Gene Expression Profiles Stratified according to Type 1 Diabetes Mellitus Susceptibility Regions

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Origin and differentiation of a sex chromosome system in Parodon hilarii (Pisces, Parodontidae). Satellite DNA, G- and C-banding

A satellite DNA sequence of Parodon hilarii ( named pPh2004) was isolated, cloned and sequenced. This satellite DNA is composed of 200 bp, 60% AT rich. In situ hybridization ( FISH) results revealed that the satellite DNA pPh2004 is located in the terminal regions of several chromosomes, forming highly evident blocks in some and punctual marks in others. The comparison between the FISH and C-banding results showed that the location of this satellite DNA coincides with that of most terminal heterochromatins. However, some regions are only marked by FISH whereas other regions are only marked by C-banding. The possible existence of more than one satellite DNA family could explain these partial differences. The in situ hybridization with the satellite DNA and the G- and C-bandings confirmed the presence of a sex chromosome system of the ZZ/ZW type in P. hilarii, as well as the correct identification of the Z chromosome in the karyotype. This chromosome displays a segment of terminal heterochromatin in the long arm, similar to the segment observed in the short arm of the W chromosome, also showing a G- banding pattern similar to that of the short arm and part of the long arm of the W chromosome. A hypothesis on the origin of the W chromosome from an ancestral chromosome similar to the Z chromosome is presented.

Large-scale transcriptome analyses reveal new genetic marker candidates of head, neck, and thyroid cancer

A detailed genome mapping analysis of 213,636 expressed sequence tags (EST) derived from nontumor and tumor tissues of the oral cavity, larynx, pharynx, and thyroid was done. Transcripts matching known human genes were identified; potential new splice variants were flagged and subjected to manual curation, pointing to 788 putatively new alternative splicing isoforms, the majority (75%) being insertion events. A subset of 34 new splicing isoforms (5% of 788 events) was selected and 23 (68%) were confirmed by reverse transcription-PCR and DNA sequencing. Putative new genes were revealed, including six transcripts mapped to well-studied chromosomes such as 22, as well as transcripts that mapped to 253 intergenic regions. In addition, 2,251 noncoding intronic RNAs, eventually involved in transcriptional regulation, were found. A set of 250 candidate markers for loss of heterozygosis or gene amplification was selected by identifying transcripts that mapped to genomic regions previously known to be frequently amplified or deleted in head, neck, and thyroid tumors. Three of these markers were evaluated by quantitative reverse transcription-PCR in an independent set of individual samples. Along with detailed clinical data about tumor origin, the information reported here is now publicly available on a dedicated Web site as a resource for further biological investigation. This first in silico reconstruction of the head, neck, and thyroid transcriptomes points to a wealth of new candidate markers that can be used for future studies on the molecular basis of these tumors. Similar analysis is warranted for a number of other tumors for which large EST data sets are available.

Acute, subacute toxicity and genotoxic effect of a hydroethanolic extract of the cashew (Anacardium occidentale L.)

Anacardium occidentale L. (Anacardiaceae), popularly known as cajueiro is a native plant to Brazil, and largely used in popular medicine to treat ulcers, hypertension and diarrhea. In the present study, acute, 30-day subacute toxicity and genotoxicity assays were carried out. The crude extract did not produce toxic symptoms in rats in doses up to 2000 mg/kg. Based on biochemical analyses of renal and hepato-biliary functions, such as the level of urea, creatinine, transaminases and alkaline phosphatase, we determined that the extract is generally tolerated by rats. This was also confirmed by hematological and histopathological exams. Genotoxicity was accessed by the Ames test in Salmonella typhimurium strains TA97, TA98, TA 100, TA 102 and by the bone marrow micronucleus test in mice. The extract was shown to induce frameshift, base pair substitution and damage to the chromosomes. However, this effect was less deleterious than the clastogenic effect of ciclophosphamide. (c) 2006 Elsevier B.V.. All rights reserved.

