Experimental models in vaccine research: malaria and leishmaniasis

Animal models have a long history of being useful tools, not only to test and select vaccines, but also to help understand the elaborate details of the immune response that follows infection. Different models have been extensively used to investigate putative immunological correlates of protection against parasitic diseases that are important to reach a successful vaccine. The greatest challenge has been the improvement and adaptation of these models to reflect the reality of human disease and the screening of vaccine candidates capable of overcoming the challenge of natural transmission. This review will discuss the advantages and challenges of using experimental animal models for vaccine development and how the knowledge achieved can be extrapolated to human disease by looking into two important parasitic diseases: malaria and leishmaniasis.

Experimental model of intervertebral disc degeneration by needle puncture in Wistar rats

Animal models of intervertebral disc degeneration play an important role in clarifying the physiopathological mechanisms and testing novel therapeutic strategies. The objective of the present study is to describe a simple animal model of disc degeneration involving Wistar rats to be used for research studies. Disc degeneration was confirmed and classified by radiography, magnetic resonance and histological evaluation. Adult male Wistar rats were anesthetized and submitted to percutaneous disc puncture with a 20-gauge needle on levels 6-7 and 8-9 of the coccygeal vertebrae. The needle was inserted into the discs guided by fluoroscopy and its tip was positioned crossing the nucleus pulposus up to the contralateral annulus fibrosus, rotated 360° twice, and held for 30 s. To grade the severity of intervertebral disc degeneration, we measured the intervertebral disc height from radiographic images 7 and 30 days after the injury, and the signal intensity T2-weighted magnetic resonance imaging. Histological analysis was performed with hematoxylin-eosin and collagen fiber orientation using picrosirius red staining and polarized light microscopy. Imaging and histological score analyses revealed significant disc degeneration both 7 and 30 days after the lesion, without deaths or systemic complications. Interobserver histological evaluation showed significant agreement. There was a significant positive correlation between histological score and intervertebral disc height 7 and 30 days after the lesion. We conclude that the tail disc puncture method using Wistar rats is a simple, cost-effective and reproducible model for inducing disc degeneration.

Effect of melatonin on the functional recovery from experimental traumatic compression of the spinal cord

Spinal cord injury is an extremely severe condition with no available effective therapies. We examined the effect of melatonin on traumatic compression of the spinal cord. Sixty male adult Wistar rats were divided into three groups: sham-operated animals and animals with 35 and 50% spinal cord compression with a polycarbonate rod spacer. Each group was divided into two subgroups, each receiving an injection of vehicle or melatonin (2.5 mg/kg, intraperitoneal) 5 min prior to and 1, 2, 3, and 4 h after injury. Functional recovery was monitored weekly by the open-field test, the Basso, Beattie and Bresnahan locomotor scale and the inclined plane test. Histological changes of the spinal cord were examined 35 days after injury. Motor scores were progressively lower as spacer size increased according to the motor scale and inclined plane test evaluation at all times of assessment. The results of the two tests were correlated. The open-field test presented similar results with a less pronounced difference between the 35 and 50% compression groups. The injured groups presented functional recovery that was more evident in the first and second weeks. Animals receiving melatonin treatment presented more pronounced functional recovery than vehicle-treated animals as measured by the motor scale or inclined plane. NADPH-d histochemistry revealed integrity of the spinal cord thoracic segment in sham-operated animals and confirmed the severity of the lesion after spinal cord narrowing. The results obtained after experimental compression of the spinal cord support the hypothesis that melatonin may be considered for use in clinical practice because of its protective effect on the secondary wave of neuronal death following the primary wave after spinal cord injury.

