Yes / Pulmonary arterial hypertension (PAH) is a cardiovascular disorder associated with enhanced proliferation and suppressed apoptosis of pulmonary arterial smooth muscle cells (PASMCs). Heterozygous mutations in the type II receptor for bone morphogenetic protein (BMPR2) underlie the majority of the inherited and familial forms of PAH. The transforming growth factor beta (TGFbeta) pathway is activated in both human and experimental models of PAH. However, how these factors exert pro-proliferative and anti-apoptotic responses in PAH remains unclear. Using mouse primary PASMCs derived from knock-in mice, we demonstrated that BMPR-II dysfunction promotes the activation of small mothers against decapentaplegia-independent mitogen-activated protein kinase (MAPK) pathways via TGFbeta-associated kinase 1 (TAK1), resulting in a pro-proliferative and anti-apoptotic response. Inhibition of the TAK1-MAPK axis rescues abnormal proliferation and apoptosis in these cells. In both hypoxia and monocrotaline-induced PAH rat models, which display reduced levels of bmpr2 transcripts, this study further indicates that the TGFbeta-MAPK axis is activated in lungs following elevation of both expression and phosphorylation of the TAK1 protein. In ex vivo cell-based assays, TAK1 inhibits BMP-responsive reporter activity and interacts with BMPR-II receptor. In the presence of pathogenic BMPR2 mutations observed in PAH patients, this interaction is greatly reduced. Taken together, these data suggest dysfunctional BMPR-II responsiveness intensifies TGFbeta-TAK1-MAPK signalling and thus alters the ratio of apoptosis to proliferation. This axis may be a potential therapeutic target in PAH.

La sclérose systémique (ScS) est une maladie auto-immune dont l’un des principaux auto-anticorps, dirigé contre la protéine centromérique B (CENP-B), est fortement associé à l’hypertension artérielle pulmonaire, l’une des causes majeures de décès dû à la ScS. L’hypertension résulte de l’occlusion progressive des vaisseaux suite à une hyperactivation des cellules musculaires lisses (CML) de la paroi vasculaire. Cependant, les facteurs responsables de ce remodelage vasculaire restent inconnus. Plusieurs études récentes ont démontré que certains auto-antigènes possèdent des fonctions biologiques additionnelles lorsqu'ils se retrouvent dans le milieu extracellulaire. En effet, une fois libérés par nécrose ou apoptose, ces auto-antigènes adoptent une activité biologique qui s'apparente à celles des cytokines et peuvent ainsi participer aux processus normaux de réparation de blessure et/ou acquérir une activité pathogène qui contribue au développement de certaines maladies auto-immunes. Nos résultats suggèrent que la CENP-B peut être ajoutée à cette liste de molécules bifonctionnelles. À l'aide des techniques d'immunofluorescence, d'ELISA cellulaire et de cytométrie en flux, nous avons démontré que la CENP-B se liait spécifiquement à la surface des CML vasculaire de l’artère pulmonaire avec une plus grande affinité pour le phénotype contractile que synthétique. Cette liaison provoquait la migration des cellules ainsi que la sécrétion de cytokines pro-inflammatoires telles que l’interleukine 6 et 8. Les mécanismes par lesquels la protéine exerçait ces effets impliquaient la phosphorylation de FAK et Src ainsi que la voie des MAP kinases, avec ERK1/2 et p38. Des études de signalisation intracellulaire effectuées à l’aide de plusieurs inhibiteurs spécifiques ainsi que des études de désensibilisation nous ont permis d’identifier le récepteur de la CENP-B en plus d’identifier les mécanismes complets de sa signalisation membranaire. Nous avons démontré que la CENP-B se liait de manière spécifique aux CML vasculaire via le récepteur de chémokine 3 (CCR3) pour ensuite transactiver le récepteur EGF, selon un mécanisme