INTRODUCTION: Diastolic heart failure (HF) i.e., HF with preserved ejection fraction (HFpEF) accounts for ~50% of all clinical HF presentations; but unlike systolic HF i.e., HF with reduced ejection fraction (HFrEF), there are no evidenced based therapies. Obesity is commonly associated with HFpEF. However, there exist a sub-group of obese patients that exhibit a higher survival rate to HFpEF as compared to average patients. Hypertension is the most important risk factor for HFpEF, with a prevalence of 60-89% reported by large controlled trials, epidemiological studies and HF registries. HFpEF morbidity and mortality rates are staggering: 50-60% 5 year mortality rate, 50% 6 month rehospitalization rate and severe clinical disability. However, there remains an incomplete mechanistic understanding about HFpEF. OBJECTIVES: We wanted to explore new pathways related to HFpEF in order to better understand the mechamisms behind its pathophysiology. To do so, we first wanted to explore the potential crosstalk between the heart and adipose tissue during HFpEF by analyzing the adipose tissue in our HFpEF model. Secondly, we sought to test the hypothesis that chronic ETA/ETB inhibition with macitentan (mac) modulates pathologic cardiac remodeling in hypertension-induced HFpEF. METHODS: Mice (20-25 g) were anesthetized, underwent uninephrectomy and received either a continuous infusion of saline (sham) or d-aldosterone (0.3 ug/hour for 4-weeks via osmotic minipumps). All mice were maintained on standard rodent chow and 1.0% sodium chloride drinking water for 4 weeks and then harvested. Second group of mice underwent the same surgical procedure and infusion. They were maintained on standard chow for 2 weeks and then each group was randomized to chow containing macitentan (30 mg/kg/day, HFpEFmac) or standard rodent chow. After 2 additional weeks, the 4 groups of mice (n=4-8/group) were harvested. Blood pressure (BP) was obtained weekly. Prior to sacrifice, body weight and echocardiography parameters (total wall thickness (TWT) and relative wall thickness (RWT)) were determined. We also obtained diastolic dysfunction parameters including deceleration time (DT), isovolumetric relaxation time (IVRT), and E/A ratio. Furthermore, we measured organ weight after harvesting the mice and obtained histological images for the adipose tissues collected. Glucose tolerance test and acute cold tolerance test were performed on HFpEF mice to determine their metabolic state. RESULTS: HFpEF mice developed hypertension, LV hypertrophy, and diastolic dysfunction. Epididymal and inguinal adipose tissue showed significantly reduced weight and adipocyte size. HFpEF mice displayed regular glucose metabolism but were not able to endure a cold tolerance test as their body temperature dropped too low. After 4 weeks, there was no difference in body weight between sham, HFpEF, shammac and HFpEFmac. As expected HFpEF increased systolic BP (117±14 vs 133±16mmHg; P=NS); macitentan did not lower systolic BP after 2 weeks in either shammac or HFpEFmac. Similarly there was no difference in systolic BP between HFpEF and HFpEFmac. Both kidney and spleen weights were increased in HFpEF but not altered by macitentan therapy. There was no change in lung congestion as measured by wet-dry lung ratio. HFpEF increased TWT (0.998±0.04 vs. 0.79±0.11 mm; P<0.01 vs. sham) and RWT (0.686± 0.10 vs. 0.476±0.05 mm; P<0.001 vs. sham) but were modulated by macitentan (HFpEF vs. HFpEFmac; P<0.05 and P<0.001, respectively). There was no difference in chamber size between HFpEF and HFpEFmac. Similarly, IVRT, DT, left ventricular ejection fraction were no different between HFpEF and and HFpEFmac. Furthermore E/A ratio was increased in HFpEF but was not affected by macitentan CONCLUSIONS: Adipose tissue collected from our HFpEF mice displayed a very different phenotype. This demonstrates that inter-tissue communication is definitely occurring between the adipose tissue and the heart. Further research is required to explore what that communication encompasses and how they can be used to improve HFpEF. Macitentan did not lower systolic BP in sham or mice with HFpEF after the development of hypertension. Diastolic dysfunction, as measured by an increased E/A ratio, was not affected by macitentan. Macitentan significantly modulated TWT and RWT after 2 weeks of therapy. It is thus plausible that macitentan may improve HFpEF by improving adverse cardiac remodeling.

