Due to the high metabolic demand and low capacity for energy storage of the brain, neurons are vitally reliant on a constant oxygen supply. Under chronic mild hypoxic conditions (10% oxygen), angiogenesis is induced in the brain in an attempt to restore tissue oxygen tension to normal levels. In brain hypoxia, vascular endothelial growth factor (VEGF) plays a critical role in angiogenesis; however, the role of its homolog placental growth factor (PlGF) is unknown. Using PlGF knockout (PlGF-/-) mice exposed to whole body hypoxia (10% oxygen) for 7, 14 and 21-days, we show that PlGF-/- animals exhibit a delay in the angiogenic response of the brain to hypoxia. PlGF-/- microvessels had a significant increase in fibrinogen accumulation and extravasation, which correlated with disruption of the tight-junction protein claudin-5. These vessels displayed large lumens, were surrounded by reactive astrocytes, lacked mural cell coverage and endothelial VEGF expression, and regressed after 21 days of hypoxia. The lack of PlGF, in combination with reduced VEGF expression levels observed in the brain of PlGF-/- animals during the first 5 days of hypoxia, is likely the cause of the delayed angiogenic response and the prothrombotic phenotype of these mice. In vitro studies conducted to analyze mechanisms involved in the impaired angiogenic phenotype and enhanced astrocytic reactivity to hypoxia of PlGF-/- animals indicated that: i) PlGF-/- mouse brain endothelial cells exhibit alterations in intracellular signaling pathways associated with sprouting (ERK1/2) and vessel branching morphogenesis (GSK-3β) and ii) PlGF-/- astrocytes overexpress VEGF receptor-2 (VEGFR-2) which through activation of the ERK1/2 signaling pathway leads to a more proliferative astrocytic phenotype. These astrocytes were more resistant to oxygen and glucose deprivation (OGD) than PlGF+/+ astrocytes, a characteristic that was shown to be independent of the classical antiapoptotic VEGFR-2-dependent PI3K/Akt pathway. The findings presented in this thesis demonstrated a critical role of PlGF in vascular remodeling in the hypoxic brain.

Due to the high metabolic demand and low capacity for energy storage of the brain, neurons are vitally reliant on a constant oxygen supply. Under chronic mild hypoxic conditions (10% oxygen), angiogenesis is induced in the brain in an attempt to restore tissue oxygen tension to normal levels. In brain hypoxia, vascular endothelial growth factor (VEGF) plays a critical role in angiogenesis; however, the role of its homolog placental growth factor (PlGF) is unknown. Using PlGF knockout (PlGF-/-) mice exposed to whole body hypoxia (10% oxygen) for 7, 14 and 21-days, we show that PlGF-/- animals exhibit a delay in the angiogenic response of the brain to hypoxia. PlGF-/- microvessels had a significant increase in fibrinogen accumulation and extravasation, which correlated with disruption of the tight-junction protein claudin-5. These vessels displayed large lumens, were surrounded by reactive astrocytes, lacked mural cell coverage and endothelial VEGF expression, and regressed after 21 days of hypoxia. The lack of PlGF, in combination with reduced VEGF expression levels observed in the brain of PlGF-/- animals during the first 5 days of hypoxia, is likely the cause of the delayed angiogenic response and the prothrombotic phenotype of these mice. In vitro studies conducted to analyze mechanisms involved in the impaired angiogenic phenotype and enhanced astrocytic reactivity to hypoxia of PlGF-/- animals indicated that: i) PlGF-/- mouse brain endothelial cells exhibit alterations in intracellular signaling pathways associated with sprouting (ERK1/2) and vessel branching morphogenesis (GSK-3β) and ii) PlGF-/- astrocytes overexpress VEGF receptor-2 (VEGFR-2) which through activation of the ERK1/2 signaling pathway leads to a more proliferative astrocytic phenotype. These astrocytes were more resistant to oxygen and glucose deprivation (OGD) than PlGF+/+ astrocytes, a characteristic that was shown to be independent of the classical antiapoptotic VEGFR-2-dependent PI3K/Akt pathway. The findings presented in this thesis demonstrated a critical role of PlGF in vascular remodeling in the hypoxic brain.

