Manipulação do genoma humano: ética e direito

A biologia molecular tem fornecido as ferramentas básicas para os geneticistas se aprofundarem nos mecanismos moleculares que influem na variação das doenças. Deve-se destacar a responsabilidade científica e moral dos pesquisadores, uma vez que os cientistas devem imaginar as consequências morais da aplicação comercial de testes genéticos, já que esse fato envolve não só o indivíduo e suas famílias, mas toda a população. Além de ser preciso, também, fazer uma reflexão sobre como essas informações do genoma humano serão utilizadas, para o bem ou mal. O objetivo desta revisão foi trazer à luz do conhecimento dados sobre características éticas da aplicação da biologia molecular, relacionando-a com os direitos do ser humano. Após análise bibliográfica, pôde-se observar que o Projeto Genoma Humano gerou várias possibilidades, como identificação de genes associados a doenças com propriedades sinergísticas, mas modificando às vezes comportamentos ao intervir geneticamente no ser humano, trazendo benefícios ou malefícios sociais. O grande desafio é decidir o que a humanidade pretende em relação a este gigantesco salto.

Nuclear and mitochondrial DNA markers in traceability of retail beef samples

Several characteristics are important in a traceability system of animal products, such as age at slaughter, breed composition, besides information of the productive chain. In general, the certification agent records information about the animals and the system which it came from, although cannot guarantee that the slaughtering, meat processing and distribution are error proof. Besides, there is a differential price, at least at the international market, based on sex and breed composition of the animals. Genetic markers allow identification of characteristics controlled in the beef cattle traceability program, as sex and breed composition, in order to correctly identify and appraise the final product for the consumer. The hypothesis of this study was that the majority beef samples retailed in the local market originate from female with a great participation of zebu breeds. Therefore, the objective of this work was to characterize retail beef samples with DNA markers that identify cattle sex and breed composition. Within 10 beef shops localized in Pirassununga, SP, Brazil, 61 samples were collected, all were genotyped as harboring Bos taurus mitochondrial DNA and 18 were positive for the Y chromosome amplification (male). For the marker sat1711b-Msp I the frequency of the allele A was 0.278 and for the marker Lhr-Hha I the frequency of the allele T was 0.417. The results of sat1711b-Msp I and Lhr-Hha I allelic frequencies are suggestive that the proportion of indicus genome compared with the taurine genome in the market meat is smaller than the observed in the Nellore breed. The procedure described in this study identified sex and subspecies characteristics of beef meat samples, with potential application in meat products certification in special as an auxiliary tool in beef cattle traceability programs.

Non-coding RNAs in schistosomes: an unexplored world

Non-coding RNAs (ncRNAs) were recently given much higher attention due to technical advances in sequencing which expanded the characterization of transcriptomes in different organisms. ncRNAs have different lengths (22 nt to >1, 000 nt) and mechanisms of action that essentially comprise a sophisticated gene expression regulation network. Recent publication of schistosome genomes and transcriptomes has increased the description and characterization of a large number of parasite genes. Here we review the number of predicted genes and the coverage of genomic bases in face of the public ESTs dataset available, including a critical appraisal of the evidence and characterization of ncRNAs in schistosomes. We show expression data for ncRNAs in Schistosoma mansoni. We analyze three different microarray experiment datasets: (1) adult worms' large-scale expression measurements; (2) differentially expressed S. mansoni genes regulated by a human cytokine (TNF-α) in a parasite culture; and (3) a stage-specific expression of ncRNAs. All these data point to ncRNAs involved in different biological processes and physiological responses that suggest functionality of these new players in the parasite's biology. Exploring this world is a challenge for the scientists under a new molecular perspective of host-parasite interactions and parasite development.

