Preclinical assessment of an anti-HTLV-1 heterologous DNA/MVA vaccine protocol expressing a multiepitope HBZ protein.

BACKGROUND: Human T-lymphotropic virus 1 (HTLV-1) is associated with the development of several pathologies and chronic infection in humans. The inefficiency of the available treatments and the challenge in developing a protective vaccine highlight the need to produce effective immunotherapeutic tools. The HTLV-1 basic leucine zipper (bZIP) factor (HBZ) plays an important role in the HTLV-1 persistence, conferring a survival advantage to infected cells by reducing the HTLV-1 proteins expression, allowing infected cells to evade immune surveillance, and enhancing cell proliferation leading to increased proviral load. METHODS: We have generated a recombinant Modified Virus Vaccinia Ankara (MVA-HBZ) and a plasmid DNA (pcDNA3.1(+)-HBZ) expressing a multiepitope protein based on peptides of HBZ to study the immunogenic potential of this viral-derived protein in BALB/c mice model. Mice were immunized in a prime-boost heterologous protocol and their splenocytes (T CD4+ and T CD8+) were immunophenotyped by flow cytometry and the humoral response was evaluated by ELISA using HBZ protein produced in prokaryotic vector as antigen. RESULTS: T CD4+ and T CD8+ lymphocytes cells stimulated by HBZ-peptides (HBZ42-50 and HBZ157-176) showed polyfunctional double positive responses for TNF-α/IFN-γ, and TNF-α/IL-2. Moreover, T CD8+ cells presented a tendency in the activation of effector memory cells producing granzyme B (CD44+High/CD62L-Low), and the activation of Cytotoxic T Lymphocytes (CTLs) and cytotoxic responses in immunized mice were inferred through the production of granzyme B by effector memory T cells and the expression of CD107a by CD8+ T cells. The overall data is consistent with a directive and effector recall response, which may be able to operate actively in the elimination of HTLV-1-infected cells and, consequently, in the reduction of the proviral load. Sera from immunized mice, differently from those of control animals, showed IgG-anti-HBZ production by ELISA. CONCLUSIONS: Our results highlight the potential of the HBZ multiepitope protein expressed from plasmid DNA and a poxviral vector as candidates for therapeutic vaccine.

Comparative Analysis of Gait Speed Estimation Using Wideband and Narrowband Radars, Thermal Camera, and Motion Tracking Suit Technologies.

Research has shown that cognitive and physical functioning of older adults can be reflected in indicators such as walking speed. While changes in cognition, mobility, or health cause changes in gait speed, often gradual variations in walking speed go undetected until severe problems arise. Discrete clinical assessments during clinical consultations often fail to detect changes in day-to-day walking speeds and do not reflect walking speeds in everyday environments, where most of the mobility issues happen. In this paper, we compare four walking speed measurement technologies to a GAITRite mat (gold standard): (1) an ultra wideband radar (covering the band from 3.3 GHz to 10 GHz), (2) a narrow band 24-GHz radar (with a bandwidth of 250 MHz), (3) a perception Neuron Motion Tracking suit, and (4) a thermal camera. Data were collected in parallel using all sensors at the same time for 10 healthy adults for normal and slow walking paces. A comparison of the sensors indicates better performance at lower gait speeds, with offsets (when compared to GAITRite) between 0.1 and 20% for the ultra wideband radar, 1.9 and 17% for the narrowband radar, 0.1 and 38% for the thermal camera, and 1.7 and 38% for the suit. This paper supports the potential of unobtrusive radar-based sensors and thermal camera technologies for ambient autonomous gait speed monitoring for contextual, privacy-preserving monitoring of participants in the community.