No description available.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第25160号 / 医博第5046号 / 新制||医||1070(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 髙橋 良輔, 教授 渡邊 直樹, 教授 林 康紀 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

No / Two different theories of migraine aura exist: In the vascular theory of Wolff, intracerebral vasoconstriction causes migraine aura via energy deficiency, whereas in the neuronal theory of Leão and Morison, spreading depression (SD) initiates the aura. Recently, it has been shown that the cerebrovascular constrictor endothelin-1 (ET-1) elicits SD when applied to the cortical surface, a finding that could provide a bridge between the vascular and the neuronal theories of migraine aura. Several arguments support the notion that ET-1¿induced SD results from local vasoconstriction, but definite proof is missing. If ET-1 induces SD via vasoconstriction/ischemia, then neuronal damage is likely to occur, contrasting with the fact that SD in the otherwise normal cortex is not associated with any lesion. To test this hypothesis, we have performed a comprehensive histologic study of the effects of ET-1 when applied topically to the cerebral cortex of halothane-anesthetized rats. Our assessment included histologic stainings and immunohistochemistry for glial fibrillary acidic protein, heat shock protein 70, and transferase dUTP nick-end labeling assay. During ET-1 application, we recorded (i) subarachnoid direct current (DC) electroencephalogram, (ii) local cerebral blood flow by laser-Doppler flowmetry, and (iii) changes of oxyhemoglobin and deoxyhemoglobin by spectroscopy. At an ET-1 concentration of 1 µM, at which only 6 of 12 animals generated SD, a microarea with selective neuronal death was found only in those animals demonstrating SD. In another five selected animals, which had not shown SD in response to ET-1, SD was triggered at a second cranial window by KCl and propagated from there to the window exposed to ET-1. This treatment also resulted in a microarea of neuronal damage. In contrast, SD invading from outside did not induce neuronal damage in the absence of ET-1 (n = 4) or in the presence of ET-1 if ET-1 was coapplied with BQ-123, an ETA receptor antagonist (n = 4). In conclusion, SD in presence of ET-1 induced a microarea of selective neuronal necrosis no matter where the SD originated. This effect of ET-1 appears to be mediated by the ETA receptor.

Eine der häufigsten kardiovaskulären Erkrankungen ist die Atherosklerose. Bei der Entstehung einer Atherosklerose spielt eine Hyperlipoproteinämie eine entscheidende Rolle. Ein weiterer Faktor für die Entstehung kardiovaskulärer Erkrankungen ist ein hoher Blutdruck. In dieser Arbeit wurde eine mögliche Interaktion zwischen Lipoproteinen und den blutdruckregulierenden Endothelin- und Renin-Angiotensin-Systemen untersucht. Weiterführende Analysen erfolgten an Rezeptoren für die Aufnahme von nLDL und oxLDL. Abschließend wurden Signalwege untersucht, die durch nLDL und oxLDL aktiviert werden. Tierexperimentielle Untersuchungen in Aorten und Herzen fettreich gefütterter Wildtyp- Mäuse unterstützen die Zellkultur-Ergebnisse einer Induzierung des Endothelin-Systems durch erhöhte Lipoproteine. Zusammenfassend zeigt diese Arbeit neue Mechanismen der Interaktion von Lipoproteinen und blutdruckregulierenden Systemen in Endothelzellen. Die Rezeptoren scheinen dabei eine Schlüsselrolle zu spielen. Dies spricht für eine Potenzierung von Hyperlipoproteinämie und Hypertonie bei der Entstehung von Herz-Kreislauf-Erkrankungen.

