Objetivo: As moléculas de adesão celular (MAC) são essenciais para a manutenção do fenótipo epitelial. Alguns estudos têm relatado associação entre as alterações de sua expressão e a carcinogênese, mas o seu papel no câncer de próstata não é claro. Nosso objetivo foi estudar o perfil de expressão de E-caderina, cateninas e integrinas em espécimes cirúrgicos de câncer de próstata e associar as suas expressões com a evolução do tumor. Avaliamos também o perfil de expressão em metástases ósseas e linfonodais, a fim de compreender a influência destes marcadores na progressão do câncer de próstata. Materiais e Métodos: Foram selecionados 111 pacientes com câncer de próstata localizado tratados com prostatectomia radical pelo mesmo cirurgião. Sessenta pacientes não apresentaram recidiva tumoral após acompanhamento médio de 123 meses. A expressão das MAC foi avaliada por imuno-histoquímica (IH) em microarranjo tecidual (TMA), contendo duas amostras de cada tumor. Empregamos análise semiquantitativa para avaliação da expressão e determinamos a associação entre a expressão de cada MAC com a recorrência do tumor após a cirurgia. Avaliamos também a expressão das MAC por IH em TMA contendo espécimes de 28 metástases ósseas e em outro TMA contendo 19 metástases linfonodais com seus 19 tumores primários correspondentes. Resultados: Nos tumores primários a análise multivariada mostrou que a expressão das integrinas 3 e 3 1 relaciona-se com recidiva da doença. Quando a expressão de 3 foi forte e a expressão de 3 1 foi positiva, as chances de recorrência foram de 3,0 e 2,5 vezes maior. Apenas 19% e 28% dos pacientes estavam livres de recidiva após seguimento médio de 123 meses, quando os tumores apresentavam forte imunoexpressão de 3 ou positiva para 3 1 respectivamente. Outras integrinas apresentaram expressão reduzida, exceto 6 que foi expressa pela maioria dos tumores primário e metástases. A E-Caderina e as cateninas não mostraram associação com o prognóstico no tumor de próstata localizado. No sítio metastático, houve perda global de expressão das MAC. Encontramos ganho de expressão com a progressão do câncer de próstata somente para a integrina 3 que mostrou forte expressão em metade das metástases ósseas e linfonodais. Encontramos forte expressão de e -catenina foi em 94% dos linfonodos e 45% Conclusões: Nossos experimentos demonstram que a expressão das integrinas 3 e 3 1 está independentemente associada à recidiva de câncer de próstata após prostatectomia radical, e que a perda das moléculas de adesão celular pode ser considerada uma característica da progressão desta neoplasia / Purpose: Cell adhesion molecules (CAM) are essential for the maintenance of epithelial phenotype. Some studies have reported correlations between abnormalities in their expression and carcinogenesis, but their role in prostate cancer is unclear. Our aim was to study the expression profile of E-cadherin, catenins and integrins in surgical specimens of prostate cancer and associate their expression with outcome. We also assessed these expressions in bone and lymph node metastases in order to understand their influence in the progression of prostate cancer. Materials and Methods: We selected 111 patients with localized prostate cancer who underwent radical prostatectomy performed by the same surgeon. Sixty patients had no tumor recurrence after a median follow-up of 123 months. The CAM expression was evaluated by immunohistochemistry in a tissue microarray (TMA) containing two samples of each tumor. A semiquantitative analysis was employed and we measured the association between the expression of CAM and tumor recurrence. We also evaluated CAM expression by immunohistochemistry in a TMA containing 28 bone metastases and in other TMA containing 19 lymph