Differences in the post-translational modifications of human papillomavirus type 6b major capsid protein expressed from a baculovirus system compared with a vaccinia virus system

Virus-like particles (VLPs) are being currently investigated in vaccines against viral infections in humans. There are different recombinant-protein-expression systems available for obtaining the necessary VLP preparation for vaccination. However, the differences in post-translational modifications of the recombinant proteins obtained and their differences in efficacy in eliciting an anti-viral response in vaccines are not well established. In this study we have compared the posttranslational modifications of human papillomavirus type-6b major capsid protein L1 (HPV 6bL1) expressed using recombinant baculovirus (rBV) in Sf9 (Spodoptera frugiperda) insect cells, with the protein expressed using recombinant vaccinia virus (rVV) in CV-1 kidney epithelial cells, Two-dimensional gel electrophoresis of biosynthetically labelled rBV-expressed HPV 6bL1 showed several post-translationally modified variants of the protein, whereas rVV-expressed HPV 6bL1 showed only a few variants. Phosphorylations were detected at threonine and serine residues for the L1 expressed from rBV compared with phosphorylation at serine residues only for the L1 expressed from rVV. HPV 6bL1 expressed using rBV incorporated [H-3]mannose and [H-3]galactose, whereas HPV 6bL1 expressed using rVV incorporated only [H-3]galactose. We conclude that post-translational modification of recombinant HPV 6bL1 can differ according to the system used for its expression. Since recombinant L1 protein is a potential human-vaccine candidate, the implication of the observed differences in post-translational modifications on immunogenicity of L1 VLPs warrants investigation.

The case for a new system for oversight of research on human subjects

The increasing emphasis on evidence-based clinical practice has thrown into sharp focus multiple deficiencies in current systems of ethical review. This paper argues that a complete overhaul of systems for ethical oversight of studies involving human subjects is now required as developments in medical, epidemiological and genetic research have outstripped existing structures for ethical supervision. It shows that many problems are now evident and concludes that sequential and piecemeal amendments to present arrangements an inadequate to address these. Ar their core present systems of ethical review still rely on the integrity and judgement of individual investigators. One possible alternative is to train and license research investigators, make explicit their responsibilities and have ethics committees devote much more of their time to monitoring research activity in order to detect those infringing the rules.

Identification of "pathologs" (disease-related genes) from the RIKEN mouse cDNA dataset using human curation plus FACTS, a new biological information extraction system

Background: A major goal in the post-genomic era is to identify and characterise disease susceptibility genes and to apply this knowledge to disease prevention and treatment. Rodents and humans have remarkably similar genomes and share closely related biochemical, physiological and pathological pathways. In this work we utilised the latest information on the mouse transcriptome as revealed by the RIKEN FANTOM2 project to identify novel human disease-related candidate genes. We define a new term patholog to mean a homolog of a human disease-related gene encoding a product ( transcript, anti-sense or protein) potentially relevant to disease. Rather than just focus on Mendelian inheritance, we applied the analysis to all potential pathologs regardless of their inheritance pattern. Results: Bioinformatic analysis and human curation of 60,770 RIKEN full-length mouse cDNA clones produced 2,578 sequences that showed similarity ( 70 - 85% identity) to known human-disease genes. Using a newly developed biological information extraction and annotation tool ( FACTS) in parallel with human expert analysis of 17,051 MEDLINE scientific abstracts we identified 182 novel potential pathologs. Of these, 36 were identified by computational tools only, 49 by human expert analysis only and 97 by both methods. These pathologs were related to neoplastic ( 53%), hereditary ( 24%), immunological ( 5%), cardio-vascular (4%), or other (14%), disorders. Conclusions: Large scale genome projects continue to produce a vast amount of data with potential application to the study of human disease. For this potential to be realised we need intelligent strategies for data categorisation and the ability to link sequence data with relevant literature. This paper demonstrates the power of combining human expert annotation with FACTS, a newly developed bioinformatics tool, to identify novel pathologs from within large-scale mouse transcript datasets.

The human temporal cortex: Characterization of neurons expressing nitric oxide synthase, neuropeptides and calcium-binding proteins, and their glutamate receptor subunit profiles

Immunocytochemical techniques were used to examine the distribution of neurons immunoreactive (-ir) for nitric oxide synthase (nNOS), somatostatin (SOM), neuropeptide Y (NPY), parvalbumin (PV), calbindin (CB) and calretinin (CH), in the inferotemporal gyros (Brodmann's area 21) of the human neocortex. Neurons that colocalized either nNOS or SOM with PV, CB or CR were also identified by double-labeling techniques. Furthermore, glutamate receptor subunit profiles (GluR1, GluR2/3, GluR2/4, GluR5/6/7 and NMDAR1) were also determined for these cells. The number and distribution of cells containing nNOS, SOM, NPY, PV, CB or CR differed for each antigen. In addition, distinct subpopulations of neurons displayed different degrees of colocalization of these antigens depending on which antigens were compared. Moreover, cells that contained nNOS, SOM, NPY, PV, GB or CR expressed different receptor subunit profiles. These results show that specific subpopulations of neurochemically identified nonpyramidal cells may be activated via different receptor subtypes. As these different subpopulations of cells project to specific regions of pyramidal calls, facilitation of subsets of these cells via different receptor subunits may activate different inhibitory circuits. Thus, various distinct, but overlapping, inhibitory circuits may act in concert in the modulation of normal cortical function, plasticity and disease.