Rapid detection of bovine coronavirus by a semi-nested RT-PCR

Bovine coronavirus (BCoV) is a member of the group 2 of the Coronavirus (Nidovirales: Coronaviridae) and the causative agent of enteritis in both calves and adult bovine, as well as respiratory disease in calves. The present study aimed to develop a semi-nested RT-PCR for the detection of BCoV based on representative up-to-date sequences of the nucleocapsid gene, a conserved region of coronavirus genome. Three primers were designed, the first round with a 463bp and the second (semi-nested) with a 306bp predicted fragment. The analytical sensitivity was determined by 10-fold serial dilutions of the BCoV Kakegawa strain (HA titre: 256) in DEPC treated ultra-pure water, in fetal bovine serum (FBS) and in a BCoV-free fecal suspension, when positive results were found up to the 10-2, 10-3 and 10-7 dilutions, respectively, which suggests that the total amount of RNA in the sample influence the precipitation of pellets by the method of extraction used. When fecal samples was used, a large quantity of total RNA serves as carrier of BCoV RNA, demonstrating a high analytical sensitivity and lack of possible substances inhibiting the PCR. The final semi-nested RT-PCR protocol was applied to 25 fecal samples from adult cows, previously tested by a nested RT-PCR RdRp used as a reference test, resulting in 20 and 17 positives for the first and second tests, respectively, and a substantial agreement was found by kappa statistics (0.694). The high sensitivity and specificity of the new proposed method and the fact that primers were designed based on current BCoV sequences give basis to a more accurate diagnosis of BCoV-caused diseases, as well as to further insights on protocols for the detection of other Coronavirus representatives of both Animal and Public Health importance.

Leptin's effect on puberty in mice is relayed by the ventral premammillary nucleus and does not require signaling in Kiss1 neurons

Studies m hum ins and rodents indicate that a minimum amount of stored energy is required for normal pubertal development The adipocyte-derived hormone leptin is a key metabolic signal to the neuroendocrine reproductive axis Humans and mice lacking leptin or the leptin receptor (LepR) (ob/ob and db/db mice, respectively) are infertile and fail to enter puberty Leptin administration to leptin-deficient subjects and ob/ob mice induces puberty and restores fertility, but the exact site or sites of leptin action are unclear Here, we found that genetic deletion of LepR selectively from hypothalamic Kiss1 neurons m mice had no effect on puberty or fertility, indicating that direct leptin signaling m Kiss1 neurons is not required for these processes However, bilateral lesions of the ventral premammillary nucleus (PMV) of ob/ob mice blunted the ability of exogenous leptin to induce sexual maturation Moreover, unilateral reexpression of endogenous LepR m PMV neurons was sufficient to induce puberty and improve fertility m female LepR-null mice This LepR reexpression also normalized the increased hypothalamic GnRH content characteristic of leptin-signaling deficiency These data suggest that the PMV is a key site for leptin's permissive action at the onset of puberty and support the hypothesis that the multiple actions of leptin to control metabolism and reproduction at e anatomically dissociated

The mitochondrial genome of the phytopathogenic basidiomycete Moniliophthora perniciosa is 109 kb in size and contains a stable integrated plasmid

We present here the sequence of the mitochondrial genome of the basidiomycete phytopathogenic hemibiotrophic fungus Moniliophthora perniciosa, causal agent of the Witches` Broom Disease in Theobroma cacao. The DNA is a circular molecule of 109103 base pairs, with 31.9 % GC, and is the largest sequenced so far. This size is due essentially to the presence of numerous non-conserved hypothetical ORFs. It contains the 14 genes coding for proteins involved in the oxidative phosphorylation, the two rRNA genes, one ORF coding for a ribosomal protein (rps3), and a set of 26 tRNA genes that recognize codons for all amino acids. Seven homing endonucleases are located inside introns. Except atp8, all conserved known genes are in the same orientation. Phylogenetic analysis based on the cox genes agrees with the commonly accepted fungal taxonomy. An uncommon feature of this mitochondrial genome is the presence of a region that contains a set of four, relatively small, nested, inverted repeats enclosing two genes coding for polymerases with an invertron-type structure and three conserved hypothetical genes interpreted as the stable integration of a mitochondrial linear plasmid. The integration of this plasmid seems to be a recent evolutionary event that could have implications in fungal biology. This sequence is available under GenBank accession number AY376688. (c) 2008 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Frozen Chicken for Wild Fish: Nutritional Transition in the Brazilian Amazon Region Determined by Carbon and Nitrogen Stable Isotope Ratios in Fingernails