node metastases with their corresponding 19 primary tumors. Results: In primary tumors, multivariate analysis showed that expression of 3 and 31 integrins was related to worse outcome. When 3 expression was strong and 31 expression was positive,the odds of recurrence were 3.0 and 2.5 fold higher. Only 19% and 28% of patients were recurrence-free in a mean follow up period of 123 months, when tumors showed strong 3 or positive 31 immuno-expression respectively. Other integrins have shown reduced expression, except 6 , which was expressed in most primary and metastatic cases. E-cadherin and catenins expressions were not associated with primary tumor outcome. At the metastatic setting, there was a global loss of CAM expression. We observed reliable gain of expression with prostate cancer progression only for integrin 3 that showed strong expression in half of bone and lymph node metastases. Interestingly, strong expression of and -catenin was observed in 94% of lymph node and 45% of bone metastases. Conclusions: We have demonstrated that the expression of integrins 3 and 31 was independently associated with recurrence after radical prostatectomy. In addition, we have shown that the loss of cell adhesion molecules can be considered a characteristic of prostate cancer progression

Introdução: In vivo ou in vitro a exposição de fibroblastos a alta concentração de glicose promove um aumento do estresse oxidativo e consequentemente prejudica a migração celular, assim como a maturação da adesão. Além disso, a elevada concentração de glicose reduz a expressão de diferentes integrinas na superfície celular devido alterações na síntese do receptor e sua reciclagem. Objetivo: Avaliar os efeitos da elevada concentração de glicose no tráfego de vesículas contendo EEA1 (endossomos primários), Rab4 (via rápida da reciclagem), Rab11 (via lenta de reciclagem) e Rab7 (endossomos de degradação) em fibroblastos NIH3T3. Métodos: células foram cultivadas em meio contendo baixa concentração de glicose (LG, 5 mM) ou em alta concentração (HG 25 mM) durante 21 dias antes de realizar os experimentos. EEA1, Rab4, Rab11 e Rab7 expressão e distribuição foram avaliados por western blotting e imunofluorescência, respectivamente. Resultados: Células expostas à alta concentração não apresentaram diferenças na expressão e distribuição das proteínas EEA1 e Rab7, enquanto a expressão de Rab11 foi reduzida em 30%. Conclusão: a alta concentração de glicose altera a via lenta da reciclagem contendo Rab11, afetando potencialmente a reciclagem de integrinas e outros receptores e a sua expressão na superfície celular. / Background: In vivo or in vitro exposure of fibroblasts to high glucose concentrations (HG) promotes oxidative stress and consequently impairs cell migration, also inhibiting adhesion maturation. Additionally, HG reduces the expression of different integrins on the cell surface, potentially due to altered receptor synthesis and recycling. Aim: to evaluate the effects of HG on the trafficking vesicles containing EEA1 (early endosomes), Rab4 (fast recycling pathway) and Rab7 (endocytic degradation pathway) on NIH3T3 fibroblasts. Methods: cells were cultured under low glucose (LG, 5 mM) or HG (25 mM) concentrations during 21 days before the assays. EEA1, Rab4 and Rab7 expression and distribution were evaluated by western blotting and immunofluorescence, respectively. Results: HG did not affect proteins EEA1 and Rab7 expression and distribution, whereas Rab11 expression was reduced by 30%. The number of vesicles containing Rab11 was also significantly reduced in HG cells. Conclusion: high glucose alters the slow recycling endocytic pathway via Rab11, potentially affecting integrins and other receptors synthesis and expression on the cell surface.