The pyramidal cell in cognition: A comparative study in human and monkey

Here we present evidence that the pyramidal cell phenotype varies markedly in the cortex of different anthropoid species. Regional and species differences in the size of, number of bifurcations in, and spine density of the basal dendritic arbors cannot be explained by brain size. Instead, pyramidal cell morphology appears to accord with the specialized cortical function these cells perform. Cells in the prefrontal cortex of humans are more branched and more spinous than those in the temporal and occipital lobes. Moreover, cells in the prefrontal cortex of humans are more branched and more spinous than those in the prefrontal cortex of macaque and marmoset monkeys. These results suggest that highly spinous, compartmentalized, pyramidal cells (and the circuits they form) are required to perform complex cortical functions such as comprehension, perception, and planning.

Human Fatalities from Cyanobacteria: Chemical and Biological Evidence for Cyanotoxins

An outbreak of acute liver failure occurred at a dialysis center in Caruaru, Brazil (8 degrees 17 'S, 35 degrees 58 'W), 134 km from Recife, the state capital of Pernambuco. At the clinic, 116 (89%) of 131 patients experienced visual disturbances, nausea, and vomiting after routine hemodialysis treatment on 13-20 February 1996. Subsequently, 100 patients developed acute liver failure, and of these 76 died. As of December 1996, 52 of the deaths could be attributed to a common syndrome now called Caruaru syndrome. Examination of phytoplankton from the dialysis clinic's water source, analyses of the clinic's water treatment system, plus serum and liver tissue of clinic patients led to the identification of two groups of cyanobacterial toxins, the hepatotoxic cyclic peptide microcystins and the hepatotoxic alkaloid cylindrospermopsin. Comparison of victims' symptoms and pathology using animal studies of these two cyanotoxins leads us to conclude that the major contributing factor to death of the dialyses patients was intravenous exposure to microcystins, specifically microcystin-YR, -LR, and -AR. From liver concentrations and exposure volumes, it was estimated that 19.5 mug/L microcystin was in the water used for dialysis treatments. This is 19.5 times the level set as a guideline for safe drinking water supplies by the World. Health Organization.

Inducible system in human hepatoma cell lines for hepatitis C virus production

We cloned the complete complementary DNA of an isolate of the hepatitis C virus, HCV-S1, into a tetra cycline-inducible expression vector and stably transfected it into two human hepatoma cell lines, Huh7 and HepG2. Twenty-six Huh7 and two HepG2-positive clones were obtained after preliminary screening. Two Huh7 (SH-7 and -9) and one HepG2 (G-19) clones were chosen for further characterisation. Expression of HCV proteins in these cells accumulated from 6 In to 4 days posttreatment. Full-length viral plus-strand RNA was detected by Northern analyses. Using RT-PCR and ribonuclease protection assay, we also detected the synthesis of minus-strand HCV RNA. Plus- and minus-strand viral RNA was still detected after treatment with actinomycin D. Indirect immunofluorescence staining with anti-E2, NS4B, and NS5A revealed that these proteins were mostly localised to the endoplasmic reticulum (ER). Culture media from tet-induced SH-9 cells was separated on sucrose density gradients and analysed for the presence of HCV RNA. Viral RNA levels peaked at two separate ranges, one with a buoyant density of 1.08 g/ml and another from 1.17 to 1.39 g/ml. Electron microscopy demonstrated the presence of subviral-like particles (approximately 20-25 nm in diameter) in the cytoplasm of SH-9 and G-19 cells, which were positively labelled by anti-HCV core antibodies. Anti-E2 antibodies strongly labelled cytoplasmic vesicular structures and some viral-like particles. Complete viral particles of about 50 nm which reacted with anti-E2 antibodies were observed in the culture media of tet-induced SH-9 cells following negative staining. Supernatant from tet-treated SH-9 cells was found to infect naive Huh7 and stable Huh7-human CD81 cells. (C) 2002 Elsevier Science (USA).

