A l’issue d’un accident nucléaire, les produits de fission de l’uranium tel les iodes radioactifs sont dispersés dans l’environnement. L’homme est susceptible d’être exposé à ces éléments majoritairement via l’inhalation d’air et/ ou l’ingestion d’aliments contaminés. L’iode 131 est connu pour être responsable de l’augmentation de l’incidence du cancer de la thyroïde. Une des contremesures pour prévenir cette pathologie est l’ingestion de dose unique de comprimés d’iodure de potassium (KI) à fin de saturer la glande thyroïde par de l’iode stable et d’éviter ainsi l’accumulation de l’iode radioactif. Les scénarios de rejets réitérés d’iodes radioactifs lors des deux accidents majeurs Tchernobyl et Fukushima ont mis en évidence les limites de cette mesure, des prises répétées de KI pour protéger dans le temps les populations s’avèrent nécessaires. Dans la littérature on dispose de peu de données clinique et préclinique sur la prise répétée d’iode stable, quant à son usage ça n’a pas été décrit. La doctrine iode ainsi que l’autorisation de mise sur le marché (AMM) du KI envisage seulement la prise unique à renouveler exceptionnellement chez la population adulte. L’iode est connu pour être un élément clé de la fonction thyroïdienne, on jouant un double rôle à la fois de substrat de régulateur de la thyroïde. Si sa présence est indispensable à la formation des hormones thyroïdienne, son excès exerce un effet inhibiteur transitoire de cette synthèse connu sous le nom de l’effet Wolff-Chaikoff. Les hormones thyroïdiennes jouent un rôle majeur dans le développement et la fonction de presque tous les organes du corps (cerveau, cœur, os...), la moindre variation de leurs niveau peut impacter l’homéostasie du corps. Ainsi, il est difficile d’appliquer la prise répétée du KI en absence connaissances biologiques et toxicologiques. Pour combler ses lacunes et proposer une solution de prophylaxie répétée en cas d’exposition réitérée, le programme de recherche français PRIODAC : PRophylaxie répétée par l’IODe stable en situation ACcidentelle (ANR/RSNR), dont fait partie cette thèse vise à exploiter les modalités d’administration répétées du KI chez toutes les tranches d’âge (in utero, adulte et âgé), et d’évaluer la toxicologie de la prise répétée de KI sur les grandes fonctions physiologiques de l’organisme. Trois modèles de rats Wistar ont fait l’objet de ce travail de thèse : le modèle de référence rat adulte (âgé de 3 mois), le modèle à risque, organisme en développement (exposé durant la gestation) et un autre modèle à risque le rat âgé (âgé de 12 mois), ces trois modèles ont reçus 8 prise consécutive de KI 1mg/kg/24h. Les effets biologiques de ce traitement pendant 8 jours ont été évalués à long-terme (30 jours post-prophylaxie). Concernant, le modèle de référence ont n’a pas observé d’impact néfaste à long-terme de la prise répétée du KI (Lebsir, Cohen et al. 2018; Lebsir, Manens et al. 2018) par contre sur les modèles à risque plusieurs effets à long-terme ont été mis en évidence. Chez la progéniture exposée in utero, la coordination motrice ainsi que l’expression de quelques gènes clés du cerveau ont été négativement modifiées par le traitement. Chez le rat âgé la biochimie clinique, l’expression de quelques gènes clés de la fonction cardiovasculaire ainsi que le système rénine-angiotensine-aldostérone ont été significativement impacté par le traitement. En conclusion, les résultats obtenus montrent l’innocuité sur le plan toxicologique du KI administré à 1mg/kg toutes les 24h pendant 8 jours chez le modèle adulte et la nocuité de ce schéma prophylactique chez les modèles à risque in utero et âgé. Ces résultats ont été communiquées à la pharmacie centrale des armées (producteur et détentrice de l’AMM) afin de servir de données d’entrée pour des études de bonne pratique de laboratoire notamment pour le modèle adulte et également, afin de contribuées à l’évolution de la doctrine de l’iode en terme de radioprotection. / Following nuclear accidents, uranium fission products such as radioactive iodines are released into the environment. Humans are likely to be exposed to these elements mainly through inhalation of air and / or ingestion of contaminated food. Iodine 131 is known to be responsible for increasing the incidence of thyroid cancer. One of the available countermeasures is the ingestion of a single dose of potassium iodide (KI) tablets to saturate the thyroid gland with stable iodine and thus prevent the uptake of the radioactive isotope.Repeated releases of radioactive iodine during the two major accidents Chernobyl and Fukushima have highlighted the weaknesses of this measure, repeated intake of KI maybe necessary. In the literature there is little clinical and preclinical data on the repeated intake of stable iodine, regarding its use it has not