métalloprotéase-dépendant qui implique le relargage du HB-EGF. Cette transactivation est un processus important dans l’activation de la voie des MAP kinases ainsi que dans la sécrétion d’IL-8 induite par la CENP-B. Finalement, nous avons démontré que les auto-anticorps anti-CENP-B pouvaient abolir cette cascade de signalisation, empêchant ainsi la CENP-B d’exercer son rôle de cytokine. L’identification de la CENP-B comme ligand du CCR3 ouvre donc plusieurs perspectives quant à l’étude du rôle pathogène des auto-anticorps anti-CENP-B dans la ScS. / CENP-B is a highly conserved, centromere associated protein and is a major autoantigen in systemic sclerosis (SSc). Anti-CENP-B autoantibodies are associated with prominent vascular manifestations such as pulmonary arterial hypertension (PAH) in the limited cutaneous subset of SSc. PAH occurs as a consequence of progressive obliteration of small arteries due to vascular smooth muscle cell dysfunction, migration and proliferation. However, the factors driving this obliteration are unknown. Earlier in vitro studies have demonstrated that some autoantigens have an additional role when they are released in the extracellular environment during the course of injurious insults resulting in cell death. Indeed, it was previously suggested that extracellular autoantigens participate in normal wound repair processes by acting like cytokines and/or chemokines and subsequently displaying pathogenic activities that contribute to the development of autoimmune diseases. Our present findings suggest that the nuclear autoantigen CENP-B can be added to this set of bifunctional molecules. The present study clearly indicates that exogenous CENP-B bound specifically to the surface of human pulmonary artery SMCs. Binding of CENP-B to SMC stimulated their migration during in vitro wound healing assays, as well as their secretion of interleukins 6 and 8. The mechanism by which CENP-B mediated these effects involved the focal adhesion kinase, Src, ERK1/2, and p38 MAPK pathways. Moreover, CENP-B released from apoptotic endothelial cells was found to bind to SMC, thus indicating a plausible in vivo source of extracellular CENP-B. Here, we also report several lines of evidence indicating that CENP-B, which has no obvious primary or secondary structural homology to chemokines, induced SMC activation by interacting with CCR3. Moreover, the present study clearly demonstrates the involvement of EGFR in CENP-B signaling leading to IL-8 secretion. Finally, anti-CENP-B autoantibodies were found to abolish this signaling pathway, thus preventing CENP-B from transactivating EGFR and exerting its cytokine-like activities toward vascular SMCs. The present study sheds new light on the possible role of extracellular CENP-B and its potent biological effects on human pulmonary artery SMCs. The identification of CENP-B as a CCR3 ligand opens up new perspectives for the study of the pathogenic role of anti-CENP-B autoantibodies.

Para avaliar a hipótese da presença de inflamação em artérias pulmonares periféricas de pacientes com hipertensão pulmonar (HP) decorrente de cardiopatias congênitas, foram quantificadas células inflamatórias através de marcação imunohistoquímica em biópsias de 26 pacientes e comparadas com 11 controles sem cardiopatia. Detectou-se quantidades semelhantes de células inflamatórias nos dois grupos, mas com predomínio de linfócitos T no grupo controle e de macrófagos jovens no grupo HP. Esses achados podem estar relacionados com a redução do estímulo dependente de macrófagos para diferenciação e maturação de linfócitos T nos cardiopatas e/ou a deficiência imunológica primária nesses pacientes / To evaluate the hypothesis of increased inflammation in peripheral pulmonary arteries from patients with pulmonary hypertension secondary to congenital cardiac shunts, we quantified the inflammatory cells with the aid of immunohystochemistry in 26 biopsies (HP group), comparing them to 11 patients with no cardiac disease. Similar quantities of inflammatory cells were observed in the two groups, with a predominance of T-lymphocytes in the controls and of young macrophages in the HP group. These findings could be related to a reduction of macrophagic stimulus to the differentiation and maturation of T-lymphocytes and/or to a primary immunological deficiency in patients with congenital cardiac shunts