Abstract Calcified aortic valve disease represents a spectrum of disease spanning from mild aortic valve sclerosis to severe aortic valve stenosis (AS), being an actively regulated disease process and showing some hallmarks of atherosclerosis. The calcified aortic valve lesion develops endothelial injury and is characterized by inflammation, lipid accumulation, renin-angiotensin system activation and fibrosis. There is no approved pharmacological treatment available in AS. This study was aimed to characterize gene expression of endothelial factors in aortic valves in patients representing different stages of calcified aortic valve disease to reveal new targets for pharmacological interventions in AS. Aortic valves obtained from 75 patients undergoing valve replacement surgery were studied. Expression of natriuretic peptides (ANP, BNP and CNP), their processing enzymes (corin and furin), natriuretic receptors (NPR-A, NPR-B and NPR-C), endothelin-1 (ET-1), endothelin converting enzyme-1 (ECE-1), endothelin receptors A and B (ETA and ETB), and apelin pathway (apelin and its receptor APJ) was characterized by reverse-transcriptase polymerase chain reaction (RT-PCR) and immunohistochemistry. AS was characterized by distinct downregulation of gene expression of CNP, its processing enzyme furin and the target receptor NPR-B. Furthermore, increased amount of ET-1 and its target receptor ETA as well as imbalance between ETA and ETB receptors and downregulated endothelial nitric oxide synthase (eNOS) gene expression were observed. Finally, gene expression of apelin and APJ receptor were significantly upregulated in stenotic valves when compared to controls in combination with disequilibrium between expression of angiotensin II receptors AT1 and AT2. The study provides a better understanding of molecular mechanisms associated with calcific aortic valve disease and suggest potential targets for novel therapeutic interventions.

Obstructive sleep apnea (OSA) is a disorder that affects a significant portion of middle-aged adult population. Patients exhibit recurring episodes of upper airway obstruction during sleep that decrease blood oxygen concentration (hypoxia) and are terminated by brief arousals. Epidemiologically, OSA has been extensively linked to cardiovascular dysfunction and is an independent risk factor for the development of hypertension. The proposed mechanism of cardiovascular dysfunction in patients is chronic sympathoexcitation and altered vascular tone, with a predominance of the vasoconstrictor endothelin-1 (ET-1) and removal of the vasodilator nitric oxide (NO). Means to reduce the effects of ET-1 and increase synthesis of NO may have beneficial effects on the cardiovascular co-morbidity commonly associated with OSA. OBJECTIVES: The major aim of this study was to assess the relative importance of circulating biomarkers of ET-1 and NO in hemodynamic function in OSA patients. Potential production of ET-1 by circulating mononuclear cells was also measured to assess their contribution to plasma ET-1 levels. Biomarker levels before and after 12 wk of continuous partial airway pressure (CPAP) therapy were used to assess standard treatment. Mild/moderate exercise training was initiated with CPAP therapy in a subgroup of OSA patients to evaluate the potential benefits of physical activity on hemodynamic function and NO and ET-1 levels. METHODS: Overall, 16 newly diagnosed OSA patients (5 female, 11 male; age 45.4 ± 2.7 yr; RDI 24.6 ± 4.0 events/hr) were selected for study. Seven apparently healthy control volunteers (5 female, 2 male; age 39.43 ± 2.6 yr) screened for OSA served as control subjects. Blood pressure was recorded over one complete day and prior to, during, and following maximal exercise testing on a cycle ergometer. Blood samples were taken prior to exercise testing and assessed for nitrate and nitrite by HPLC and for big endothelin-1 and ET-1 by ELISA. Relative gene expression of preproendothelin-1 was measured by real-time RT-PCR. Following initial testing, patients were