Performance assessment of the single photon emission microscope: high spatial resolution SPECT imaging of small animal organs

The single photon emission microscope (SPEM) is an instrument developed to obtain high spatial resolution single photon emission computed tomography (SPECT) images of small structures inside the mouse brain. SPEM consists of two independent imaging devices, which combine a multipinhole collimator, a high-resolution, thallium-doped cesium iodide [CsI(Tl)] columnar scintillator, a demagnifying/intensifier tube, and an electron-multiplying charge-coupling device (CCD). Collimators have 300- and 450-µm diameter pinholes on tungsten slabs, in hexagonal arrays of 19 and 7 holes. Projection data are acquired in a photon-counting strategy, where CCD frames are stored at 50 frames per second, with a radius of rotation of 35 mm and magnification factor of one. The image reconstruction software tool is based on the maximum likelihood algorithm. Our aim was to evaluate the spatial resolution and sensitivity attainable with the seven-pinhole imaging device, together with the linearity for quantification on the tomographic images, and to test the instrument in obtaining tomographic images of different mouse organs. A spatial resolution better than 500 µm and a sensitivity of 21.6 counts·s-1·MBq-1 were reached, as well as a correlation coefficient between activity and intensity better than 0.99, when imaging 99mTc sources. Images of the thyroid, heart, lungs, and bones of mice were registered using 99mTc-labeled radiopharmaceuticals in times appropriate for routine preclinical experimentation of <1 h per projection data set. Detailed experimental protocols and images of the aforementioned organs are shown. We plan to extend the instrument's field of view to fix larger animals and to combine data from both detectors to reduce the acquisition time or applied activity.

Investigation of cytotoxic and mutagenic effects of Malpighia glabra L. (barbados cherry) fruit pulp and vitamin C on plant and animal test systems

Fruits are important sources of nutrients in human diet, and Barbados Cherry (Malpighia glabra L.) is of particular interest due to its high content of antioxidants. Diets rich in fruits and vegetables protect individuals against diseases and cancer, but excessive intake of vitamins may act as pro-oxidant and generate changes in DNA. To evaluate the effect of different in natura (BAN) and frozen (BAF) Barbados Cherry pulp concentrations and synthetic vitamin C in liquid form (VC) on the chromosome level and the cell cycle division, root meristeme cells of Allium cepa L. and bone marrow cells of Wistar rats Rattus norvegicus, were used as test system. In Allium cepa L., BAN, at the highest concentration (0.4 mg.mL-1) and BAF, at the lowest concentration (0.2 mg.mL-1), inhibited cell division, and there was recovery of cell division after the recovery period in water only for BAN. In the Wistar rats, all treatments with Barbados Cherry, either acute or subchronic, were not cytotoxic or mutagenic; only the highest concentration of VC increased significantly the rate of chromosomal abnormalities. The data obtained are important to reinforce the use of Barbados Cherry fruit in the diet.

Use of the spray chilling method to deliver hydrophobic components: physical characterization of microparticles

Food industry has been developing products to meet the demands of increasing number of consumers who are concerned with their health and who seek food products that satisfy their needs. Therefore, the development of processed foods that contain functional components has become important for this industry. Microencapsulation can be used to reduce the effects of processing on functional components and preserve their bioactivity. The present study investigated the production of lipid microparticles containing phytosterols by spray chilling. The matrices comprised mixtures of stearic acid and hydrogenated vegetable fat, and the ratio of the matrix components to phytosterols was defined by an experimental design using the mean diameters of the microparticles as the response variable. The melting point of the matrices ranged from 44.5 and 53.4 ºC. The process yield was melting point dependent; the particles that exhibited lower melting point had greater losses than those with higher melting point. The microparticles' mean diameters ranged from 13.8 and 32.2 µm and were influenced by the amount of phytosterols and stearic acid. The microparticles exhibited spherical shape and typical polydispersity of atomized products. From a technological and practical (handling, yield, and agglomeration) points of view, lipid microparticles with higher melting point proved promising as phytosterol carriers.

Use of Moringa oleífera Lamarck leaf extract as sugarcane juice clarifier: effects on clarifed juice and sugar

The objective of this study was to evaluate the effect of Moringa oleifera Lam. leaf extract on the sedimentation of impurities in the treatment of sugarcane juice and the effects on sugar quality and on the clarified juice. The experimental design used was a 4x2 factorial arrangement with four replications. The main treatments performed included the extracted original sugarcane juice, the synthetic polyelectrolyte (Flomex 9076), the leaf extract, and a control. The secondary treatments consisted of the sugarcane varieties RB92579 and RB867515. The clarification process used was simple defecation, in which the flocculating agents and the juice, limed and heated, were poured simultaneously into a decanter. The microbiological and chemico-technological characteristics of the extracted and clarified juices were evaluated. The clarified juice was concentrated up to 60° Brix (syrup) and subjected to boiling in a pilot pan using seeds to perform the graining: The sugar was recovered by centrifugation and analyzed for microbiological and chemico-technological characteristics. It was concluded that the use of the Moringa oleifera Lam. leaves extract resulted in a better quality of clarified juice and sugar.