been described. The iodine doctrine as well as the Marketing Authorization (MA) of the KI considers only the single taking, to renew exceptionally in the adult population. Iodine is known to be a key component of thyroid function, playing a dual role as both a substrate and a regulator of the thyroid. If its presence is essential for the synthesis of thyroid hormones, its excess exerts a transient inhibitory effect on this synthesis known as the Wolff-Chaikoff effect. Thyroid hormones affect the development and function of almost all organs of the body (brain, heart, bone ...), the slightest variation in their level can impact the homeostasis of the body. Hence, the toughness of applying repeated KI intake in the absence of biological and toxicological data. To fill this gap and find a solution in the event of repeated exposure, the French research program PRIODAC: repeated potassium iodide prophylaxis in accidental situation (ANR / RSNR), of which this thesis is part, aims to define the modalities of repeated administration of KI in all age groups (in utero, adult and elderly), and evaluate the biological consequences on the the body’s major physiological functions. Three models of Wistar rats were used in this work: the reference model adult rat (3 months), the first sensitive model offspring (exposed during gestation) and another sensitive model the older rat (12 months ), these three models received 8 consecutive intake of KI 1mg / kg / 24h. And the effects were assessed at long-term (30 days post-prophylaxis). On the reference model, there was no long-term adverse impact of repeated KI intake (Lebsir, Cohen et al. 2018; Lebsir, Manens et al. 2018). On the other hand, sensitive models reveal several long-term effects; the offspring exhibited impaired motor coordination and variation of the expression of some key brain genes. And in the elderly rat urinary biochemistry, expression of some key genes of the cardiovascular function, as well as the renin-angiotensin-aldosterone system were significantly impacted by the treatment. In conclusion, the results obtained show the toxicological safety of KI administered at 1 mg / kg every 24 hours for 8 days in the adult model, and the harm of this prophylactic scheme in sensitive models in utero and elderly. These results were sent to the French central pharmacy of armies and will serve as input data for good laboratory practice studies that eventually will contribute to the evolution of the iodine doctrine and KI MA.

Les espèces réactives de l’oxygène ont initialement été identifiées comme des produits délétères dérivés du métabolisme aérobie. Il est maintenant accepté que ces espèces sont produites de manière régulée par des enzymes et interviennent dans des fonctions cellulaires telles que la défense immunitaire, la signalisation intracellulaire, la biosynthèse des hormones et la modification de matrice extracellulaire. Les NADPH (Nicotinamide Adénine Dinucléotide Phosphate) oxydases (NOX) forment une famille d’enzymes transmembranaires capables de former de l’anion superoxyde (O2.-) par transfert d’électrons du NADPH à l’oxygène moléculaire (O2). DUOX1 et DUOX2 sont deux des sept membres composant cette famille qui génèrent directement de l’H2O2 comme produit de réduction de l’O2.<p>Initialement clonés à partir de la thyroïde dans notre laboratoire, les ADNc codant pour les protéines DUOX ont été identifiées dans d’autres tissus, comme par exemple la prostate ou l’épithélium respiratoire où DUOX1 est majoritaire. DUOX2 se retrouve également dans les glandes salivaires, dans la muqueuse rectale et tout le long du tractus digestif. D’autre part, un orthologue de DUOX, appelé Udx1, a été identifié en 2004 au niveau de la membrane ovocytaire chez l’oursin. Dans chacun de ces tissus, l’H2O2 produit par les protéines joue un rôle clef. <p>Le mécanisme d’activation de DUOX dans tous ces tissus n’a été identifié que récemment. En effet, pour être exprimé sous forme active à la surface cellulaire, les protéines DUOX nécessitent un facteur de maturation spécifique. Ces facteurs, appelés DUOXA1 et 2 pour « DUOX activator », suivent l’expression tissulaire de leur DUOX respectif. Nous avons montré que la région COOH-terminale de DUOXA1 est responsable de l’activité génératrice d’H2O2 de DUOX1. DUOX2 peut produire de l’H2O2 ou de l’O2.