L’intervention de Fontan est une chirurgie palliative pour les cardiopathies congénitales univentriculaire. A long terme, la défaillance cardiaque, l’hypertension artérielle pulmonaire(HTAP) et l’entéropathie exsudative peuvent mener à l’échec tardif du Fontan.Une mise au point clinique a été realisée en se focalisant secondairement sur les patients adultes.L’intérêt de la cavo bipulmonaire (CBP), étape intermédiaire au Fontan, dans certains cas de retours veineux pulmonaires anormaux totaux a été décrite. Afin d’affiner de futures études cliniques, nous avons discuté de l’intérêt du Brain NatriureticPeptide en post opératoire de Fontan. Nous avons démontré la fiabilité d’une nouvelle technologiede l’oxyme´trie de pouls chez des enfants cyanosés et la non fiabilité d’un nouveau dispositifd’impédancemétrie pour le monitorage non invasif du de´bit cardiaque en comparaison avecun cathéter artériel pulmonaire.Un modèle animal viable et chronique de la CBP a été développé. Il a montré que le flux pulsatileantérograde empêche la formation de fistules artério-veineuses pulmonaires (FAVP), améliorel’hématose et atténue le développement de l’HTAP soutenant cliniquement le maintien de ce fluxlors de la création d’une CBP. Par ailleurs, la perte de pulsatilité, en diminuant le contrainte de cisaillement, entraîne unediminution de l’eNOS synthase et une réponse altérée vaso-relaxante dépendante de l'endothélium artérielle pulmonaire. La micropulsatilité limite ces effets délétères. Ces résultats impliquent desthérapies potentielles contre l’HTAP du Fontan, en maintenant un débit pulmonaire accessoire et la modulation pharmaceutique vaso-relaxante non endothélium-dépendante / The Fontan surgery is a palliative surgery for univentricular congenital heart disease. On the longterm, heart failure, pulmonary arterial hypertension (PAH) and exudative enteropathy may lead tothe Fontan failure.A clinical adjustment has been realized by focusing secondarily on adult patients. The advantage, insome cases of total anomalous pulmonary venous return, of the bidirectional cavopulmonary (BCP),which is an intermediate stage of Fontan, has been described.In order to prepare future clinical studies, we have discussed about Brain Natriuretic Peptide inpostoperative Fontan surgery. We have demonstrated the reliability of a new technology of pulseoxymetry in cyanotic children and the unreliability of a new device of impedance for non-invasivemonitoring of cardiac outpout in comparison with pulmonary arterial catheter.A viable chronic animal model of bidirectional cavo-pulmonary (BCP) has been developed. Itshowed that the antegrade pulsatile flow prevents the formation of pulmonary arteriovenous fistulas(PAVMs) ,improves hematosis and attenuates the development of PAH clinically supporting themaintenance of this flow when creating a BCP.Moreover, the loss of pulsatility while reducing the shear stress causes a decrease in e-NOSsynthase and a dependent vaso-relaxing altered response of the pulmonary arterial endothelium. Themicropulsatility limits these deleterious effects. These results imply potential therapies against thePAH of Fontan surgery, maintaining an accessory pulmonary blood flow and the pharmaceuticalvaso-relaxing non-endothelium dependent modulation.