Cloning, expression and characterization of SeM protein of Streptococcus equi subsp. equi and evaluation of its use as antigen in an indirect ELISA / Clonagem, expressão e caracterização de proteína SeM de Streptococcus equi subsp. equi e avaliação de sua utilização como antígeno em um ELISA indireto

Strangles is an economically important horse disease caused by Streptococcus equi subsp. equi. The diagnosis can be confirmed either directly by bacterial isolation and PCR or by ELISA, which is an indirect method based on the detection of serum antibodies. The aim of this study was to clone, express and characterize the SeM protein of Streptococcus equi subsp. equi, evaluate its use as antigen in indirect ELISA and determine its performance to distinguish sera of negative, vaccinated and positive animals. This was initially performed by cloning the gene encoding the SeM protein and its expression in Escherichia coli. Subsequently, the protein produced was characterized and used as antigen in ELISA. Serum samples for evaluation were taken from 40 negative foals, 46 horses vaccinated with a commercial vaccine against strangles and 46 horses diagnosed with the disease. The test showed high specificity and sensitivity, allowing discrimination between negative and positive, positive and vaccinated animals, and vaccinated animals and negative sera. Thus, it was concluded that the protein produced rSeM, which can be used as antigen for disease diagnosis, and the described ELISA might be helpful to evaluate the immune status of the herd...

A adenite equina é uma enfermidade economicamente importante de equinos, causada por Streptococcus equi subsp. equi. Seu diagnóstico pode ser confirmado de forma direta, por meio de isolamento bacteriano e de PCR, ou de forma indireta, por meio de ELISA, método baseado na detecção de anticorpos séricos. O objetivo deste estudo foi clonar, expressar e caracterizar a proteína SeM de Streptococcus equi subsp. equi, avaliar sua utilização como antígeno em um ELISA indireto e determinar a capacidade do teste de distinguir soros de animais negativos, vacinados e positivos. Para tal, foi inicialmente realizada a clonagem do gene que codifica para a proteína SeM e sua expressão em Escherichia coli. Posteriormente, a proteína produzida foi caracterizada e utilizada como antígeno em um teste de ELISA indireto. Para avaliação do teste, foram utilizadas amostras de soro de 40 potros negativos, de 46 equinos vacinados com uma vacina comercial contra adenite equina e de 46 equinos com diagnóstico da doença. O teste demonstrou alta sensibilidade e especificidade, permitindo discriminar entre soros negativos e positivos, positivos e de animais vacinados, e negativos e de animais vacinados. Assim, conclui-se que a proteína rSeM produzida pode ser usada como antígeno para o diagnóstico da enfermidade e que o ELISA descrito pode ser útil para avaliar o estado imunológico do rebanho...

Transmission of intra-cellular genetic information: a system proposal.

One of the great challenges of the scientific community on theories of genetic information, genetic communication and genetic coding is to determine a mathematical structure related to DNA sequences. In this paper we propose a model of an intra-cellular transmission system of genetic information similar to a model of a power and bandwidth efficient digital communication system in order to identify a mathematical structure in DNA sequences where such sequences are biologically relevant. The model of a transmission system of genetic information is concerned with the identification, reproduction and mathematical classification of the nucleotide sequence of single stranded DNA by the genetic encoder. Hence, a genetic encoder is devised where labelings and cyclic codes are established. The establishment of the algebraic structure of the corresponding codes alphabets, mappings, labelings, primitive polynomials (p(x)) and code generator polynomials (g(x)) are quite important in characterizing error-correcting codes subclasses of G-linear codes. These latter codes are useful for the identification, reproduction and mathematical classification of DNA sequences. The characterization of this model may contribute to the development of a methodology that can be applied in mutational analysis and polymorphisms, production of new drugs and genetic improvement, among other things, resulting in the reduction of time and laboratory costs.

Caffeine triggers behavioral and neurochemical alterations in adolescent rats.

Caffeine is the psychostimulant most consumed worldwide but concerns arise about the growing intake of caffeine-containing drinks by adolescents since the effects of caffeine on cognitive functions and neurochemical aspects of late brain maturation during adolescence are poorly known. We now studied the behavioral impact in adolescent male rats of regular caffeine intake at low (0.1mg/mL), moderate (0.3mg/mL) and moderate/high (1.0mg/mL) doses only during their active period (from 7:00 P.M. to 7:00 A.M.). All tested doses of caffeine were devoid of effects on locomotor activity, but triggered anxiogenic effects. Caffeine (0.3 and 1mg/mL) improved the performance in the object recognition task, but the higher dose of caffeine (1.0mg/mL) decreased the habituation to an open-field arena, suggesting impaired non-associative memory. All tested doses of caffeine decreased the density of glial fibrillary acidic protein and synaptosomal-associated protein-25, but failed to modify neuron-specific nuclear protein immunoreactivity in the hippocampus and cerebral cortex. Caffeine (0.3-1mg/mL) increased the density of brain-derived neurotrophic factor (BDNF) and proBDNF density as well as adenosine A1 receptor density in the hippocampus, whereas the higher dose of caffeine (1mg/mL) increased the density of proBDNF and BDNF and decreased A1 receptor density in the cerebral cortex. These findings document an impact of caffeine consumption in adolescent rats with a dual impact on anxiety and recognition memory, associated with changes in BDNF levels and decreases of astrocytic and nerve terminal markers without overt neuronal damage in hippocampal and cortical regions.

