Establishment of animal models to analyze the kinetics and distribution of human tumor antigen-specific CD8⁺ T cells.

Many patients develop tumor antigen-specific T cell responses detectable in peripheral blood mononuclear cells (PBMCs) following cancer vaccine. However, measurable tumor regression is observed in a limited number of patients receiving cancer vaccines. There is a need to re-evaluate systemically the immune responses induced by cancer vaccines. Here, we established animal models targeting two human cancer/testis antigens, NY-ESO-1 and MAGE-A4. Cytotoxic T lymphocyte (CTL) epitopes of these antigens were investigated by immunizing BALB/c mice with plasmids encoding the entire sequences of NY-ESO-1 or MAGE-A4. CD8(+) T cells specific for NY-ESO-1 or MAGE-A4 were able to be detected by ELISPOT assays using antigen presenting cells pulsed with overlapping peptides covering the whole protein, indicating the high immunogenicity of these antigens in mice. Truncation of these peptides revealed that NY-ESO-1-specific CD8(+) T cells recognized D(d)-restricted 8mer peptides, NY-ESO-181-88. MAGE-A4-specific CD8(+) T cells recognized D(d)-restricted 9mer peptides, MAGE-A4265-273. MHC/peptide tetramers allowed us to analyze the kinetics and distribution of the antigen-specific immune responses, and we found that stronger antigen-specific CD8(+) T cell responses were required for more effective anti-tumor activity. Taken together, these animal models are valuable for evaluation of immune responses and optimization of the efficacy of cancer vaccines.

The size and distribution of donations: Effects of numbers of potential recipients

Whereas much literature exists on choice overload, little is known about effects of numbers of alternatives in donation decisions. How do these affect both the size and distribution of donations? We hypothesize that donations are affected by the reputation of recipients and increase with their number, albeit at a decreasing rate. Allocations to recipients reflect different concepts of fairness equity and equality. Both may be employed but, since they differ in cognitive and emotional costs, numbers of recipients are important. Using a cognitive (emotional) argument, distributions become more uniform (skewed) as numbers increase. In a survey, respondents indicated how they would donate lottery winnings of 50 Euros. Results indicated that more was donated to NGO s that respondents knew better. Second, total donations increased with the number of recipients albeit at a decreasing rate. Third, distributions of donations became more skewed as numbers increased. We comment on theoretical and practical implications.

Nonrandom Distribution of Cryptic Repeating Triplets of Purines and Pyrimidines (RNY)(n) in gp120 of HIV Type1.

Abstract We have analyzed purine (R) and pyrimidine (Y) codon patterns in variable and constant regions of HIV-1 gp120 in seven patients infected with different HIV-1 subtypes and naive to antiretroviral therapy. We have calculated the relative frequency of each in-frame codon RNY, YNR, RNR, and YNY (N=any nucleotide) in variable and constant regions of gp120, in the sequence within indels and at indels' flanking sites. Our data show that hypervariable regions V1, V2, V4, and V5 are characterized by the presence of long stretches of RNY codons constituting the majority of the sequence portion within insertions/deletions. In full-length gp120 and within inserted/deleted fragments the number of AVT (V=A, C, G) codons did not exceed 50% of the total RNY codons. RNY strings in variable regions spanned up to 21 codons and were always in frame. In contrast, RNY strings in constant regions were mostly out of frame and their length was limited to five codons. The frequency of the codon RNY was found to be significantly higher in variable regions (p<0.0001; t-test), within indels, and at indels' flanking sites (p<0.0001; χ(2) test). Analysis of the distribution of RNY strings equal to or longer than five codons in the full genome of HXB2 also shows that these sequences are mostly out of frame, unless they contain a potential N-glycosylation site or an asparagine. These data suggest that cryptic repeats of RNY may play a role in the genesis of multiple base insertions and deletions in hypervariable regions of gp120.

Geographic distribution of hepatitis C virus genotypes in Brazil

Brazil is a country of continental dimension with a population of different ethnic backgrounds. Thus, a wide variation in the frequencies of hepatitis C virus (HCV) genotypes is expected to occur. To address this point, 1,688 sequential samples from chronic HCV patients were analyzed. HCV-RNA was amplified by the RT-PCR from blood samples collected from 1995 to 2000 at different laboratories located in different cities from all Brazilian States. Samples were collected in tubes containing a gel separator, centrifuged in the site of collection and sent by express mail in a refrigerated container to Laboratório Bioquímico Jardim Paulista, São Paulo, SP, Brazil. HCV- RNA was extracted from serum and submitted to RT and nested PCR using standard procedures. Nested PCR products were submitted to cycle sequencing reactions without prior purification. Sequences were analyzed for genotype determination and the following frequencies were found: 64.9% (1,095) for genotype 1, 4.6% (78) for genotype 2, 30.2% (510) for genotype 3, 0.2% (3) for genotype 4, and 0.1% (2) for genotype 5. The frequencies of HCV genotypes were statistically different among Brazilian regions (P = 0.00017). In all regions, genotype 1 was the most frequent (51.7 to 74.1%), reaching the highest value in the North; genotype 2 was more prevalent in the Center-West region (11.4%), especially in Mato Grosso State (25.8%), while genotype 3 was more common in the South (43.2%). Genotypes 4 and 5 were rarely found and only in the Southeast, in São Paulo State. The present data indicate the need for careful epidemiological surveys throughout Brazil since knowing the frequency and distribution of the genotypes would provide key information for understanding the spread of HCV.