Objectives: Amazonian populations are experiencing dietary changes characteristic of the nutrition transition. However, the degree of change appears to vary between urban and rural settings. To investigate this process, we determined carbon and nitrogen stable isotope ratios in fingernails and dietary intake of Amazonian populations living along a rural to urban continuum along the Solimoes River in Brazil. Methods: Carbon and nitrogen stable isotope ratios were analyzed from the fingernails of 431 volunteer subjects living in different settings ranging from rural villages, small towns to urban centers along the Solimoes River. Data from 200 dietary intake surveys were also collected using food frequency questionnaires and 24-h recall interviews in an effort to determine qualitative aspects of diet composition. Results: Fingernail delta(13)C values (mean standard deviation) were -23.2 +/- 1.3, 20.2 +/- 1.5, and 17.4 +/- 1.3 parts per thousand and delta(15)N values were 11.8 +/- 0.6, 10.4 +/- 0.8, and 10.8 +/- 0.7 parts per thousand for those living in rural villages, small towns, and major cities, respectively. We found a gradual increase in the number of food items derived from C(4) plant types (meat and sugar) and the replacement of food items derived from C(3) plant types (fish and manioc flour) with increasing size of urban centers. Conclusion: Increasing urbanization in the Brazilian Amazon is associated with a significant change in food habits with processed and industrialized products playing an increasingly important role in the diet and contributing to the nutrition transition in the region. Am. J. Hum. Biol. 23:642-650, 2011. (C) 2011 Wiley-Liss, Inc.

Frequency Domain Connectivity Identification: An Application of Partial Directed Coherence in fMRI

Functional magnetic resonance imaging (fMRI) has become an important tool in Neuroscience due to its noninvasive and high spatial resolution properties compared to other methods like PET or EEG. Characterization of the neural connectivity has been the aim of several cognitive researches, as the interactions among cortical areas lie at the heart of many brain dysfunctions and mental disorders. Several methods like correlation analysis, structural equation modeling, and dynamic causal models have been proposed to quantify connectivity strength. An important concept related to connectivity modeling is Granger causality, which is one of the most popular definitions for the measure of directional dependence between time series. In this article, we propose the application of the partial directed coherence (PDC) for the connectivity analysis of multisubject fMRI data using multivariate bootstrap. PDC is a frequency domain counterpart of Granger causality and has become a very prominent tool in EEG studies. The achieved frequency decomposition of connectivity is useful in separating interactions from neural modules from those originating in scanner noise, breath, and heart beating. Real fMRI dataset of six subjects executing a language processing protocol was used for the analysis of connectivity. Hum Brain Mapp 30:452-461, 2009. (C) 2007 Wiley-Liss, Inc.

Risk Profiles and Penetrance Estimations in Multiple Endocrine Neoplasia Type 2A Caused by Germline RET Mutations Located in Exon 10

Multiple endocrine neoplasia type 2 is characterized by germline mutations in RET. For exon 10, comprehensive molecular and corresponding phenotypic data are scarce. The International RET Exon 10 Consortium, comprising 27 centers from 15 countries, analyzed patients with RET exon 10 mutations for clinical-risk profiles. Presentation, age-dependent penetrance, and stage at presentation of medullary thyroid carcinoma (MTC), pheochromocytoma, and hyperparathyroidism were studied. A total of 340 subjects from 103 families, age 4-86, were registered. There were 21 distinct single nucleotide germline mutations located in codons 609 (45 subjects), 611 (50), 618 (94), and 620 (151). MTC was present in 263 registrants, pheochromocytoma in 54, and hyperparathyroidism in 8 subjects. Of the patients with MTC, 53% were detected when asymptomatic, and among those with pheochromocytoma, 54%. Penetrance for MTC was 4% by age 10, 25% by 25, and 80% by 50. Codon-associated penetrance by age 50 ranged from 60% (codon 611) to 86% (620). More advanced stage and increasing risk of metastases correlated with mutation in codon position (609-620) near the juxtamembrane domain. Our data provide rigorous bases for timing of premorbid diagnosis and personalized treatment/prophylactic procedure decisions depending on specific RET exon 10 codons affected. Hum Mutat 32:51-58, 2011. (C) 2010 Wiley-Liss, Inc.

