Argument: Hypertension is a main contributing risk factor to many cardiovascular diseases and may be the cause or the result of cardiovascular dysfunction. Black Africans, especially, suffer from hypertension because of lifestyle changes that occur during westernisation, which may lead to sympatho-adrenal hyperactivity. Vascular endothelial growth factor-A (VEGF-A) and angiopoietin-2 (Ang-2) are regulators of angiogenesis and are significantly up regulated during states of vascular dysfunction. Levels of angiogenic factors are unknown for African people and may not be the same as levels thus far reported for Caucasians. Aims: The aim of this study is firstly, to determine whether differences exist regarding the levels of VEGF-A and Ang-2 in urbanised compared to rural black Africans and secondly, to determine whether increased levels of VEGF-A and Ang-2 factors are related to hypertension in black Africans. Methodology: This is a sub study that is based upon the Prospective Urban and Rural Epidemiological (PURE) study. Apparently healthy, fasting African men and women (N=272, aged 35 to 50 years) from the North-West province of South Africa were selected by a medical doctor to participate in this study. Groups were stratified according to gender and urbanisation status based upon information derived from sociodemographic questionnaires. Cardiovascular parameters (Omron HEM-757), pulse wave velocity (PWV) (Compiler SP), plasma angiogenic factor levels (ELISA) and anthropometric measures were determined. An independent t-test and Pearson Chi-square test were used to compare urban and rural data, followed by an analysis of covariance (ANCOVA) while correcting for confounders (age, body mass index, physical activity and tobacco usage). ANCOVAs (corrected for confounders) were applied where hypertensive and normotensive groups were compared within the whole group and urbanised groups. Correlations, correcting for confounders, between cardiovascular variables and angiogenic factors were determined within the whole group and urbanised groups. Results and conclusion: Plasma VEGF-A values for all black Africans were very low while the ANG-2 levels were elevated compared to control values for Caucasians (normotensive and hypertensive) in literature. Urbanised men were more overweight and indicated a higher incidence of hypertension (42.47%) and elevated VEGF-A levels, but lower Ang-2 levels compared to rural men. Urbanised women were generally overweight, physically less active and smoked less, but indicated higher diastolic blood pressure (BP), VEGF-A levels and lower PWV compared with their rural counterparts. Ang-2 levels indicate a negative relationship to diastolic BP data in rural women. No relationships between hypertensive individuals and high angiogenic factor levels were uncovered. Conclusive evidence suggested that angiogenic factor levels were affected more by urbanisation than by the state of hypertension. If low levels of VEGF-2 occur, ANG-2 stimulation and properties may be altered, thereby switching ANG-2 from an anti-angiogenic to a pro-angiogenic molecule, inferring blood vessel destabilisation and vascular dysfunction, such as is observed in hypertensive urbanised men. Higher ANG-2 levels may result in Tie-2 receptor down regulation, hence causing VEGF-A levels to be lower. Further study is needed to ascertain this mechanism since Tie-2 receptor activity was not determined in this study. / Thesis (M.Sc. (Physiology))--North-West University, Potchefstroom Campus, 2009.

