Identification des bases génétiques des myopathies à multi-minicores avec ou sans cardiomyopathie

Thèse réalisée en cotutelle avec l'Université Pierre et Marie Curie, Paris 6(UPMC, Paris, France).

Myostatin regulates cardiomyocyte growth through modulation of Akt signaling

Myostatin is a highly conserved, potent negative regulator of skeletal muscle hypertrophy in many species, from rodents to humans, although its mechanisms of action are incompletely understood. Transcript profiling of hearts from a genetic model of cardiac hypertrophy revealed dramatic upregulation of myostatin, not previously recognized to play a role in the heart. Here we show that myostatin abrogates the cardiomyocyte growth response to phenylephrine in vitro through inhibition of p38 and the serine - threonine kinase Akt, a critical determinant of cell size in many species from drosophila to mammals. Evaluation of male myostatin-null mice revealed that their cardiomyocytes and hearts overall were slightly smaller at baseline than littermate controls but exhibited more exuberant growth in response to chronic phenylephrine infusion. The increased cardiac growth in myostatin-null mice corresponded with increased p38 phosphorylation and Akt activation in vivo after phenylephrine treatment. Together, these data demonstrate that myostatin is dynamically regulated in the heart and acts more broadly than previously appreciated to regulate growth of multiple types of striated muscle.

CHANGES OF GENE EXPRESSION IN ELECTRICALLY STIMULATED AND CONTRALATERAL RAT SOLEUS MUSCLES

In this study we investigate the effect of a single session of high-intensity contractions on expression of pleiotropic genes and, in particular, those genes associated with metabolism in soleus muscle from electrically stimulated (ES) and contralateral (CL) limbs. The right limbs of male Wistar rats were submitted to contractions by 200-ms trains of electrical stimulation at 100-Hz frequency with pulses of 0.1 ms (voltage 24 3 V) delivered each second for 1 hour. Soleus muscles were isolated 1 hour after contraction, and gene expression was analyzed by a macroarray technique (Atlas Toxicology 1.2 Array; Clontech Laboratories). Electrical stimulation increased expression in 92 genes (16% of the genes present in the membrane). Sixty-six genes were upregulated in both ES and CL soleus muscles, and expression of 26 genes was upregulated in the ES muscle only. The most altered genes were those related to stress response and metabolism. Electrical stimulation also raised expression of transcription factors, translation and posttranslational modification of proteins, ribosomal proteins, and intracellular transducers/effectors/modulators. The results indicate that a single session of electrical stimulation upregulated expression of genes related to metabolism and oxidative stress in soleus muscle from both ES and CL limbs. These findings may indicate an association with tissue hypertrophy and metabolic adaptations induced by physical exercise training not only in the ES but also in the CL non-stimulated muscle, suggesting a cross-education phenomenon. Muscle Nerve 40: 838-846, 2009

Deciphering the role of the electrostatic interactions in the alpha-tropomyosin head-to-tail complex

Skeletal alpha-tropomyosin (Tm) is a dimeric coiled-coil protein that forms linear assemblies under low ionic strength conditions in vitro through head-to-tail interactions. A previously published NMR structure of the Tin head-to-tail complex revealed that it is formed by the insertion of the N-terminal coiled-coil of one molecule into a cleft formed by the separation of the helices at the C-terminus of a second molecule. To evaluate the contribution of charged residues to complex stability, we employed single and double-mutant Tm fragments in which specific charged residues were changed to alanine in head-to-tail binding assays, and the effects of the mutations were analyzed by thermodynamic double-mutant cycles and protein-protein docking. The results show that residues K5, K7, and D280 are essential to the stability of the complex. Though D2, K6, D275, and H276 are exposed to the solvent and do not participate in intermolecular contacts in the NMR structure, they may contribute to head-to-tail complex stability by modulating the stability of the helices at the Tm termini.

Congenital fistulae of the lower lip in Van der Woude syndrome: A histomorphological study

Objective: To evaluate a significant number of cases of fistulae of the lower lip with the light microscope in order to investigate the histological variation and to attempt to define the histopathological pattern of the lesion.Methods: Congenital fistulae of the lower lips of 17 patients with Van der Woude syndrome were analyzed by light microscopy,Results and Conclusion: the walls of most of the fistulae consisted of stratified nonkeratinized squamous epithelium and a lamina propria of dense connective tissue with areas of lymphohistiocytic inflammatory infiltrates. Bundles of striated muscle fibers, blood vessels, nerves, adipose tissue, and mixed acinar glands were observed. These glands surrounded the entire wall of the lesion, and their excretory ducts opened into the lumen of the fistula, explaining the clinical observation of elimination of mucous secretion through the opening of the fistula, Two microform cases of fistula were analyzed that presented only a depression in the epithelium at the site corresponding to the opening of the fistula.