Due to the high metabolic demand and low capacity for energy storage of the brain, neurons are vitally reliant on a constant oxygen supply. Under chronic mild hypoxic conditions (10% oxygen), angiogenesis is induced in the brain in an attempt to restore tissue oxygen tension to normal levels. In brain hypoxia, vascular endothelial growth factor (VEGF) plays a critical role in angiogenesis; however, the role of its homolog placental growth factor (PlGF) is unknown. Using PlGF knockout (PlGF-/-) mice exposed to whole body hypoxia (10% oxygen) for 7, 14 and 21-days, we show that PlGF-/- animals exhibit a delay in the angiogenic response of the brain to hypoxia. PlGF-/- microvessels had a significant increase in fibrinogen accumulation and extravasation, which correlated with disruption of the tight-junction protein claudin-5. These vessels displayed large lumens, were surrounded by reactive astrocytes, lacked mural cell coverage and endothelial VEGF expression, and regressed after 21 days of hypoxia. The lack of PlGF, in combination with reduced VEGF expression levels observed in the brain of PlGF-/- animals during the first 5 days of hypoxia, is likely the cause of the delayed angiogenic response and the prothrombotic phenotype of these mice. In vitro studies conducted to analyze mechanisms involved in the impaired angiogenic phenotype and enhanced astrocytic reactivity to hypoxia of PlGF-/- animals indicated that: i) PlGF-/- mouse brain endothelial cells exhibit alterations in intracellular signaling pathways associated with sprouting (ERK1/2) and vessel branching morphogenesis (GSK-3β) and ii) PlGF-/- astrocytes overexpress VEGF receptor-2 (VEGFR-2) which through activation of the ERK1/2 signaling pathway leads to a more proliferative astrocytic phenotype. These astrocytes were more resistant to oxygen and glucose deprivation (OGD) than PlGF+/+ astrocytes, a characteristic that was shown to be independent of the classical antiapoptotic VEGFR-2-dependent PI3K/Akt pathway. The findings presented in this thesis demonstrated a critical role of PlGF in vascular remodeling in the hypoxic brain.

