969 resultados para restriction fragment length polymorphism


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Context. - Hodgkin lymphoma is a neoplastic disease in which the immune system plays a major role in its pathogenesis. Interleukin 10 ( IL-10), an immunosuppressive cytokine actively produced in patients with Hodgkin lymphomas, favors the survival of the Hodgkin/Reed-Sternberg cells. Individual variations in IL-10 levels may be due, in part, to the presence of single nucleotide polymorphisms in the IL10 gene promoter.Objective. - To evaluate whether particular single nucleotide polymorphisms in the IL10 gene are found more frequently in Hodgkin lymphoma cases associated with Epstein-Barr virus infection.Design. - the identification of single nucleotide polymorphisms at positions -1082 and -819/-592 in the IL10 gene was performed by polymerase chain reaction and restriction length fragment polymorphisms analysis in 65 cases of Hodgkin lymphoma and 50 cases of reactive benign follicular lymphoid hyperplasia ( non-Hodgkin lymphoma control group).Results. - the frequency of the genotype GG at position -1082 was found to be significantly higher in patients with Epstein-Barr virus-positive Hodgkin lymphoma compared with Epstein-Barr virus-negative cases.Conclusions. - the results suggest that the presence of specific single nucleotide polymorphisms in the IL10 gene, notably those associated with high IL-10 production, may play a role in the susceptibility to Epstein-Barr virus -positive Hodgkin lymphoma development.

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This investigation was carried out to study the influence of early qualitative feed restriction and environmental rearing temperature on long bone development in broiler. Energy and protein restriction reduced femur width and humerus weight, but did not affect tibia parameters. Broilers kept at cold environmental temperature showed reduced femur, tibia and humerus length and tibia weight, but the calculated density was not affected by rearing temperature. These findings suggest that qualitative feed restriction and environmental temperature influenced the normal long bone growth; however, bone weight/bone length index (calculated density) was not affected by rearing temperature. (c) 2007 Elsevier Ltd. All rights reserved.

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

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Studies have demonstrated that nutrient deficiency during pregnancy or in early postnatal life results in structural abnormalities in the offspring hippocampus and in cognitive impairment. In an attempt to analyze whether gestational protein restriction might induce learning and memory impairments associated with structural changes in the hippocampus, we carried out a detailed morphometric analysis of the hippocampus of male adult rats together with the behavioral characterization of these animals in the Morris water maze (MWM). Our results demonstrate that gestational protein restriction leads to a decrease in total basal dendritic length and in the number of intersections of CA3 pyramidal neurons whereas the cytoarchitecture of CA1 and dentate gyrus remained unchanged. Despite presenting significant structural rearrangements, we did not observe impairments in the MWM test. Considering the clear dissociation between the behavioral profile and the hippocampus neuronal changes, the functional significance of dendritic remodeling in fetal processing remains undisclosed. © 2012 ISDN.

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This study was aimed to evaluate the influence of vitamin D (VD) deficiency on cardiac metabolism, morphology, and function. Thus, we investigated the relationship of these changes with the length of the nutrient restriction. Male weanling Wistar rats were allocated into 4 groups: C2 (n=24), animals were fed an AIN-93G diet with 1000 IU VD/kg of chow and were kept under fluorescent light for 2 months; D2 (n=22), animals were fed a VD-deficient AIN-93G diet and were kept under incandescent light for 2 months; C4 (n=21) animals were kept in the same conditions of C2 for 4 months; and D4 (n=23) animals were kept in the same conditions of D2 for 4 months. Biochemical analyses showed lower β-hydroxyacyl coenzyme-A dehydrogenase activity and higher lactate dehydrogenase activity in VD-deficient animals. Furthermore, VD deficiency was related to increased cytokines release, oxidative stress, apoptosis, and fibrosis. Echocardiographic data showed left ventricular hypertrophy and lower fractional shortening and ejection fraction in VD-deficient animals. Difference became evident in the lactate dehydrogenase activity, left ventricular weight, right ventricle weight, and left ventricular mass after 4 months of VD deficiency. Our data indicate that VD deficiency is associated with energetic metabolic changes, cardiac inflammation, oxidative stress, fibrosis and apoptosis, cardiac hypertrophy, left chambers alterations, and systolic dysfunction. Furthermore, length of the restriction influenced these cardiac changes.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

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Crown-root fractures account for 5% of all fractures in permanent teeth and can involve enamel, dentin, and cementum. Depending on whether there is pulpal involvement, these problems may be classified as complicated (which are more common) or noncomplicated. The treatment depends on the level of the fracture line, root length and/or morphology, and esthetic needs. Several treatment strategies are available for esthetic and functional rehabilitation in crown-root fractures. Adhesive tooth fragment reattachment is the most conservative restorative option when the tooth fragment is available and the biological width has no or minimal violation. This article reports a case of an uncomplicated crown-root fracture in the permanent maxillary right central incisor of a young patient who received treatment with adhesive tooth fragment reattachment, preserving the anatomic characteristics of the fractured tooth after periodontal intervention. The fracture line of the fragment had an unusual shape, starting on the palatal side and extending to the buccal side subgingivally. After 7 years, the attached coronal fragment remained in position with good esthetics, as well as clinical and radiographic signs of pulpal vitality, periodontal health, and root integrity, thus indicating success.