Orientador: Sergio Luis Felisbino / Resumo: O câncer de próstata (PCa) é o segundo tipo de câncer mais frequente e tem a segunda maior taxa de morbidade e mortalidade entre os homens. A cultura de células prostáticas LNCaP e PC3 tem sido utilizada para investigar possíveis alvos e vias de sinalização celular importantes para o crescimento e progressão do CaP. Trabalhos anteriores do nosso laboratório demonstraram que a presença de fibronectina no meio de cultura altera significativamente o padrão de expressão gênica dessas células. Também, bacteriófagos recentemente usados como agentes terapêuticos no tratamento de vários tipos de câncer, diretamente ou portadores de moléculas antitumorais, incluindo câncer de próstata, foram relatados. Estudos anteriores mostraram que bacteriófagos podem interagir com proteínas de membrana de células de mamíferos, incluindo integrinas que reconhecem sequências de RGD em proteínas virais, bem como fibronectina e outras moléculas da matriz extracelular. O objetivo deste projeto foi caracterizar a interação de três bacteriófagos com as duas linhagens celulares epiteliais prostáticas LNCaP e PC3. Estas duas linhas celulares foram cultivadas em seus meios de cultura, nos quais foram adicionados bacteriófagos com concentrações definidas. As células foram coletadas após 24 horas de tratamento. A expressão gênica de genes relacionados as vias integrinas ITGA5, ITGAV, ITGB1, ITGB3, ITGB5, AKT, PI3K, MAPK1, MAPK3, HSP27, HSP90, receptor de androgênio AR, STAT3, PGC1A foi analisada por qPCR. A a... (Resumo completo, clicar acesso eletrônico abaixo) / Doutor

Melanoma primário da mucosa oral (MPMO) é um tumor raro e agressivo. Estudos recentes demonstraram uma correlação entre o aumento da invasão tumoral e o fenótipo metastático com uma alteração no padrão de expressão das moléculas de adesão. Neste estudo analisamos a expressão de integrinas e imunoglobulinas nos melanomas primários da mucosa oral e relacionamos os resultados com os parâmetros clínicos. As análises imunoistoquímicas dos padrões de expressão destas moléculas foram realizadas em 35 casos de melanomas primários da mucosa oral, e os resultados foram correlacionados com características clínicas e histológicas. Observou-se que a subunidade beta-4 de integrina foi negativa em casos com invasão vascular. A presença de integrina beta-3 e de CD166 (ALCAM) estavam estatisticamente associadas à extensa invasão vascular (p < 0,05). A menor expressão de CD54 (ICAM) foi marginalmente relacionada a casos com necrose extensa, enquanto a maioria dos casos com doença metastática foi negativa para CD66 (CEACAM). Conclusão: padrões alterados de expressão de moléculas de adesão, principalmente integrinas e imunoglobulinas, podem participar da patogênese e do desenvolvimento dos melanomas primários da mucosa oral / Primary oral mucosal melanoma is a rare and an aggressive tumor. Recent studies have demonstrated the correlation among increased tumor invasion, the metastatic phenotype and altered adhesion molecule expression profiles. The present study analyzed the expression of integrins and immunoglobulin-like adhesion molecules in oral mucosal melanomas and correlated results with clinical parameters. Immunohistochemical analyses of their expression patterns were performed on thirty-five cases of primary oral mucosal melanomas. The results were correlated with clinical and histological features of the cohort. The beta-4 subunit of integrin was negative and this was related with vascular invasion. Positivity of integrin beta-3 and CD166 (ALCAM) was statistically associated with extensive vascular invasion (p < 0.05). Lower expression of CD54 (ICAM) was associated with cases with extensive necrosis. Most cases with metastatic disease were negative for CD66 (CEACAM). Conclusion: Altered patterns of adhesion molecule expression, mainly integrins and immunoglobulin-like proteins, may participate in the pathogenesis and outcome of primary oral mucosal melanomas

A conversão da proteína príon celular (PrPc) em sua isoforma anormal PrPsc está associada a uma série de doenças neurodegenerativas, genericamente designadas por doenças priônicas. Embora a literatura tenha enfatizado o estudo do PrPsc e o mecanismo de propagação das doenças de príon, pouco tem sido feito para o entendimento do papel fisiológico do PrPc. Em 1997 nosso grupo descreveu um receptor/ligante para o PrPc utilizando o princípio da hidropaticidade complementar. Neste trabalho isolamos e identificamos este ligante de PrPc como sendo a STI-1 (Stress Inducible Protein-1). In vitro, a STI-1interage com o PrPc de maneira específica, saturável e com alta afinidade (Kd=8x10-8M). Paralelamente, mostramos que o PrPc se liga ao domínio RNIAEIIKDI da laminina (Ln) (Kd=2x10-8M). O bloqueio de PrPc na superfície de neurônios hipocampais de embriões de ratos e camundongos, reduziu a neuritogênese induzida por Ln. Além disso, neurônios provenientes de animais PrP -/- são incapazes de estender neuritos sobre o peptídeo RNIAEIIKDI, sugerindo que o PrPc é o único receptor celular para este domínio da Ln. Estes dados indicam que a interação PrPc-Ln seja relevante nos fenômenos de adesão e diferenciação neuronais. A caracterização das interações PrPc-Ln e PrPc-STI-1 representa contribuições importantes para a elucidação do papel biológico do PrPc. / Conversion of the cellular prion protein (PrPc) to its abnormal isoform PrPsc is associated with some neurodegenerative and fatal diseases called prion diseases. Although the literature has been emphasizing the mechanism of PrPsc conversion and illness propagation, little attention has been given to the PrPc physiological role. In 1997, our group described a PrPc receptor/ligand based on the complementary hydropathy theory. Herein, we identify the PrPc receptor/ligand as STI-1, the Stress Inducible Protein-1. In vitro studies showed that STI-1 is a specific, saturable and high affinity ligand for PrPc (Kd=8x10-8M). In parallel, we demonstrated that PrPc interacts with RNIAEIIKDI domain of laminin (Ln) (Kd=2x10-8M). The blockage of PrPc, both from embryonic rats and mice hippocampal neuros, inhibited Ln-induced neurite outgrowth. In addition, neurons from PrPc null mice are unable to extend neurites on RNIAEIIKDI, suggesting that PrPc is the unique cellular receptor for this Ln domain. These data indicate that PrPc-Ln interaction is relevant for neuronal adhesion and differentiation. The characterization of PrPc-Ln and PrPc-STl-1 interactions represents important contributions for the elucidation of the PrPc physiological role.