La Bronchopneumopathie Chronique Obstructive (BPCO) associe inflammation et hypoxie, vraisemblablement à l’origine d’altérations du tissu musculaire des patients dont l'état est corrélé au pronostic vital. Bien que justifiée, la réhabilitation par l'exercice n'est pas efficace dans 1/3 des cas, sans que les mécanismes à son origine n’aient été identifiés. Cette non-réponse pourrait impliquer un contrôle épigénétique de l'expression des gènes en lien avec la croissance musculaire via leurs profils d'acétylation. Pour étudier les effets de l'exercice sur le muscle BPCO, nous avons développé un modèle murin d'hypertrophie musculaire induite utilisé dans un contexte d'hypoxie (HC) et d'inflammation pulmonaire chronique (IP). Dans ce contexte, en comparant soléaire et plantaire, il apparaît que l’hypertrophie du soléaire est altérée par l’IP alors que celle du plantaire est fortement inhibée par l’HC. L'administration d'un inhibiteur des Bromodomaines et domaines Extra-Terminaux (i-BET), éléments impliqués dans la lecture des niveaux d’acétylations des histones, restaure leurs capacités d'hypertrophie. Cette altération de la croissance musculaire est associée à des perturbations des voies de protéosynthèse (Akt, S6k1, Erk) et de protéolyse (MuRF-1) muscles dépendantes. Dans le soléaire, l’acétylation des lysines des histones H3 et H4 est augmentée par l’IP ainsi que la transcription d’Histones Déacétylases (HDAC) est diminuée. Ces modifications associées à la restauration de l’hypertrophie du soléaire par l’i-BET malgré la présence d’une IP, suggèrent fortement l’implication des mécanismes épigénétiques dans l‘inhibition de la croissance musculaire. L’augmentation de l’acétylation des histones H3 et H4 du plantaire soumis à une HC tend à confirmer l'implication de mécanismes épigénétiques dans l’altération de la réponse de ce muscle. Toutefois des travaux supplémentaires seraient nécessaires pour confirmer cette hypothèse. Pour finir, les mesures de paramètres hémodynamiques cardiaques nous ont montrés que l’i-BET était également à l’origine d’une diminution de l’Hypertension Artérielle Pulmonaire (HTAP), de l’hypertrophie du ventricule droit et de l’hématocrite lors de l’exposition de nos animaux à une HC.Ces travaux suggèrent que la non-réponse au réentrainement d’un tiers des patients BPCO pourrait être liée à la présence d’une IP associée à une hypoxémie. La restauration des capacités adaptatives du muscle par un traitement à l’iBET pourrait constituer une perspective thérapeutique prometteuse, permettant à ces patients de retirer les importants bénéfices d’une telle prise en charge. Toutefois, la réduction de l’hématocrite de nos animaux traités avec l’i BET en HC, implique une prise en charge parallèle à, ce traitement pour maintenir l’adaptation à l’HC des patients BPCO hypoxémiques. / The Chronic Obstructive Pulmonary Disease (COPD) associates inflammation with hypoxia, likely causing deterioration of muscle tissue whose status is tightly correlated to vital prognosis. Although justified due by its anabolic effects, rehabilitation through exercise which progressively became a key medical care in COPD is inefficient in 1/3 of the patients. This non-response could involve epigenetic control of gene expression via alterations of the acetylation profile induced by pulmonary Inflammation (PI) and Chronic Hypoxia (CH). To study this, we used a murine model of either soleus or plantaris muscle hypertrophy induced by a functional overload, in PI and CH conditions. Results show that soleus hypertrophy is diminished by PI while plantaris hypertrophy is inhibited by CH. These specific responses are associated with alterations in proteosynthesis (Akt, S6k1, Erk) and proteolysis (MuRF-1) in a muscle-dependent manner. With PI, acetylation of lysines of histones H3 and H4 is increased in soleus muscle while transcription of Histone Deacetylases (HDACs) is decreased. Inhibiting the proteins in charge of reading the acetylations (BET) is able to restore the hypertrophic capacities of the soleus when exposed to PI, reinforcing the hypothesis of an involvement of epigenetic regulatory mechanisms in the problem of muscle response to a hypertrophic stimulus. Moreover, the use of the BET inhibitor (i-BET) prevents the development of pulmonary arterial hypertension, of the right ventricle hypertrophy and the increase in hematocrit in animals exposed to CH. Restoring the muscle adaptive capacities using i-BET led us to consider new promising therapeutic perspectives in COPD patients who present limited or no response to exercise rehabilitation.

The mortality of cardiovascular diseases remains high and it likely tends to increase in the future. Although many ways how to increase the resistance against myocardial ischemia- reperfusion damage have been described, few of them were transferred into clinical practice. Cardioprotective effect of chronic hypoxia has been described during 60s of the last century. Its detailed mechanism has not been elucidated, but a number of components has been identified. One of these components presents protein kinase C (PKC). The role of PKC was described in detail in the mechanism of ischemic preconditioning, but its involvement in the mechanism of cardioprotection induced by chronic hypoxia remains unclear. One reason is the amount of PKC isoforms, which have often contradictory effects, and the diversity of hypoxic models used. The most frequently mentioned isoforms in connection with cardioprotection are PKCδ and PKCε. The aim of my thesis was to analyze changes in these PKC isoforms at two different cardioprotective models of hypoxia - intermittent hypobaric (IHH) and continuous normobaric hypoxia (CNH). We also examined the target proteins of PKCδ and PKCε after the adaptation to IHH, which could be involved in the mechanism of cardioprotection. These included proteins associated with apoptosis and...

