Expression of VCAN and its receptors in canine mammary carcinomas with or without myoepithelial proliferation.

The proteoglycan versican (VCAN) plays a complex role in cancer. The expression of this molecule has been related to invasion and progression in malignant mixed tumors, such as carcinoma in mixed tumors (CMT) of the canine mammary gland. In addition, its interaction with surface cell receptors EGFR, HER-2 and CD44 in malignant epithelial cells may be responsible for proliferation and cellular motility in early stages of cancer. We comparatively evaluated the expression of this proteoglycan and its receptors in in situ and invasive areas of simple carcinomas (SC) and CMT to investigate similarities and differences between these histological types. Immunohistochemistry was performed with anti-VCAN, anti-CD44, anti-EGFR and anti-HER-2 antibodies in 32 cases of SC or CMT. VCAN was highly expressed in stroma adjacent to invasive areas in SC and CMT. CMTs presented comparatively higher expression of VCAN in stroma adjacent to in situ and in invasive areas than in corresponding areas in SCs. In CMT, EGFR and HER-2 expressions were higher in situ compared to invasive areas. In contrast, increased CD44 and EGFR expression was found in invasive areas in SC compared to CMT. These results indicate that versican expression is similarly associated with invasiveness in SC and CMT, however higher levels were seen in CMT suggesting that the presence of myoepithelial proliferation in this tumor type participates in stromal composition and promoting an increase in the expression of versican.

Association of whole mtDNA, an NADPH G11914A variant, and haplogroups with high physical performance in an elite military troop.

Physical performance is a multifactorial and complex trait influenced by environmental and hereditary factors. Environmental factors alone have been insufficient to characterize all outstanding phenotypes. Recent advances in genomic technologies have enabled the investigation of whole nuclear and mitochondrial genome sequences, increasing our ability to understand interindividual variability in physical performance. Our objective was to evaluate the association of mitochondrial polymorphic loci with physical performance in Brazilian elite military personnel. Eighty-eight male military personnel who participated in the Command Actions Course of the Army were selected. Total DNA was obtained from blood samples and a complete mitochondrial genome (mtDNA) was sequenced using Illumina MiSeq platform. Twenty-nine subjects completed the training program (FINISHED, 'F'), and fifty-nine failed to complete (NOT_FINISHED, 'NF'). The mtDNA from NF was slightly more similar to genomes from African countries frequently related to endurance level. Twenty-two distinct mtDNA haplogroups were identified corroborating the intense genetic admixture of the Brazilian population, but their distribution was similar between the two groups (FST=0.0009). Of 745 polymorphisms detected in the mtDNA, the position G11914A within the NADPH gene component of the electron transport chain, was statistically different between F and NF groups (P=0.011; OR: 4.286; 95%CI: 1.198-16.719), with a higher frequency of the G allele in group F individuals). The high performance of military personnel may be mediated by performance-related genomic traits. Thus, mitochondrial genetic markers such as the ND4 gene may play an important role on physical performance variability.

Association of whole mtDNA, an NADPH G11914A variant, and haplogroups with high physical performance in an elite military troop

Physical performance is a multifactorial and complex trait influenced by environmental and hereditary factors. Environmental factors alone have been insufficient to characterize all outstanding phenotypes. Recent advances in genomic technologies have enabled the investigation of whole nuclear and mitochondrial genome sequences, increasing our ability to understand interindividual variability in physical performance. Our objective was to evaluate the association of mitochondrial polymorphic loci with physical performance in Brazilian elite military personnel. Eighty-eight male military personnel who participated in the Command Actions Course of the Army were selected. Total DNA was obtained from blood samples and a complete mitochondrial genome (mtDNA) was sequenced using Illumina MiSeq platform. Twenty-nine subjects completed the training program (FINISHED, 'F'), and fifty-nine failed to complete (NOT_FINISHED, 'NF'). The mtDNA from NF was slightly more similar to genomes from African countries frequently related to endurance level. Twenty-two distinct mtDNA haplogroups were identified corroborating the intense genetic admixture of the Brazilian population, but their distribution was similar between the two groups (FST=0.0009). Of 745 polymorphisms detected in the mtDNA, the position G11914A within the NADPH gene component of the electron transport chain, was statistically different between F and NF groups (P=0.011; OR: 4.286; 95%CI: 1.198-16.719), with a higher frequency of the G allele in group F individuals). The high performance of military personnel may be mediated by performance-related genomic traits. Thus, mitochondrial genetic markers such as the ND4 gene may play an important role on physical performance variability.