stratified into either a standard therapy group (nCPAP) or a standard therapy group with a mild/moderate intensity aerobic training regimen (nCPAP+Ex). Baseline testing was repeated following 12 wk of treatment. Statistical significance was set at p < 0.05 a priori. RESULTS: 24 hr ambulatory systolic and diastolic blood pressure were elevated in OSA patients vs. control subjects (systolic: 128.9 ± 3.8 mmHg vs. 108.8 ± 1.3 mmHg, respectively; diastolic: 97.5 ± 2.0 mmHg vs. 82.1 ± 1.9 mmHg, respectively). OSA patients experienced significant elevations in systolic (OSA 209.7 ± 5.7 mmHg; Control 174.5 ± 6.2 mmHg) and mean arterial pressures (OSA 125.8 ± 3.2 mmHg; Control 109.05 ± 4.5 mmHg) at peak exercise. No differences in nitrate, nitrite, or big endothelin-1 were noted. Plasma endothelin-1 concentrations were below assay detection limit. Big endothelin-1 levels were significantly correlated with BMI in both OSA patients (r=0.955; p=0.001) and control subjects (r=0.799; p=0.045). Relative gene expression of preproendothelin-1 was not elevated in OSA patients (0.40 ± 0.20 fold increase over control subjects). Group nCPAP usage was above minimum therapeutic threshold, but was non-uniform in both groups, with an overall range of 182 to 495 min mean usage per night. A mild/moderate exercise training program failed to elicit a training response through standard hemodynamic or cardiopulmonary indices. Plasma nitrite levels rose from 55.3 ± 4.7 μg/ml to 71.0 ± 7.6 μg/ml in the nCPAP group. CONCLUSIONS: Moderate OSA is associated with elevated blood pressure at rest and during exercise stress that bears no relationship to circulating biomarkers of NO and ET-1 or immune preproendothelin production in patients without diagnosed hypertension. nCPAP therapy failed to elicit significant improvements in hemodynamic function, with or without moderate exercise. Plasma nitrite levels rose following nCPAP therapy, indicating a possible increase in basal nitric oxide formation. Higher intensity exercise regimens may be needed to elicit the positive benefits of exercise training in OSA patients without significant cardiovascular dysfunction. / Ph. D.

Die Arbeit sollte klären, ob die pulmonalprotektiven Effekte von AT III bei LPS-induziertem ARDS auch auf einer Stimulation der pulmonalvaskulären PGI2-Freisetzung beruhen. Die Freisetzung von Big ET-1 und ET-1 unter septischen Bedingungen sollte quantifiziert sowie mögliche Effekte von AT III auf diese Freisetzung untersucht werden. Dabei wurde das Modell der isolierten Rattenlunge verwendet. Die Perfusion der Lunge mit LPS führte zu einer Steigerung der Kon-zentration von 6-Keto-PGF1(, dem stabilen Metaboliten von PGI2, auf das 1,6fache und der Konzentration von TxB2, dem stabilen Metaboliten von TxA2, auf das 2,9fache gegenüber der Kontrollgruppe. Die Konzentration von ET-1 erhöhte sich unter LPS auf das 1,6fache, während der Big ET-1 Spiegel konstant blieb. Die Gabe von AT III hatte keinen Effekt auf die Freisetzung von PGI2 und TxA2. Die kombinierte Gabe von LPS und AT III wirkte ebenso wie die Gabe von LPS allein. Die Konzentrationen von Big ET-1 und ET-1 erhöhten sich unter 2 U/ml AT III auf das 1,7- bzw. 1,2fache und unter 5 U/ml AT III auf das 1,6- bzw. 1,3fache gegen-über den Kontrollen. Die kombinierte Gabe von LPS und AT III führte zu einem signifikant höheren Big ET-1-Spiegel vom 2,6fachen des Basalwertes, während sich die Konzentration von ET-1 nicht von der unter LPS bzw. AT III allein unterschied. Die Gabe von Cicaprost, einem stabilen synthetischen PGI2-Analogon, beeinflußte weder die basale noch die durch 2 U/ml AT III und 50 µg/ml LPS stimulierte Big-ET-1- und ET-1-Freisetzung. Nicardipin, ein Blocker der L-Typ-Kalzium-Kanäle, Heparin und N-Acetyl-Heparin, ein nicht an AT III bindendes Heparin, antagonisierten jeweils den stimulierenden Effekt von AT III auf die Big-ET-1- und ET-1-Freisetzung komplett. Staurosporin, ein Proteinkinase C-Inhibitor und Genistein, ein Tyrosinkinase-Inhibitor hatten keinen Effekt auf die durch AT III stimulierte Big-ET-1- und