-. L’extrémité NH2-terminale de DUOXA2 est critique dans cette activité et détermine le type de dérivé oxygéné produit. Dans notre système, DUOXA2 n’est pas détecté à la surface cellulaire, sauf en cas de modification de son extrémité amino-terminale par l’addition d’un épitope. DUOXA1 peut être exprimé à la membrane plasmique mais sa présence n’est pas nécessaire au sein du complexe formé avec DUOX pour que ce dernier soit actif. Les facteurs de maturation jouent donc un rôle de protéine chaperonne, induisant la maturation et la translocation d’une protéine DUOX active à la surface cellulaire. <p>Dans la thyroïde, l’H2O2 produit par les protéines DUOX constitue le cofacteur de la thyroperoxydase catalysant l’oxydation de l’iode et le couplage de sa forme oxydée sur des résidus tyrosines de la thyroglobuline menant in fine à la synthèse des hormones thyroïdiennes T3 et T4 et leur relarguage dans la circulation sanguine. Plusieurs mutations dans le gène DUOX2 ont déjà été décrites chez des patients atteints de dyshormonogenèse transitoire ou permanente. Nous avons mis en évidence qu’une inactivation totale de la protéine DUOX2 était compatible avec un état hypothyroïdien peu sévère et transitoire, indiquant l’intervention probable de DUOX1 dans la synthèse des hormones thyroïdiennes. Le défaut génétique identifié est composé d’une délétion génomique partielle d’un allèle associée à une mutation faux-sens (G1518S) sur l’autre allèle du patient hypothyroïdien. Cette mutation, située dans le site catalytique de l’enzyme, mène à une abolition de l’activité de l’enzyme qui est néanmoins exprimée partiellement à la surface cellulaire. <p>Les messagers des DUOX ont été identifiés récemment dans les tractus digestif et respiratoire. Le rôle joué par l’H2O2 dans ces tissus semble avant tout être un rôle de défense contre les micro-organismes en mettant en jeu la lactoperoxydase oxydant le thiocyanate en composé bactéricide actif. Nous avons montré que l’H2O2 produit par DUOX exerce un effet répulsif sur les bactéries. En effet, l’invasion de cellules CHO exprimant de manière stable DUOX2 et DUOXA2 par Salmonella Typhimurium est diminuée lorsque la production d’H2O2 de ces cellules est stimulée. Cet effet répulsif constituerait un rôle primordial pour DUOX au niveau des muqueuses respiratoire et digestive.<p>Lors de la fertilisation, une explosion respiratoire a lieu et de l’H2O2 est produit. Cet H2O2 fourni à l’ovoperoxydase permettrait la formation d’une enveloppe rigide autour de l’ovocyte, bloquant ainsi l’entrée de spermatozoïdes surnuméraires. Ce phénomène a été largement étudié dans l’ovocyte d’oursin, dans lequel la NADPH oxydase responsable de la production d’H2O2 a été caractérisée: il s’agit de Udx1, l’orthologue de DUOX. Chez les mammifères, le phénomène existe mais le mécanisme est en grande partie inconnu. Nous avons montré que les ARNm des DUOX sont exprimés dans l’ovocyte humain ;ceci nous permet d’émettre l’hypothèse que l’inhibition de la polyspermie chez l’homme pourrait être similaire à celle de l’oursin. <p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

Achievement of endothernic capacity is vital for independence from ambient temperature changes, sustained activity, optimal biochemical reactions and optimization of parental care. During early avian development, the core tenets of transition from ectothermy to endothermy are development of metabolic capacity (oxygen consumption, mitochondrial bioenergetics), enhanced cardiovascular function (heart rate and cardiac output), pulmonary ventilation and thermogenic capacity. Thyroid hormones, particularly T3, are key metabolic regulators of basal metabolism, thermogenesis, pulmonary ventilation and mitochondrial respiration. Thyroid hormone fluctuation patterns during both precocial and altricial avian endothermic transition suggest a prominent role in maturation of endothermy, cardiovascular, respiratory and skeletal muscle physiology. This body of work explores effects of T3 manipulations in two avian species: the precocial Pekin duck and the altricial Red-winged Blackbird. Increased plasma T3 during late incubation resulted in increased cardiac mass, elevated resting and intrinsic heart rate, intrinsic mean arterial pressure, increased cholinergic tone and blunted alpha-adrenergic tone in the precocial Pekin duck. In both Pekin duck and Red-winged blackbird, plasma T3 levels correlated with changes in the trajectory of endothermic ontogeny, systemic oxygen consumption, thermogenesis, maturation of pulmonary ventilatory function, altered growth and effects on skeletal and cardiac mitochondrial bioenergetics. These observations support the role of thyroid hormones as metabolic and developmental regulators at the time of attainment of endothermy during the perinatal period in precocial and altricial avian species. Insights into the role of thyroid hormone as a metabolic and development regulator at the time of avian endothermic attainment provide a more thorough understanding of metabolic and physical transitions a hatchling bird must undergo to reach the adult endothermic phenotype. Such insights also deepen understanding of the complex role thyroid hormones play in homeostasis and offer implications about the evolutionary history of endothermic capacity.