Inflammatory bone diseases like rheumatoid arthritis and periodontal disease lead to increased bone loss in the inflamed areas. The multinucleated bone resorbing cells, the osteoclasts, present in these diseases are larger than normal, and these larger osteoclasts (10+ nuclei) resorb more bone and more often than smaller osteoclasts (2-5 nuclei). Thus, the focus of this thesis was to determine if there are differences in mechanisms of osteoclast activation between large and small osteoclasts. Experiments using authentic rabbit osteoclasts and RAW 264.7-derived osteoclasts revealed differences in the expression of a number of activating factors; with large osteoclasts expressing higher levels of activating receptors (RANK, IL-1RI, TNFR1 and integrins αv and β3), as well as enzymes involved in cellular resorption, while small osteoclasts expressed higher levels of an alleged fusion receptor and the inhibitory receptor, IL-1RII. Further studies revealed that large osteoclasts more readily responded to stimulation by IL-1 compared to small osteoclasts and at lower concentrations suggesting this is a result of their higher expression of activating receptors. Differences in responses to the IL-1 isoforms, IL-1α and IL-1β, were also seen in large osteoclasts: IL-1α generated more large osteoclasts over the course of differentiation, while IL-1β induced changes in cell morphology and in the induction of integrin β3 phosphorylation. These observations suggested that differences in osteoclast responses are induced by IL-1α and IL-1β and it led to the hypothesis that there are differences in signaling between large and small osteoclasts. To elucidate differences in signaling mechanisms a signaling pathway microarray was used which revealed higher expression of Vegfa in large compared to small osteoclasts. Osteoclast differentiation with RANKL increased Vegfa gene expression in a time-dependent manner and VEGF-A secretion was elevated in populations enriched for large osteoclasts. Furthermore, mechanistic studies with inhibitors of transcription factors involved in differentiation revealed that RANKL-mediated Vegfa expression in large osteoclasts was regulated by the NF-κB pathway via induction of Hif1α. These results support the hypothesis that signaling differences exist between large and small osteoclasts and implicates VEGF-A in osteoclast hyperactivity in inflammatory conditions.

Endometrial cancer is one of the most significant gynaecological malignancies that affect women from New Zealand and the rest of the world. One of the critical stages in the development of a tumour is the onset of hypoxia. The malignancy responds by having raised levels of Hypoxia Inducible Factor (HIF) that in turn induces increased production of Vascular Endothelial Growth Factor (VEGF). VEGF is a potent angiogenic factor that will mediate vascular supply of nutrients and oxygen to the developing tumour. The aim of this study was to investigate whether two compounds found in extracts of plant materials, Resveratrol (Resveratrol) and Epigallocatechin gallate (EGCG), altered the levels of VEGF in the supernatant of cultured endometrial cancer cells. Resveratrol is a phytoalexin that is found in many foods, such as grapes, nuts and berries, as well as in high concentrations in some red wines. 100 µM of resveratrol was added to cell cultures for 24 hours. VEGF levels in the supernatant were then analysed using ELISA. Resveratrol was found to have significant inhibitory effects in both primary endometrial cancer cell cultures and immortalised endometrial cancer cell cultures. Resveratrol was also shown to reverse the increase in VEGF caused by the hypoxia mimic cobalt chloride (CoCl₂). Epigallocatechin gallate (EGCG) is an antioxidant catechin extracted from green tea. The effect of EGCG was analysed using the same method as for resveratrol. 100 µM of EGCG was also shown to have a significant inhibitory effect on the level of VEGF in the supernatant of cultured endometrial cancer cells, as well as reducing the effect of CoCl₂. These results suggest that selected food compounds, resveratrol and EGCG, can reduce VEGF levels by inhibiting HIF. Further investigation This may have anti-tumour effects in women with endometrial cancer.

Submitted by Diogo Misoguti (diogo.misoguti@gmail.com) on 2016-07-01T14:40:17Z No. of bitstreams: 1 cp113141.pdf: 8930595 bytes, checksum: 9aee50a9eae23a11edd665ba24b81a77 (MD5) / Approved for entry into archive by Diogo Misoguti (diogo.misoguti@gmail.com) on 2016-07-01T14:40:54Z (GMT) No. of bitstreams: 1 cp113141.pdf: 8930595 bytes, checksum: 9aee50a9eae23a11edd665ba24b81a77 (MD5) / Made available in DSpace on 2016-07-01T14:40:54Z (GMT). No. of bitstreams: 1 cp113141.pdf: 8930595 bytes, checksum: 9aee50a9eae23a11edd665ba24b81a77 (MD5) Previous issue date: 2009