Isolation and pathogenicity of Toxoplasma gondii in naturally infected (rustic farm) pigs in southern Brazil.

This study reported a serological test for Toxoplasma gondii infection in 100 pigs from 58 rural farms in the state of Rio Grande do Sul, Brazil. Thirty-six pigs were seropositive (IFAT≥1:64). Bioassays were performed for all 36 seropositive pigs, and 17 isolates were obtained (47.2%). Seven of these isolates (41.2%) were highly pathogenic to mice, as clinical signs of acute infection were observed, and tachyzoites were found in the peritoneal exudates, livers, and lungs. The remaining 10 isolates were able to establish a chronic infection in mice, therefore, they were not highly virulent. The results of this study indicate that pork is a potential source of T. gondii transmission to humans.

Anxiolytic-like effects of a new 1-N substituted analog of melatonin in pinealectomized rats.

In spite of the wide variety of drugs available for treating anxiety, this disorder continues to represent a worldwide health problem that is classified within the first 10 causes of disability. Therefore, the search continues for new antianxiety agents, particularly those not related to benzodiazepines. Even though melatonin has been prescribed as an anxiolytic drug, its use is currently limited due to its short half-life and photo-sensitivity, among other disadvantages. The present study explores the antianxiety properties of a new 1-N substituted melatonin analog, M3C, in pinealectomized rats submitted to two behavioral tests (the cumulative burying behavior paradigm and the elevated plus-maze). Results from both tests show that M3C is effective as an anxiolytic-like agent, at doses lower than any other melatonin analog previously reported. The blocking of these actions by luzindole together with the available data suggests that the anxiolytic properties of M3C are mediated by MT1 and MT2 receptors.

E-cadherin loss rather than beta-catenin alterations is a common feature of poorly differentiated thyroid carcinomas.

AIMS: To investigate the immunohistochemical and molecular genetic features of the cadherins/catenins complex in thyroid carcinoma based on the hypothesis that poorly differentiated carcinoma of the thyroid represents an intermediate step between well-differentiated and undifferentiated carcinomas. METHODS AND RESULTS: Immunohistochemistry for E-, P- and N-cadherins and alpha-, beta- and gamma-catenins was performed in a series of 17 cases of poorly differentiated carcinoma of the thyroid. All cases showed absence of membranous expression of E-cadherin with no aberrant expression of P- or N-cadherins; regarding catenins there was heterogeneous loss of expression with membranous immunolocalization of the three catenins in most cases. Molecular analysis of the E-cadherin gene and exon 3 of the beta-catenin gene was also performed by polymerase chain reaction/single-strand conformation polymorphism and sequencing. No mutations in either gene were detected in any case. CONCLUSIONS: In contrast to previous reports, our results suggest that loss of E-cadherin rather than beta-catenin mutation is the crucial event in determining the differentiation 'level' of thyroid carcinomas.

Microsatellite instability, mitochondrial DNA large deletions, and mitochondrial DNA mutations in gastric carcinoma.