Indução de resposta inflamatória sistêmica e espessamento de artérias subepicárdicas em um modelo animal de uremia

A disfunção renal é um fator de risco para doença cardiovascular (DCV). Estudos experimentais controlados que possam analisar o impacto da disfunção renal no sistema cardiovascular, isolando esses fatores relacionados à uremia dos fatores de risco tradicionais, que são altamente prevalentes na população com doença renal crônica (DRC), ainda são escassos. OBJETIVO: Analisar o impacto cardiovascular em ratos com disfunção renal, analisando biomarcadores de risco cardiovascular e a histologia das artérias subepicárdicas desses animais. MÉTODOS: Estudo experimental envolvendo trinta ratos machos Wistar, divididos em dois grupos. Um grupo foi submetido à ablação renal e o outro grupo SHAM (grupo controle) à manipulação do pedículo renal. Ambos os grupos foram acompanhados por oito semanas, período em que foram feitas dosagens de ureia, fósforo e do fator de necrose tumoral alfa (TNF-α). Lâminas obtidas de cortes do miocárdio foram confeccionadas para análise das características das arteríolas subepicárdicas. RESULTADOS: O grupo DRC apresentou níveis elevados de uréia e fósforo em relação ao grupo SHAM. Já os níveis de TNF-α, em todas as análises, foram indetectáveis nos animais do grupo SHAM, em contraste com o grupo DRC, onde se observaram elevados níveis de TNF-α (p < 0,05). A espessura da camada média dos vasos subepicárdicos do grupo DRC foi significativamente maior do que em relação ao grupo SHAM (p = 0,011). CONCLUSÃO: A indução de disfunção renal determinou alterações em biomarcadores de risco cardiovascular e um aumento na espessura dos vasos subepicárdicos estudados em comparação aos animais com função renal normal.

Accelerating food safety and quality control: The use of antibody engineering techniques in the detec-tion of low-molecular weight food contaminants

The increased awareness and evolved consumer habits have set more demanding standards for the quality and safety control of food products. The production of foodstuffs which fulfill these standards can be hampered by different low-molecular weight contaminants. Such compounds can consist of, for example residues of antibiotics in animal use or mycotoxins. The extremely small size of the compounds has hindered the development of analytical methods suitable for routine use, and the methods currently in use require expensive instrumentation and qualified personnel to operate them. There is a need for new, cost-efficient and simple assay concepts which can be used for field testing and are capable of processing large sample quantities rapidly. Immunoassays have been considered as the golden standard for such rapid on-site screening methods. The introduction of directed antibody engineering and in vitro display technologies has facilitated the development of novel antibody based methods for the detection of low-molecular weight food contaminants. The primary aim of this study was to generate and engineer antibodies against low-molecular weight compounds found in various foodstuffs. The three antigen groups selected as targets of antibody development cause food safety and quality defects in wide range of products: 1) fluoroquinolones: a family of synthetic broad-spectrum antibacterial drugs used to treat wide range of human and animal infections, 2) deoxynivalenol: type B trichothecene mycotoxin, a widely recognized problem for crops and animal feeds globally, and 3) skatole, or 3-methyindole is one of the two compounds responsible for boar taint, found in the meat of monogastric animals. This study describes the generation and engineering of antibodies with versatile binding properties against low-molecular weight food contaminants, and the consecutive development of immunoassays for the detection of the respective compounds.