La couleur est un trait biologique essentiel et très variable qui permet à un organisme d'être adapté à son environnement biotique et abiotique. De multiples gènes influent sur la couleur ; leur évolution a été étudiée ici chez les Vertébrés en lien avec les duplications de génome survenues dans cette lignée . Alors que l'essentiel des gènes sont perdus lors de la rediploïdisation suivant une duplication totale de génome, ces travaux montrent que les gènes de la pigmentation appartiennent à des fa milles de gènes qui sont en moyenne plus retenues que le reste du génome . Ceci est particulièrement vrai pour les poissons Téléostéens, cette expansion du répertoire de gènes de pigmentation pouvant être en lien avec la grande diversité de cellules pigmenta ires et de patrons de pigmentation observés chez ces animaux . Par ailleurs, l'étude de la pigmentation d'un organisme modèle émergent, le poisson -clown Amphiprion oellaris, a été réalisée dans le cadre de cette thèse . La nature cellulaire de la bande blanche de ce poisson a été déterminée et a permis l'identification - d'un nouveau gène présent presque exclusivement chez les poissons Actinoptérygiens , probablement acquis par transfert horizontal, et impliqué dans le développement d'un type de cellules pigmentaires, les iridophores . Enfin, l'acquisition de la pigmentation au cours du développement du poisson -clown A. ocellaris, un poisson corallien au cycle de développement marqué par une métamorphose, a été étudiée en rapport avec la signalisation thyroïdienne, qui joue un rôle majeur au cours du développement des Vertébrés . / Color is a key biological trait that allows individuals to adapt to their biotic and abiotic environment, and many genes have been shown to play an essential role in color acquisition in vertebrates. In this work, the consequences on this specifie set of genes of the whole-genome duplications that occurred during vertebrate evolution were investigated. Whereas most genes a re lost after a w hole- genome duplication event, this work shows that pigmentation genes and other genes within their families have been more retained compared to the average observed genome retention. This was particularly the case for the teleost-specific whole-genome duplications . Hence, this high retention rate could  be related to the high pigmentation diversity observed in the teleost fish lineage, both  in terms of patterns and of pigment cell types . In addition, during this PhD thesis, pigmentation in an emerging model organism, the clownfish Amphiprion ocellaris, was investigated. The white color in clownfish was shawn to be due to specifie pigment cells,  iridophores, and new iridophore genetic markers were detected . One such marker is almost exclusively restricted to ray-finned fish and could have been acquired through a horizontal gene transfer event that occurred at the onset of the evolution of this lineage. Last, in this work, the genetic basis of pigmentation acquisition was scrutinized during the development of clownfish A. ocellar is, and especially during metamorphosis - a major life history transition  in coral  reef  fish . In particular, the  deployment  during  development  of thyroid  hormone signaling  which is a key pathway in metamorphosis regulation in vertebrates  was examined.

O presente estudo avaliou o papel do receptor AT1 de Angiotensina II no desenvolvimento da hipertrofia dos cardiomiócitos promovida pelo T3, bem como a participação dos mecanismos intracelulares deflagrados pelo receptor AT1 neste modelo de hipertrofia cardíaca. O silenciamento do receptor AT1 com RNA de interferência preveniu totalmente o desenvolvimento da hipertrofia dos cardiomiócitos induzida pelo T3. Os cardiomiócitos tratados com T3 demonstraram uma rápida ativação da via da Akt/GSK-3/mTOR, a qual foi atenuada ou prevenida pelo silenciamento do receptor AT1. Ainda, a expressão de Angiotensina I/II no lisado celular e a expressão do receptor AT1 foram rapidamente aumentados pelo T3. Esses dados demonstram pela primeira vez que o receptor AT1 é um mediador crítico da hipertrofia dos cardiomiócitos induzida pelo T3, bem como para a ativação da via da Akt, sugerindo que a via Ang I/II-AT1-Akt/GSK-3/mTOR corresponde a um potencial mediador dos efeitos tróficos exercidos pelo T3 nessas células. / The present study investigated the role of Angiotensin type 1 receptor (AT1R) in T3-induced cardiomyocyte hypertrophy, as well as the participation of the intracellular mechanisms mediated by AT1R in this cardiac hypertrophy model. The AT1R silencing using small interfering RNA totally prevented the development of T3-induced cardiomyocyte hypertrophy. The cardiomyocytes treated with T3 demonstrated a rapid activation of Akt/GSK-3/mTOR signaling pathway, which was attenuated or prevented by the AT1R silencing. In addition, local Angiotensin I/II (Ang I/II) levels and the AT1R expression were rapidly increased by T3 treatment. These data demonstrate for the first time that the AT1R is a critical mediator to the T3-induced cardiomyocyte hypertrophy, as well as to the activation of the Akt signaling, suggesting that the Ang I/II-AT1R-Akt/GSK-3/mTOR pathway corresponds to a potential mediator of the trophic effect exerted by T3 in cardiomyocytes.