Mitochondrial DNA (mtDNA) large deletions and mtDNA mutations have been demonstrated in various types of human cancer. The relationship between the occurrence of such alterations and the nuclear microsatellite instability (MSI) status of the neoplastic cells remains controversial. In an attempt to clarify the situation in gastric carcinoma, we studied, by PCR/SSCP and sequencing, five mitochondrial genes and two D-loop regions in 32 gastric carcinomas that had been previously screened for MSI and mitochondrial common deletion. MtDNA alterations were detected in 26 carcinomas (81%). All the mtDNA mutations, which occurred mainly in the D-loop and ND1 and ND5 genes, were transitions. D-loop alterations (insertions and/or deletions) were not significantly associated with mutations in the coding regions. There was a trend towards an inverse relationship between the occurrence of mitochondrial common deletion and mtDNA mutations. No significant relationship was observed between MSI status and mtDNA mutations, whereas the mitochondrial common deletion appeared to be almost exclusively restricted to MSI-negative tumors. The latter finding--almost no gastric carcinoma with MSI-positive phenotype has large deletions of mtDNA--needs to be confirmed in a larger series and in tumors from other organs.

Abnormalities of the E-cadherin/catenin adhesion complex in classical papillary thyroid carcinoma and in its diffuse sclerosing variant.

The cadherin/catenin complex regulates cellular adhesion and motility and is believed to function as an invasion suppressor system. Several studies have identified alterations in the expression profiles of those molecules in different histotypes of thyroid carcinoma. The diffuse sclerosing variant (DSV) of papillary thyroid carcinoma (PTC) is a rare, highly invasive variant of PTC in which an impairment of cell-cell adhesion may play a major role. In an attempt to progress in the understanding of the clinicopathological features of DSV, this study examined eight cases of DSV, 18 cases of classical PTC and a control group of normal thyroid by immunohistochemistry (E-, P- and N-cadherins and beta-, gamma- and alpha-catenins). The E-cadherin gene was also studied by polymerase chain reaction/single strand conformation polymorphism (PCR/SSCP) and methylation-specific PCR (MSP). In contrast to classical PTC, which showed heterogeneous loss of E-cadherin expression, in almost every case of DSV a pronounced reduction was observed in its membranous expression, accompanied by a relocation to the cytoplasm. Inactivation of the E-cadherin/catenin complex appears to occur in DSV via two different pathways: E-cadherin alteration either through mutation (one out of the eight cases) or through methylation of the E-cadherin gene promoter (three out of five cases); and beta- and/or gamma-catenin alterations (three of the eight cases). Methylation of the E-cadherin gene promoter, abnormalities of E-cadherin expression and alterations of gamma-catenin were also detected in classical PTC. In DSV, as in classical PTC, there is neo-expression of P-cadherin in areas of squamous metaplasia and no N-cadherin expression. In conclusion, abnormalities of the E-cadherin/catenin complex appear to be more pronounced in DSV than in classical PTC. It remains to be shown whether or not such differences are associated with the more aggressive behaviour of DSV compared with classical PTC.

Defective E-cadherin/catenin complexes in human cancer.

Cancer is caused by a series of genomic changes leading directly or indirectly to disturbance of growth, differentiation and tissue integrity. Genomic, transcriptional or posttranscriptional alterations of E-cadherin/catenin complexes that are implicated in various steps of cancer development comprise mutational inactivation, transcriptional downregulation of E-cadherin sometimes accompanied by upregulation of N-cadherin, proteolysis of E-cadherin and posttranslational stabilisation of beta-catenin and plakoglobin. The E-cadherin/catenin complex serves not only cell-cell adhesion but also transduces signals to the nucleus and to the cytoskeleton, either directly or through its connections with multiple other complexes. We review here the expression of E-cadherin/catenin in human cancers, emphasising methods of observation and prognostic interpretation of results. This is illustrated in thyroid lesions from the benign follicular adenoma to the extremely malignant anaplastic carcinoma. The eye is an organ largely neglected by students of cadherins and catenins. The implication of a variety of members of these molecular families in the embryonic development of the eye strongly suggests that disturbances of cadherin/catenin complexes are crucial also in the development of ocular tumours.

Aortic stenosis. Gender influence on left ventricular geometry and function in patients under 70 years of age.