Due to the high metabolic demand and low capacity for energy storage of the brain, neurons are vitally reliant on a constant oxygen supply. Under chronic mild hypoxic conditions (10% oxygen), angiogenesis is induced in the brain in an attempt to restore tissue oxygen tension to normal levels. In brain hypoxia, vascular endothelial growth factor (VEGF) plays a critical role in angiogenesis; however, the role of its homolog placental growth factor (PlGF) is unknown. Using PlGF knockout (PlGF-/-) mice exposed to whole body hypoxia (10% oxygen) for 7, 14 and 21-days, we show that PlGF-/- animals exhibit a delay in the angiogenic response of the brain to hypoxia. PlGF-/- microvessels had a significant increase in fibrinogen accumulation and extravasation, which correlated with disruption of the tight-junction protein claudin-5. These vessels displayed large lumens, were surrounded by reactive astrocytes, lacked mural cell coverage and endothelial VEGF expression, and regressed after 21 days of hypoxia. The lack of PlGF, in combination with reduced VEGF expression levels observed in the brain of PlGF-/- animals during the first 5 days of hypoxia, is likely the cause of the delayed angiogenic response and the prothrombotic phenotype of these mice. In vitro studies conducted to analyze mechanisms involved in the impaired angiogenic phenotype and enhanced astrocytic reactivity to hypoxia of PlGF-/- animals indicated that: i) PlGF-/- mouse brain endothelial cells exhibit alterations in intracellular signaling pathways associated with sprouting (ERK1/2) and vessel branching morphogenesis (GSK-3β) and ii) PlGF-/- astrocytes overexpress VEGF receptor-2 (VEGFR-2) which through activation of the ERK1/2 signaling pathway leads to a more proliferative astrocytic phenotype. These astrocytes were more resistant to oxygen and glucose deprivation (OGD) than PlGF+/+ astrocytes, a characteristic that was shown to be independent of the classical antiapoptotic VEGFR-2-dependent PI3K/Akt pathway. The findings presented in this thesis demonstrated a critical role of PlGF in vascular remodeling in the hypoxic brain.

L’hypertension pulmonaire (HTP) est la principale pathologie de la circulation pulmonaire et a un très mauvais pronostic. Elle se caractérise par une hyperréactivité et un remodelage des petites artères pulmonaires (AP) entraînant une augmentation progressive des résistances vasculaires pulmonaires, qui, ultimement, aboutit à une insuffisance cardiaque droite et au décès du patient. Il est admit que le calcium joue un rôle très important aussi bien dans les mécanismes de remodelage que dans l’hyperréactivité des AP observés dans l’HTP. Dans le présent travail, nous avons étudié l’expression et le rôle d’une famille particulière de canaux calciques, les TRPV, dans les AP de rats contrôles (normoxiques) et souffrant d’hypertension pulmonaire (rats hypoxiques chroniques et traités à la monocrotaline). Nous montrons que (1) les canaux TRPV1, V2 et V4 sont exprimés dans les AP et que cette expression est augmentée au cours de l’HTP ; (2) la stimulation de ces canaux par des agonistes spécifiques induit une augmentation de la concentration calcique intracellulaire dans les cellules musculaires lisses (CML) ; (3) le récepteur à la ryanodine de type 2 (RRy 2) du réticulum sarcoplasmique est impliqué dans la voie de signalisation dépendante de TRPV4 et que son expression est également augmentée au cours de l’HTP ; (4) les canaux TRPV1 et TRPV4 sont impliqués dans la migration des CML, processus fondamental du remodelage ; (5) les contractions induites par l’activation de TRPV2 et TRPV4 dans les AP de rats hypertendus sont significativement diminuées par la streptomycine, un inhibiteur des canaux SAC (stretch activated channels). Ce travail démontre donc l’implication des canaux TRPV à la fois dans l’hyperréactivté et le remodelage des AP. De nouveaux traitements ciblant les canaux TRPV pourraient constituer une approche thérapeutique innovante de l’hypertension pulmonaire. / Pulmonary hypertension (PH)) is the primary pathology of the pulmonary circulation and has a very bad prognostic. This disease is characterized by a hyperreactivity and remodelling of small pulmonary arteries (PA) leading to a progressive increase in pulmonary vascular resistance which ultimately leads to right heart failure and death of the patient. It is admitted that calcium plays an important role both in the mechanisms of remodelling and in the hyperresponsiveness of PA observed in PH. In the present work, we studied the expression and the role of a particular family of calcium channels, TRPV channels, in PA from control rats (normoxic) and pulmonary hypertensive rats (chronically hypoxic and monocrotaline-treated rats). We show that (1) TRPV1, V2 and V4 channels are expressed in the PA and that their expression are increased in PH; (2) stimulation of these channels by specific agonists induces an increase in the intracellular calcium concentration in smooth muscle cells (SMC), (3) the ryanodine receptor type 2 (RRy2) of the sarcoplasmic reticulum is involved in the TRPV4-dependent signaling pathway and its expression is also increased in PH, (4) TRPV1 and TRPV4 channels are involved in the migration of SMC, the fundamental process of remodelling, (5) contractions induced by activation of TRPV2 and TRPV4 in the PA from hypertensive rats are significantly decreased by streptomycine, an inhibitor of stretch activated channels (SAC). This work thus demonstrates the involvement of TRPV channels in both the hyperreactivity and remodelling of PA. New treatments targeting TRPV channels could be an innovative therapeutic approach for pulmonary hypertension.