Mechatronic system for ABO human blood typing

In medical emergency situations, when a patient needs a blood transfusion, the universal blood type O− is administered. This procedure may lead to the depletion of stock reserves of O− blood. Nowadays, there is no commercial equipment capable of determining the patient's blood type in situ, in a fast and reliable process. Human blood typing is usually performed through the manual test, which involves a macroscopic observation and interpretation of the results by an analyst. This test, despite of having a fast response time, may lead to human errors, which sometimes can be fatal to the patient. This paper presents the development of an automatic mechatronic prototype for determining human blood typing (ABO and Rh systems) through image processing techniques. The prototype design takes into account the characteristics of reliability of analysis, portability, and response time allowing the system to be used in emergency situations. The developed prototype performs blood and reagents mixture acquires the resultant image and processes the data (based on image processing techniques) to determine the sample blood type. It was tested in a laboratory, using cataloged samples of blood types, provided by the Portuguese Institute of Blood and Transplantation. Hereafter, it is expected to test and validate the prototype in clinical environments.

Effect of technical parameters on dose and image quality in a computed radiography system

The discovery of X-rays was undoubtedly one of the greatest stimulus for improving the efficiency in the provision of healthcare services. The ability to view, non-invasively, inside the human body has greatly facilitated the work of professionals in diagnosis of diseases. The exclusive focus on image quality (IQ), without understanding how they are obtained, affect negatively the efficiency in diagnostic radiology. The equilibrium between the benefits and the risks are often forgotten. It is necessary to adopt optimization strategies to maximize the benefits (image quality) and minimize risk (dose to the patient) in radiological facilities. In radiology, the implementation of optimization strategies involves an understanding of images acquisition process. When a radiographer adopts a certain value of a parameter (tube potential [kVp], tube current-exposure time product [mAs] or additional filtration), it is essential to know its meaning and impact of their variation in dose and image quality. Without this, any optimization strategy will be a failure. Worldwide, data show that use of x-rays has been increasingly frequent. In Cabo Verde, we note an effort by healthcare institutions (e.g. Ministry of Health) in equipping radiological facilities and the recent installation of a telemedicine system requires purchase of new radiological equipment. In addition, the transition from screen-films to digital systems is characterized by a raise in patient exposure. Given that this transition is slower in less developed countries, as is the case of Cabo Verde, the need to adopt optimization strategies becomes increasingly necessary. This study was conducted as an attempt to answer that need. Although this work is about objective evaluation of image quality, and in medical practice the evaluation is usually subjective (visual evaluation of images by radiographer / radiologist), studies reported a correlation between these two types of evaluation (objective and subjective) [5-7] which accredits for conducting such studies. The purpose of this study is to evaluate the effect of exposure parameters (kVp and mAs) when using additional Cooper (Cu) filtration in dose and image quality in a Computed Radiography system.

Hand gesture recognition system based in computer vision and machine learning : applications on human-machine interaction

Sendo uma forma natural de interação homem-máquina, o reconhecimento de gestos implica uma forte componente de investigação em áreas como a visão por computador e a aprendizagem computacional. O reconhecimento gestual é uma área com aplicações muito diversas, fornecendo aos utilizadores uma forma mais natural e mais simples de comunicar com sistemas baseados em computador, sem a necessidade de utilização de dispositivos extras. Assim, o objectivo principal da investigação na área de reconhecimento de gestos aplicada à interacção homemmáquina é o da criação de sistemas, que possam identificar gestos específicos e usálos para transmitir informações ou para controlar dispositivos. Para isso as interfaces baseados em visão para o reconhecimento de gestos, necessitam de detectar a mão de forma rápida e robusta e de serem capazes de efetuar o reconhecimento de gestos em tempo real. Hoje em dia, os sistemas de reconhecimento de gestos baseados em visão são capazes de trabalhar com soluções específicas, construídos para resolver um determinado problema e configurados para trabalhar de uma forma particular. Este projeto de investigação estudou e implementou soluções, suficientemente genéricas, com o recurso a algoritmos de aprendizagem computacional, permitindo a sua aplicação num conjunto alargado de sistemas de interface homem-máquina, para reconhecimento de gestos em tempo real. A solução proposta, Gesture Learning Module Architecture (GeLMA), permite de forma simples definir um conjunto de comandos que pode ser baseado em gestos estáticos e dinâmicos e que pode ser facilmente integrado e configurado para ser utilizado numa série de aplicações. É um sistema de baixo custo e fácil de treinar e usar, e uma vez que é construído unicamente com bibliotecas de código. As experiências realizadas permitiram mostrar que o sistema atingiu uma precisão de 99,2% em termos de reconhecimento de gestos estáticos e uma precisão média de 93,7% em termos de reconhecimento de gestos dinâmicos. Para validar a solução proposta, foram implementados dois sistemas completos. O primeiro é um sistema em tempo real capaz de ajudar um árbitro a arbitrar um jogo de futebol robótico. A solução proposta combina um sistema de reconhecimento de gestos baseada em visão com a definição de uma linguagem formal, o CommLang Referee, à qual demos a designação de Referee Command Language Interface System (ReCLIS). O sistema identifica os comandos baseados num conjunto de gestos estáticos e dinâmicos executados pelo árbitro, sendo este posteriormente enviado para um interface de computador que transmite a respectiva informação para os robôs. O segundo é um sistema em tempo real capaz de interpretar um subconjunto da Linguagem Gestual Portuguesa. As experiências demonstraram que o sistema foi capaz de reconhecer as vogais em tempo real de forma fiável. Embora a solução implementada apenas tenha sido treinada para reconhecer as cinco vogais, o sistema é facilmente extensível para reconhecer o resto do alfabeto. As experiências também permitiram mostrar que a base dos sistemas de interação baseados em visão pode ser a mesma para todas as aplicações e, deste modo facilitar a sua implementação. A solução proposta tem ainda a vantagem de ser suficientemente genérica e uma base sólida para o desenvolvimento de sistemas baseados em reconhecimento gestual que podem ser facilmente integrados com qualquer aplicação de interface homem-máquina. A linguagem formal de definição da interface pode ser redefinida e o sistema pode ser facilmente configurado e treinado com um conjunto de gestos diferentes de forma a serem integrados na solução final.