Structural and functional evidence that a B chromosome in the characid fish Astyanax scabripinnis is an isochromosome

Astyanax scabripinnis possesses a widespread polymorphism for metacentric B chromosomes as large as the largest chromosome pair in the A complement. on the basis of C-banding pattern, it was hypothesized that these B chromosomes are isochromosomes that have arisen by means of centromere misdivision and chromatid nondisjunction. In the present paper we test this hypothesis by analysing (i) the localization of a repetitive DNA sequence on both B chromosome arms, and (ii) synaptonemal complex formation, in order to test the functional homology of both arms. Genomic DNA digested with KpnI and analysed by gel electrophoresis showed fragments in a ladder-like pattern typical of tandemly repetitive DNA. These fragments were cloned and their tandem organization in the genome was confirmed. A 51-bp long consensus sequence, which was AT-rich (59%) and contained a variable region and two imperfect reverse sequences, was obtained. Fluorescence in situ hybridization (FISH) localized this repetitive DNA into noncentromeric constitutive heterochromatin which encompasses the terminal region of some acrocentric chromosomes, the NOR region, and interstitial polymorphic heterochromatin in chromosome 24. Most remarkably, tandem repeats were almost symmetrically placed in the two arms of the B chromosome, with the exception of two additional small clusters proximally located on the slightly longer arm. Synaptonemal complex (SC) analysis showed 26 completely paired SCs in males with 1B. The ring configuration of the B univalent persisting until metaphase I suggests that the two arms formed chiasmata. All these data provided strong support for the hypothesis that the B chromosome is an isochromosome.

Accentuated polymorphism of heterochromatin and nucleolar organizer regions in Astyanax scabripinnis (Pisces, Characidae): tools for understanding karyotypic evolution

Astyanax scabripinnis has been considered a species complex because it presents high karyotypic and morphological variability among its populations. In this work, individuals of two A. scabripinnis populations from different streams in the same hydrographic basin were analyzed through C-banding and AgNOR. Although they present distinct diploid numbers, they show meta and submetacentric chromosome groups highly conserved (numerically and morphologically). Other chromosomal characteristics are also shared by both populations, as the pattern of constitutive heterochromatin distribution (large blocks in the telomeric regions of subtelocentric and acrocentric chromosomes) and some nucleolar chromosomes. Inter-individual variations both in the number and size of heterochromatic blocks, and in the number and localization of NORs were verified in the studied populations, characterizing them as polymorphics for these regions. The mechanisms involved in the dispersion of heterochromatin and NORs through the karyotypes, as well as the possible events related to the generation of polymorphism of those regions are discussed. Furthermore, relationships between these populations and within the context of the scabripinnis complex are also approached.

Cytogenetic characterization of F1, F2 and backcross hybrids of the Neotropical catfish species Pseudoplatystoma corruscans and Pseudoplatystoma reticulatum (Pimelodidae, Siluriformes)

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

First report of a B chromosome in a natural population of Astyanax altiparanae (Characiformes, Characidae)

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

Cytogenetic markers as diagnoses in the identification of the hybrid between Piauçu (Leporinus macrocephalus) and Piapara (Leporinus elongatus)

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

Chromosome polymorphism of heterochromatin and nucleolar regions in two populations of the fish Astyanax bockmanni (Teleostei: Characiformes)

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

Cytogenetic characterization of the strongly electric Amazonian eel, Electrophorus electricus (Teleostei, Gymnotiformes), from the Brazilian rivers Amazon and Araguaia

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

Análise citogenética de células-tronco derivadas do subendotélio da veia umbilical humana

Mesenchymal stem cells (MSCs) are known as a population of multi-potential cells able to proliferate and differentiate into multiple mesodermal tissues including bone, cartilage, muscle, ligament, tendon, fat and stroma. Several applications of the study of EC can be emphasized the therapeutic techniques such as guided bone regeneration by implantation of EC in the affected site, without the need for bone grafts, using titanium as a vehicle. The process of cryopreservation is essential for the maintenance of cell cultures, since the cell line is frozen, it can be maintained in liquid nitrogen for an indefinite period and then thawed for therapeutic or experimental purposes. The aim of this study was to isolate a population of MSCs derived from the subendothelium of the umbilical vein human (MSCs-SUVH) to assess cytogenetic analysis by the possibility of appearance of chromosomal changes in two different situations: MSCs-SUVH regarding the process of cryopreservation and MSCs-SUVH grown on the surface of titanium. Flow cytometry analysis revealed that, this cell population was positive for the markers CD29, CD73 and CD90, but there was no expression of hematopoietic lineage markers, such as CD14, CD34 and CD45 and demonstrated capacity for osteogenic differentiation. The chromosomes obtained from the primary culture of MSCs-SUVH were analyzed by GTW banding technique, and results are described as guidelines to ISCN 2005. There was not the emergence of clonal chromosomal changes in the MSCs-SUVH in different situations analyzed. However one of the strings presented a balanced paracentric inversion, probably a cytogenetic constitutional alterations, which was present before and after the experimental situations that the MSCs-SUVH was submitted