The use of relaxation therapy as an adjunct to traditional psoriasis therapy /

This research investigated the impact of stress management and relaxation techniques on psoriasis. It had a dual purpose to see if stress management and relaxation techniques, as an adjunct to traditional medical treatment, would improve the skin condition of psoriasis. In addition it attempted to provide psoriasis patients with a sense of control over their illness by educating them about the connection between mind and body through learning stress management and relaxation techniques. The former purpose was addressed quantitatively, while the latter was addressed qualitatively. Using an experimental design, the quantitative study tested the efficacy of stress management and relaxation techniques on 38 dermatological patients from St. John's, Newfoundland. The study which lasted ten weeks, suggested a weak relationship between psoriasis and stress. These relationships were not statistically significant. The qualitative data were gathered through unstructured interviews and descriptive/interpretative analysis was used to evaluate them. Patients in the experimental group believed in the mind body connection as it related to their illness and stress. The findings also showed that the patients believed that the stress reduction and relaxation techniques improved their quality of life, their level of psoriasis, and their ability to live with the condition. Based on the contradictory nature of the findings, further research is needed. It is posited that replication of this study would be vastly improved by increasing the sample size to increase the possibility of significant findings. As wel~ increasing the length of time for the experiment would control for the possibility of a lag effect. Finally, the study looked at linear relationships between stress and psoriasis. Further study should ascertain whether the relationship might be nonlinear

Attitudes of second year diploma nurses in three community colleges toward the use of computers in the nursing role /

The effectiveness of various kinds of computer programs is of concern to nurse-educators. Using a 3x3 experimental design, ninety second year diploma student nurses were randomly selected from a total population at three community colleges in Ontario. Data were collected via a 20-item valid and reliable Likert-type questionnaire developed by the nursing profession to measure perceptions of nurses about computers in the nursing role. The groups were pretested and posttested at the beginning and end of one semester. Subjects attending College A group received a computer literacy course which comprised word processing with technology awareness. College B students were exposed to computer-aided instruction primarily in nursing simulations intermittently throughout the semester. College C subjects maintained their regular curriculum with no computer involvement. The student's t-test (two-tailed) was employed to assess the attitude scores data and a one-way analysis of variance was performed on the attitude scores. Posttest analysis revealed that there was a significant difference (p<.05) between attitude scores on the use of computers in the nursing role between College A and C. No significant differences (p>.05) were seen between College B and A in posttesting. Suggestions for continued computer education of diploma student nurses are provided.

Elaborative interrogation as a learning technique for students with learning disabilities

This study examined whether or not students with learning disabilities could effectively use a question and answer strategy known as elaborative interrogation. This technique involved students answering why they thought facts based on familiar animal stories were true. Thirty students from a provincial demonstration high school (for students with learning disabilities) were assigned to one of two study conditions, (a) elaborative interrogation or (b) reading for understanding. Three students, one from the experimental condition and two from the control did not complete the study. Both conditions required that the students learn 36 facts concerning six familiar animals. Immediately following the study session the students completed a free-recall test, a matched association test and a questionnaire regarding their perceived difficulty of the animal stories. After 30 days a matched association test was completed. The oneway ANOVA, 2 x 2 split plot ANOVA and Tukey's Honestly Significant Test were used to determine significance. There was no significant difference in the two conditions for free recall retention. There were significant differences in the elaborative interrogation condition for the immediate matched association test and for the 30-day matched association test. The probability of the students' responses in the elaborative interrogation were measured to determine the effects of adequate responses on long-term retention. It was found that the adequate responses were more likely to promote retention than inadequate responses. In conclusion, long-term retention of factual information was significantly better in the elaborative interrogation condition in comparison to the reading for understanding control. For future research, the dependent measure, free recall should be given both verbally and in written format. In addition, extra time should be allowed for processing of the new information to occur.

Utilitarianism and Buddist ethics: a comparative approach to the ethics of animal research

This thesis explores the comparison utilitarianism and Buddhist ethics as they can be applied to animal research. It begins by examining some of the general discussions surrounding the use of animals in research. The historical views on the moral status of animals, the debate surrounding their use in animals, as well as the current 3R paradigm and its application in Canadian research are explored. The thesis then moves on to expound the moral system of utilitarianism as put forth by Jeremy Bentham and John Stuart Mill, as well as contemporary additions to the system. It also looks at the basics of Buddhist ethics well distinguishing the Mahayana from the Therevada. Three case studies in animal research are used to explore how both systems can be applied to animal research. It then offers a comparison as to how both ethical systems function within the field of animal research and explores the implications in their application on its practice.