ET-1-Freisetzung. SCHLUßFOLGERUNGEN: Das für den protektiven Effekt des AT III bei ARDS verantwortlich gemachte PGI2 scheint nichtpulmonalen Ursprungs zu sein. Eine PGI2-mediierte Hemmung der pulmonalen ET-1-Sekretion war nicht zu beobachten und scheint somit nicht am protektiven Effekt des AT III beim septischen ARDS beteiligt zu sein. Der beobachtete stimulierende Effekt des AT III auf die Freisetzung der pulmonalen Endotheline ist von möglicher pathophysiologischer Relevanz, da er die erwähnte protektive Wirkung des AT III mit hoher Wahrscheinlichkeit abschwächt. Dieser stimulierende Effekt des AT III scheint dabei an der intakten Rattenlunge weder von der Proteinkinase C noch von Tyrosinkinasen vermittelt zu sein. Weiterhin ist festzustellen, daß die stimulierende Wirkung des AT III auf die pulmonalvaskuläre Freisetzung von Big ET-1 und ET-1 von einem Kalziumeinstrom durch L-Typ-Kalzium-Kanäle und damit von der intrazellulären Kalziumkonzentration abhängig ist. Wie die gleiche Wirksamkeit von Heparin und N-Azetyl-Heparin zeigt, erfordert die Blockade des AT-III-Effektes durch die Heparine keine direkte Bindung an AT III, was auf die zusätzliche Rolle der intrazellulären Kalziumfreisetzung über IP3 hinweist. / The aim of the present study was to clarify if the pulmonary protective effects of AT III in LPS-induced ARDS can be attributed to a stimulation of the pulmonary vascular release of PGI2. The pulmonary vascular release of big ET-1 and ET-1 under septic conditions and the possible influence of AT III was to be investigated. To this end, we used the model of the isolated perfused rat lung. Exposure of the lung to LPS increased the release of 6-Keto-PGF1(, the stable metabolite of PGI2, 1.6fold and the production of TxB2, the stable metabolite of TxA2, 2.9fold compared with control lungs. The release of ET-1 increased 1.6fold under LPS, whereas the concentration of big ET-1 was unchanged. The application of AT III had no effect on the release of PGI2 and TxA2. The effects following combined application of LPS and AT III were similar to the effects of LPS alone. Compared with controls, AT III, at 2 U/ml, increased the perfusate levels of big ET-1 and ET-1 1.7fold and 1.2fold, respectively; the administration of 5 U/ml AT III raised big ET-1 and ET-1 1.6fold and 1.3fold, respectively. Combined application of LPS and AT III resulted in a 2.6fold rise of big ET-1 levels compared with controls, whereas concentrations of ET-1 did not differ from those in the presence of LPS or AT III alone. Cicaprost, a stable PGI2 analogue, affected neither the basal nor the AT III plus LPS-stimulated release of big ET-1 and ET-1. Nicardipin, an L-type calcium channel blocker, heparin and N-acetyl heparin, a heparin derivative devoid of AT III affinity, each antagonized completely the AT III-stimulated increase in big ET-1 and ET-1 levels. Staurosporin, an inhibitor of protein kinase C, and genistein, an inhibitor of tyrosine kinases, did not influence the AT III effects on endothelins. CONCLUSIONS: In ARDS, the well-known rise in plasma PGI2 in response to AT III obviously originates from non-pulmonary sources. PGI2 does not suppress the pulmonary ET-1 secretion; therefore, this mechanism seems not involved in the AT III-induced lung protection during septic ARDS. The AT III-mediated stimulation of the release of pulmonary endothelins is of potential pathophysiological relevance, because it may blunt the protective effects of AT III in ARDS. In the intact rat lung, this stimulatory effect of AT III is mediated neither by protein kinase C nor by tyrosine kinases. Moreover, the observed effect of AT III on pulmonary endothelins is based on calcium influx through L-type calcium channels and depends on the intracellular calcium activity. The equipotency of heparin and N-acetyl heparin in inhibiting the AT III action demonstrates that direct binding of AT III is not essential for the blocking effect of heparins. This fact points to additional involvement of an IP3-dependent intracellular calcium release.