Recovery & identification of human Y-STR DNA from immatures of chrysomya albiceps (Diptera: Calliphoridae). Simulation of sexual crime investigation involving victim corpse in state of decay.

The number of sexual crimes in Brazil, as in several other countries, is very high. In many of these crimes the women raped are murdered and their bodies are found days later, in an advanced state of decomposition, with intense cadaverous fauna. Forensic Entomology studies insects and other arthropods that can be used in the expert analysis of various types of crimes. Diptera, the order of insects that comprises the two-winged or true flies, represents one of the largest known groups of insects and is the principal source of cadaveric entomofauna. Members of its Calliphoridae family are observed in cadavers in all phases of decomposition. The retrieval and identification of human Y-STR DNA from the gastrointestinal tract of Calliphoridae species Chrysomya albiceps maggots and pupae can provide a good tool for the gathering of evidence in sexual crime investigations involving rape and death, in which the abandoned victim's body is found in a putrefied state. In this study, the animal model used was a female pig, Sus scrofa, which was sacrificed in a forested area with three shots from a 0.40 calibre Taurus pistol, and inoculated with semen to its anal and vaginal regions, simulating rape and homicide. During decomposition, 20-80 maggots were collected every 24 h and preserved in 70 % alcohol, totalling 289 maggots and 157 pupae (446 immatures) over a period of 14 days (336 h) of decomposition. Each maggot was then dissected for removal of the digestive tract, which was placed in extraction buffer. The molecular phase proceeded with extraction, quantification, amplification and capillary electrophoresis of samples, testing 16 STR loci of the Y chromosome. It was possible to establish a partial Y-STR DNA profile, with the amplification of up to eight sites, by considering a combination of the samples taken at hours 144 h, 168 h, 192 h, 216 h, 240 h, 288 h, 312 h and 336 h.

Generation of patient-specific induced pluripotent stem cell lines from one patient with Jervell and Lange-Nielsen syndrome, one with type 1 long QT syndrome and two healthy relatives.

Four human iPSC cell lines (one Jervell and Lange-Nielsen Syndrome, one Long QT Syndrome-type 1 and two healthy controls) were generated from peripheral blood obtained from donors belonging to the same family. CytoTune™-iPS 2.0 Sendai Reprogramming Kit (containing OCT3/4, KLF4, SOX2 and cMYC as reprogramming factors) was used to generate all cell lines. The four iPSCs have normal karyotype, express pluripotency markers as determined by RT-PCR and flow cytometry and differentiated spontaneously in vitro into cells of the three germ layers, confirming their pluripotent capacity.

Evaluating DNA evidence in a genetically complex population.

In forensic genetics, the likelihood ratio (LR), measuring the value of DNA profile evidence, is computed from a database of allele frequencies. Here, we address the choice of database and adjustments for population structure and sample size in the context of Brazil. The Brazilian population underwent a complex process of colonization, migration and mating, which created an admixed genetic composition that makes it difficult to obtain an appropriate database for a given case. National databases are now available, as well as databases for many Brazilian states. However, those databases are not statistically random samples, and state boundaries may not accurately reflect the sub-structuring of genetic diversity. We compared the LR calculated using the relevant state-specific database with the statistics calculated when a national database and when international databases were used. We evaluated two methods of adjustment for population structure, due to Wright [13] and Balding and Nichols [14]. We also considered two adjustments for database sample size: the Balding size bias correction [15] and a minimum allele frequency [16]. Our results show that the use of a national database with the Balding and Nichols adjustment and θ = 0.002 generated lower LR values than did the state-specific database in more than 50% of the profiles simulated using the state-based allele frequencies, while θ = 0.01 produced lower LRs for more than 90% of the profiles. We conclude that the utilization of a national database for Brazilian cases can be justified in association with the appropriate adjustment for population structure.