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SERA5 is regarded as a promising malaria vaccine candidate of the most virulent human malaria parasite Plasmodium falciparum. SERA5 is a 120 kDa abundantly expressed blood-stage protein containing a papain-like protease. Since substantial polymorphism in blood-stage vaccine candidates may potentially limit their efficacy, it is imperative to fully investigate polymorphism of the SERA5 gene (sera5). In this study, we performed evolutionary and population genetic analysis of sera5. The level of inter-species divergence (kS = 0.076) between P. falciparum and Plasmodium reichenowi, a closely related chimpanzee malaria parasite is comparable to that of housekeeping protein genes. A signature of purifying selection was detected in the proenzyme and enzyme domains. Analysis of 445 near full-length P. falciparum sera5 sequences from nine countries in Africa, Southeast Asia, Oceania and South America revealed extensive variations in the number of octamer repeat (OR) and serine repeat (SR) regions as well as substantial level of single nucleotide polymorphism (SNP) in non-repeat regions (2562 bp). Remarkably, a 14 amino acid sequence of SERA5 (amino acids 59-72) that is known to be the in vitro target of parasite growth inhibitory antibodies was found to be perfectly conserved in all 445 worldwide isolates of P. falciparum evaluated. Unlike other major vaccine target antigen genes such as merozoite surface protein-1, apical membrane antigen-1 or circumsporozoite protein, no strong evidence for positive selection was detected for SNPs in the non-repeat regions of sera5. A biased geographical distribution was observed in SNPs as well as in the haplotypes of the sera5 OR and SR regions. In Africa, OR- and SR-haplotypes with low frequency (<5%) and SNPs with minor allele frequency (<5%) were abundant and were mostly continent-specific. Consistently, significant genetic differentiation, assessed by the Wright's fixation index (FST) of inter-population variance in allele frequencies, was detected for SNPs and both OR- and SR-haplotypes among almost all parasite populations. The exception was parasite populations between Tanzania and Ghana, suggesting frequent gene flow in Africa. The present study points to the importance of investigating whether biased geographical distribution for SNPs and repeat variants in the OR and SR regions affect the reactivity of human serum antibodies to variants. (C) 2011 Elsevier Ltd. All rights reserved.

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Abstract Background Identification of nontuberculous mycobacteria (NTM) based on phenotypic tests is time-consuming, labor-intensive, expensive and often provides erroneous or inconclusive results. In the molecular method referred to as PRA-hsp65, a fragment of the hsp65 gene is amplified by PCR and then analyzed by restriction digest; this rapid approach offers the promise of accurate, cost-effective species identification. The aim of this study was to determine whether species identification of NTM using PRA-hsp65 is sufficiently reliable to serve as the routine methodology in a reference laboratory. Results A total of 434 NTM isolates were obtained from 5019 cultures submitted to the Institute Adolpho Lutz, Sao Paulo Brazil, between January 2000 and January 2001. Species identification was performed for all isolates using conventional phenotypic methods and PRA-hsp65. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing a 441 bp fragment of hsp65. Phenotypic evaluation and PRA-hsp65 were concordant for 321 (74%) isolates. These assignments were presumed to be correct. For the remaining 113 discordant isolates, definitive identification was based on sequencing a 441 bp fragment of hsp65. PRA-hsp65 identified 30 isolates with hsp65 alleles representing 13 previously unreported PRA-hsp65 patterns. Overall, species identification by PRA-hsp65 was significantly more accurate than by phenotype methods (392 (90.3%) vs. 338 (77.9%), respectively; p < .0001, Fisher's test). Among the 333 isolates representing the most common pathogenic species, PRA-hsp65 provided an incorrect result for only 1.2%. Conclusion PRA-hsp65 is a rapid and highly reliable method and deserves consideration by any clinical microbiology laboratory charged with performing species identification of NTM.

