Mécanismes d'action des immunoglobulines intraveineuses dans un modèle murin de la maladie d'Alzheimer

Affectant près de 36 millions de personnes dans le monde, la maladie d’Alzheimer (MA) est une préoccupation grandissante de santé publique. La présence en post-mortem de plaques amyloïdes et d’enchevêtrements neurofibrillaires constitue un trait caractéristique de la pathologie. Malgré les récents progrès en recherche, il n'existe toujours pas de traitement efficace pour cette maladie. Au cours des dernières années, les immunoglobulines intraveineuses (IgIV) ont démontré un certain potentiel thérapeutique pour la MA. Le présent travail vise à déterminer les mécanismes d’action par lesquels les IgIV exercent leurs effets bénéfiques dans un modèle murin de la MA. Globalement, mes travaux de recherche ont permis de constater l’absence d’effet des IgIV sur les plaques amyloïdes et les enchevêtrements neurofibrillaires et ont plutôt identifié l’inhibition de l’activation de la microglie comme mécanisme possible des IgIV, permettant ainsi une baisse de l’inflammation au cerveau et une augmentation de la neurogénèse.Affecting nearly 36 million people worldwide, Alzheimer's disease (AD) is a growing public health concern. AD is characterized by a progressive and irreversible slow deterioration of cognitive functions. The presence of postmortem amyloid plaques and neurofibrillary tangles is a hallmark of the disease. Despite recent advances in research, there is still no effective treatment for this disease. In recent years, intravenous immunoglobulin (IVIg) has shown some therapeutic potential for AD. The present work aims to determine the mechanisms of action by which IVIg exerts its beneficial effects in a mouse model of AD. Overall, my work has identified the inhibition of microglia activation as a possible mechanism of IVIg, allowing a decrease in brain inflammation and an increase in neurogenesis

Interactions entre les cellules tumorales et stromales dans le microenvironnement du cancer de la vessie

Les fibroblastes associés au cancer (CAF) constituent le type cellulaire le plus abondant du microenvironnement tumoral. In vivo, les tumeurs les plus agressives corrèlent avec un enrichissement en CAF et une matrice extracellulaire plus dense. En effet, les interactions dynamiques et réciproques entre les cellules cancéreuses et les CAF favoriseraient la progression tumorale. Cependant, les molécules impliquées dans ces interactions ainsi que leurs effets sur le devenir de la tumeur et sur le remodelage du microenvironnement sont mal connus. Or, mieux définir et comprendre les mécanismes moléculaires impliqués dans cette interaction est crucial afin de pouvoir développer de nouvelles cibles thérapeutiques. Ainsi, nous avons étudié les interactions entre les cellules cancéreuses et les cellules stromales dans le microenvironnement du cancer de la vessie. Les exosomes sont une classe de vésicules extracellulaires d’origine endocytique de 30 à 200 nm de diamètre. Ils sont sécrétés par tous les types cellulaires et constituent, entre autres, un moyen de communication intercellulaire en transportant protéines, lipides et ARN d’une cellule à l’autre. Les cellules cancéreuses sécrètent une grande quantité d’exosomes et ces derniers joueraient un rôle dans la modulation du microenvironnement tumoral, notamment en activant les fibroblastes sains en CAF. Les travaux présentés dans cette thèse ont permis de démontrer que les exosomes sécrétés par les cellules cancéreuses sont internalisés par les fibroblastes vésicaux sains et qu’ils favorisent la prolifération de ces derniers. De plus, les exosomes dérivés de cellules cancéreuses activent les fibroblastes sains en CAF grâce au TGFβ qu’ils contiennent. La neutralisation du TGFβ par des anticorps spécifiques confirme ces résultats. Une fois activés, les CAF augmentent la prolifération, la migration et l’invasion des cellules cancéreuses via une sécrétion soutenue de la molécule IL-6. D’ailleurs, le blocage de la voie de signalisation de l’IL-6 renverse les effets observés sur les cellules cancéreuses. Nous avons également démontré que les CAF diminuent la sensibilité des cellules cancéreuses à la mitomycine C. Enfin, les CAF remodèlent la matrice extracellulaire du microenvironnement tumoral notamment par une sécrétion accrue des protéines oncofétales ténascine C et EDA-fibronectine, ainsi qu’une activité LOX-1 et MMP augmentée. Par ailleurs, la matrice extracellulaire générée par les CAF favorise la transition épithéliomésenchymateuse des cellules urothéliales saines en inhibant le marqueur épithelial Ecadhérine au profit du marqueur mésenchymateux N-cadhérine. Ainsi, une communication étroite et complexe entre les cellules cancéreuses et les CAF favorise la progression tumorale. En secrétant des facteurs solubles à activité protumorale et des protéines de la matrice extracellulaire, les CAF favorisent la prolifération, l’invasion et la chimiorésistance des cellules cancéreuses. Globalement, nos travaux soutiennent l'idée que l’inhibition de la transdifférenciation des fibroblastes sains en CAF est une cible thérapeutique de choix dans le développement de nouveaux anticancéreux.Cancer-associated fibroblasts (CAFs) are the most abundant cell type of the tumor microenvironment. In vivo, aggressive tumors correlate with an enrichment of CAFs and a denser extracellular matrix. Indeed, the dynamic and reciprocal interactions between tumor cells and CAFs promote tumor progression. However, the molecules involved in these interactions, as well as their effects on the fate of the tumor and the remodeling of the microenvironment are poorly known. However, better define and understand the molecular mechanisms of this interaction is crucial to develop new treatments. Thus, we studied interactions between tumor cells and stromal cells in the microenvironment of bladder cancer. Exosomes are a class of extracellular vesicles with of endocytic origin measuring 30 to 200 nm in diameter. They are secreted by all types of cells and constitute, among others, a means of intercellular communication by transporting proteins, lipids and RNA from one cell to another. Cancer cells secrete a large amount of exosomes and these exert a role in the modulation of the tumor microenvironment, notably by activating healthy fibroblasts in CAFs. The work presented in this thesis has demonstrated that the exosomes secreted by cancer cells are internalized by vesical fibroblasts and promote their proliferation. In addition, exosomes derived from cancer cells activate healthy fibroblasts in CAFs using the TGFβ that they transport. The neutralization of TGFβ by specific antibodies confirms these results. Once activated, CAFs increase the proliferation, migration and invasion of cancer cells via a sustained secretion of the IL-6 molecule. Moreover, the blocking the IL-6 signaling pathway reverses the effects observed in cancer cells. We have also demonstrated that CAFs decrease the sensitivity of cancer cells to mitomycin C. Finally, CAFs remodel the extracellular matrix of the tumor microenvironment notably by an increased secretion of tenascin C and EDA-fibronectin oncofetal proteins, as well as a LOX-1 and MMP increased activity. In addition, the extracellular matrix generated by CAFs promotes the epithelio-mesenchymal transition of healthy urothelial cells by inhibiting the epithelial marker E-cadherin in favor of the mesenchymal marker N-cadherin. Thus, a close and complex communication between the cancer cells and the CAFs increases tumor progression. By secreting soluble factors with a pro-tumor activity and extracellular matrix proteins, CAFs promote the proliferation, invasion and chemoresistance of cancer cells. Overall, our work supports the idea that the inhibition of the transdifferentiation of healthy fibroblasts into CAFs is a therapeutic target of choice in the development of novel anticancer drugs

