were found to overlap with high certainty. Most of the samples analyzed in this study were isolated from adjacent areas linked by rivers. Conclusion: This study revealed the existence of several dozen regional variants associated with vampire bats in Brazil, with the distribution patterns of these variants found to be affected by mountain ranges and rivers. These results suggest that epidemiological characteristics of vampire bat-related rabies appear to be associated with the topographical and geographical characteristics of areas where cattle are maintained, and the factors affecting vampire bat ecology.

FAM5C Contributes to Aggressive Periodontitis

Aggressive periodontitis is characterized by a rapid and severe periodontal destruction in young systemically healthy subjects. A greater prevalence is reported in Africans and African descendent groups than in Caucasians and Hispanics. We first fine mapped the interval 1q24.2 to 1q31.3 suggested as containing an aggressive periodontitis locus. Three hundred and eighty-nine subjects from 55 pedigrees were studied. Saliva samples were collected from all subjects, and DNA was extracted. Twenty-one single nucleotide polymorphisms were selected and analyzed by standard polymerase chain reaction using TaqMan chemistry. Non-parametric linkage and transmission distortion analyses were performed. Although linkage results were negative, statistically significant association between two markers, rs1935881 and rs1342913, in the FAM5C gene and aggressive periodontitis (p = 0.03) was found. Haplotype analysis showed an association between aggressive periodontitis and the haplotype A-G (rs1935881-rs1342913; p = 0.009). Sequence analysis of FAM5C coding regions did not disclose any mutations, but two variants in conserved intronic regions of FAM5C, rs57694932 and rs10494634, were found. However, these two variants are not associated with aggressive periodontitis. Secondly, we investigated the pattern of FAM5C expression in aggressive periodontitis lesions and its possible correlations with inflammatory/immunological factors and pathogens commonly associated with periodontal diseases. FAM5C mRNA expression was significantly higher in diseased versus healthy sites, and was found to be correlated to the IL-1 beta, IL-17A, IL-4 and RANKL mRNA levels. No correlations were found between FAM5C levels and the presence and load of red complex periodontopathogens or Aggregatibacter actinomycetemcomitans. This study provides evidence that FAM5C contributes to aggressive periodontitis.

A Genome-Wide Association Study of Upper Aerodigestive Tract Cancers Conducted within the INHANCE Consortium

Genome-wide association studies (GWAS) have been successful in identifying common genetic variation involved in susceptibility to etiologically complex disease. We conducted a GWAS to identify common genetic variation involved in susceptibility to upper aero-digestive tract (UADT) cancers. Genome-wide genotyping was carried out using the Illumina HumanHap300 beadchips in 2,091 UADT cancer cases and 3,513 controls from two large European multi-centre UADT cancer studies, as well as 4,821 generic controls. The 19 top-ranked variants were investigated further in an additional 6,514 UADT cancer cases and 7,892 controls of European descent from an additional 13 UADT cancer studies participating in the INHANCE consortium. Five common variants presented evidence for significant association in the combined analysis (p <= 5 x 10(-7)). Two novel variants were identified, a 4q21 variant (rs1494961, p = 1 x 10(-8)) located near DNA repair related genes HEL308 and FAM175A (or Abraxas) and a 12q24 variant (rs4767364, p = 2 x 10(-8)) located in an extended linkage disequilibrium region that contains multiple genes including the aldehyde dehydrogenase 2 (ALDH2) gene. Three remaining variants are located in the ADH gene cluster and were identified previously in a candidate gene study involving some of these samples. The association between these three variants and UADT cancers was independently replicated in 5,092 UADT cancer cases and 6,794 controls non-overlapping samples presented here (rs1573496-ADH7, p = 5 x 10(-8); rs1229984-ADH1B, p = 7 x 10(-9); and rs698-ADH1C, p = 0.02). These results implicate two variants at 4q21 and 12q24 and further highlight three ADH variants in UADT cancer susceptibility.

Revisiting clinical trials on glycemic control and cardiovascular risk

The most relevant clinical trials, assessing the role of glycemic control in reducing cardiovascular risk, are examined. The UKPDS was the first to address this issue. More recent trials (ACCORD, ADVANCE and VADT) are controversial and evidences did not support that strict glycemic control (reflected by normal glycated hemoglobin) exclusively is sufficient to reduce cardiovascular risk in complicated individuals with long-term type 2 diabetes mellitus. Some possible reasons for controversies are included.

Genetic Variation among Major Human Geographic Groups Supports a Peculiar Evolutionary Trend in PAX9

A total of 172 persons from nine South Amerindian, three African and one Eskimo populations were studied in relation to the Paired box gene 9 (PAX9) exon 3 (138 base pairs) as well as its 5' and 3' flanking intronic segments (232 bp and 220 bp, respectively) and integrated with the information available for the same genetic region from individuals of different geographical origins. Nine mutations were scored in exon 3 and six in its flanking regions; four of them are new South American tribe-specific singletons. Exon3 nucleotide diversity is several orders of magnitude higher than its intronic regions. Additionally, a set of variants in the PAX9 and 101 other genes related with dentition can define at least some dental morphological differences between Sub-Saharan Africans and non-Africans, probably associated with adaptations after the modern human exodus from Africa. Exon 3 of PAX9 could be a good molecular example of how evolvability works.