Human embryonic stem cells are pluripotent cells that have indefinite replicative potential and ability to differentiate into derivatives of three germ layers. HESCs are conventionally derived and grown on mitotically inactivated mouse embryonic fibroblasts and there are some alternative feeder types of human origin that have been used to replenish hESCs while trying to prevent cross-species contamination. The trophic factors that are secreted by the feeders are found to be important for long-term pluripotency but there are also supportive culture systems for hESCs lacking feeder cells which might suggest that not only the interactions with the feeders affect the behaviour of hESCs but also the components of the niche may take part in the decision of self-renewal or differentiation. Extracellular matrix components are known to exert their stimulatory or inhibitory effects by localising cells into a specific microenvironment in natural niches but have been relatively little investigated for hESCs. The aim of this study was to investigate ECM components which might have a role in the maintenance of hESCs. I have first investigated human placental stromal fibroblasts and immortalised human placental stromal fibroblasts for the support hESC pluripotency as an anlternative feeder type to conventional mouse embryonic fibroblasts. Secondly, the matrices derived from hPSFs and ihPSFs were assessed for their ability to support hESC pluripotency. Tandem mass spectrometry was used to identify ECM components released by human feeders in order to characterise the range of extracellular matrix proteins that support the growth of self-renewing hESCs. The majority of the molecules was shared between the cell types irrespective of hPSF cell derived matrix was not being supportive for hESC pluripotency, with some ECM components being unique ihPSFs. Collagen VI, tenascin C and versican were tested for hESC attachment and as substrates for feeder-free culture system in order to develop an optimised feeder-free system. Furthermore, integrin receptor profile of different hESC lines was also determined in order to identify the mechanisms of substrate attachment. Integrin attachment was shown to be vital for hESC engagement to fibronectin and vitronectin in feeder-free systems. The components of the integrin signalling machinery were identified in hESCs and the significance of integrin-mediated signalling in hESC self-renewal was demonstrated by blocking integrin β1 on fibronectin and integrin aVβ5 on vitronectin. Moreover, intracellular signalling mediator c-Src was shown to involve in ECMregulated signalling by affecting the phosphorylation of Focal Adhesion Kinase. Inhibition of Src led to a decrease in the expression of pluripotency-associated markers. Finally, the effects of growth factor supplementation on the maintenance of pluripotency in defined feeder-free conditions were studied by withdrawal of growth factors and blocking FGF Receptors. FGF-2 was shown to be essential for long-term self-renewal while the effects on pluripotency deteriorated in the absence of both FGF-2 and Activin A. Taken together this project highlighted the importance of substrate attachment and growth factors on the regulation of hESC self-renewal.