O hormônio tireoidiano (HT) é essencial para o desenvolvimento, maturação e metabolismo ósseos, enquanto que o sistema nervoso simpático (SNS) é, também, um potente regulador do remodelamento ósseo. Demonstrou-se que SNS regula negativamente a massa óssea, agindo via receptores ?2-adrenérgicos (?2-AR), expressos em osteoblastos. O nosso grupo demonstrou que os receptores ?2 adrenérgicos (?2-AR) também medeiam ações do SNS no esqueleto e que são expressos em osteoblastos, osteócitos, condrócitos e osteoclastos. Considerando-se que o HT interage com o SNS para regular uma série de processos fisiológicos, e que o excesso de HT e a ativação do SNS causam perda de massa óssea, levantamos a hipótese de que há interação entre o HT com o SNS para regular a massa óssea. Estudos do nosso grupo vêm sustentando essa hipótese, uma vez que camundongos com inativação gênica dos receptores beta2-AR apresentam resistência à osteopenia induzida por doses tóxicas de HT. Considerando-se, ainda, que a interação do HT com o SNS em vários tecidos e/ou órgãos depende da sinalização beta2 adrenérgica, o presente estudo teve como objetivo avaliar se a interação do HT com o SNS para regular a morfofisiologia óssea envolve o beta2-AR. Para tanto, estudamos o efeito de 10x e 20x a dose fisiológica de triiodotironina (3,5ug ou 7.0ug de T3/100g de massa corporal/dia, respectivamente), por 90 dias, na microaquitetura óssea e em parâmetros biomecânicos do fêmur de camundongos com inativação gênica do beta2-AR (beta2-AR-/-), e nos seus respectivos Selvagens (Selv), os camundongos da linhagem FVB. Como esperado, o tratamento com T3 promoveu efeitos deletérios na microarquitetura trabecular das fêmeas Selv, enquanto alguns desses efeitos foram mais brandos ou inexistentes nos animais beta2-AR-/-, revelando resistência do osso trabecular dos animais knockout (KO) aos efeitos deletérios da tireotoxicose. Em contraste, a microarquitetura femoral dos camundongos machos beta2-AR-/- se mostrou mais sensível aos efeitos deletérios da tireotoxicose, em relação aos respectivos Selv. Quanto ao osso cortical femoral, vimos que o tratamento com T3 aumentou o perímetro endosteal e a área medular nos animais Selv machos e fêmeas, mas não nos animais beta2-AR-/-, o que sugere que o T3 promove reabsorção óssea endosteal no osso cortical, em um mecanismo que depende da via de sinalização do beta2-AR. Vimos, ainda, que o tratamento com T3 causou reduções significativas na carga máxima, tenacidade, rigidez e resiliência do fêmur dos camundongos fêmeas Selv. Em contraste, nenhum desses parâmetros biomecânicos foi afetado pelo tratamento com T3 no fêmur das fêmeas KO, evidenciando, mais uma vez, uma resistência desses animais aos efeitos deletérios da tireotoxicose no tecido ósseo. Por outro lado, os camundongos machos Selv e KO se mostraram resistentes aos efeitos deletérios do tratamento com T3 sobre os parâmetros biomecânicos do fêmur, sugerindo a participação de fatores sexuais na interação do HT com o SNS para regular a morfofisiologia óssea. Em conjunto, os achados do presente estudo corroboram a hipótese de que o HT interage com o SNS através da via dos receptores beta2 adrenérgicos para regular a morfofisiologia óssea, especialmente em fêmeas e no osso cortical / Thyroid hormone (TH) is essential for bone development, maturation and metabolism, while the sympathetic nervous system (SNS) is also a potent regulator of bone remodeling. SNS has been shown to negatively regulate bone mass, acting via beta2-adrenergic (beta2-AR) receptors expressed in osteoblasts. Our group demonstrated that alpha2-adrenergic (alpha2-AR) receptors also mediate SNS actions in the skeleton and are expressed in osteoblasts, osteocytes, chondrocytes and osteoclasts. Considering that TH interacts with the SNS to regulate a series of physiological processes, and that the excess of TH and the activation of the SNS cause loss of bone mass, we hypothesize that there is interaction between TH and the SNS to regulate the bone mass. Studies of our group have supported this hypothesis, since mice with gene inactivation of alpha2-AR present resistance to the osteopenia induced by toxic doses of TH. Considering that the TH-SNS interaction in various tissues and/or organs depends on beta2-adrenergic signaling, the present study aimed to evaluate whether the interaction