Heteroduplex formation and S1 digestion for mapping alternative splicing sites

The identification of alternatively spliced transcripts has contributed to a better comprehension of developmental mechanisms, tissue-specific physiological processes and human diseases. Polymerase chain reaction amplification of alternatively spliced variants commonly leads to the formation of heteroduplexes as a result of base pairing involving exons common between the two variants. S1 nuclease cleaves single-stranded loops of heteroduplexes and also nicks the opposite DNA strand. In order to establish a strategy for mapping alternative splice-prone sites in the whole transcriptome, we developed a method combining the formation of heteroduplexes between 2 distinct splicing variants and S1 nuclease digestion. For 20 consensuses identified here using this methodology, 5 revealed a conserved splice site after inspection of the cDNA alignment against the human genome (exact splice sites). For 8 other consensuses, conserved splice sites were mapped at 2 to 30 bp from the border, called proximal splice sites; for the other 7 consensuses, conserved splice sites were mapped at 40 to 800 bp, called distal splice sites. These latter cases showed a nonspecific activity of S1 nuclease in digesting double-strand DNA. From the 20 consensuses identified here, 5 were selected for reverse transcription-polymerase chain reaction validation, confirming the splice sites. These data showed the potential of the strategy in mapping splice sites. However, the lack of specificity of the S1 nuclease enzyme is a significant obstacle that impedes the use of this strategy in large-scale studies.

Alternative splicing enriched cDNA libraries identify breast cancer-associated transcripts

Background: Alternative splicing (AS) is a central mechanism in the generation of genomic complexity and is a major contributor to transcriptome and proteome diversity. Alterations of the splicing process can lead to deregulation of crucial cellular processes and have been associated with a large spectrum of human diseases. Cancer-associated transcripts are potential molecular markers and may contribute to the development of more accurate diagnostic and prognostic methods and also serve as therapeutic targets. Alternative splicing-enriched cDNA libraries have been used to explore the variability generated by alternative splicing. In this study, by combining the use of trapping heteroduplexes and RNA amplification, we developed a powerful approach that enables transcriptome-wide exploration of the AS repertoire for identifying AS variants associated with breast tumor cells modulated by ERBB2 (HER-2/neu) oncogene expression. Results: The human breast cell line (C5.2) and a pool of 5 ERBB2 over-expressing breast tumor samples were used independently for the construction of two AS-enriched libraries. In total, 2,048 partial cDNA sequences were obtained, revealing 214 alternative splicing sequence-enriched tags (ASSETs). A subset with 79 multiple exon ASSETs was compared to public databases and reported 138 different AS events. A high success rate of RT-PCR validation (94.5%) was obtained, and 2 novel AS events were identified. The influence of ERBB2-mediated expression on AS regulation was evaluated by capillary electrophoresis and probe-ligation approaches in two mammary cell lines (Hb4a and C5.2) expressing different levels of ERBB2. The relative expression balance between AS variants from 3 genes was differentially modulated by ERBB2 in this model system. Conclusions: In this study, we presented a method for exploring AS from any RNA source in a transcriptome-wide format, which can be directly easily adapted to next generation sequencers. We identified AS transcripts that were differently modulated by ERBB2-mediated expression and that can be tested as molecular markers for breast cancer. Such a methodology will be useful for completely deciphering the cancer cell transcriptome diversity resulting from AS and for finding more precise molecular markers.

Genome-wide SNP genotyping highlights the role of natural selection in Plasmodium falciparum population divergence

Background: The malaria parasite Plasmodium falciparum exhibits abundant genetic diversity, and this diversity is key to its success as a pathogen. Previous efforts to study genetic diversity in P. falciparum have begun to elucidate the demographic history of the species, as well as patterns of population structure and patterns of linkage disequilibrium within its genome. Such studies will be greatly enhanced by new genomic tools and recent large-scale efforts to map genomic variation. To that end, we have developed a high throughput single nucleotide polymorphism (SNP) genotyping platform for P. falciparum. Results: Using an Affymetrix 3,000 SNP assay array, we found roughly half the assays (1,638) yielded high quality, 100% accurate genotyping calls for both major and minor SNP alleles. Genotype data from 76 global isolates confirm significant genetic differentiation among continental populations and varying levels of SNP diversity and linkage disequilibrium according to geographic location and local epidemiological factors. We further discovered that nonsynonymous and silent (synonymous or noncoding) SNPs differ with respect to within-population diversity, interpopulation differentiation, and the degree to which allele frequencies are correlated between populations. Conclusions: The distinct population profile of nonsynonymous variants indicates that natural selection has a significant influence on genomic diversity in P. falciparum, and that many of these changes may reflect functional variants deserving of follow-up study. Our analysis demonstrates the potential for new high-throughput genotyping technologies to enhance studies of population structure, natural selection, and ultimately enable genome-wide association studies in P. falciparum to find genes underlying key phenotypic traits.