Evaluation of InnoTyper® 21 in a sample of Rio de Janeiro population as an alternative forensic panel.

The use of bi-allelic markers such as retrotransposable element insertion polymorphisms or Innuls (for insertion/null) can overcome some limitations of short tandem repeat (STR) loci in typing forensic biological evidence. This study investigated the efficiency of the InnoTyper® 21 Innul markers in an urban admixed population sample in Rio de Janeiro (n = 40) and one highly compromised sample collected as evidence by the Rio de Janeiro police. No significant departures from Hardy-Weinberg equilibrium were detected after the Bonferroni correction (α' ≈ 0.05/20, p < 0.0025), and no significant linkage disequilibrium was observed between markers. Assuming loci independence, the cumulative random match probability (RMP) was 2.3 × 10-8. A lower mean Fis value was obtained for this sample population compared with those of three North American populations (African-American, Southwest Hispanic, US Caucasian). Principal component analysis with the three North American populations and one from 21 East Asian population showed that African Americans segregated as an independent group while US Caucasian, Southwest Hispanic, East Asian, and Rio de Janeiro populations are in a single large heterogeneous group. Also, a full Innuls profile was produced from an evidence sample, despite the DNA being highly degraded. In conclusion, this system is a useful complement to standard STR kits.

DNA-based identification of forensically important species of Sarcophagidae (Insecta: Diptera) from Rio de Janeiro, Brazil.

Sarcophagidae, or flesh flies, are of great importance in forensic entomology, but their effective application requires precise taxonomic identification, which relies almost exclusively on characteristics of the male genitalia. Given that female flies and larvae are most abundant in animal carcasses or on corpses, precise morphological identification can be difficult; therefore, DNA sequencing can be an additional tool for use in taxonomic identification. This paper analyzes part of the mitochondrial cytochrome c oxidase subunit I (COI) gene from three Sarcophagidae species of forensic importance in the City of Rio de Janeiro: Oxysarcodexia fluminensis, Peckia chrysostoma, and Peckia intermutans. COI fragments of 400 bp from 36 specimens of these three species were sequenced. No intraspecific differences were found among specimens of O. fluminensis, but P. chrysostoma and P. intermutans each had two haplotypes, ranging from 0 to 0.7%. The interspecific divergence was 8.5-11.6%, corroborating previously reported findings.

The heritable path of human physical performance: from single polymorphisms to the "next generation".

Human physical performance is a complex multifactorial trait. Historically, environmental factors (e.g., diet, training) alone have been unable to explain the basis of all prominent phenotypes for physical performance. Therefore, there has been an interest in the study of the contribution of genetic factors to the development of these phenotypes. Support for a genetic component is found with studies that shown that monozygotic twins were more similar than were dizygotic twins for many physiological traits. The evolution of molecular techniques and the ability to scan the entire human genome enabled association of several genetic polymorphisms with performance. However, some biases related to the selection of cohorts and inadequate definition of the study variables have complicated the already difficult task of studying such a large and polymorphic genome, often resulting in inconsistent results about the influence of candidate genes. This review aims to provide a critical overview of heritable genetic aspects. Novel molecular technologies, such as next-generation sequencing, are discussed and how they can contribute to improving understanding of the molecular basis for athletic performance. It is important to ensure that the large amount of data that can be generated using these tools will be used effectively by ensuring well-designed studies.

Evaluation of a 49 InDel Marker HID panel in two specific populations of South America and one population of Northern Africa.