of TH with the SNS to regulate the bone morphophysiology involves beta2- AR. Therefore, we studied the effect of 10x and 20x the physiological dose of triiodothyronine (3.5ug or 7.0ug of T3/100g body mass/day, respectively), for 90 days, in the bone microarchitecture and biomechanical parameters of the femur mice with beta2-AR gene inactivation (beta2-AR-/-), and of their respective Wild-type (WT) controls, the FVB lineage mice. As expected, T3 treatment promoted deleterious effects on the trabecular microarchitecture of the WT females, while some of these effects were milder or nonexistent in beta2-AR-/- animals, revealing trabecular bone resistance of knockout (KO) animals to the deleterious effects of thyrotoxicosis. In contrast, the femoral microarchitecture of the male beta2-AR-/- mice was more sensitive to the deleterious effects of thyrotoxicosis, in relation to the respective WT animals. Regarding to the femoral cortical bone, we saw that T3 treatment increased the endosteal perimeter and the medullary area both male and female WT animals, but not in the beta2-AR-/- mice, suggesting that T3 promotes endosteal bone resorption in the cortical bone, in a mechanism that depends on the alpha2-AR signaling pathway. We also found that treatment with T3 caused significant reductions in the maximum load, tenacity, stiffness and resilience of femurs of the WT female mice. In contrast, none of these biomechanical parameters was affected by T3 treatment in the KO females, demonstrating again resistance of these animals to the deleterious effects of thyrotoxicosis on bone tissue. On the other hand, WT and KO male mice were resistant to the deleterious effects of T3 treatment on the biomechanical parameters of the femur, suggesting the participation of sexual factors in the interaction of HT with the SNS to regulate bone morphophysiology. Taken together, the findings of the present study corroborate the hypothesis that TH interacts with the SNS through the beta2 adrenergic receptor pathway to regulate bone morphophysiology, especially in females and cortical bone

Níveis elevados de hormônios tireoidianos (HTs) são comumente associados à ativação do sistema renina angiotensina local e ao desenvolvimento da hipertrofia cardíaca. O envolvimento do receptor de angiotensina II tipo 1 (AT1R) nos efeitos hipertróficos dos HTs fora descrito previamente. No entanto, os mecanismos subjacentes a essa interação ainda são desconhecidos. O AT1R pertence à família dos receptores acoplados à proteína G e, portanto, promove a transdução de sinal por mecanismos dependentes e independentes de proteína G. Recentemente, a sinalização dependente de beta-arrestinas (independente de proteína G) tem sido descrita por contribuir com a resposta hipertrófica em diferentes modelos experimentais. Assim, no presente estudo investigou-se o envolvimento da sinalização dependente de beta-arrestinas nos efeitos hipertróficos dos HTs, mediados pelo AT1R, bem como a participação de ERK&#189; nesse processo. Culturas primárias de cardiomiócitos foram estimuladas com T3 (triiodotironina; 15nM) para indução da hipertrofia. O tratamento dos cardiomiócitos com T3 por tempos rápidos (5-30 min) resultou na ativação transiente de ERK&#189;, a qual foi parcialmente atenuada quando da administração de Losartan (1&#181;M), antagonista do AT1R. A contribuição de ERK&#189; na hipertrofia dos cardiomiócitos foi verificada através do uso de PD98059 (20&#181;M), inibidor de MEK&#189;, o qual preveniu a transcrição de marcadores hipertróficos. Ensaios de imunoprecipitação revelaram o aumento da interação entre AT1R e beta-arrestina 2 sob estímulo do T3, sugerindo o recrutamento de beta-arrestina 2 e, possível, internalização do AT1R. Através de ensaios de imunofluorescência e fracionamento subcelular, foi demonstrado que o T3 estimula a translocação do AT1R, amentando sua expressão no núcleo dos cardiomiócitos. Além disso, tanto a ativação de ERK&#189; quanto a hipertrofia cardiomiocítica mostraram-se sensíveis à inibição da endocitose, a qual foi avaliada através de Concanavalina A (0,5&#181;g/ml). Ensaios de silenciamento gênico por RNA de interferência foram eficientes em demonstrar o envolvimento de beta-arrestina 2 na ativação de ERK&#189; e na hipertrofia cardiomiocítica induzida por T3. Desta forma, os resultados evidenciam