Made available in DSpace on 2016-08-29T15:37:23Z (GMT). No. of bitstreams: 1 tese_5526_MARIA APARECIDA DA SILVA.pdf: 3362386 bytes, checksum: 9e40b51a695b6e8a58969240cbd6e47c (MD5) Previous issue date: 2012-02-27 / A hematúria enzoótica bovina (HEB) é causada pela ingestão crônica de Pteridium aquilinum e caracteriza-se pela presença de sangue na urina e desenvolvimento de lesões na bexiga, sendo responsável por grandes perdas econômicas. A intoxicação por esta planta também pode ocorrer em humanos. Objetivou-se avaliar lesões de bexigas de animais com HEB na região sul do Espírito Santo. Para isto, foram avaliadas 350 bexigas de bovinos em matadouro frigorífico e, destas, selecionadas 46 que apresentavam lesões macroscópicas e/ou hematúria. Amostras de cada bexiga foram fixadas em formol a 10% submetidas ao processamento histológico de rotina e classificadas histomorfologicamente. A imunoistoquímica foi realizada com anti-vimentina, anti-citoqueratina, anti-CD31, anti-VEGF e anti-uroplaquina apenas nas 26 bexigas que revelaram neoplasia. Lesões não neoplásicas foram observadas em 100% das amostras e neoplásicas em 56,52%. A presença de neoplasias foi significativa (p<0,05) na porção caudal da bexiga. As neoplasias encontradas foram carcinoma urotelial; carcinoma in situ, adenocarcinoma, hemangioma, mixoma e hemangiossarcoma. Houve maior frequência de displasia, metaplasia de células claras, inflamação e espessamento vascular em bexigas com neoplasia. A expressão de citoqueratina foi significativa (p<0,05) nas neoplasias epiteliais e vimentina nas mesenquimais. A marcação da uroplaquina III diferiu nos diversos tipos neoplásicos e revelou-se típica e atípica enquanto que a do CD31 foi significativa (p<0,05) nas neoplasias mesenquimais vasculares. Houve diferença significativa (p<0,05) na quantidade de vasos extratumorais marcados pelo VEGF entre os mixomas e adenocarcinomas e nos vasos intratumorais marcados por CD31 e VEGF nos diferentes tipos tumorais. Houve correlação positiva entre os vasos extra e intratumorais nos hemangiomas, hemangiossarcomas e mixomas marcados pelo CD31; nos hemangiomas, mixomas e adenocarcinomas marcados pelo VEGF; entre a expressão de vimentina e CD31 e entre citoqueratina e uroplaquina. Conclui-se que bexigas de bovinos com HEB apresentam lesões não neoplásicas e neoplásicas, isoladas ou associadas. Os biomarcadores auxiliam na diferenciação da histogênese das neoplasias epiteliais e mesenquimais vasculares. Uroplaquina demonstrou-se efetiva para a avaliação da integridade urotelial e os marcadores vasculares (CD31 e VEGF) para a integridade endotelial e para o prognóstico.

Orientador: Jose Eduardo Tanus dos Santos / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-14T19:43:36Z (GMT). No. of bitstreams: 1 Muniz_JaquelineJoice_M.pdf: 1631017 bytes, checksum: 07857094129c2a3044916bda7b45c0d5 (MD5) Previous issue date: 2009 / Resumo: O Fator de Crescimento Endotelial Vascular (VEGF, ou também conhecido como VEGF-A), é uma glicoproteína homodimérica de 45KDa que é produzida principalmente em células endoteliais em condições de hipóxia. VEGF leva a proliferação, migração e sobrevivência de células endoteliais; desenvolve um papel importante na regulação da permeabilidade vascular e angiogênese, tanto fisiológica como fisiopatológica, além de outras atividades biológicas, como vasodilatação, vasculogênese e homeostase vascular. Enquanto muitos SNPs (polimorfismos de base única) estão presentes no gene do VEGF, três SNPs clinicamente significantes localizados na região promotora do gene (C-2578A, G-1154A e G-634C) têm sido associados a várias doenças cardiovasculares. Porém associações inconsistentes entre esses polimorfismos e doenças cardiovasculares têm sido encontradas. Essas diferenças podem ser uma conseqüência da diversidade étnica, que leva a uma distribuição desproporcional dos variantes do gene do VEGF entre grupos étnicos. Embora alguns estudos sugiram que haja uma diferença na distribuição destes SNPs, nenhum estudo prévio analisou esta hipótese em populações miscigenadas. Nós analisamos a distribuição destes três SNPs em 175 brancos e 185 negros brasileiros, avaliamos também a distribuição haplotípica e a associação entre as variantes desses SNPs. O alelo C-2578 e o alelo G-1154 foram mais comuns em indivíduos negros do que brancos (71% e 61%, respectivamente), enquanto para o SNP G-634C não houve diferença quanto a frequência alélica e genotípica entre os dois