Thèse de doctorat en Sciences : Biologie cellulaire et physiologie
Sous la direction de Sergueï Fetissov.
Soutenue en 2015
à Rouen , dans le cadre de École doctorale Normande de biologie intégrative, santé, environnement (Mont-Saint-Aignan, Seine-Maritime) , en partenariat avec TC2N, Trans Channel neuroscience network, Normandy, Rouen, France (laboratoire) et de IRIB, Institut for research and innovation in biomedicine, Normandy (Rouen) (laboratoire) .
Ghrelin is 28 amino acid peptide hormone produced mainly in the stomach that was discovered in 1999. After its release in the general circulation, ghrelin reaches its peripheral and central targets simulating food intake, adiposity, and modulating emotions. Because of its involvement in numerous functions that are deregulated in anorexia and obesity, ghrelin has been extensively studied in these disorders. It has been established that plasma levels of ghrelin are increased during conditions of chronic starvation, such as in patients with anorexia nervosa, but are low or norma; during conditions of positive energy baiance, such as in obese patients. However, it is not known whether such changes of ghrelin levels are related to the number of preproghrelin mRNA-expressing cells in the stomach. Thus, in this thesis, we investigated if circulating ghrelin levels are associated with the number of ghrelin mRNA-producing cells revealed by in situ hybridization (ISH), in several animal models of energy imbalance including: mice with diet-induced obesity (D10), leptin-deficient obese (ob/ob) mice, mice with activity-based anorexia (ABA) and food restriction as well as in methotrexate (MIX) chemotherapy-induced anorexia in rats. We found that plasma concentrations of ghrelin were not directly related to the number of preproghrelin mRNA-expressing cells in obese mice and MTX-treated rats. Only ABA and food-restricted mice showed an increased number of preproghrelin mRNA-expressing cells in the stomach. In contrast, this number was decreased in ob/ob but unchanged in D1O mice. In MTX-treated anorectic rats increased number of ghrelin-producing cells was not accompanied by increased plasma ghrelin. We also investigated possible presence of a central source of ghrelin, using ISH and imrriunohistochemistry in normal conditions and during chronic starvation. No solid evidence was found for ghrelin expression in the hypothalamus of rodents in any of the experimental models. Furthermore, to study other possible mechanisms involved in the modulation of plasma levels of ghrelin in these animal models, we measured plasma levels of ghrelin-reactive autoantibodies. Indeed, after its release, plasma ghrelin is protected from degradation by ghrelin-reactive immunoglobulins (Ig). Ghrelin-reactive IgG were characterized by increased affinity in both models of obesity and were found at lower levels in plasma but were increased in the hypothalamus, suggesting that they may enhance the transportation of the peptide from plasma to its central targets. Furthermore, after an acute stress, ghrelin was increased in plasma of some lean mice but not in obese mice, and ghrelin-reactive IgG tended to be lower in plasma but increased in the hypothalamus of lean mice while remaining unchanged in obese mice. Thus, different stress-induced changes of plasma ghrelin and ghrelin-reactive IgG in obese mice show that altered ghrelin's response to stress may characterize the obesity phenotype. MTX strongly decreased the production of ghrelin-reactive IgG, which low levels may compromise ghrelin's stability, potentially contributing to low active ghrelin ratio, anorexia and anxiety observed in MIX-treated rats. Finally, we investigated if ghrelin-reactive Ig are associated with anxiety, depression, stress-induced cortisol response and anti-viral antibodies in a general population of adolescents. Serum levels of ghrelin-reactive autoAbs showed weak correlations with anxiety and stress-response in girls suggesting that they may not be important regulators of stress and emotions in healthy individuals. However, strong correlations with influenza A and B antibodies were observed suggesting that production of ghrelin-reactive autoAbs could be influenced by viral infections and should be further studied in patients with anxiety and depression. In conclusion, this work further characterize the ghrelin system in animal models of anorexia and obesity, in particular by providing, for the first time, the quantitative analysis of preproghrelin mRNA-expressing cells by ISH. It shows that plasmatic levels of ghrelin are not always directly associated with the number of preproghrelin-expressing cells in the stomach suggesting involvement of other mechanisms possibly interfering with ghrelin stability. Moreover, a deficient stress-induced response of ghrelin release in obese animals may contribute to the mechanisms of stress-related disorders such as anxiety. Our work further supports a role of ghrelin-reactive IgG in ghrelin signaling, in particular, their low production in animals with anorexia but increased affinity in animal models of obesity suggests that they may contribute to the ghrelin's orexigenic role, and hence, can be considered as a new therapeutic target in anorexia and obesity.
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