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The Adriatic Sea is considered a feeding and developmental area for Mediterranean loggerhead turtles, but this area is severely threatened by human impacts. In the Adriatic Sea loggerhead turtles are often found stranded or floating, but they are also recovered as by-catch from fishing activities. Nevertheless, information about population structuring and origin of individuals found in the Adriatic Sea are still limited. Cooperation with fishermen and a good network of voluntary collaborators are essential for understanding their distribution, ecology and for developing conservation strategies in the Adriatic Sea. In this study, a comparative analysis of biometric data and DNA sequence polymorphism of the long fragment of the mitochondrial control region was carried out on ninety-three loggerheads recovered from three feeding areas in the Adriatic Sea: North-western, North-eastern and South Adriatic. Differences in turtles body sizes (e.g. Straight Carapace Length) among the three recovery areas and relationship between SCL and the type of recovery were investigated. The origin of turtles from Mediterranean rookeries and the use of the Adriatic feeding habitats by loggerheads in different life-stages were assessed to understand the migratory pathway of the species. The analysis of biometric data revealed a significant difference in turtle sizes between the Southern and the Northern Adriatic. Moreover, size of captured turtles resulted significantly different from the size of stranded and floating individuals. Actually, neritic sub-adults and adults are more affected by incidental captures than juveniles because of their feeding behavior. The Bayesian mixed-stock analysis showed a strong genetic relationship between the Adriatic aggregates and Mediterranean rookeries, while a low pro¬portion of individuals of Atlantic origin were detected in the Adriatic feeding grounds. The presence of migratory pathways towards the Adriatic Sea due to the surface current system was reinforced by the finding of individuals bearing haplotypes endemic to the nesting populations of Libya, Greece and Israel. A relatively high contribution from Turkey and Cyprus to the Northwest and South Adriatic populations was identified when the three sampled areas were analyzed independently. These results have to be taken in account in a conservative perspective, since coastal hazards, affecting the population of turtles feeding in the Adriatic Sea may also affect the nesting populations of the Eastern Mediterranean with a unique genetic pattern.

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Hereditary hair length variability in mice and dogs is caused by mutations within the fibroblast growth factor 5 (FGF5) gene. The aim of this study was to evaluate the feline FGF5 orthologue as a functional candidate gene for the long hair phenotype in cats, which is recessive to short hair. We amplified the feline FGF5 cDNA and characterised two alternatively spliced transcripts by RT-PCR. Comparative cDNA and genomic DNA sequencing of long- and short-haired cats revealed four non-synonymous polymorphisms in the FGF5 coding sequence. A missense mutation (AM412646:c.194C>A) was found in the homozygous state in 25 long-haired Somali, Persian, Maine Coon, Ragdoll and crossbred cats. Fifty-five short-haired cats had zero or one copy of this allele. Additionally, we found perfect co-segregation of the c.194C>A mutation within two independent pedigrees segregating for hair length. A second FGF5 exon 1 missense mutation (AM412646:c.182T>A) was found exclusively in long-haired Norwegian Forest cats. The c.182T>A mutation probably represents a second FGF5 mutation responsible for long hair in cats. In addition to the c.194C>A mutation, a frameshift mutation (AM412646:c.474delT) was found with a high frequency in the long-haired Maine Coon breed. Finally, a missense mutation (AM412646:c.475A>C) was also associated with the long-haired phenotype in some breeds. However, as one short-haired cat was homozygous for this polymorphism, it is unlikely that it has a functional role in the determination of hair length.

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CONTEXT: A polymorphism of the GH receptor (GHR) gene resulting in genomic deletion of exon 3 (GHR-d3) has been associated with responsiveness to GH therapy. However, the data reported so far do vary according to the underlying condition, replacement dose, and duration of the treatment. OBJECTIVE, DESIGN: The aim of this study was to analyze the impact of the GHR genotypes in terms of the initial height velocity (HV) resulting from treatment and the impact upon adult height in patients suffering from severe isolated GH deficiency. CONTROLS, PATIENTS, SETTING: A total of 181 subjects (peak stimulated GHlength/full-length genotype (P<0.05). A GHR-d3 allele dose-dependent effect was found for both HV SDS (r=0.72) and height gain (r=0.77). However, there was no significant difference in final adult height and height SDS according to the exon-3 genotypes. CONCLUSIONS: Our results indicate that in patients with severe isolated GH deficiency, although the GHR genotype might play a role in GH responsiveness, at least at the beginning of treatment, there is no effect on final height.