The majority of STR loci are not ideal for the analysis of forensic samples with degraded and/or low template DNA. One alternative to overcome these limitations is the use of bi-allelic markers, which have low mutation rates and shorter amplicons. Human identification (HID) InDel marker panels have been described in several countries, including Brazil. The commercial kit available is, however, mostly suitable for Europeans, with lower discrimination power for other population groups. Recently, a combination of 49 InDel markers used in four different ethnic groups in the USA has been shown to be more informative than another panel from Portugal, already tested in a Rio de Janeiro sample. However, these 49 InDels have yet to be applied to other admixed or isolated populations. We assessed the efficiency of this panel in two urban admixed populations (Rio de Janeiro, Brazil; Tripoli, Libya) and one isolated Native Brazilian community. All markers are in Hardy-Weinberg equilibrium (HWE) after the Bonferroni correction, and no Linkage disequilibrium was detected. Assuming loci independence and no substructure effect, cumulative RMP was 2.7×10(-18), 1.5×10(-20), and 4.5×10(-20) for Native Brazilian, Rio de Janeiro, and Tripoli populations, respectively. The overall Fst value was 0.05512. Rio de Janeiro and Tripoli showed similar admixture levels, however for Native Brazilians one parental cluster represented over 60 % of the total parental population. We conclude that this panel is suitable for HID on these urban populations, but is less efficient for the isolated group.

The association of ACE gene D/I polymorphism with cardiovascular risk factors in a population from Rio de Janeiro.

Our aim was to determine the frequencies of the angiotensin-converting enzyme (ACE) gene alleles D and I and any associations to cardiovascular risk factors in a population sample from Rio de Janeiro, Brazil. Eighty-four adults were selected consecutively during a 6-month period from a cohort subgroup of a previous large cross-sectional survey in Rio de Janeiro. Anthropometric data and blood pressure measurements, echocardiogram, albuminuria, glycemia, lipid profile, and ACE genotype and serum enzyme activity were determined. The frequency of the ACE*D and I alleles in the population under study, determined by PCR, was 0.59 and 0.41, respectively, and the frequencies of the DD, DI, and II genotypes were 0.33, 0.51, and 0.16, respectively. No association between hypertension and genotype was detected using the Kruskal-Wallis method. Mean plasma ACE activity (U/mL) in the DD (N = 28), DI (N = 45) and II (N = 13) groups was 43 (in males) and 52 (in females), 37 and 39, and 22 and 27, respectively; mean microalbuminuria (mg/dL) was 1.41 and 1.6, 0.85 and 0.9, and 0.6 and 0.63, respectively; mean HDL cholesterol (mg/dL) was 40 and 43, 37 and 45, and 41 and 49, respectively, and mean glucose (mg/dL) was 93 and 108, 107 and 98, and 85 and 124, respectively. A high level of ACE activity and albuminuria, and a low level of HDL cholesterol and glucose, were found to be associated with the DD genotype. Finally, the II genotype was found to be associated with variables related to glucose intolerance.

The association of ACE gene D/I polymorphism with cardiovascular risk factors in a population from Rio de Janeiro

Our aim was to determine the frequencies of the angiotensin-converting enzyme (ACE) gene alleles D and I and any associations to cardiovascular risk factors in a population sample from Rio de Janeiro, Brazil. Eighty-four adults were selected consecutively during a 6-month period from a cohort subgroup of a previous large cross-sectional survey in Rio de Janeiro. Anthropometric data and blood pressure measurements, echocardiogram, albuminuria, glycemia, lipid profile, and ACE genotype and serum enzyme activity were determined. The frequency of the ACE*D and I alleles in the population under study, determined by PCR, was 0.59 and 0.41, respectively, and the frequencies of the DD, DI, and II genotypes were 0.33, 0.51, and 0.16, respectively. No association between hypertension and genotype was detected using the Kruskal-Wallis method. Mean plasma ACE activity (U/mL) in the DD (N = 28), DI (N = 45) and II (N = 13) groups was 43 (in males) and 52 (in females), 37 and 39, and 22 and 27, respectively; mean microalbuminuria (mg/dL) was 1.41 and 1.6, 0.85 and 0.9, and 0.6 and 0.63, respectively; mean HDL cholesterol (mg/dL) was 40 and 43, 37 and 45, and 41 and 49, respectively, and mean glucose (mg/dL) was 93 and 108, 107 and 98, and 85 and 124, respectively. A high level of ACE activity and albuminuria, and a low level of HDL cholesterol and glucose, were found to be associated with the DD genotype. Finally, the II genotype was found to be associated with variables related to glucose intolerance.