Levantamento cariotípico em espécies de peixes marinhos costeiros de fundo arenoso (Osteichthypes, Perciformes)

Cytogenetics analyses in fish are important because they compose a private group among the vertebrates, occupying a central position in the animal evolution. The Perciforms Order, dominant in the marine and freshwater environment, it constitutes a model potentially useful in the genetic evaluation of populations, as well as in the understanding of its evolutionary processes. In spite of this, cytogenetics studies in this great group is scarce, above all for the inhabitants of sandy bottom and pelagics habits. The present work proposed to contribute for the cytogenetic characterization of nine species of fish marine of sandy bottom of the coast of Rio Grande do Norte (Brazil), identifying the evolutionary patterns related to the karyotype in these species and the existence of filogenetics affinities between them and other Perciformes. The animals were collected in the beaches of the Redinha, Ponta Negra and Búzios (Coast of Rio Grande do Norte) and in Saint Peter and Saint Paul Archipelago. Later on they were submitted to the cytogenetics technical that consist of mitotic estimulation, obtaining of mitotics chromosomes, proceeded by techniques of conventional coloration (Giemsa) and chromosomic bands (Ag-RONs and C band). Diploid number and fundamental number equal to 48 were observed in most of the species: Menticirrhus americanus, Ophioscion punctatissimus, Pareques acuminatus (Sciaenidae); Chloroscombrus chrysurus (Carangidae); Echeneis sp. 2 (Echeneidae); Archosargus probatocephalus (Sparidae) and Orthopristis ruber (Haemulidae). Trachinotus goodei (NF=52) (Carangidae) and Echeneis sp. 1 (Echeneidae) (NF=54) presented variation in NF, staying constant a diploid number equal to 48. RONs was situated in pericentromeric position in whole the scianids, and in the species Echeneis sp. 2 (22° pair), O. ruber and A. probatocephalus (1° pair), coinciding with great heterocromatics blocks in M. americanus (1° pair), P. acuminatus (2° pairl) and O. ruber (1° pair). RONs was also located in the telomeric area of the short arm of the 5° and 11° acrocentrics pairs in T. goodei, 4° and 19° pairs of C. chrysurus, 1° pair (sm) of Echeneis sp. 1. The C band detected centromeric blocks in most of the chromosomes of the species of Sciaenidae, Carangidae and Echeneidae, with great blocks in A. probatocephalus (4° pair). Heterocromatic blocks in telomeric areas in submetacentrics of Echeneis sp. 1, and pericentromerics in M. americanus (1° and 8° pairs), O. punctatissimus (1° pair) and P. acuminatus (2° pair) were also observed. It is noticed a marked conservatism cromossomic in the species of the family Scianidae and Haemulidae in what says respect to the number of acrocentrics chromosomes and the location of RONs. Even so it is outstanding the presence of heterocromatinization events during the karyotypic evolution of this family. Already in the families Sparidae and Carangidae, the obtained results reaffirm examples of small variations structural resultants of inversion and translocation Robertsonian, as important mechanisms of diversification karyotipical, as well as a pattern numerical evolutionary conserved, also observed in representatives of Echeneidae of Atlantic in relation to Pacific. The presence of RONs multiple, observed in the species T. goodei and C. chrysurus seems to represent a character derived in the family Carangidae. The results for the species O. ruber and A. probatocephalus suggest the presence of possible geographical or climatic barriers among populations of NE of Brazil in relationship the one of the SE

Padrões cariotípicos em tetraodontiformes (Osteichthypes) :redução genômica e mecanismos de diversificação