OBJECTIVE: To verify if adaptive left ventricle (LV) characteristics are also present in individuals under 70 years of age with severe aortic stenosis (AS). METHODS: The study comprised 40 consecutive patients under 70 years of age with AS and no associated coronary artery disease, referred for valve surgery. Out of the 40 patients, 22 were men and 18 women, and the mean age was 49.8 +/- 14.3 years. Cardiac symptoms, presence of systemic hypertension (SH), functional class according to the New York Heart Association (NYHA), and valve lesion etiology were considered. LV cavity dimensions, ejection fraction (EF), fractional shortening (FS), mass (MS), and relative diastolic thickness (RDT) were examined by Doppler echocardiography. RESULTS: Fourteen (63.6%) men and 11 (61.6%) women were classified as NYHA class III/IV (p = 0.70). There was no difference in the frequency of angina, syncope or dyspnea between genders. The incidence of SH was greater in women than in men (10 versus 2, p = 0.0044). Women had a smaller LV end-diastolic diameter index (32.1 +/- 6.5 x 36.5 +/- 5.3 mm/m2, p = 0.027), LV end-systolic diameter index (19.9 +/- 5.9 x 26.5 +/- 6.4 mm/m2, p = 0.0022) and LV mass index (MS) (211.4 +/- 71.1 x 270.9 +/- 74.9 g/m2, p = 0.017) when compared with men. EF (66.2 +/- 13.4 x 52.0 +/- 14.6%, p = 0.0032), FS (37.6 +/- 10.7 x 27.9 +/- 9.6%, p = 0.0046) and RDT (0.58 +/- 0.22 x 0.44 +/- 0.09, p = 0.0095) were significantly greater in women than in men. CONCLUSION: It is the patient gender rather than age that influences left ventricular adaptive response to AS.

Myxoma of the mitral valve.

Only rarely do myxomas originate from the mitral valve. This is the report of a 49-year-old woman presenting with congestive heart failure. The diagnosis of an intracardiac tumor involving the anterior cuspid of the mitral valve was made by transesophageal echocardiography. The patient underwent surgery for tumor resection and plasty of the valve was made with reconstruction and preservation of the valve. The diagnosis of myxoma was confirmed by histology. This is the 23rd case of myxoma of the mitral valve reported in the literature.

[Coronary artery dissection without acute myocardial infarction]. / Dissecção de Artéria coronária sem infarto agudo do miocárdio.

A 57 year-old-man with acute aortic dissection (DeBakey type I) who developed right coronary artery dissection without acute myocardial infarction. He was successful surgically treated and became asymptomatic.

[Myocardial revascularization in left coronary trunk lesion in patients over 65 years old]. / Cirurgia de revascularização miocárdica na lesão do tronco da coronária esquerda em pacientes acima de 65 anos de idade.

PURPOSE: To study the short and long-term follow-up of patients with left main coronary artery disease (LMCAD) and age over 65 years, by comparing the results with patients under 65 years-old. METHODS: Twenty-two patients with LMCAD and mean age of 69 +/- 3.5 years (group I) were underwent isolated coronary artery bypass grafting (CABG) and compared to 31 patients with LMCAD, mean age of 54 +/- 7 years (group II), who also underwent isolated CABG. The life-table Kaplan-Meyer method was used to estimate the post-operative survival. The chi-square and Student "t" test were used when necessary. RESULTS: Despite higher operative mortality in group I (9.1% x 3.2%), the difference was statistically not significant. The operative morbidity was similar in both groups. Actuarial survival at 4 years was 85% in group I and 95% in group II. Actuarial survival free of cardiac events was 69% in group II and 75% in group II. CONCLUSION: The CABG is well tolerated and had low morbidity and acceptable mortality in old patients with LMCAD. The long-term survival in these patients was very similar to the younger patients.

Miocardiopatia periparto: Tratamento imunossupressor / Peripartum cardiomyopathy: Immunosupressor therapy

O caso de uma mulher parda de 26 anos de idade que desenvolveu síndrome de insuficiência cardíaca congestiva (ICC) na primeira semana de puerpério de sua primeira gestaçäo é relatado. Refratária às medidas terapêuticas intensivas habituais, a síndrome de ICC respondeu ao uso da associaçäo de ciclofosfamida e prednisona. Chama-se a atençäo para a possibilidade de melhora da miocardiopatia periparto que cursa com sinais de miocardite ou que torna refratária às medidas terapêuticas habituais com uso ddrogas imunossupressoras.