Systematic Management of Variability in UML-based Software Product Lines

This paper presents SMarty, a variability management approach for UML-based software product lines (PL). SMarty is supported by a UML profile, the SMartyProfile, and a process for managing variabilities, the SMartyProcess. SMartyProfile aims at representing variabilities, variation points, and variants in UML models by applying a set of stereotypes. SMartyProcess consists of a set of activities that is systematically executed to trace, identify, and control variabilities in a PL based on SMarty. It also identifies variability implementation mechanisms and analyzes specific product configurations. In addition, a more comprehensive application of SMarty is presented using SEI's Arcade Game Maker PL. An evaluation of SMarty and related work are discussed.

Crystallization and preliminary structural analysis of the giant haemoglobin from Glossoscolex paulistus at 3.2 angstrom

Glossoscolex paulistus is a free-living earthworm encountered in south-east Brazil. Its oxygen transport requirements are undertaken by a giant extracellular haemoglobin, or erythrocruorin (HbGp), which has an approximate molecular mass of 3.6 MDa and, by analogy with its homologue from Lumbricus terrestris (HbLt), is believed to be composed of a total of 180 polypeptide chains. In the present work the full 3.6 MDa particle in its cyanomet state was purified and crystallized using sodium citrate or PEG8000 as precipitant. The crystals contain one-quarter of the full particle in the asymmetric unit of the I222 cell and have parameters of a = 270.8 angstrom, b = 320.3 angstrom and c = 332.4 angstrom. Diffraction data were collected to 3.15 angstrom using synchrotron radiation on beamline X29A at the Brookhaven National Laboratory and represent the highest resolution data described to date for similar erythrocruorins. The structure was solved by molecular replacement using a search model corresponding to one-twelfth of its homologue from HbLt. This revealed that HbGp belongs to the type I class of erythrocruorins and provided an interpretable initial electron density map in which many features including the haem groups and disulfide bonds could be identified.

Validation of dot blot hybridization and denaturing high performance liquid chromatography as reliable methods for TP53 codon 72 genotyping in molecular epidemiologic studies

Background: Mutations in TP53 are common events during carcinogenesis. In addition to gene mutations, several reports have focused on TP53 polymorphisms as risk factors for malignant disease. Many studies have highlighted that the status of the TP53 codon 72 polymorphism could influence cancer susceptibility. However, the results have been inconsistent and various methodological features can contribute to departures from Hardy-Weinberg equilibrium, a condition that may influence the disease risk estimates. The most widely accepted method of detecting genotyping error is to confirm genotypes by sequencing and/or via a separate method. Results: We developed two new genotyping methods for TP53 codon 72 polymorphism detection: Denaturing High Performance Liquid Chromatography (DHPLC) and Dot Blot hybridization. These methods were compared with Restriction Fragment Length Polymorphism (RFLP) using two different restriction enzymes. We observed high agreement among all methodologies assayed. Dot-blot hybridization and DHPLC results were more highly concordant with each other than when either of these methods was compared with RFLP. Conclusions: Although variations may occur, our results indicate that DHPLC and Dot Blot hybridization can be used as reliable screening methods for TP53 codon 72 polymorphism detection, especially in molecular epidemiologic studies, where high throughput methodologies are required.

Multifractal regime transition in a modified minority game model

The search for more realistic modeling of financial time series reveals several stylized facts of real markets. In this work we focus on the multifractal properties found in price and index signals. Although the usual minority game (MG) models do not exhibit multifractality, we study here one of its variants that does. We show that the nonsynchronous MG models in the nonergodic phase is multifractal and in this sense, together with other stylized facts, constitute a better modeling tool. Using the structure function (SF) approach we detected the stationary and the scaling range of the time series generated by the MG model and, from the linear (non-linear) behavior of the SF we identified the fractal (multifractal) regimes. Finally, using the wavelet transform modulus maxima (WTMM) technique we obtained its multifractal spectrum width for different dynamical regimes. (C) 2009 Elsevier Ltd. All rights reserved.

Effect of physical exercise on risk factors for chronic diseases in obeses women

This study aimed to evaluate the effects of physical exercise on body weight reduction. For 12 weeks, 22 obese women (BMI>30 kg/m(2)) were submitted to a physical exercise program. At the beginning and at the final of the program there were evaluated: BMI, waist (WC) and hip circumferences (HC), and waist-hip ratio (WHR); body composition by DEXA; hemoglobin and erythroctye, total cholesterol, HDL and LDL, triacylglycerol and blood glucose; aerobic power. At the final of the program, aerobic power, hemoglobin and erythrocyte values were significantly increased, confirming the physical training effects. Related to anthropometric values, only the visceral fat (WC, HC and WHR) were reduced. The exercise shows to be an important supporting in the body weight loss program, not exactly promoting body weight loss, but lowering risk factors to develop chronic diseases.