grupos étnicos. O haplótipo contendo os alelos C-2578/G-1154/G-634 foi o mais comum em ambos grupos étnicos e também o mais freqüente em negros comparados a brancos. O haplótipo contendo os alelos C-2578/A-1154/C-634 e o haplótipo contendo os alelos C-2578/A-1154/G-634 foram mais frequentes em indivíduos brancos do que em negros. Estes resultados mostraram diferenças na distribuição de variantes genéticas e haplótipos do gene do VEGF relevantes clinicamente quando brancos e negros são comparados. Estas diferenças podem explicar, no mínimo em parte, os resultados inconsistentes em estudos de associação desses SNPs com doenças cardiovasculares e, as diferenças interétnicas na suscetibilidade a doenças cardiovasculares / Abstract: Vascular Endothelial Growth Factor (VEGF, or also known as VEGF-A) is a homodimeric glycoprotein of 45kDa produced mostly in endothelial cells in hypoxic conditions. VEGF leads to proliferation, migration and survival of endothelial cells, plays an important role in regulating vascular permeability and angiogenesis, both physiological and pathophysiological and other biological characteristics such as vasodilation, vasculogenesis and vascular homeostasis. While many Single Nucleotides Polimporphisms (SNPs) are present in the VEGF gene, three clinically significant SNPs in the promoter region of the gene (C-2578A, G-1154A and G-634C) have been associated with cardiovascular diseases, however inconsistent associations have been found between these polymorphisms and cardiovascular diseases. These differences may be a consequence of ethnic diversity, which leads to a distinct distribution of VEGF gene variants between ethnic groups. Although some studies suggest difference in the distribution of these SNPs, no previous study has examined this hypothesis in admixed populations. We examined the distribution of these three SNPs in 175 white and 185 black Brazilian subjects, we have also evaluated haplotype distribution and the association between variants of these SNPs. The C-2578 and G-1154 alleles were more frequent in black subjects than in white (71% and 61% respectively), while for the G-634C SNP no differences in allele and genotype frequencies were found between the two ethnic groups. The haplotype containing the alleles C-2578/G-1154/ G-634 was the most common in both ethnic groups and was more common in blacks compared to whites. The haplotype containing the alleles C-2578/A-1154/C-634 and the haplotype containing the alleles C-2578/A-1154/G-634 were more frequent in whites compared to blacks. These results showed differences in the distribution of clinically relevant genetic variants and haplotypes of VEGF gene when whites and blacks are compared. These differences may explain, at least in part, the inconsistent results in studies of association of these SNPs with cardiovascular disease, and ethnic disparities in susceptibility to cardiovascular disease / Mestrado / Mestre em Farmacologia

Les cellules osseuses évoluent in vivo sur des matrices extracellulaires principalement formées de collagène de type I, dont le degré de minéralisation varie au cours du remodelage osseux. Le minéral de l'os, de structure apatitique, a été montré comme potentialisant les activités et modifiant la forme des cellules ostéoblastiques. Dans le but de comprendre les effets du micro-environnement matriciel sur les évènements précurseurs à la phase de formation, nous avons émis l'hypothèse que ces modifications morphologiques pouvaient expliquer en elles-mêmes l'augmentation de l'activité descellules ostéoblastiques, par augmentation de leur mécano-sensibilité, et que ce changement de préhension environnementale pouvait moduler la réponse aux stimulations mécaniques les plus fréquemment observées in vivo, à savoir les micro-vibrations. Nous avons montré que sur les matériaux de collagène minéralisé ACC (Apatite Collagen Complex), les pré-ostéoblastes de la lignée MC3T3-E1 synthétisaient une matrice riche en ostéopontine, fibronectine et facteurs angiogéniques, de façon concomitante à une augmentation dépendante de la quantité de minéral de leur adhérence et de leur migration. Nous avons de plus observé une augmentation de la mécano-sensibilité (expression et turn-over augmentés des adhésions focales) des pré-ostéoblastes sur ACC. Finalement, nous avons établi que la réponse aux stimuli vibratoires était positive sur des matériaux non minéralisés (information) et négative sur ACC (stress) par rapports aux supports non stimulés, ce que nous avons interprété comme une hypersensibilité mécanique cellulairelors de la culture sur ACC. L'ensemble de ces données nous a montré que les