La sclérose systémique (ScS) est une maladie auto-immune dont l’un des principaux auto-anticorps, dirigé contre la protéine centromérique B (CENP-B), est fortement associé à l’hypertension artérielle pulmonaire, l’une des causes majeures de décès dû à la ScS. L’hypertension résulte de l’occlusion progressive des vaisseaux suite à une hyperactivation des cellules musculaires lisses (CML) de la paroi vasculaire. Cependant, les facteurs responsables de ce remodelage vasculaire restent inconnus. Plusieurs études récentes ont démontré que certains auto-antigènes possèdent des fonctions biologiques additionnelles lorsqu'ils se retrouvent dans le milieu extracellulaire. En effet, une fois libérés par nécrose ou apoptose, ces auto-antigènes adoptent une activité biologique qui s'apparente à celles des cytokines et peuvent ainsi participer aux processus normaux de réparation de blessure et/ou acquérir une activité pathogène qui contribue au développement de certaines maladies auto-immunes. Nos résultats suggèrent que la CENP-B peut être ajoutée à cette liste de molécules bifonctionnelles. À l'aide des techniques d'immunofluorescence, d'ELISA cellulaire et de cytométrie en flux, nous avons démontré que la CENP-B se liait spécifiquement à la surface des CML vasculaire de l’artère pulmonaire avec une plus grande affinité pour le phénotype contractile que synthétique. Cette liaison provoquait la migration des cellules ainsi que la sécrétion de cytokines pro-inflammatoires telles que l’interleukine 6 et 8. Les mécanismes par lesquels la protéine exerçait ces effets impliquaient la phosphorylation de FAK et Src ainsi que la voie des MAP kinases, avec ERK1/2 et p38. Des études de signalisation intracellulaire effectuées à l’aide de plusieurs inhibiteurs spécifiques ainsi que des études de désensibilisation nous ont permis d’identifier le récepteur de la CENP-B en plus d’identifier les mécanismes complets de sa signalisation membranaire. Nous avons démontré que la CENP-B se liait de manière spécifique aux CML vasculaire via le récepteur de chémokine 3 (CCR3) pour ensuite transactiver le récepteur EGF, selon un mécanisme métalloprotéase-dépendant qui implique le relargage du HB-EGF. Cette transactivation est un processus important dans l’activation de la voie des MAP kinases ainsi que dans la sécrétion d’IL-8 induite par la CENP-B. Finalement, nous avons démontré que les auto-anticorps anti-CENP-B pouvaient abolir cette cascade de signalisation, empêchant ainsi la CENP-B d’exercer son rôle de cytokine. L’identification de la CENP-B comme ligand du CCR3 ouvre donc plusieurs perspectives quant à l’étude du rôle pathogène des auto-anticorps anti-CENP-B dans la ScS. / CENP-B is a highly conserved, centromere associated protein and is a major autoantigen in systemic sclerosis (SSc). Anti-CENP-B autoantibodies are associated with prominent vascular manifestations such as pulmonary arterial hypertension (PAH) in the limited cutaneous subset of SSc. PAH occurs as a consequence of progressive obliteration of small arteries due to vascular smooth muscle cell dysfunction, migration and proliferation. However, the factors driving this obliteration are unknown. Earlier in vitro studies have demonstrated that some autoantigens have an additional role when they are released in the extracellular environment during the course of injurious insults resulting in cell death. Indeed, it was previously suggested that extracellular autoantigens participate in normal wound repair processes by acting like cytokines and/or chemokines and subsequently displaying pathogenic activities that contribute to the development of autoimmune diseases. Our present findings suggest that the nuclear autoantigen CENP-B can be added to this set of bifunctional molecules. The present study clearly indicates that exogenous CENP-B bound specifically to the surface of human pulmonary artery SMCs. Binding of CENP-B to SMC stimulated their migration during in vitro wound healing assays, as well as their secretion of interleukins 6 and 8. The mechanism by which CENP-B mediated these effects involved the focal adhesion kinase, Src, ERK1/2, and p38 MAPK pathways. Moreover, CENP-B released from apoptotic endothelial cells was found to bind to SMC, thus indicating a plausible in vivo source of extracellular CENP-B. Here, we also report several lines of evidence indicating that CENP-B, which has no obvious primary or secondary structural homology to chemokines, induced SMC activation by interacting with CCR3. Moreover, the present study clearly demonstrates the involvement of EGFR in CENP-B signaling leading to IL-8 secretion. Finally, anti-CENP-B autoantibodies were found to abolish this signaling pathway, thus preventing CENP-B from transactivating EGFR and exerting its cytokine-like activities toward vascular SMCs. The present study sheds new light on the possible role of extracellular CENP-B and its potent biological effects on human pulmonary artery SMCs. The identification of CENP-B as a CCR3 ligand opens up new perspectives for the study of the pathogenic role of anti-CENP-B autoantibodies.