An fMRI Normative Database for Connectivity Networks Using One-Class Support Vector Machines

The application of functional magnetic resonance imaging (fMRI) in neuroscience studies has increased enormously in the last decade. Although primarily used to map brain regions activated by specific stimuli, many studies have shown that fMRI can also be useful in identifying interactions between brain regions (functional and effective connectivity). Despite the widespread use of fMRI as a research tool, clinical applications of brain connectivity as studied by fMRI are not well established. One possible explanation is the lack of normal pattern, and intersubject variability-two variables that are still largely uncharacterized in most patient populations of interest. In the current study, we combine the identification of functional connectivity networks extracted by using Spearman partial correlation with the use of a one-class support vector machine in order construct a normative database. An application of this approach is illustrated using an fMRI dataset of 43 healthy Subjects performing a visual working memory task. In addition, the relationships between the results obtained and behavioral data are explored. Hum Brain Mapp 30:1068-1076, 2009. (C) 2008 Wiley-Liss. Inc.

Bioimpedance Analysis: Should it be Used in Morbid Obesity?

Objectives: Questions about reliability of bioimpedance analysis (BIA) in morbidly obese subjects have curtailed its use in this setting, but metabolic implications might reignite the debate. In a prospective study, it was aimed to analyze anthropometric and clinical associations. Methods: Bariatric candidates (n = 94) with or without metabolic syndrome were consecutively investigated. Age was 34.9 +/- 10.4 years (68.1% females), and BMI was 40.8 +/- 4.6 kg m(-2). Methods included single-frequency BIA, anthropometrics, inflammatory indices, and general biochemical profile. Results: Body composition results (water, fat) in females, but not in males, were entirely consistent with the literature. In both genders good association was observed with anthropometrics (BMI, waist circumference), inflammatory indices (ferritin, C-reactive protein) and general biochemical variables. Anthropometric measurements also displayed comparable associations. Multivariate tests including the two sets of measurements indicated no predominance of one method over the other, one complementing the other as metabolic marker. Conclusions: BIA limitations were mostly relevant for males, not females. Despite such discrepancies, good associations with anthropometry were demonstrated for both genders. Correlations with liver enzymes, and indices of protein, carbohydrate, and lipid metabolism could be demonstrated. BIA deserves more investigations concerning liver steatosis and ongoing inflammation, and it could contribute as well, synergistically with anthropometry, to monitor weight loss, body fat shifts, and metabolic risk. Am. J. Hum. Biol. 23: 420-422, 2011. (c) 2011 Wiley-Liss, Inc.

The Cholinergic System in Mild Cognitive Impairment and Alzheimer`s Disease: An In Vivo MRI and DTI study

Few studies have investigated in vivo changes of the cholinergic basal forebrain in Alzheimer`s disease (AD) and amnestic mild cognitive impairment (MCI), an at risk stage of AD. Even less is known about alterations of cortical projecting fiber tracts associated with basal forebrain atrophy. In this study, we determined regional atrophy within the basal forebrain in 21 patients with AD and 16 subjects with MCI compared to 20 healthy elderly subjects using deformation-based morphometry of MRI scans. We assessed effects of basal forebrain atrophy on fiber tracts derived from high-resolution diffusion tensor imaging (DTI) using tract-based spatial statistics. We localized significant effects relative to a map of cholinergic nuclei in MRI standard space as determined from a postmortem brain. Patients with AD and MCI subjects showed reduced volumes in basal forebrain areas corresponding to anterior medial and lateral, intermediate and posterior nuclei of the Nucleus basalis of Meynert (NbM) as well as in the diagonal band of Broca nuclei (P < 0.01). Effects in MCI subjects were spatially more restricted than in AD, but occurred at similar locations. The volume of the right antero-lateral NbM nucleus was correlated with intracortical projecting fiber tract integrity such as the corpus callosum, cingulate, and the superior longitudinal, inferior longitudinal, inferior fronto-occipital, and uncinate fasciculus (P < 0.05, corrected for multiple comparisons). Our findings suggest that a multimodal MRI-DTI approach is supportive to determine atrophy of cholinergic nuclei and its effect on intracortical projecting fiber tracts in AD. Hum Brain Mapp 32: 1349-1362, 2011. (C) 2010 Wiley-Liss, Inc.