Distribution of human immunodeficiency virus type 1 subtypes in the state of Amazonas, Brazil, and subtype C identification

Few studies have reported the molecular epidemiological characterization of HIV-1 in the Northern region of Brazil. The present study reports the molecular and epidemiological characterization of 31 HIV-1 isolates from blood donors from the State of Amazonas who donated blood between April 2006 and March 2007. Serum/plasma samples from all donors were screened for HIV antibodies by ELISA and the results confirmed by Western blot analysis. Genomic DNA was extracted from the buffy coat using the Super Quik-Gene-DNA Isolation kit. Nested PCR was performed on the env, gag, and pol regions of HIV-1 using the Gene Amp PCR System 9700. Sequencing reactions were performed using the inner PCR primers and the DYEnamic™ ET Dye Terminator Kit, and phylogenetic analysis was performed using the gag, pol, and env gene sequences. We collected samples from 31 blood donors who tested positive for HIV-1 in confirmatory experiments. The male:female ratio of blood donors was 3.4:1, and the mean age was 32.4 years (range: 19 to 61 years). Phylogenetic analysis showed that subtype B is the most prevalent among Northern Brazilian HIV-1-seropositive blood donors. One HIV-1 subtype C and one circulating recombinant form (CRF_BF) of HIV-1 were identified in the State of Amazonas. This is the first study showing the occurrence of a possible "homogenous" subtype C in this region of Brazil. This finding could contribute to a better characterization of the HIV-1 strains that circulate in the country.

SCaFoS: a tool for Selection, Concatenation and Fusion of Sequences for phylogenomics

Affiliation: Département de Biochimie, Université de Montréal

An analytical study of the power distribution pattterns of silica waveguides with high paraboloic cylindrical deformation

An attempt is made to determine the relative power distribution in a step-index parabolic cylindrical waveguide (PCW) with high deformation across the direction of propagation. The guide is assumed to be made of silica. The scalar field approximation is employed for the analysis under which a vanishing refractive-index (RI) difference in the waveguide materials is considered. Further, no approximation for folds- is used in the analytical treatment. Due to the geometry of such waceguides, PCWs lose the well-defined modal discreteness, and a kind of mode bunching is observed instead, which becomes much more prominent in PCWs with high bends. However, with the increase in cross-sectional size, the mode-bunching tendency is slightly reduced. The general expressions for power in the guiding and nonguiding sections are obtained, and the fractional power patterns in all of the sections are presented for PCWs of various cross-sectional dimensions. It is observed that the confinement of power in the core section is increased for PCWs of larger cross-sectional size. Moreover, a fairly uniform distribution of power is seen over the modes having intermediate values of propagation constants

Sequence analysis and distribution of an IS3-like insertion element isolated from Salmonella enteritidis

The nucleotide sequence of a 3 kb region immediately upstream of the sef operon operon of Salmonella enteritidis was determined. A 1230 base pair insertion sequence which shared sequence identity (> 75%) with members of the IS3 family was revealed. This element, designated IS1230, had almost identical (90% identity) terminal inverted repeats to Escherichia coli IS3 but unlike other IS3-like sequences lacked the two characteristic open reading frames which encode the putative transposase. S. enteritidis possessed only one copy of this insertion sequence although Southern hybridisation analysis of restriction digests of genomic DNA revealed another fragment located in a region different from the sef operon which hybridised weakly which suggested the presence of an IS1230 homologue. The distribution of IS1230 and IS1230-like elements was shown to be widespread amongst salmonellas and the patterns of restriction fragments which hybridised differed significantly between Salmonella serotypes and it is suggested that IS1230 has potential for development as a differential diagnostic tool.