Para avaliar a hipótese da presença de inflamação em artérias pulmonares periféricas de pacientes com hipertensão pulmonar (HP) decorrente de cardiopatias congênitas, foram quantificadas células inflamatórias através de marcação imunohistoquímica em biópsias de 26 pacientes e comparadas com 11 controles sem cardiopatia. Detectou-se quantidades semelhantes de células inflamatórias nos dois grupos, mas com predomínio de linfócitos T no grupo controle e de macrófagos jovens no grupo HP. Esses achados podem estar relacionados com a redução do estímulo dependente de macrófagos para diferenciação e maturação de linfócitos T nos cardiopatas e/ou a deficiência imunológica primária nesses pacientes / To evaluate the hypothesis of increased inflammation in peripheral pulmonary arteries from patients with pulmonary hypertension secondary to congenital cardiac shunts, we quantified the inflammatory cells with the aid of immunohystochemistry in 26 biopsies (HP group), comparing them to 11 patients with no cardiac disease. Similar quantities of inflammatory cells were observed in the two groups, with a predominance of T-lymphocytes in the controls and of young macrophages in the HP group. These findings could be related to a reduction of macrophagic stimulus to the differentiation and maturation of T-lymphocytes and/or to a primary immunological deficiency in patients with congenital cardiac shunts

Ce travail a été consacré à l’étude non invasive de la circulation pulmonaire normale par mise en œuvre de l’échocardiographie Doppler. <p>En intégrant les mesures obtenues dans une approche physiopathologique, et en exploitant les nouvelles possibilités d’échocardiographes portables, techniquement performants, nous avons analysé les effets d’un inhibiteur de la phosphodiestérase-5 et d’une prostacycline, pour tenter d’en identifier d’éventuels effets introtropes intrinsèques, nous avons exploré le concept de réserve vasculaire pulmonaire comme facteur limitant de l’aptitude aérobie et indice potentiel d’une atteinte vasculaire pulmonaire précoce, et obtenu des résultats préliminaires permettant d’identifier une hypertension artérielle pulmonaire (HTAP) latente. Nos principaux résultats peuvent être résumés comme suit :<p>1. Chez le sujet sain, en normoxie ou dans un modèle expérimental d’HTAP induite par l’inhalation d’un mélange gazeux hypoxique, le sildenafil per os ou l’epoprostenol par voie intraveineuse, à des doses utilisées en clinique pour le traitement de l’HTAP, améliorent les indices de la fonction ventriculaire droite en proportion de leurs effets vasodilatatoires pulmonaires, sans effets inotropes intrinsèques détectables.<p>2. La consommation d’oxygène maximale du sujet sain augmente en raison directe de son volume capillaire pulmonaire (calculé à partir de sa capacité de diffusion pour l’oxyde nitrique et le monoxyde de carbone) et en raison inverse de sa résistance vasculaire pulmonaire, non seulement en altitude, mais aussi au niveau de la mer. Ce résultat suggère qu’une plus grande réserve vasculaire pulmonaire est propice aux efforts aérobiques intenses, probablement par moindre postcharge ventriculaire droite.<p>3. Des mesures réalisées chez un petit nombre de sujets suggèrent que la distensibilité vasculaire pulmonaire, calculée à partir d’une relation débit-pression vasculaire pulmonaire, est typiquement réduite chez des porteurs asymptomatiques de la mutation BMPR2, qui est actuellement le facteur de risque le plus élevé connu de l’HTAP. La mutation BMPR2 pourrait aussi être associée à une réactivité vasculaire pulmonaire accrue à l’hypoxie. <p>Nos résultats suggèrent indirectement que l’échocardiographie Doppler, de repos ou de stress, pourrait être davantage développée dans la mise au point de patients à risque d’HTAP./<p><p>Novel advances in echocardiography offer the opportunity to reliably characterize pulmonary circulation in terms of pressure-flow relationship, and to better understand the coupling of right ventricular (RV) function with normal and abnormal pulmonary hemodynamics. Moreover, when combined with the measurement of pulmonary capillary blood volume, this renewed methodological approach may help to understand the concept of pulmonary vascular reserve as a limiting factor of exercise capacity and potential sensitive marker of early vascular disease.