Cardiac energy metabolism is the one of the most complex system in the body. To sustain life, but also to respond quickly to any sudden changes (e.g. running, emotional stress), the heart has developed a unique ability and has become a metabolic "omnivore". At physiological conditions, long chain fatty acids (LCFAs) present the major energetic source for the adult myocardium. However, the cardiac energy metabolism may be compromised during pathophysiological states. One of the most dangerous is, undoubtedly, ischaemia-reperfusion injury with its acute form, myocardial infarction. However, the adaptation to chronic hypoxia has been known for decades for its cardioprotective effect against I/R. Changes of cardiac energy metabolism induced by the adaptation have not been fully explored and the system conceals still too many secrets. This thesis has aimed to determine how adaptation to chronic hypoxia affects the cardiac metabolism of the rat LVs in the following set-ups: 1. The effect of chronic normobaric hypoxia (CNH; 3 weeks, 5500m) during a brief I/R protocol in vitro on the protein kinase B/hexokinase (Akt/HK) pathway, including the expression and phosphorylation of Akt, the expression and localization of HK, the expression of mitochondrial creatine kinase (mtCKS), and the level of Bcl-2 family...

Endogenous cardiac protection against acute ischemia/reperfusion injury can by increased by cardiac adaptation to various forms of chronic hypoxia. Chronic hypoxia induces a large variety of adaptive changes in the myocardium that could be considered as protective, but the exact mechanism of increased ischemic tolerance is unknown. Different studies suggest that catecholamine release and their effect on -adrenergic signaling after adaptation to chronic hypoxia contributes to cardioprotection. In this study we focused on characterization of -adrenergic receptors ( -ARs) in the myocardium of rats after adaptation to three different hypoxic conditions: 1. intermittent normobaric hypoxia - INH/R (23 h hypoxia, 1 h reoxygenation), 2. intermittent normobaric hypoxia - INH (8 h hypoxia, 16 h normoxia), 3. continuous normobaric hypoxia - CNH (24 h hypoxia). We compared how each hypoxic model affects the total number of -adrenergic receptors and proportion of individual subtypes ( 1-and 2-ARs) in the left and right ventricles compared control normoxic rats. The INH model had apparently no effect on -ARs in either ventricles. On the other hand, adaptation to INH/R and CNH was accompanied by a significant decrease (by about 25%) in the total number of -adrenergic receptors in the right ventricles. Our present...

Cardiac energy metabolism is the one of the most complex system in the body. To sustain life, but also to respond quickly to any sudden changes (e.g. running, emotional stress), the heart has developed a unique ability and has become a metabolic "omnivore". At physiological conditions, long chain fatty acids (LCFAs) present the major energetic source for the adult myocardium. However, the cardiac energy metabolism may be compromised during pathophysiological states. One of the most dangerous is, undoubtedly, ischaemia-reperfusion injury with its acute form, myocardial infarction. However, the adaptation to chronic hypoxia has been known for decades for its cardioprotective effect against I/R. Changes of cardiac energy metabolism induced by the adaptation have not been fully explored and the system conceals still too many secrets. This thesis has aimed to determine how adaptation to chronic hypoxia affects the cardiac metabolism of the rat LVs in the following set-ups: 1. The effect of chronic normobaric hypoxia (CNH; 3 weeks, 5500m) during a brief I/R protocol in vitro on the protein kinase B/hexokinase (Akt/HK) pathway, including the expression and phosphorylation of Akt, the expression and localization of HK, the expression of mitochondrial creatine kinase (mtCKS), and the level of Bcl-2 family...