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Coronary heart disease (CHD) is the leading cause of death in the United States. Recently, renin-angiotensin system (RAS) was found associated with atherosclerosis formation, with angiotensin II inducing vascular smooth muscle cell growth and migration, platelet activation and aggregation, and stimulation of plasminogen activator inhibitor-1. Angiotensin II is converted from angiotensin I by angiotensin I-converting enzyme (ACE) and this enzyme is mainly genetically determined. The ACE gene has been assigned to chromosome 17q23 and an insertion/deletion (I/D)polymorphism has been characterized by the presence/absence of a 287 bp fragment in intron 16 of the gene. The two alleles form three genotypes, namely, DD, ID and II and the DD genotype has been linked to higher plasma ACE levels and cell ACE activity.^ In this study, the association between the ACE I/D polymorphism and carotid artery wall thickness measured by B-mode ultrasound was investigated in a biracial sample, and the association between the gene and incident CHD was investigated in whites and if the gene-CHD association in whites, if any, was due to the gene effect on atherosclerosis. The study participants are from the prospective Atherosclerosis Risk in Communities (ARIC) Study, including adults aged 45 to 65 years. The present dissertation used a matched case-control design for studying the associations of the ACE gene with carotid artery atherosclerosis and an unmatched case-control design for the association of the gene with CHD. A significant recessive effect of the D allele on carotid artery thickness was found in blacks (OR = 3.06, 95% C.I: 1.11-8.47, DD vs. ID and II) adjusting for age, gender, cigarette smoking, LDL-cholesterol and diabetes. No similar associations were found in whites. The ACE I/D polymorphism is significantly associated with coronary heart disease in whites, and while stratifying data by carotid artery wall thickness, the significant associations were only observed in thin-walled subgroups. Assuming a recessive effect of the D allele, odds ratio was 2.84 (95% C.I:1.17-6.90, DD vs. ID and II) and it was 2.30 (95% C.I:1.22-4.35, DD vs. ID vs. II) assuming a codominant effect of the D allele. No significant associations were observed while comparing thick-walled CHD cases with thin-walled controls. Following conclusions could be drawn: (1) The ACE I/D polymorphism is unlikely to confer appreciable increase in the risk of carotid atherosclerosis in US whites, but may increases the risk of carotid atherosclerosis in blacks. (2) ACE I/D polymorphism is a genetic risk factor for incident CHD in US whites and this effect is separate from the chronic process of atherosclerosis development. Finally, the associations observed here are not causal, since the I/D polymorphism is in an intron, where no ACE proteins are encoded. ^

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This research characterized a serologically indistinguishable form of HLA-DR1 that: (1) cannot stimulate some DR1-restricted or specific T-lymphocyte clones; (2) displays an unusual electrophoretic pattern on two dimensional gels; and (3) is marked by a polymorphic restriction site of the alpha gene. Inefficient stimulation of some DR1-restricted clones was a property of DR1$\sp{+}$ cells that shared HLA-B14 on the same haplotype and/or were carriers of 21-hydroxylase (21-OH) deficiency. Nonclassical 21-OH deficiency frequently demonstrates genetic linkage with HLA-B14;DR1 haplotypes and associates with duplications of C4B and one 21-OH gene. Cells having both stimulatory (DR1$\sb{\rm n}$) and nonstimulatory (DR1$\sb{\rm x}$) parental haplotypes did not mediate proliferation of these clones. However, heterozygous DR1$\sb{\rm x}$, 2 and DR1$\sb{\rm x}$, 7 cells were efficient stimulators of DR2 and DR7 specific clones, respectively, suggesting that a trans acting factor may modify DR1 alleles or products to yield a dominant DR1$\sb{\rm x}$ phenotype. Incompetent stimulator populations did not secrete an intercellular soluble or contact dependent suppressor factor nor did they express interleukin-2 receptors competing for T-cell growth factors. Two dimensional gel analysis of anti-DR immunoprecipitates revealed, in addition to normal DR$\alpha$ and DR$\beta$ chains, a 50kD species from DR1$\sb{\rm x}$ but not from the majority of DR1$\sb{\rm n}$ or non-DR1 cells. The 50kD structure was stable under reducing conditions in SDS and urea, had antigenic homology with DR, and dissociated after boiling into 34kD and 28kD peptide chains apparently identical with DR$\alpha$ and DR$\beta$ as shown by limited digest peptide maps. N-linked glycosylation and sialation of DRgp50 appeared to be unchanged from normal DR$\alpha$ and DR$\beta$. Bg1II digestion and $DR\alpha$ probing of DR1$\sb{\rm x}$ genomic DNA revealed a 4.5kb fragment while DR1$\sb{\rm n}$ DNA yielded 3.8 and 0.76kb fragments; all restriction sites mapped to the 3$\sp\prime$ untranslated region of $DR\alpha$. Collectively, these data suggest that DRgp50 represents a novel combinatorial association between constitutive chains of DR that may interfere with or compete for normal T cell receptor recognition of DR1 as both an alloantigen and restricting element. Furthermore, extensive chromosomal abnormalities previously mapped to the class III region of B14;DR1 haplotypes may extend into the adjacent class II region with consequent intrusion on immune function. ^