o envolvimento da sinalização dependente de beta-arrestina 2 na ativação de ERK&#189;, através do AT1R, a qual contribui com a hipertrofia cardiomiocítica promovida pelo T3. / Elevated levels of thyroid hormones (THs) are commonly associated with activation of the local renin angiotensin system and the development of cardiac hypertrophy. The involvement of the angiotensin II receptor type 1 (AT1R) in the hypertrophic effects of the THs was previously described. However, the mechanisms underlying this interaction are still unknown. AT1R belongs to the G-protein coupled receptor family and promotes its signal transduction by G-protein dependent and independent mechanisms. Recently, beta-arrestin signaling (G-protein independent) has been described as contributing to the hypertrophic response in different experimental models. Thus, the present study investigated the involvement of beta-arrestin signaling in the hypertrophic effects of THs mediated by AT1R, as well as the participation of ERK&#189; in this process. Primary cardiomyocytes cultures were stimulated with T3 (triiodothyronine; 15nM) for the induction of hypertrophy. Cardiomyocytes acutely treated with T3 (5-30 min) resulted in transient activation of ERK&#189;, which was partially attenuated upon Losartan (1&#181;M) administration, an AT1R antagonist. The contribution of ERK&#189; to cardiomyocyte hypertrophy was verified by using PD98059 (20&#181;M), a MEK&#189; inhibitor, which prevented the transcription of hypertrophic markers. Immunoprecipitation assays revealed increased interaction between AT1R and beta-arrestin 2 under T3 stimulation, suggesting the recruitment of beta-arrestin 2 and, possibly, the internalization of AT1R. Through immunofluorescence and subcellular fractionation assays, T3 has been shown to stimulate AT1R translocation, enhancing its expression in the cardiomyocyte nucleus. In addition, both ERK&#189; activation and cardiomyocyte hypertrophy were sensitive to the inhibition of endocytosis, which was assessed by Concanavalin A (0.5&#181;g/ml). Interfering RNA assays were efficient in demonstrating the involvement of beta-arrestin 2 in ERK&#189; activation and in T3-induced cardiomyocyte hypertrophy. Therefore, the results evidenced the involvement of beta-arrestin-2-dependent signaling in the activation of ERK&#189;, through the AT1R, which contributes to the cardiomyocyte hypertrophy promoted by T3.

No coração, doses elevadas de hormônio tireoideano (T3) por tempo prolongado promove hipertrofia cardíaca. Os mecanismos envolvidos neste processo necessitam de maior esclarecimento. Analisou-se dados de um ensaio de microarray de tecido cardíaco de ratos submetidos a hipertireoidismo experimental. O algoritmo MAS5 foi mais eficiente para processamento dos dados. Identificou-se os filamentos grossos, banda M e discos intercalares como hotspots de atuação do T3. A T-Caderina apresentou aumento transitório nos níveis de mRNA e proteicos sob efeito do T3. Estudo de imunofluorescência evidenciou marcação para T-Caderina próxima à membrana plasmática de cardiomiócitos. Com 24 horas de tratamento com T3, observamos aumento global e difuso de marcação para T-Caderina. Obeservou-se marcação nuclear para T-Caderina. Portanto, é possível que a T-caderina possa estar envolvida no processo de hipertrofia cardíaca. Todavia, para verificar essa possibilidade, são necessários mais estudos. / Cardiac hypertrophy is observed in response to long-term hyperthyroidism. The molecular basis of cardiac hypertrophy induced by hyperthyroidism remains to be determined. Using microarray approach, the gene expression profile of heart tissue from rats submitted to hyperthyroidism were analysed. MAS5 were found to be the best for our low-level analysis. Sarcomeric hotspots such as thick-filaments, M-band and intercalated disks under thyroid hormone (T3) treatment were identified. T3 induced transient mRNA and protein levels of T-Cadherin, a interecalated disks member. T-Cadherin were observed next to plasmatic membrane on immunofluorescence analysis. On 24 hours group, diffuse cytoplasmic T-Cadherin staining were evident. Another interesting aspect was T-Cadherin nuclear staining in all groups. Moreover, T-Cadherin possibly play role in T3-induced cardiac hypertrophy. However further studies are needed to verify this possibility.