Staining methods applied to glycol methacrylate embedded tissue sections

The use of glycol methacrylate (GMA) avoids some technical artifacts, which are usually observed in paraffin-embedded sections, providing good morphological resolution. on the other hand, weak staining have been mentioned during the use of different methods in plastic sections. In the present study, changes in the histological staining procedures have been assayed during the use of staining and histochemical methods in different GMA-embedded tissues.Samples of tongue, submandibular and sublingual glands, cartilage, portions of respiratory tract and nervous ganglion were fixed in 4% formaldehyde and embedded in glycol methacrylate. The sections of tongue and nervous ganglion were stained by H&E. Picrosirius, Toluidine Blue and Sudan Black B methods were applied, respectively, for identification of collagen fibers in submandibular gland, sulfated glycosaminoglycans in cartilage (metachromasia) and myelin lipids in nervous ganglion. Periodic Acid-Schiff (PAS) method was used for detection of glycoconjugates in submandibular gland and cartilage while AB/PAS combined methods were applied for detection of mucins in the respiratory tract. In addition, a combination of Alcian Blue (AB) and Picrosirius methods was also assayed in the sublingual gland sections.The GMA-embedded tissue sections showed an optimal morphological integrity and were favorable to the staining methods employed in the present study. In the sections of tongue and nervous ganglion, a good contrast of basophilic and acidophilic structures was obtained by H&E. An intense eosinophilia was observed either in the striated muscle fibers or in the myelin sheaths in which the lipids were preserved and revealed by Sudan Black B. In the cartilage matrix, a strong metachromasia was revealed by Toluidine Blue in the negatively-charged glycosaminoglycans. In the chondrocytes, glycogen granules were intensely positive to PAS method. Extracellular glycoproteins were also PAS positive in the basal membrane and in the region occupied by the lamina externa and reticular fibers surrounding each smooth muscle cells of the blood vessels. In the epithelial cells of the respiratory tract, acid and neutral mucins were histochemically detected by AB and PAS methods, respectively. Moreover, granules containing acid and neutral mucins were revealed in purple by AB and PAS concomitantly. In the sublingual gland sections, a distinct affinity of acid mucins by AB (in turquoise-blue) and collagen fibers by Picrosirius (in red) was obtained when these methods were combined. Although some routine dyes used in paraffin sections have showed a weak stain in historesin sections, our results showed that different dyes could be applied in GMA sections if modified staining procedures were assayed. Therefore, appropriate staining contrast and, thus, detection of one or different substances in a same section can be acquired in association to the good morphological resolution provided by GMA. (C) 2003 Elsevier Ltd. All rights reserved.

Structure of the posthepatic septum and its influence on visceral topology in the tegu lizard, Tupinambis merianae (Teiidae : Reptilia)

The posthepatic septum (PHS) divides the body cavity of Tupinambis merianae into two parts: the cranial one containing the lungs and liver and the caudal one containing the remaining viscera. The PHS is composed of layers of collagenous fibers and bundles of smooth muscle, neither of which show systematic orientation, as well as isolated blood vessels, lymphatic vessels, and nerves. Striated muscle of the abdominal wall does not invade the PHS. The contractions of the smooth muscles may stabilize the pleurohepatic cavity under conditions of elevated aerobic needs rather than supporting breathing on a breath-by-breath basis. Surgical removal of the PHS changes the anatomical arrangement of the viscera significantly, with stomach and intestine invading the former pleurohepatic cavity and reducing the space for the lungs, Thus, the PHS is essential to maintain the visceral topography in Tupitionibis. J. Morphol. 258:151-157, 2003. (C) 2003 Wiley-Liss. Inc.