Cancer-associated fibroblasts induce epithelial–mesenchymal transition of bladder cancer cells through paracrine IL-6 signalling

Abstract Background Cancer-associated fibroblasts (CAFs), activated by tumour cells, are the predominant type of stromal cells in cancer tissue and play an important role in interacting with neoplastic cells to promote cancer progression. Epithelial-mesenchymal transition (EMT) is a key feature of metastatic cells. However, the mechanism by which CAFs induce EMT program in bladder cancer cells remains unclear. Methods To investigate the role of CAFs in bladder cancer progression, healthy primary bladder fibroblasts (HFs) were induced into CAFs (iCAFs) by bladder cancer-derived exosomes. Effect of conditioned medium from iCAFs (CM iCAF) on EMT markers expression of non-invasive RT4 bladder cancer cell line was determined by qPCR and Western blot. IL6 expression in iCAFs was evaluated by ELISA and Western blot. RT4 cell proliferation, migration and invasion were assessed in CM iCAF +/− anti-IL6 neutralizing antibody using cyQUANT assay, scratch test and transwell chamber respectively. We investigated IL6 expression relevance for bladder cancer progression by querying gene expression datasets of human bladder cancer specimens from TCGA and GEO genomic data platforms. Results Cancer exosome-treated HFs showed CAFs characteristics with high expression levels of αSMA and FAP. We showed that the CM iCAF induces the upregulation of mesenchymal markers, such as N-cadherin and vimentin, while repressing epithelial markers E-cadherin and p-ß-catenin expression in non-invasive RT4 cells. Moreover, EMT transcription factors SNAIL1, TWIST1 and ZEB1 were upregulated in CM iCAF-cultured RT4 cells compared to control. We also showed that the IL-6 cytokine was highly expressed by CAFs, and its receptor IL-6R was found on RT4 bladder cancer cells. The culture of RT4 bladder cancer cells with CM iCAF resulted in markedly promoted cell growth, migration and invasion. Importantly, inhibition of CAFs-secreted IL-6 by neutralizing antibody significantly reversed the IL-6-induced EMT phenotype, suggesting that this cytokine is necessary for CAF-induced EMT in the progression of human bladder cancer. Finally, we observed that IL6 expression is up-regulated in aggressive bladder cancer and correlate with CAF marker ACTA2. Conclusions We conclude that CAFs promote aggressive phenotypes of non-invasive bladder cancer cells through an EMT induced by the secretion of IL-6

Cancer-Associated Fibroblasts in a 3D Engineered Tissue Model Induce Tumor-like Matrix Stiffening and EMT Transition

A tumor microenvironment is characterized by its altered mechanical properties. However, most models remain unable to faithfully recreate the mechanical properties of a tumor. Engineered models based on the self-assembly method have the potential to better recapitulate the stroma architecture and composition. Here, we used the self-assembly method based on a bladder tissue model to engineer a tumor-like environment. The tissue-engineered tumor models were reconstituted from stroma-derived healthy primary fibroblasts (HFs) induced into cancer-associated fibroblast cells (iCAFs) along with an urothelium overlay. The iCAFs-derived extracellular matrix (ECM) composition was found to be stiffer, with increased ECM deposition and remodeling. The urothelial cells overlaid on the iCAFs-derived ECM were more contractile, as measured by quantitative polarization microscopy, and displayed increased YAP nuclear translocation. We further showed that the proliferation and expression of epithelial-to-mesenchymal transition (EMT) marker in the urothelial cells correlate with the increased stiffness of the iCAFs-derived ECM. Our data showed an increased expression of EMT markers within the urothelium on the iCAFs-derived ECM. Together, our results demonstrate that our tissue-engineered tumor model can achieve stiffness levels comparable to that of a bladder tumor, while triggering a tumor-like response from the urothelium