The integrins play a crucial role in cancer cell proliferation, migration, differentiation, survival and angiogenesis. It has been shown that integrin expression is positively correlated to cancer dissemination, this suggests targeting selected integrins as an anti-metastatic strategy. The aim of this study is to investigate the effect of novel antagonists of α5β1, αvβ3 and αvβ5 integrins on cancer cell migration, a key process in tumour cell dissemination. Immunohistochemistry was used to evaluate the expression of α5, αv, β3 and β5 integrin subunits in prostate cancer tissues. Furthermore the expression of these integrin subunits in tumour and normal human head and neck tissues was compared. The expression profile of these integrin subunits in established human cancer cell lines was subsequently evaluated using immunodetection methods in cells and xenograft tumour samples. The effect of integrin inhibition on cell migration was then assessed using neutralizing antibodies against αvβ3, αvβ5, and α5β1 integrins in the scratch-wound healing assay. This assay was then used to evaluate the potential of novel small molecule integrin antagonists in preventing tumour cell migration. In H & N tissues, αvβ3, αvβ5 and α5β1 integrins are extensively expressed in tumour tissues but weakly expressed in normal tissue from the same patient. Further, prostate cancer tissues expressed variable levels of αvβ3, αvβ5 and α5β1 integrins. αvβ3 and αvβ5 integrins were expressed in variable levels in OSC-19, PC-3, DU145, DLD-1, HT-29, HUVEC, MCF-7, MCF-7ADR and M14 human tumour cell lines and in OSC-19, PC-3, HT-29 and MCF-7 xenografts. α5β1 integrin was expressed in all cell lines and xenografts except in MCF-7 cell line and HT-29 cell line and xenograft. Overall, the expression was elevated in xenografts compared to the corresponding cultured cells. Based on the expression profile and ability of cells to migrate, three cell lines (DLD-1 colon, DU145 prostate and OSC-19 HNSCC) were selected as models to further evaluate the potential of novel small molecule integrin antagonists to inhibit cell migration. The cell lines were characterized by using neutralizing antibodies against αvβ3, αvβ5, and α5β1 integrins to determine which of these three integrins were primarily involved in tumour cell migration. In DLD-1 and DU145, blocking αvβ5 and αvβ3 significantly inhibited migration, whilst the migration of OSC-19 was 50% inhibited by a multi-integrin inhibitor combination. Among the antagonists, ICT9055 and ICT9072 significantly decreased DLD-1 cell migration by 70% and 60% respectively while ICT9023, ICT9024, and ICT9026 significantly decreased DU145 cell migration by 60%, 60% and 50% respectively. The findings suggest that single integrin inhibition is not sufficient to prevent cell migration whereas dual or multiple inhibition is more effective. Two novel anti-migratory agents were identified in colon cancer and three in prostate cancer which would warrant further investigation.

Melanoma primário da mucosa oral (MPMO) é um tumor raro e agressivo. Estudos recentes demonstraram uma correlação entre o aumento da invasão tumoral e o fenótipo metastático com uma alteração no padrão de expressão das moléculas de adesão. Neste estudo analisamos a expressão de integrinas e imunoglobulinas nos melanomas primários da mucosa oral e relacionamos os resultados com os parâmetros clínicos. As análises imunoistoquímicas dos padrões de expressão destas moléculas foram realizadas em 35 casos de melanomas primários da mucosa oral, e os resultados foram correlacionados com características clínicas e histológicas. Observou-se que a subunidade beta-4 de integrina foi negativa em casos com invasão vascular. A presença de integrina beta-3 e de CD166 (ALCAM) estavam estatisticamente associadas à extensa invasão vascular (p < 0,05). A menor expressão de CD54 (ICAM) foi marginalmente relacionada a casos com necrose extensa, enquanto a maioria dos casos com doença metastática foi negativa para CD66 (CEACAM). Conclusão: padrões alterados de expressão de moléculas de adesão, principalmente integrinas e imunoglobulinas, podem participar da patogênese e do desenvolvimento dos melanomas primários da mucosa oral / Primary oral mucosal melanoma is a rare and an aggressive tumor. Recent studies have demonstrated the correlation among increased tumor invasion, the metastatic phenotype and altered adhesion molecule expression profiles. The present study analyzed the expression of integrins and immunoglobulin-like adhesion molecules in oral mucosal melanomas and correlated results with clinical parameters. Immunohistochemical analyses of their expression patterns were performed on thirty-five cases of primary oral mucosal melanomas. The results were correlated with clinical and histological features of the cohort. The beta-4 subunit of integrin was negative and this was related with vascular invasion. Positivity of integrin beta-3 and CD166 (ALCAM) was statistically associated with extensive vascular invasion (p < 0.05). Lower expression of CD54 (ICAM) was associated with cases with extensive necrosis. Most cases with metastatic disease were negative for CD66 (CEACAM). Conclusion: Altered patterns of adhesion molecule expression, mainly integrins and immunoglobulin-like proteins, may participate in the pathogenesis and outcome of primary oral mucosal melanomas