Differential distribution of some extracellular matrix fibers in an experimentally denervated rat megaileum

Absence of enteric neurons is associated with thickening of the intestinal muscularis externa in Chagas' disease. The thickening is due to hyperplasia and hypertrophy of the smooth muscle cells and increased extracellular matrix components. The influence of the nervous system on the structure of the smooth muscle cells and its associated matrix has been poorly investigated. An experimental model of denervation of the ileum in rats was performed by application of the surfactant agent benzalkonium chloride that selectively destroys the myenteric plexus. Three months later, ileal tissue samples were obtained and studied by histochemistry and transmission electron microsocopy. Sham operated rats were used as controls. The diameter of collagen fibrils was evaluated in electron micrographs. The histopathological analysis showed thickening of the muscular layer. The thin and weakly arranged collagen and reticulin fibers surrounding the smooth muscle cells, observed in control cases by Picrosirius polarization (PSP) stain method, corresponded to a population of loosely packed thin collagen fibrils of uniform diameters (mean = 29.16 nm) at the ultrastructural level. In contrast, the thick and strongly birefringent fibers around the muscle cells, observed in the treated group, stained by PSP, corresponded to densely packed thicker fibrils with large variation in diameter (mean = 39.41 nm). Comparison of the data demonstrated statistically significant difference between the groups suggesting that the replacement of loosely arranged reticulin fibers by fibrous tissue (with typical collagen fiber), may alter the biomechanical function resulting in impairment of muscular contraction. (c) 2007 Elsevier Ltd. All rights reserved.

Distribution of the Bari-I transposable element in stable hybrid strains between Drosophila melanogaster and Drosophila simulans and in Brazilian populations of these species

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

Distribution of Cr, Ni, Cu, Cd and Pb in different molecular size of humic fractions extracted from water and sediment samples collected at Anhumas reservoir - Araraquara/SP

In this work was studied the distribution of Cr, Ni, Cu, Cd and Pb in humic fractions with different molecular size. The HS were extracted from waters (AHS), surface sediments (HESS), interface water sediment (HSIS) and bottom sediment (HSBS) collected in the Anhumas surface water collection reservoir, located in the district of Araraquara - São Paulo State Brazil. The humic substances were extracted by procedures recommended by International Humic Substances Society (IHSS). After purification by dialysis, the humic substances were fractionated using a multistage tangential flow ultrafiltration system. The fractionation patterns of HS characterized a mass distribution relatively uniform among the fractions with different molecular sizes, with larger values in the fractions F-2 (20.8%) and F-4 (23.8%), Except for the ions Pb(II) and Cu(II), which presented relatively higher concentrations in the fractions F-2 and F-4, respectively. In general, chromium, nickel, cadmium and lead have similar distributions in the five fractions with larger and medium molecular sizes (F-1 to F-5). With relation to the mass distributions in the different humic substances fractions extracted from sediment samples collected at three depth, they presented 42-48% of HS in the fractions with larger molecular sizes (F-1 and F-2), 29-31% in the middle fractions (F-3 and F-4) and 13-20% in the fractions with smaller molecular sizes (F-5 and F-6). In general, the metallic ions presented distributions similar among the respective fractions F-1 to F-6, Exceptions for Pb(II) and M(II) in surface sediment with concentrations relatively smaller in the fractions F-2 and F-4, respectively,

Geographical distribution of vampire bat-related cattle rabies in Brazil

Seventy-seven rabies virus (RV) isolates originating from Brazilian cattle were genetically characterized. Partial nucleoprotein gene sequences of these isolates were phylogenetically and geographically analyzed. Cattle isolates, which clustered with the vampire bat-related RV group, were further subdivided into nine genetic subgroups. These subgroups were distributed widely in lowland regions, with some subgroups separated from each other by mountain ranges. In addition, separation of the groups in mountainous regions was correlated with altitude. These results indicate that cattle rabies is derived from several regionally-defined variants, which suggests that its geographical distribution is related to that of the vampire bat population.

Intrachromosomal distribution of telomeric repeats in Eumops glaucinus and eumops perotis (Molossidae, Chiroptera)

The location of chromosomal telomeric repeats (TTAGGG)(n) was investigated in two species of the Molossidae family, Eumops glaucinus and Eumops perotis. The diploid chromosome number (2n) is 40 in E. glaucinus and 48 in E. perotis and the fundamental numbers (FN) are 64 and 58, respectively. It has been suggested that the E. glaucinus karyotype has evolved from the E. perotis karyotype through Robertsonian fusion events. In the present study, the telomeric sequences were detected at the termini of chromosomes in both species. In addition, E. glaucinus also displayed telomeric repeats in centromeric and pericentromeric regions in almost all biarmed chromosomes. Conversely, in E. perotis pericentromeric signals were only observed in two biarmed chromosomes. In both E. glaucinus and E. perotis, such telomeric sequences were observed as part of the heterochromatin. The interstitial sites of telomeric sequences suggest that they are remnants of telomeres of ancestral chromosomes that participated in the fusion event.

Origin and distribution of AT-rich repetitive DNA families in Triatoma infestans (Heteroptera)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)