Mécanismes de protection endothéliale induits par l'apoA-I : identification d'une nouvelle voie de signalisation impliquant la PI3K bêta en aval de l'ecto-F1-ATPase

Des taux élevés de Lipoprotéines de Haute Densité (HDL) sont considérés comme protecteurs dans les pathologies cardiovasculaires et présentent un intérêt thérapeutique évident dans la prévention de la resténose, complication majeure des traitements de l'athérosclérose. En effet, en plus de leurs effets sur le métabolisme du cholestérol, ces lipoprotéines semblent jouer un rôle bénéfique important sur la paroi vasculaire. Ces effets peuvent être expliqués d'une part par l'effet anti-inflammatoire des HDL mais également par un effet direct de ces lipoprotéines sur l'endothélium. En effet, il a été montré que les HDL étaient capables de faire proliférer et migrer les cellules endothéliales et de les protéger contre l'apoptose. Toutefois les mécanismes moléculaires mis en jeu dans ces processus restent mal connus à l'heure actuelle. Une partie semble pouvoir être attribuée à la partie lipidique des HDL, capable d'activer les voies de signalisation PI3K/Akt. Toutefois, d'autres études ont montré que la partie protéique des HDL était capable de mimer ces effets, suggérant l'implication d'autres mécanismes moléculaires. Notre équipe s'intéresse depuis plusieurs années au complexe enzymatique membranaire F1-ATPase, capable de générer de l'ADP à la surface cellulaire, comme récepteur de haute affinité pour l'apolipoprotéine A-I (apoA-I), composante protéique majeure des HDL. Nous avons récemment pu montrer que ce complexe était capable d'induire la prolifération et la survie des cellules endothéliales en culture. Dans ce travail, nous nous sommes attachés à déterminer les voies de signalisation impliquées dans la prolifération des cellules endothéliales, induite par l'activation de l'ecto-F1-ATPase par l'apoA-I libre. Mon projet de thèse a permis de déterminer, in vitro, la voie de signalisation mise en jeu dans la prolifération des cellules endothéliales (HUVEC). Ainsi, mes travaux ont permis d'identifier que l'apoA-I purifiée est capable, de la même manière que les HDL, de stimuler la voie PI3K/Akt. En utilisant un inhibiteur naturel de la F1-ATPase, nous avons pu montrer que cette voie est totalement dépendante de la F1-ATPase. D'autre part, nos travaux ont révélé un rôle clé de l'isoforme beta des PI3K dans cette voie de signalisation conduisant à la prolifération des cellules endothéliales. L'ensemble de ces données devrait permettre, non seulement de mieux comprendre les mécanismes moléculaires mis en jeu lors de la cicatrisation artérielle, mais également de proposer de nouvelles cibles thérapeutiques dans la prévention de l'athérosclérose et de la resténose.High Density Lipoproteins (HDL) are known to have beneficial functions on the arterial wall, such as endothelial cell proliferation and survival, through activation of the PI3K/Akt signaling pathway. ApoA-I, the main structural apolipoprotein of HDL, was previously reported to bind to ecto-F1-ATPase, an enzymatic complex related to the mitochondrial ATP synthase and expressed at the cell surface of different cell types such as hepatocytes or endothelial cells. ApoA-I binding to ecto-F1-ATPase activates extracellular ATP hydrolysis into ADP, which promotes endothelial cell survival. In this study, we aimed to investigate the signaling pathway involved downstream ecto-F1-ATPase activation by apoA-I and leading to endothelial cell proliferation. We first observed that both HDL and lipid-free apoA-I could induce Akt phosphorylation in endothelial cells (HUVEC). Inhibitors of F1-ATPase activity such as inhibitor factor 1 (IF1) or oligomycin, could specifically inhibit apoA-I-induced Akt phosphorylation, suggesting that this Akt signaling pathway strictly depend on ecto-F1-ATPase activation by apoA-I. The use of specific inhibitors and silencing of PI3K isoforms evidenced the beta isoform of PI3K (PI3K beta) as the major signaling molecule mediating Akt phosphorylation and cell proliferation in apoA-I- and HDL-stimulated endothelial cells. Altogether, these results demonstrate that PI3K beta plays a major role in endothelial cell proliferation downstream ecto-F1-ATPase activation by apoA-I and suggest that activation of these pathway could enhance vascular endothelial protection