Seroprevalence of human parvovirus B19 in a suburban population in São Paulo, Brazil

OBJETIVO: Analisar a prevalência de anticorpos IgG ao parvovírus humano B19. MÉTODOS: Estudo transversal em uma comunidade de subúrbio de São Paulo, Brasil, de novembro 1990 a janeiro de 1991. Amostras aleatórias (N=435) e representativas de soro foram coletadas de crianças sadias a partir de 15 dias de idade e de adultos com até 40 anos. Os anticorpos IgG ao parvovírus humano B19 foram detectados pelo teste ELISA. RESULTADOS: A prevalência de anticorpos IgG ao parvovírus B19 foi de 87 por cento dos recém-nascidos. A prevalência de anticorpos IgG de origem materna decaiu exponencialmente até o 19o mês de idade. Baixa prevalência de anticorpos foi observada nos primeiros quatro anos de vida, aumentando até 72 por cento no grupo etário de 31-40 anos. A idade média de aquisição da primeira infecção nesta comunidade é de 21 ± 7 anos. A idade ótima para se vacinar as crianças desta comunidade com uma vacina hipotética é de um ano de idade. CONCLUSÕES: A prevalência de anticorpos IgG ao parvovírus B19 foi alta entre recém-nascidos e no grupo etário 31-40 anos. A análise por estrutura etária mostrou padrão similar aos estudos prévios relacionados à baixa prevalência de infecção em crianças que aumenta com a idade.

Seroprevalence of human parvovirus B19 in a suburban population in São Paulo, Brazil.

OBJECTIVE: To analyze the prevalence of IgG antibodies to human parvovirus B19. METHODS: Cross-sectional study in a suburban community in São Paulo, Southeastern Brazil, between November 1990 and January 1991. Randomly selected (N=435) representative samples of sera were collected from healthy children older than 15 days old and adults up to 40 years old. IgG antibodies were detected using ELISA. RESULTS: High prevalence of IgG antibodies to B19 parvovirus was found in 87% of newborns. The prevalence of maternally derived IgG antibodies exponentially plunged up to the 19th month of age. Low prevalence of antibodies was found in the first 4 years of life, increasing up to 72% in those aged 31-40 years. It was estimated that the average age of first infection in this population is 21 +/- 7 years old and the optimal age for vaccination with a hypothetical vaccine would be 1 year of age. CONCLUSIONS: Parvovirus B19 IgG antibody prevalence was high in newborns and those aged 31-40 years. The analysis by age groups showed a pattern similar to that found in previous studies, i.e., low prevalence of infection in children that increases with age.

Analysis of renin-angiotensin-aldosterone system gene polymorphisms in resistant hypertension.

Essential hypertension is a disease multifactorially triggered by genetic and environmental factors. The contribution of genetic polymorphisms of the renin-angiotensin-aldosterone system and clinical risk factors to the development of resistant hypertension was evaluated in 90 hypertensive patients and in 115 normotensive controls living in Southwestern Brazil. Genotyping for insertion/deletion of angiotensin-converting enzyme, angiotensinogen M235T, angiotensin II type 1 receptor A1166C, aldosterone synthase C344T, and mineralocorticoid receptor A4582C polymorphisms was performed by PCR, with further restriction analysis when required. The influence of genetic polymorphisms on blood pressure variation was assessed by analysis of the odds ratio, while clinical risk factors were evaluated by logistic regression. Our analysis indicated that individuals who carry alleles 235-T, 1166-A, 344-T, or 4582-C had a significant risk of developing resistant hypertension (P < 0.05). Surprisingly, when we tested individuals who carried the presumed risk genotypes A1166C, C344T, and A4582C we found that these genotypes were not associated with resistant hypertension. However, a gradual increase in the risk to develop resistant hypertension was detected when the 235-MT and TT genotypes were combined with one, two or three of the supposedly more vulnerable genotypes - A1166C (AC/AA), C344T (TC/TT) and A4582C (AC/CC). Analysis of clinical parameters indicated that age, body mass index and gender contribute to blood pressure increase (P < 0.05). These results suggest that unfavorable genetic renin-angiotensin-aldosterone system patterns and clinical risk variables may contribute to increasing the risk for the development of resistant hypertension in a sample of the Brazilian population.