Real-Time Visual Ground-Truth System for Indoor Robotic Applications

The robotics community is concerned with the ability to infer and compare the results from researchers in areas such as vision perception and multi-robot cooperative behavior. To accomplish that task, this paper proposes a real-time indoor visual ground truth system capable of providing accuracy with at least more magnitude than the precision of the algorithm to be evaluated. A multi-camera architecture is proposed under the ROS (Robot Operating System) framework to estimate the 3D position of objects and the implementation and results were contextualized to the Robocup Middle Size League scenario.

Calibration Method for Underwater Visual Ground-Truth System

This work presents an automatic calibration method for a vision based external underwater ground-truth positioning system. These systems are a relevant tool in benchmarking and assessing the quality of research in underwater robotics applications. A stereo vision system can in suitable environments such as test tanks or in clear water conditions provide accurate position with low cost and flexible operation. In this work we present a two step extrinsic camera parameter calibration procedure in order to reduce the setup time and provide accurate results. The proposed method uses a planar homography decomposition in order to determine the relative camera poses and the determination of vanishing points of detected lines in the image to obtain the global pose of the stereo rig in the reference frame. This method was applied to our external vision based ground-truth at the INESC TEC/Robotics test tank. Results are presented in comparison with an precise calibration performed using points obtained from an accurate 3D LIDAR modelling of the environment.

Visualization and interaction in a simulation system for flood emergencies

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau de Mestre em Engenharia Informática

Morphological changes in the digestive system of 93 human immunodeficiency virus positive patients: an autopsy study

Involvement of the digestive system in patients with acquired immunodeficiency syndrome (AIDS) is frequent and many changes in these patients are diagnosed only at autopsy. There are few studies of autopsy with detailed analysis of this system and only one was conducted in Brazil. We evaluated each segment of the digestive system in 93 consecutive autopsies of patients infected with human immunodeficiency virus (HIV) and the importance of these lesions to death. Of these, 90 (96.8%) patients had AIDS. We reviewed medical records, autopsy reports and histological sections from tongue to rectum stained with hematoxylin-eosin. When necessary, we analyzed special stains and immunohistochemistry to investigate infections. There was damage to the digestive system in 73 (78.5%) cases. The most common infections were candidiasis (42%), cytomegalovirus (29%), histoplasmosis (11.8%), toxoplasmosis (9.7%) and mycobacterial infection (9.7%). Malignancies were rare, present in four (4.3%) cases (two Kaposi's sarcoma, one adenocarcinoma and one metastatic embryonal carcinoma). All segments showed lesions: tongue (48.6%), esophagus (44.8%), stomach (44.7%), colon (43.2%) and small intestine (28.9%). The lesions found were immediate cause of death in five (5.4%) cases. In another 36 (38.7%) cases the basic disease was systemic and also compromised the digestive system.

Evaluation of WGA and Concanavalin A (Con A) lectin as biomarkers of hepatosplenic schistosomiasis in human biopsies with no evidence of egg-granuloma system

Introduction: Colonic lesions are predominant in patients with schistosomiasis. However, carbohydrate alterations in colonic schistosomiasis remain unclear. Lectin-ligands allow us to identify changes in the saccharide patterns of cells. Methods: Biopsies of descending and rectosigmoid colon of patients were submitted to WGA and Con A lectin histochemistry. Results: WGA stained stroma and gland cells of descending colon and rectosigmoid tissues in a granular strong cytoplasmatic pattern in schistosomiasis specimens differing from normal control and Con A failing to recognize all samples analyzed. Conclusions: WGA ligands are expressed differently in patients with hepatosplenic schistosomiasis and no evidence of egg-granuloma system.