<p>Integrins are receptors presented on most cells. By binding ligand they can generate signalling pathways inside the cell. Those pathways are a linkage to proteins in the cytosol. It is known that tumor cells can survive and proliferate in the absence of a solid support while normal cells need to be bound to ligand. To understand why tumour cells act that way, we first have to know how ligand-binding to integrins affect the cell. This research field includes studies on activation of proteins by integrins and the following protein-protein interactions.</p><p>The part of the research that I did, focused on the activation of PI3K by integrins and the question whether Ras is included in that pathway. I also studied the conformation changes of the integrins and tried to identify factors which regulate these changes.</p><p>Known is that Ras can activate PI3K. But we wanted to know if this is a step in the activation of PI3K by integrins. So if this would be a fact then Ras must be activated by integrins.</p><p>To see if integrins could activate Ras I did a pull down assay. GTP loaded Ras was isolated through its affinity for Raf. Only when Ras is in its activated state then it is GTP loaded, otherwise it is GDP loaded. In the experiment we also compared the β1A and the β1B splice variants. As result we could see that both splice variants probably can activate Ras. By blotting with anti-PI3K antibody we looked if PI3K had bound to Ras but no clear result could be obtained.</p><p>Integrins presented on blood cells are mostly in the inactive state while adherent cells have integrins which are mostly in the active state. PI3K has been shown, for blood cells, to be involved in the conformation regulation of integrins. Possibly, there is a positive circle that for blood cells just has to be switched on. It could be that the integrins in adherent cells are active because the cells are adhesive. By being adhesive, PI3K is activated. PI3K may then activate the integrins, through which the integrins stay in the active state. This circle could be broken at two points: we could inhibit PI3K or we could make the cells un-adhesive. I analysed this in cell attachment assay and by binding of conformation-specific integrin antibodies in FACScan. From the results we could not find any evidence that the whole idea around the positive circle is correct. Surprisingly we saw that the integrin value at the surface decrease if you add PI3K inhibitor. This could be due to distribute recirculation of integrins from the cytoplasm to the cell surface.</p><p>β1- and β3-integrins are both widely spread, but no functional difference could be shown already. Previous results suggest that there is a difference between migrations of those two types. To ensure this suggestion I did a wound assay. Hereby I compared the migration of different cell types, with different integrins on their surface and on different ligands.</p>

Les organismes vivants sont en intime relation avec leur environnement et sont constamment influencés par de nombreux facteurs chimiques et physiques. Parmi les facteurs physiques présents dans notre environnement, les forces mécaniques, y compris la gravité, les radiations, dont les ultraviolets, et les champs électromagnétiques constituent les trois pôles principaux de nos travaux de doctorat. Des outils biologiques, cellulaires et moléculaires ont été développés afin dévaluer le rôle des RhoGTPases dans les altérations morphologiques, prolifératives et phénotypiques induites par la perte du vecteur gravité au cours de vols spatiaux. Au cours du vol de la capsule spatiale inhabitée FOTON-M3, nous avons pu mettre en évidence que la suppression de Rac1 par ARN interférentiel permettait de contrecarrer les effets délétères de la microgravité sur larchitecture du cytosquelette suggérant que cette molécule de signalisation participe à la réception et à la réactivité à la gravité. RhoA et Cdc42 ne semblent pas impliqués. Nous avons également développé un modèle expérimental dinduction de flux calcique par des peptides mimétiques de la matrice extracellulaire activant les intégrines destiné à être expérimenté