Analysis of renin-angiotensin-aldosterone system gene polymorphisms in resistant hypertension

Essential hypertension is a disease multifactorially triggered by genetic and environmental factors. The contribution of genetic polymorphisms of the renin-angiotensin-aldosterone system and clinical risk factors to the development of resistant hypertension was evaluated in 90 hypertensive patients and in 115 normotensive controls living in Southwestern Brazil. Genotyping for insertion/deletion of angiotensin-converting enzyme, angiotensinogen M235T, angiotensin II type 1 receptor A1166C, aldosterone synthase C344T, and mineralocorticoid receptor A4582C polymorphisms was performed by PCR, with further restriction analysis when required. The influence of genetic polymorphisms on blood pressure variation was assessed by analysis of the odds ratio, while clinical risk factors were evaluated by logistic regression. Our analysis indicated that individuals who carry alleles 235-T, 1166-A, 344-T, or 4582-C had a significant risk of developing resistant hypertension (P < 0.05). Surprisingly, when we tested individuals who carried the presumed risk genotypes A1166C, C344T, and A4582C we found that these genotypes were not associated with resistant hypertension. However, a gradual increase in the risk to develop resistant hypertension was detected when the 235-MT and TT genotypes were combined with one, two or three of the supposedly more vulnerable genotypes - A1166C (AC/AA), C344T (TC/TT) and A4582C (AC/CC). Analysis of clinical parameters indicated that age, body mass index and gender contribute to blood pressure increase (P < 0.05). These results suggest that unfavorable genetic renin-angiotensin-aldosterone system patterns and clinical risk variables may contribute to increasing the risk for the development of resistant hypertension in a sample of the Brazilian population.

Personalized diagnosis by cached solutions with hypertension as a study model.

Statistical modeling of links between genetic profiles with environmental and clinical data to aid in medical diagnosis is a challenge. Here, we present a computational approach for rapidly selecting important clinical data to assist in medical decisions based on personalized genetic profiles. What could take hours or days of computing is available on-the-fly, making this strategy feasible to implement as a routine without demanding great computing power. The key to rapidly obtaining an optimal/nearly optimal mathematical function that can evaluate the "disease stage" by combining information of genetic profiles with personal clinical data is done by querying a precomputed solution database. The database is previously generated by a new hybrid feature selection method that makes use of support vector machines, recursive feature elimination and random sub-space search. Here, to evaluate the method, data from polymorphisms in the renin-angiotensin-aldosterone system genes together with clinical data were obtained from patients with hypertension and control subjects. The disease "risk" was determined by classifying the patients' data with a support vector machine model based on the optimized feature; then measuring the Euclidean distance to the hyperplane decision function. Our results showed the association of renin-angiotensin-aldosterone system gene haplotypes with hypertension. The association of polymorphism patterns with different ethnic groups was also tracked by the feature selection process. A demonstration of this method is also available online on the project's web site.

Personalized diagnosis by cached solutions with hypertension as a study model

Statistical modeling of links between genetic profiles with environmental and clinical data to aid in medical diagnosis is a challenge. Here, we present a computational approach for rapidly selecting important clinical data to assist in medical decisions based on personalized genetic profiles. What could take hours or days of computing is available on-the-fly, making this strategy feasible to implement as a routine without demanding great computing power. The key to rapidly obtaining an optimal/nearly optimal mathematical function that can evaluate the [quot ]disease stage[quot ] by combining information of genetic profiles with personal clinical data is done by querying a precomputed solution database. The database is previously generated by a new hybrid feature selection method that makes use of support vector machines, recursive feature elimination and random sub-space search. Here, to evaluate the method, data from polymorphisms in the renin-angiotensin-aldosterone system genes together with clinical data were obtained from patients with hypertension and control subjects. The disease [quot ]risk[quot ] was determined by classifying the patients' data with a support vector machine model based on the optimized feature; then measuring the Euclidean distance to the hyperplane decision function. Our results showed the association of renin-angiotensin-aldosterone system gene haplotypes with hypertension. The association of polymorphism patterns with different ethnic groups was also tracked by the feature selection process. A demonstration of this method is also available online on the project's web site.