A Voxel-Based Morphometry Study of Frontal Gray Matter Correlates of Impulsivity

Impulsivity is a personality trait exhibited by healthy individuals, but excessive impulsivity is associated with some mental disorders. Lesion and functional, neuroimaging Studies indicate that the ventromedial prefrontal region (VMPFC), including the orbitofrontal cortex (OFC), anterior cingulate cortex (ACC) and medial prefrontal cortex, and the amygdala may modulate impulsivity and aggression. However, no morphometric study has examined the association between VMPFC and impulsivity. We hypothesized that healthy subjects with high impulsivity would have smaller volumes in these brain regions compared with those with low impulsivity. Sixty-two healthy Subjects were Studied (age 35.4 +/- 12.1 years) using a 1.5-T MRI system. The Barratt impulsiveness scale (BIS) was used to assess impulsivity. Images were processed using an optimized voxel-based morphometry (VBM) protocol. We calculated the correlations between BIS scale scores and the gray matter (GM) and white matter (WM) volumes of VMPFC and amygdala. GM volumes of the left and right OFC were inversely correlated with the BIS total score (P = 0.04 and 0.02, respectively). Left ACC GM Volumes had a tendency to be inversely correlated with the BIS total score (P = 0.05. Right OFC GM Volumes were inversely correlated with BIS nonplanning impulsivity, and left OFC GM volumes were inversely correlated with motor impulsivity. There were no significant WM volume correlations with impulsivity. The results Of this morphometry Study indicate that small OFC volume relate to high impulsivity and extend the prior finding that the VMPFC is involved in the circuit modulating impulsivity. HUM Brain Mapp 30:1188-1195, 2009. (C) 2008 Wiley-Liss, Inc.

Assessing Interethnic Admixture Using an X-Linked Insertion-Deletion Multiplex

In this study, a PCR multiplex was optimized, allowing the simultaneous analysis of 13 X-chromosome Insertion/deletion polymorphisms (INDELs). Genetic variation observed in Africans, Europeans, and Native Americans reveals high inter-population variability. The estimated proportions of X-chromosomes in an admixed population from the Brazilian Amazon region show a predominant Amerindian contribution (congruent to 41%), followed by European (congruent to 32%) and African (congruent to 27%) contributions. The proportion of Amerindian contribution based on X-linked data is similar to the expected value based on mtDNA and Y-chromosome information. The accuracy for assessing interethnic admixture, and the high differentiation between African, European, and Native American populations, demonstrates the suitability of this INDEL set to measure ancestry proportions in three-hybrid populations, as it is the case of Latin American populations. Am. J. Hum. Biol. 21:707-709, 2009. (C) 2009 Wiley-Liss, Inc.

Cultural Consonance, a 5HT2A Receptor Polymorphism, and Depressive Symptoms: A Longitudinal Study of Gene x Culture Interaction in Urban Brazil

In this study in urban Brazil we examine, as a predictor of depressive symptoms, the interaction between a single nucleotide polymorphism in the 2A receptor in the serotonin system (-1438G/A) and cultural consonance in family life, a measure of the degree to which an individual perceives her family as corresponding to a widely shared cultural model of the prototypical family. A community sample of 144 adults was followed over a 2-year-period. Cultural consonance in family life was assessed by linking individuals` perceptions of their own families with a shared cultural model of the family derived from cultural consensus analysis. The -1438G/A polymorphism in the 2A serotonin receptor was genotyped using a standard protocol for DNA extracted from leukocytes. Covariates included age, sex, socioeconomic status, and stressful life events. Cultural consonance in family life was prospectively associated with depressive symptoms. In addition, the interaction between genotype and cultural consonance in family life was significant. For individuals with the A/A variant of the -1438G/A polymorphism of the 2A receptor gene, the effect of cultural consonance in family life on depressive symptoms over a 2-year-period was larger (beta = -0.533, P < 0.01) than those effects for individuals with either the G/A (beta = -0.280, P < 0.10) or G/G (beta = -0.272, P < 0.05) variants. These results are consistent with a process in which genotype moderates the effects of culturally meaningful social experience on depressive symptoms. Am. J. Hum. Biol. 21:91-97, 2009. (C) 2008 Wiley-Liss, Inc.