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Importance de la S-nitrosation des récepteurs cérébrovasculaires de l’angiotensine II

Abstract : Stroke is the second leading cause of death worldwide, the development of new therapeutics is thus urgent. Two actors play a major role in the regulation of cerebral circulation: nitric oxide (NO) and the renin-angiotensin system (RAS). The first chapter of this manuscript focuses on NO, its role and its signaling pathways. We present the available NO donors as well as those in development. Dinitrosobucillamine, a new NO donor developed in our team, is evaluated in vitro and in vivo. The second part of the introduction focuses on RAS and its preponderant role in blood pressure maintenance and cerebrovascular regulation. We present the angiotensin II (AngII) receptors, AT1 and AT2 responsible for vasoconstriction and vasodilation of cerebral arteries, respectively. Finally, the last part presents the regulation of AngII receptors by NO, in particular through S-nitrosation of the receptors, the covalent bound between NO and cysteine residues. We present the work of Leclerc, showing that exposure of cells overexpressing AT1 to NO causes a decrease in AngII affinity for AT1 (Leclerc et al., 2006). The second chapter is devoted to the experimental studies. The objective of this work is to study the importance of AngII receptor S-nitrosation at the cerebrovascular level. First, we address the current problematic concerning the nonspecificity of anti-AT1 antibodies. We show here that the new monoclonal anti-AT1 antibody, which is supposed to be more specific, does not recognize AT1 in western blot and immunofluorescence, making its use impossible. We then make a pharmacological demonstration of S-nitrosation effects on AngII receptors. We show that exposure to NO donors (S-nitrosoglutathione or sodium nitroprusside) specifically abolishes AT1-mediated vasoconstriction compared to other vasoconstrictors sharing or not its signaling pathway. Moreover, this exposure also abolishes AT1-mediated AngII-independent myogenic tone, suggesting an alteration on the receptor itself. We also show that this effect (i) does not depend on endogenous NO, (ii) is mediated by S-nitrosation rather than by the cGMP/sGC pathway. Finally, we study AT1 internalization by flow cytometry on a heterologous model of AT1 expression. Our results show that GSNO does not alter AT1 cell surface localization and does not prevent its internalization, indicating that the ß-arrestin pathway is not impacted by nitrosation
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https://tel.archives-ouvertes.fr/tel-01868468
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Submitted on : Wednesday, September 5, 2018 - 2:49:15 PM
Last modification on : Tuesday, October 13, 2020 - 10:47:05 AM
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  • HAL Id : tel-01868468, version 1

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Marie Lynda Bouressam. Importance de la S-nitrosation des récepteurs cérébrovasculaires de l’angiotensine II. Médecine humaine et pathologie. Université de Lorraine, 2018. Français. ⟨NNT : 2018LORR0067⟩. ⟨tel-01868468⟩

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