modifications de mécanique cellulaire de préostéoblastes cultivés sur ACC engendraient une fonctionalisation spécifique ressemblant à celle observée in vivo dans la ligne cémentante, indispensable à la formation osseuse. D'autre part, les modifications de mécano-sensibilité observées sur ACC, en faisant un support mécano-mimétique et nous amenant à la comparaison du comportement cellulaire observé avec les ostéocytes, pourraient en elles-mêmes expliquer le dépôt matriciel spécifique et la réception modifiée aux signaux vibratoires. Dans notre but ultime de création d'un modèle de remodelage osseux in vitro, les paramètres physicochimiques matriciels osseux et l'établissement de cocultures seront à prendre en compte / Bone cells interact in vivo with extracellular matrices mainly formed of type-I collagen, for which the mineral content changes during the bone remodeling cycle. Bone mineral, which is apatitic in nature, was shown to respectively increase and alter the activity and form of osteoblasts. In order to study the micro-environmental effects of the matrix on the preliminary steps of bone formation, it was hypothesized that these morphological alterations could explain the increased activity of the osteoblastic cells by enhancing their mecano-sensibility. This altered mechano-sensibility could in turn modify the osteoblastic cells’ response to the widely perceived micro-vibrations in vivo. It was demonstrated that, on the collagen-mineralized materials ACC (Apatite Collagen Complex), MC3T3-E1 pre-osteoblastic cells formed a matrix rich in osteopontin, fibronectin and angiogenic factors. At the same time, an increase in cell adhesion and migration dependent on the mineral content was seen. We also observed an enhanced mechano-sensibility (increased focal adhesion gene expression and turn-over) when cells were cultured on ACC. Furthermore, it was found that the vibratory stimuli response was up-regulated on non-mineralized materials (information) and downregulated on ACC (stress) vs. non-stimulated substrates. This observation was interpreted as a hypersensitization to environmental cues on ACC. Taken together, these data have demonstrated that pre-osteoblastic cell mechanic alterations on ACC give rise to a specific functionalization mimicking what is observed in vivo in the cement line required for bone-formation. ACC-related mechano-sensibility changes, which render ACC a mechanomimetic substrate and lead us to compare the observed cell behavior with osteocytes, could explain the specific matrix deposition and altered response to vibrations. The final goal of establishing a model for in vitro bone remodeling can only be fulfill by considering physico-chemical parameters of the bonematrix and cocultures

Background: Lung function is a highly heritable trait. So far there is limited knowledge of the genetic factors that influence lung function. Vascular endothelial growth factor A (VEGF-A) is expressed in the lung at high levels and is known to play a role in angiogenesis and lung remodelling, both in utero and throughout life. A candidate gene study was carried out in order to investigate the role of variants within the VEGF-A gene in determining lung function in childhood and adult life.Methods: Using available longitudinal data previously collected for an unselected birth cohort (Manchester Asthma and Allergy Study-MAAS) the relationship between lung function and single-nucleotide polymorphisms (SNPs) in VEGF-A was assessed. Replication studies were performed in cross-sectional studies of adults from Manchester and children with asthma from Croatia, in whom FEV1/FVC ratio was measured using spirometry. The potential functional roles of two consistently associated SNPs were then further investigated. Finally, using the genome-wide data generated in the discovery cohort (MAAS) I assessed why associations between VEGF-A and lung function had not been reported in recent genome-wide association studies of lung function.Results: Two VEGF-A SNPs, rs10434 and rs3025028, were significantly associated with lung function at multiple ages in a discovery population (MAAS). Subjects with a GG genotype for either SNP had significantly diminished lung function compared to subjects with other genotypes. These findings were replicated in two additional populations (631 parents of children participating in MAAS and in 410 Croatian children with physician-diagnosed asthma aged 6-18 years). SNP rs10434 is located in the 3’UTR and based on its location I hypothesised that it may affect mRNA stability. No significant difference in the rate of VEGF-A mRNA degradation was found between GG and the