The Tetraodontiformes order is composed for about 400 species of fish, distributed in ten families, with circuntropical distribution. The morphologic diversity of each family reflects in, part, the different levels of specialization. This group represents an ancestry after-Perciformes and constitutes the last branch of the diffusion of the Teleósteos, occupying a position of prominence. The phylogenetics relationships of the Tetraodontiformes exist diverse works examining and, in all, these families are recognized as groups brothers, being Diodontidae next to Tetraodontidae and Balistidae next to Monacanthidae. Although it possesss a representative number of species, the works involving of the families Balistidae and Monacanthidae are few exemplary, especially species of oceanic islands. In this work cytogenetic studies in five species had been analyzed Cantherhines macrocerus, Cantherhines pullus (Monacanthidae), Melichthys niger (Balistidae), Sphoeroides testudíneus (Tetraodontidae) and Chilomycterus antennatus (Diodontidae); through conventional coloration, Ag-NORs and C banding. Ahead of the different karyological trends of evolution presented by the Tetraodontiformes, the present work also searched to verify the relation existence enters the total size of the chromosomes with the amount of DNA in these groups of Tetraodontiformes. For such, they had been correlated the total size of the chromosomes of these species, with values of content of available DNA in literature. The cytogenetics analyses for the species C.macrocerusJ C.pullus (Monacanthidae) and M.niger (Balistidae), had disclosed 40 chromosomes, all acrocentrics. All possess only one pair of NORs and pericentromeric heterochromatin. For S.testudíneus the found dyploid number was equal 2n=46, with NF=78 (16m+18sm+8st+4a), while that for C.antennatus it possesss 2n=50, with NF=76 (4m+22st+24a). Both species possess simple NORs and pericentromeric heterochromatin blocks. In M.niger, the presence of positive marking (heterochromatin and NOR) in the secondary constriction in the second chromosomic pair suggesting the occurrence of a rearrangement, possibly a fusing involving these homologous ones, indicating that these events had been important for the establishment of the karyological history of this group. A maintenance of the chromosomic constancy found in the populations of C.macrocerus (Monacanthidae) and S.testudineus (Tetraodontidae) perhaps if must for the aiding of the gene flow through oceanic chains. These data contrast with the differentiated kinds of chromosomes of C.antennatus between the Northeast coast and Southeastern, suggesting that the ecological standards of each species, added to the conditions of the marine environment, can be responsible for the karyological delineation of each species. The found characteristics for the species C.macrocerus, C.pullus, M.niger, S.testudineus and C.antennatus add it the available data for other species of Tetraodontiformes. From the data gotten in the present study, it can be inferred that the DNA content possesss direct relation with the total length of the chromosomes

Diversificação cariotípica em cinco espécies de moréias do Atlântico Ocidental (Anguilliformes)

The Anguiliformes is constituted by 15 families, 141 sorts and 737 species. In this group eight families possess at least one karyotyped species, where a prevalence of karyotypes with 2n=38 is evidenced chromosomes and high NF, apparently basal for the Anguiliformes. The only family who shows a different karyotypic pattern from the others is the Muraenidae family. In this, of the eight species already described, all of them present 2n=42 chromosomes. Despite the dimension of this Order, few species present karyotypics descriptions. In the present work, a species of Ophichthidae, Myrichthys ocellatus (2n=38, 8m+14sm+10st+6a, NF=70) and three species of Muraenidae, Enchelycore nigricans (2n=42, 6m+8sm+12st+16a, NF=68), Gymnothorax miliaris (2n=42, 14m+18sm+10st, NF=84), Gymnothorax vicinus (2n=42, 8m+6sm+28a, NF=56) and Muraena pavonina (2n=42, 6m+4sm+32a, NF=52), collected in the coast of the Rio Grande do Norte state, Saint Peter and Paul Rocks and in the coast of Bahia state were analyzed. Mitotics chromosomes had been gotten through mitotic stimulation with yeasts. Among the analyzed species, it is observed the presence of characteristic large metacentric chromosomic pairs (≅10µm). As for the structural standard, heterochromatics regions in these species in centromeric position of the majority of the chromosomic pairs and simple ribosomal sites had been evidenced. For the Ophichthidae family, the gotten data corroborate the hypothesis of karyotypic diversification mediated by the occurrence of pericentrics inversions and robertsonians rearrangements, while in the Muraenidae, the identification of larger chromosomic values (2n=42), suggests derived karyotypes, possibly caused by possible chromosomic fissions