Elastin-derived peptides potentiate atherosclerosis through the immune Neu1-PI3Kγ pathway

Aims Elastin is degraded during vascular ageing and its products, elastin-derived peptides (EP), are present in the human blood circulation. EP binds to the elastin receptor complex (ERC) at the cell surface, composed of elastin-binding protein (EBP), a cathepsin A and a neuraminidase 1. Some in vitro functions have clearly been attributed to this binding, but the in vivo implications for arterial diseases have never been clearly investigated. Methods and results Here, we demonstrate that chronic doses of EP injected into mouse models of atherosclerosis increase atherosclerotic plaque size formation. Similar effects were observed following an injection of a VGVAPG peptide, suggesting that the ERC mediates these effects. The absence of phosphoinositide 3-kinase γ (PI3Kγ) in bone marrow-derived cells prevented EP-induced atherosclerosis development, demonstrating that PI3Kγ drive EP-induced arterial lesions. Accordingly, in vitro studies showed that PI3Kγ was required for EP-induced monocyte migration and ROS production and that this effect was dependent upon neuraminidase activity. Finally, we showed that degradation of elastic lamellae in LDLR−/− mice fed an atherogenic diet correlated with atherosclerotic plaque formation. At the same time, the absence of the cathepsin A-neuraminidase 1 complex in cells of the haematopoietic lineage abolished atheroma plaque size progression and decreased leucocytes infiltration, clearly demonstrating the role of this complex in atherogenesis and suggesting the involvement of endogenous EP. Conclusion Altogether, this work identifies EP as an enhancer of atherogenesis and defines the Neuraminidase 1/PI3Kγ signalling pathway as a key mediator of this function in vitro and in viv

Elastin-derived peptides potentiate atherosclerosis through the immune Neu1-PI3Kγ pathway.

International audienceAIMS: Elastin is degraded during vascular ageing and its products, elastin-derived peptides (EP), are present in the human blood circulation. EP binds to the elastin receptor complex (ERC) at the cell surface, composed of elastin-binding protein (EBP), a cathepsin A and a neuraminidase 1. Some in vitro functions have clearly been attributed to this binding, but the in vivo implications for arterial diseases have never been clearly investigated. METHODS AND RESULTS: Here, we demonstrate that chronic doses of EP injected into mouse models of atherosclerosis increase atherosclerotic plaque size formation. Similar effects were observed following an injection of a VGVAPG peptide, suggesting that the ERC mediates these effects. The absence of phosphoinositide 3-kinase γ (PI3Kγ) in bone marrow-derived cells prevented EP-induced atherosclerosis development, demonstrating that PI3Kγ drive EP-induced arterial lesions. Accordingly, in vitro studies showed that PI3Kγ was required for EP-induced monocyte migration and ROS production and that this effect was dependent upon neuraminidase activity. Finally, we showed that degradation of elastic lamellae in LDLR(-/-) mice fed an atherogenic diet correlated with atherosclerotic plaque formation. At the same time, the absence of the cathepsin A-neuraminidase 1 complex in cells of the haematopoietic lineage abolished atheroma plaque size progression and decreased leucocytes infiltration, clearly demonstrating the role of this complex in atherogenesis and suggesting the involvement of endogenous EP. CONCLUSION: Altogether, this work identifies EP as an enhancer of atherogenesis and defines the Neuraminidase 1/PI3Kγ signalling pathway as a key mediator of this function in vitro and in vivo