Die hier im Rahmen einer kumulativen Habilitation vorgelegten Arbeiten fassen die wichtigsten Ergebnisse des Autors zum Thema "Neue Mediatoren in der Pathophysiologie der Herzinsuffizienz" zusammen. Folgende neurohumorale Faktoren waren dabei Gegenstand klinischer und experimenteller Untersuchungen: Relaxin, Urotensin-II, Endothelin-1 und Adrenomedullin. Das wichtigste Ergebnis der klinischen und experimentellen Untersuchungen ist die Charakterisierung des Schwangerschaftshormones Relaxin als kompensatorisch wirksamer Mediator und Gegenspieler des Vasokonstriktors Endothelin-1. Aufgrund des Spektrums der biologischen Eigenschaften von Relaxin - funktioneller Endothelin-1-Antagonismus, Vasodilatation, Fibrosehemmung, Pro-Angiogenese, Föderung der glomerulären Filtration und Abschwächung renaler Vasokonstriktoreneffekte - erscheint das Konzept einer therapeutischen Nutzung des Peptides naheliegend. Von besonderem Interesse könnten wegen des ausgeprägten Endothelinantagonismus und der anti-fibrotischen Eigenschaften die Effekte von Relaxin bei pulmonalvaskulärer Hypertonie sein. Bezüglich der Stellung von Urotensin-II in der Pathophysiologie der Herzinsuffizienz sprechen die gewonnenen klinischen Daten zunächst nicht für eine signifikante Rolle des Peptides. Diese Fragestellung und auch die Frage nach der physiologischen Bedeutung von Urotensin-II sind derzeit Gegenstand einer sehr kontroversen wissenschaftlichen Debatte, so dass weitere und umfangreichere Studien zur endgültigen Klärung nötig sind. Schließlich wurde in einem Flusskammermodell erstmalig der bei Herzinsuffizienz typischerweise erhöhte pulmonalvaskuläre Druck als Regulator der pulmonalendothelialen Mediatorsynthese identifiziert, was klinische Daten zur pulmonalen Freisetzung von Endothelin-1 und Adrenomedullin bestätigt und ergänzt. Diese Befunde sollten Anlaß sein, nun die Signaltransduktion ("Mechanotransduktion") druckinduzierter Prozesse zu untersuchen, welche im Gegensatz zur Transduktion scherabhängiger Vorgänge bisher kaum Gegenstand von Forschungsarbeiten war. / This work comprises a summary of the author s experimental and clinical results regarding "Novel mediators in the pathophysiology of heart failure", including investigations on relaxin, endothelin-1, adrenomedullin, and urotensin-II. In this context, identification of the pregnancy hormone relaxin as compensatory mediator and counterplayer to the vasoconstrictor endothelin-1 represents the most intriguing and important result of these studies. In spite of the spectrum of biological properties of relaxin - functional antagonism towards endothelin-1, vasodilation, inhibition of fibrosis, promotion of angiogenesis, stimulation of glomerular filtration and mitigation of renal vasoconstrictor effects - the therapeutical use of relaxin in heart failure seems to be a compelling concept. Given the pronounced functional endothelin antagonism and the profound anti-fibrotic effects of relaxin its use in pulmonary hypertension may be of particular interest. With regard to the relevance of urotensin-II the clinical data presented here do not confirm the view that this peptide plays a significant role in heart failure. However, this point as well as the physiological importance of urotensin-II are currently subject to a controversial scientific debate; therefore, additional studies are necessary to unravel these questions. Finally, using a novel flowchamber model, pulmonary vascular pressure - typically elevated in heart failure - was characterized as regulator of pulmonary endothelial mediator synthesis. These findings corroborate and extend clinical data showing pulmonary vascular release of endothelin-1 and adrenomedullin in patients with heart failure. Based on these results it appears rewarding to investigate signaling mechanisms of pressure-related vascular processes ("mechanotransduction"), which are poorly understood at present.