<p><p>In the present work we used a model of hypoxic pulmonary vasoconstriction to analyse the effects of two targeted therapies of pulmonary arterial hypertension (PAH) on the RV function. We showed that the beneficial effects of these drugs are mainly driven by a decrease in RV afterload and not an enhanced myocardial inotropic state. Whether this is transposable to abnormal RV-arterial coupling in PAH patients remains to be investigated.<p><p>Echocardiography may be useful to explore the pulmonary vascular reserve as an important limiting factor of exercise capacity. We showed that a higher pulmonary vascular reserve, defined by a decreased PVR and increased lung diffusing capacity, allows for an improved aerobic exercise capacity (as assessed by a higher peak oxygen consumption), at a lower ventilatory cost, at sea level and at high altitude. <p><p>Stress echocardiography may detect an abnormal pulmonary vasoreactivity. We showed that asymptomatic relatives of patients suffering from idiopathic pulmonary arterial hypertension, and who carry a bone morphogenetic protein receptor type 2 mutation (BMPR2) present with a decreased pulmonary vascular distensibility and an enhanced pulmonary vasoreactivity to hypoxia, which are identifiable by echocardiography examination. However, the predictive value of these findings is not known. <p><p>Thus echocardiography may represent, in experienced and dedicated hands, a noninvasive, safe, widely available, applicable at the bed-side as well as in extreme environment (e.g. high altitudes), less expensive alternative for the evaluation of the pulmonary circulation, either by the interrogation of pressure-flow relationship (stress echocardiography), by the investigation of the right ventricle global and regional function in relation to its afterload (standard and Tissue Doppler Imaging), or by a combined approach with the measurement of lung diffusing capacity (DLNO / DLCO) to assess the pulmonary vascular reserve.<p><p>The present data are encouraging for further development and implementation of echocardiography for the detection, but also the diagnosis and follow-up of patients with pulmonary hypertension.<p><p> / Doctorat en Sciences médicales / info:eu-repo/semantics/nonPublished

Magnetic resonance images (MRI) are acquired by sampling the current of induced electromotiveforce (EMF). EMF is induced due to flux of the net magnetic field from coherent nuclear spins with intrinsic magnetic dipole moments. The spins are excited by (non-ionizing) radio frequency electromagnetic radiation in conjunction with stationary and gradient magnetic fields. These images reveal detailed internal morphological structures as well as enable functional assessment of the body that can help diagnose a wide range of medical conditions. The aim of this project was to unwrap phase contrast cine magnetic resonance images, targeting the great vessels. The maximum encoded velocity (venc) is limited to the angular phase range [-π, π] radians. This may result in aliasing if the venc is set too low by the MRI personnel. Aliased images yield inaccurate cardiac stroke volume measurements and therefore require acquisition retakes. The retakes might be avoided if the images could be unwrapped in post-processing instead. Using computer vision, the angular phase of flow measurements as well as the angular phase of retrospectively wrapped image sets were unwrapped. The performances of three algorithms were assessed, Laplacian algorithm, sequential tree-reweighted message passing and iterative graph cuts. The associated energy formulation was also evaluated. Iterative graph cuts was shown to be the most robust with respect to the number of wraps and the energies correlated with the errors. This thesis shows that there is potential to reduce the number of acquisition retakes, although the MRI personnel still need to verify that the unwrapping performances are satisfactory. Given the promising results of iterative graph cuts, next it would be valuable to investigate the performance of a globally optimal surface estimation algorithm.