au cours de vols paraboliques. Au cours de nos travaux visant à évaluer les effets biologiques des champs électromagnétiques, nous avons observé que les EMF de très basse fréquence (450µT-50Hz) naffectent ni les signaux calciques induits par des concentrations élevées de sérum ou des peptides mimétiques de la matrice extracellulaire, ni lexpression des gènes régulés par les UV-B. Ils sont cependant capables de soutenir les oscillations calciques induites par une concentration sub-optimale de sérum, sans toutefois réguler de manière évidente les voies de signalisation contrôlant la prolifération. Lirradiation par les UV-B dun grand nombre de cellules, primaires, immortalisées et tumorales induit, par épissage alternatif du préARNm du VEGF-A, lexpression dun nouveau variant, le VEGF111. Celui-ci est constitué de la combinaison des exons 1-4 et de lexon 8. Cette nouvelle forme de VEGF-A contient donc les sites de fixation aux VEGF-R1 et R-2 et est pro-angiogène in vitro sur les cellules endothéliales et les cellules souche embryonnaires et in vivo chez la souris. Labsence des exons 6 et 7 codant pour la liaison aux protéines de la matrice extracellulaire lui confère une diffusibilité tissulaire. Une de ses caractéristiques remarquable est sa résistance à la dégradation en raison de labsence du site de clivage par la plasmine et les MMPs. Ce nouveau variant est également induit par diverses substances génotoxiques dont les agents chimiothérapeutiques. Les mécanismes régulant lexpression du VEGF111 dépendent des voies de signalisation ATM/ATR, p53 et MAPKinases. La double personnalité de ce nouveau facteur pro-angiogène, néfaste par son induction potentielle au cours de traitements anti-cancéreux mais bénéfique par son utilisation dans le traitement de pathologies ischémiques particulièrement pertinente en cas dactivités protéolytiques élevées, ouvre un champ considérable dinvestigations. ----------------------------------------------------------------------------------------------------- Living organisms interact with their environment and are constantly influenced by various chemical and physical factors. Among the physical factors present in our environment, mechanical forces, including gravity, radiations, among which ultraviolet radiations, and electromagnetic fields constitute the three main poles of our research. Biological, cellular and molecular tools have been developed with the aim to evaluate the role of RhoGTPases in the morphological, proliferative and phenotypic alterations induced by the loss of gravitational field experienced during space flight. During the flight of the unmanned FOTON-M3 capsule, we have demonstrated that the suppression of Rac1 by small interference RNA was able to counteract the deleterious effects of microgravity on the cytoskeleton architecture. This suggests that this signaling molecule participates to the reception and reaction to gravity. RhoA and Cdc42 do not seem to be implicated. We have also developed an experimental model of induction of intracellular calcium ions fluxes by mimetic peptides of the extracellular matrix activating integrins to be used in parabolic flights. During our investigations aimed at evaluating the biological effects of electromagnetic fields, we observed that EMF of very low-frequency (450µT-50Hz) do not affect neither the calcium signals induced by high concentrations of serum or extracellular matrix mimetic peptides, nor the expression of genes regulated by UV-B. They are however able to sustain calcium oscillations induced by a sub-optimal concentration of serum but without disturbing the cellular proliferation rate. Irradiation by UV-B of a large number of cells, primary, immortalized and tumoral, induces, by alternative splicing of the VEGF-A pre-mRNA, the expression of a new variant, the VEGF111. This isoform is made of a combination of exons 1-4 and exon 8. This new VEGF-A variant contains therefore the binding sites to VEGF-R1 and R-2 and proved to be proangiogenic in vitro for endothelial and ES cells and in vivo in mice. The absence of exons 6 and 7 coding for the heparin binding sites confers it with tissue diffusibility. One of its striking characteristics is its resistance to degradation due to the absence of the cleavage site by plasmin and MMPs. This new variant is also induced by a series of genotoxic agents, including chemotherapeutic drugs. The mechanisms controlling the VEGF111 expression depend on the ATM/ATR, p53 and MAPKinases signaling pathways. The dual faces of VEGF111, detrimental by its potential induction during anti-cancer therapy but beneficial by its use for managing ischemic pathologies, mostly relevant when associated with high proteolytic activities, opens a considerable field of investigations.