Le système endothéline (ET) est activé en condition d’hypertension pulmonaire (HTP). L’efficacité des antagonistes des récepteurs à l’ET a clairement été démontrée et a menée à l’approbation clinique de tels antagonistes dans le traitement de l’hypertension artérielle pulmonaire (HTAP). Toutefois, il existe présentement un important débat opposant l’utilisation d’un antagoniste sélectif des récepteur ETA à l’utilisation d’un antagoniste double ETA/ETB dans le traitement de cette pathologie. Bien que nous sachions que le système ET est activé et contribue à l’HTAP, les modifications locales de ce système induites par la pathologie, particulièrement au niveau des artères de résistance pulmonaires, demeurent inconnues. De plus, l’impact de ces modifications sur la réponse pharmacologique aux divers antagonistes des récepteurs à l’ET (sélectifs versus double) est d’une importance capitale. Ainsi, le but de la première étude de cette thèse était d’évaluer les modifications potentielles de la pharmacologie du système ET au niveau des artères de résistance pulmonaires induites par l’HTAP. Dans cette étude, nous avons démontré qu’en condition contrôle l’antagoniste sélectif ETA et l’antagoniste double n’ont eu aucun effet sur la réponse vasoconstrictrice à l’ET-1. Toutefois, en condition d’HTAP, les antagonistes sélectif et double ont tous deux été en mesure de réduire la vasoconstriction pulmonaire induite par l’ET-1. Une diminution importante de l’expression génique du récepteur ETB pourrait être à l’origine de cette modification du profil pharmacologique des antagonistes. Une meilleure compréhension des rôles joués par les récepteurs ETA et ETB au niveau des artères de résistance pulmonaires pourrait permettre l’optimisation des traitements de l’HTAP. Ainsi, le but de la deuxième étude était d’évaluer les effets d’un traitement antisens ex vivo dirigé contre l’ARNm des récepteurs ETA et ETB dans la vasoconstriction des artères de résistance pulmonaires induite par l’ET-1. Dans cette étude, nous avons démontré dans un premier temps que les récepteurs ETA et ETB pouvaient former des dimères au niveau des artères de résistance pulmonaires. De plus, nous avons observé qu’une réduction de l’expression protéique du R-ETA entraînait une potentialisation de la vasoconstriction ETB dépendante suggérant ainsi qu’en condition contrôle, le récepteur ETA aurait un effet inhibiteur sur la vasoconstriction pulmonaire induite par la stimulation du récepteur ETB. Les effets délétères de l’ET-1 sur la circulation pulmonaire sont bien connus, toutefois seules quelques études ont porté leur attention sur l’implication de l’ET-3 dans l’HTAP. Ainsi, le but de la troisième étude était d’évaluer l’implication potentielle de l’ET-3 dans l’HTAP. Dans cette étude, nous avons démontré qu’il était nécessaire en condition contrôle de bloquer simultanément les récepteurs ETA et ETB afin de réduire la réponse vasoconstrictrice pulmonaire à l’ET-3. En condition d’HTAP, nous avons observé une augmentation non-significative des concentrations plasmatiques d’ET-3 ainsi qu’une modification du profil pharmacologique des antagonistes des récepteurs à l’ET. En effet, l’utilisation de l’antagoniste sélectif ETA ou de l’antagoniste double était dans les deux cas en mesure de réduire la vasoconstriction pulmonaire à l’ET-3. Les résultats de ces trois études suggèrent qu’il est préférable d’utiliser un antagoniste double dans le traitement de l’HTAP. En effet, (1) en condition d’HTAP, l’utilisation d’un antagoniste double est aussi efficace que l’utilisation d’un antagoniste sélectif ETA; (2) les récepteurs ETA et ETB peuvent former des dimères au niveau des artères de résistance pulmonaires et (3) le récepteur ETB joue un rôle prédominant