Disfunções da glândula tireóide são, frequentemente, associadas a manifestações cardiovasculares e, em situações de hipertireoidismo, o coração hipertrofia. A hipertrofia cardíaca (HC) consiste em uma resposta adaptativa caracterizada pelo aumento de síntese de proteínas estruturais. O Sistema Ubiquitina Proteassoma (UPS) corresponde ao principal mecanismo de proteólise intracelular e crescentes evidências sugerem seu envolvimento no desenvolvimento da HC. O objetivo do presente estudo foi avaliar a modulação do UPS no tecido cardíaco de animais submetidos ao hipertireoidismo. Os resultados referentes ao aumento da atividade e expressão do proteassoma (PT) cardíaco apresenta-se mais contundente no grupo tratado por 7 dias, período em que a HC já encontra-se estável. Ao término de 14 e 21 dias, a modulação desse sistema tende à normalização. Os resultados obtidos atestam evidências da literatura que sugerem o aumento da atividade do PT cardíaco como resposta compensatória ao aumento de síntese proteica. / Thyroid gland disorders are often associated with cardiovascular events and hyperthyroidism state promotes cardiac hypertrophy (CH). CH consists in adaptive response characterized by increased synthesis of structural proteins. The Ubiquitin Proteasome System (UPS) is the major mechanism of intracellular proteolysis and increased evidences suggest its involvement in the development of CH. The aim of this study was to evaluate the modulation of UPS in cardiac tissue of animals subjected to hyperthyroidism. The results related to the increased proteasome (PT) activity and expression in the heart was more accentuated in the group treated for 7 days, when the CH process finds stable. At the end of 14 and 21 days of hyperthyroidism, the modulation of cardiac UPS achieves standard values. These results suggest an increased activity of cardiac PT as a compensatory response to protein synthesis induced by thyroid hormones.

Objetivo: Descrever características biológicas e ambientais associadas a habilidades motoras finas, medidas por diferentes níveis de conduta visual em lactentes de 1 e 3 meses. Desenho de estudo: 82 díades mãe-bebê foram recrutadas durante consultas pré-natais ou imediatamente após o nascimento e visitadas duas vezes para a coleta de dados (25-40 e 85-100 dias após o nascimento). Durante as visitas foram coletados fatores maternos (idade materna, escolaridade, tipo de parto, número de consultas pré-natais, tipo de amamentação e tabagismo), características biológicas do bebê (gênero, idade gestacional, nascimento, peso, circunferência da cabeça, Apgar), testes motores infantis (através da Escala de Avaliação do Comportamento Visuomotor Infantil e Alberta Infant Motor Scale/AIMS), perfil psicológico materno (através da Escala de Depressão Pós-natal de Edimburgo/EPDS e Escala de Ansiedade de Hamilton), variáveis ambientais (medidas pelo Affordance in the home environment for development scale/AHEMD), cuidados maternos (avaliado através do Coding Interacting Behavior/CIB), bem como a coleta de leite (medição de ácidos graxos totais, proteínas e cortisol) e sangue materno (medição de hormônios séricos e interleucinas). Resultados: 51 crianças foram testadas em tarefas de conduta visual e outras medidas. As mães das crianças com baixos escores de fixação visual apresentaram maiores níveis de proteína no leite materno aos 3 meses. Quanto ao perfil metabólico materno, as mães das crianças que apresentaram melhores escores de fixação visual apresentaram melhores níveis séricos de T4 (no primeiro mês) e prolactina (no terceiro mês). Conclusão: O desenvolvimento neuromotor precoce do bebê, especialmente as habilidades visuais e de motricidade fina, estão intimamente associados aos fatores biológicos maternos (fatores metabólicos maternos e composição do leite materno). / Objective: To describe biological and environmental characteristics associated with fine motor skills measured by different levels of visual tracking in infants of 1 and 3 months. Study design: 82 mother-infant dyads were recruited during prenatal consultations or immediately after birth and visited at two times for data collection (25-40 and 85-100 days after birth). During the visits were collected maternal factors (maternal age, education level, type of delivery, number of antenatal consultations, type of breastfeeding and smoking), biological characteristics of the baby (gender, gestational age, birth, weight, head circumference, Apgar), infant motor tests (through the Child Visuomotor Behavior Rating Scales and Alberta Infant Motor Scale/AIMS), maternal psychological profile (through the Edinburgh Postnatal Depression Scale/EPDS and Hamilton Anxiety Scale), environmental variables (measured by the Affordance in the home environment for development scale/AHEMD), maternal care (evaluated through Coding Interacting Behavior/CIB), as well as milk collection (measurement of total fatty acids, proteins and cortisol) and maternal blood (measurement of serum hormones and interleukins). Results: 51 children were tested on visual tracking tasks and other measures. The mothers of children with low visual fixation scores presented higher levels of protein in breastmilk at 3 months. Regarding the maternal metabolic profile, the mothers of the children who presented better visual conduct scores had better serum levels of T4 (in the first month) and prolactin (in the third month). Conclusion: Early neuromotor development of the baby, especially the visual and fine motor skills, are closely associated with maternal biological characteristics (metabolic factors and composition of breast milk).