AA homozygotes. SNP rs3025028 is an intronic SNP in a close proximity to the splice site involved in alternative splicing which generates two different isoforms of VEGF-A; I therefore tested the hypothesis that a change of base at this position could affect the splicing mechanism and cause a change in the ratio of the isoforms. Western blot analysis was used to demonstrate a difference in the ratio of the splice variants VEGF-A165b and total VEGF-A165 (relative to a reference sample) between genotype groups. The VEGF-A165b/panVEGF-A165 ratio was significantly higher at birth (cord plasma), in school-age children and in adults amongst CC compared to GG homozygotes at rs3025038 (p<0.03). Finally, the genome-wide data for the discovery cohort showed that the region containing VEGF-A was not well targeted by either genotyped or imputed SNPs in genome-wide arrays. Conclusion: Evidence was provided to demonstrate that variants within the VEGF-A gene are significantly associated with lung function in both children and adults. Furthermore, data was presented to support a functional role for one of the SNPs (rs3025028). I investigated why associations between VEGF-A and lung function had not previously been reported in recent GWAS and concluded that the region containing VEGF-A was poorly covered by all of the currently available arrays.

The extracellular matrix (ECM), as the native cellular substrate, provides necessary mechanical and biological signals to cells. Cells exert forces in the nanonewton range, which when applied over time can strain extracellular matrix fibers until breakage. Cells and tissues inherently interact mechanically with their surrounding matrix, so tissue engineering materials would benefit from the ability to fully exploit mechanical-biochemical interactions to enhance integration with the human body. In this work, I developed an increased understanding of ECM fiber mechanical and mechano-biochemical properties. First, I generated novel composite ECM fibers that can be used to study combinations of ECM proteins in a controlled way. I determined how protein composition impacts mechanical properties of novel single ECM fibers in a hydrated state and showed how mechanical properties can be tuned through composition. Next, I assayed for strain and heparin-sensitive allosteric binding of ligands to fibronectin and fibrin fibers, and determined that the binding of two key growth factors is impacted by strain and heparin. Finally, I investigated the impact of fiber strain, heparin-pretreatment, and growth factor interactions on endothelial cell migration. The novel contributions of this project are the generation of new composite extracellular matrix fiber types with tunable mechanical properties, as well as the identification of extracellular matrix protein mechanosensitive and heparin-sensitive interactions with growth factors and their impact on endothelial cell migration, which could be used to aid in the design of protein-based biomaterials for cardiovascular applications. / 2024-05-23T00:00:00Z

This body of work encompasses two studies: the collection of canine tears via a novel polyester<br />rod and the comparison of VEGF-A concentrations in tears from dogs with normal and<br />vascularized corneas. The first study used polyester rods for tear collection in dogs. Fluid volume and VEGF recovery characteristics, as well as potential binding of VEGF to the rod, were determined. Tears were harvested from normal dogs using rods and glass capillary tubes. Tears were assayed for tear film VEGF using a commercial canine VEGF sandwich ELISA kit. Dilutions of VEGF standard were wicked into the rods or drawn into capillary tubes, eluted, and assayed. Percent volume recovery is adequate for polyester rods as is percent VEGF recovery. VEGF is detectable in normal canine tears.The second study harvested tear samples from eyes of dogs with vascularizing corneal disease, as well as the contralateral unaffected eye of unilaterally diseased dogs, and normal dogs. Vascularization scores were assigned to diseased eyes and tear film VEGF concentration was assayed as above. Mean tear film VEGF concentration of diseased eyes did not differ from control eyes, and was not correlated with disease process, extent of vascularization, or other parameters. Tear film VEGF in unaffected eyes was significantly higher than control and vascularized eyes. Canine tear film VEGF exceeds biologically active concentrations, but does not correlate with state of corneal vascularization. VEGF-related control of corneal vascularization may be mediated by other proangiogenic factors. / Master of Science