dans la vasoconstriction pulmonaire, il semble donc essentiel de bloquer simultanément les récepteurs ETA et ETB afin d’inhiber la réponse vasoconstrictrice induite par l’ET. Mots-clés: endothéline-1, endothéline-3, artère de résistance pulmonaire, récepteur vasculaire, antagoniste des récepteurs à l’ET, dimérisation, phosphorothioate, hypertension artérielle pulmonaire / The endothelin (ET) system is activated in pulmonary arterial hypertension (PAH); indeed, increased plasma levels of ET-1 were detected in patients with various forms of PAH and in various experimental models. The therapeutic value of pharmacological blockade of ET receptors has been demonstrated in various animal models and led to the current approval and continued development of these drugs for the therapy of human PAH. Whether the net effect of either selective ETA receptor blockade or combined ETA/ETB receptor blockade provides greater therapeutic benefit remains a subject of debate. Although the ET system contributes to PAH, we currently incompletely comprehend which local modifications of this system occur as a consequence of PAH, particularly in small resistance arteries, and how this could affect the pharmacological response to ET receptor antagonists. The purpose of the first study was therefore to evaluate potential modifications of the pharmacology of the ET system in rat pulmonary resistance arteries from monocrotaline-induced PAH. Our results reveal striking changes in pulmonary vasculature sensitivity to ET receptor antagonism in PAH that may be related to a reduction in ETB receptor expression. A better understanding of the exact role played by both ETA and ETB receptors on pulmonary resistance arteries might contribute to optimization of PAH treatments. Therefore the aim of the second study was to clarify the role played by both ETA and ETB receptors in ET-1 induced pulmonary vasoconstriction using an antisense (AS) oligonucleotide ex vivo treatment. In this study we have demonstrated that ETA and ETB receptors can form heterodimers in pulmonary resistance arteries. Furthermore, suppression of ETA receptors potentiated the response to ET-1 suggesting that in control condition the ETA receptor has an inhibitory action on the ET-1 induced pulmonary vasoconstriction induced by the stimulation of the ETB receptor. Although the deleterious effects of ET-1 on the pulmonary circulation are well established, only a few studies have focused on ET-3 in PAH. Therefore, the purpose of the last study was to evaluate the potential implication of ET-3 in MCT-induced PAH and evaluate the roles of ETA and ETB receptors on ET-3-induced pulmonary vascular reactivity. In control condition, the use of a combination of both ETA and ETB receptor antagonists is necessary to reduce the ET-3 induced pulmonary vasoconstriction. In PAH, we found an increased ET-3 plasma levels and a modification of the pharmacological profile of ET receptor antagonists. Indeed, the use of either the ETA receptor antagonist or the dual antagonist was able to reduce the ET-3 response. The results from these three studies suggest that it is preferable to use a dual antagonist in the treatment of PAH. Indeed, (1) in PAH the use of a dual antagonist is as effective as the use of a selective ETA receptor antagonist (2) ETA and ETB receptors can form heterodimers in pulmonary resistance arteries and (3) ETB receptor plays an important role in the ET-1 induced pulmonary vasoconstriction, suggesting that it is necessary to block both receptors to reduce the ET-1 induced pulmonary vasoconstriction. Keywords: endothelin-1, endothelin-3, pulmonary resistance artery, receptor, endothelin receptor antagonist, dimerisation, phosphorothioate pulmonary arterial hypertension