"Re-huminization": the role of human-animal similarity in predicting prejudice towards immigrants and non-human animals

Research implies that there ~ay be an association between attitudes toward margil1alized human outgroups and non-human animals. Very few studies, however, have specifically tested this relation empirically. The general purpose of the present research was to determine if such a relation exists and if perceptions of human-animal similarity avail as a common predictor of both types of attitudes. Ideological orientations associated with prejudiced attitudes (Social Dominance Orientation, Right-Wing Authoritarianism, and Universal Orientation) were also examined as individual differences in predicting perceptions of human-animal similarity. As predicted, people who endorsed prejudiced attitudes toward human outgroups (Study 1) and immigrants in particular (Studies 2 and 3), were more likely to endorse prejudiced attitudes toward non-human animals. In Study 2, perceptions that humans are superior (versus similar) to other animals directly predicted higher levels of prejudice toward non-human animals, whereas the effect of human superiority beliefs on immigrant prejudice was mediated by dehumanization. In other words, greater perceptions of humans as superior (versus similar) to other animals "allowed for" greater dehumanization of immigrants, which in turn resulted in heightened immigrant prejudice. Furthermore, people higher in Social Dominance Orientation or Right-Wing Authoritarianism were particularly likely to perceive humans as superior (versus similar) to other animals, whereas people characterized by a greater Universal Orientation were more likely to perceive humans and non-human animals as similar. Study 3 examined whether inducing perceptions of human-animal similarity through experimental manipulation would lead to more favourable attitudes toward non-human animals and immigrants. Participants were randomly assigned to read one of four 11 editorials designed to highlight either the similarities or differences between humans and other animals (i.e., animals are similar to humans; humans are similar to animals;~~nimals are inferior to humans; humans are superior to animals) or to a neutral control condition. Encouragingly, when animals were described as similar to humans, prejudice towards non-human animals and immigrants was significantly lower, and to some extent this finding was also true for people naturally high in prejudice (i.e., high in Social Dominance Orientation or Right-Wing Authoritarianism). Inducing perceptions that nonhuman animals are similar to humans was particularly effective at reducing the tendency to dehumanize immigrants ("re-humanization"), lowering feelings of personal threat regarding one's animal-nature, and at increasing inclusive intergroup representations and empathy, all of which uniquely accounted for the significant decreases in prejudiced attitudes. Implications for research, theory and prejudice interventions are considered.

Experimental and Computational Studies on Copper and Hydrobenzoin Derivatives in Catalytic Asymmetric Reactions

The use of theory to understand and facilitate catalytic enantioselective organic transformations involving copper and hydrobenzoin derivatives is reported. Section A details the use of theory to predict, facilitate, and understand a copper promoted amino oxygenation reaction reported by Chemler et al. Using Density Functional Theory (DFT), employing the hybrid B3LYP functional and a LanL2DZ/6-31G(d) basis set, the mechanistic details were studied on a N-tosyl-o-allylaniline and a [alpha]-methyl-[gamma]-alkenyl sulfonamide substrate. The results suggest the N-C bond formation proceeds via a cisaminocupration, and not through a radical-type mechanism. Additionally, the origin of diastereoselection observed with [alpha]-methyl-[gamma]-alkenyl sulfonamide arises from avoidance of unfavourable steric interactions between the methyl substituent and the N -protecting group. Section B details the computationally guided, experimental investigation of two hydrobenzoin derivatives as ligands/ catalysts, as well as the attempted synthesis of a third hydrobenzoin derivative. The bis-boronic acid derived from hydrobenzoin was successful as a Lewis acid catalyst in the Bignielli reaction and the Conia ene reaction, but provided only racemic products. The chiral diol derived from hydrobenzoin successfully increased the rate of the addition of diethyl zinc to benzaldehyde in the presence of titanium tetraisopropoxide, however poor enantioinduction was obseverved. Notably, the observed reactivity was successfully predicted by theoretical calculations.