A morphological and ultrastructural study of the paracloacal (Scent) glands of the marsupial Metachirus nudicaudatus Geoffroy, 1803

Morphological and ultrastructural features of the paracloacal glands of Metachirus nudicaudatus are described. Two pairs of glands, one on the right and the other on the left of the anal canal, are formed, each consisting of a major and a minor portion. Their wall is made up of three layers: a mucosal, a muscular and an external capsule. The inner one is a mucosa the epithelium of which contains holocrine cells characterized by lipid droplets and intermediate filaments. The surrounding vascular lamina propria contains flattened tubular apocrine glands whose epithelial cells contain abundant endoplasmic reticulum, prominent Golgi complexes and numerous secretory granules. The middle layer is formed by skeletal striated muscle and the outer (third layer) consists of dense connective tissue. Each gland originates from a single duct. Transverse sections show that each duct, except in the female major gland, is in fact formed by a duct system. One of these ducts comes from the central cavity, lined by holocrine epithelium, and the others result from the branched tubular glands of the lamina propria.

Efeitos do tratamento oral com sulfato de vanadila em ratos diabeticos jovens

This work intends to evaluate the effects of oral vanadyl treatment (VOSO 4, 1 mg/mL) in young streptozotocin-diabetic rats during 19 and 29 days. In several times of treatment the rats were monitored to determine body weight, food and water intakes, glycemia, and the urinary excretion of glucose and urea. The animals were killed in the 19(th) and 29(th) days, and the glycemia level was determined again, as well as the weight of pancreas, muscles (Soleus and Extensor digitorum longus - EDL) and adipose tissues (epididymal and retroperitoneal). The results showed that the treatment of young diabetic rats with VOSO 4 promotes the reduction of hyperglycemia (p < 0.01), food (p < 0.01) and water intakes (p < 0.05) and body weight (p < 0.05). Neither the tissues and pancreas weights nor the urinary urea level of the treatment group varied in comparison to the control group. In conclusion, the vanadyl treatment in the studied period is able to reduce the main metabolic alterations often found in diabetes. These data are very useful and important for the future experiments to verify the effects of vanadyl sulfate on muscle protein metabolism in diabetic rats.

Histoenzymology and morphometry of the masticatory muscles of tufted capuchin monkey (Cebus apella Linnaeus, 1758)

Samples of the anterior and posterior regions of the masseter and temporal muscles and of the anterior belly of the digastric muscle of 4 adult male tufted capuchin monkeys (Cebus apella) were removed and stained with HE and submitted to the m-ATPase reaction (with alkaline and acid preincubation) and to the NADH-TR and SDH reactions. The results of the histoenzymologic reactions were similar, except for acid reversal which did not occur in fibers of the fast glycolytic (FG) type in the mandibular locomotor muscles. FG fibers had a larger area and were more frequent in all regions studied. No significant differences in frequency or area of each fiber type were detected, considering the anterior and posterior regions of the masseter and temporal muscles. The frequency of fibers of the fast oxidative glycolytic (FOG) and slow oxidative (SO) types and of FOG area differed significantly between the anterior belly of the digastric muscle and the mandibular locomotor muscle. The predominance of fast twitch (FG and FOG) fibers and the multipenniform and bipenniform internal architecture of the masseter and temporal muscles, respectively, are characteristics that permit the powerful bite typical of tufted capuchin monkeys.

Static lung compliance and body pressures in Tupinambis merianae with and without post-hepatic septum

The surgical removal of the post-hepatic septum (PHS) in the tegu lizard, Tupinambis merianae, significantly reduces resting lung volume (VLr) and maximal lung volume (VLm) when compared with tegus with intact PHS. Standardised for body mass (MB), static lung compliance was significantly less in tegus without PHS. Pleural and abdominal pressures followed, like ventilation, a biphasic pattern. In general, pressures increased during expiration and decreased during inspiration. However, during expiration pressure changes showed a marked intra- and interindividual variation. The removal of the PHS resulted in a lower cranio-caudal intracoelomic pressure differential, but had no effect on the general pattern of pressure changes accompanying ventilation. These results show that a perforated PHS that lacks striated muscle has significant influence on static breathing mechanics in Tupinambis and by analogy provides valuable insight into similar processes that led to the evolution of the mammalian diaphragm. © 2003 Elsevier Science B.V. All rights reserved.

Dietary restriction and fibre supplementation: Oxidative stress and metabolic shifting for cardiac health

Dietary modification ought to be the first line of strategy in prevention of the development of cardiac disease. The purpose of this study was to investigate whether dietary restriction, dietary-fibre-enriched diet, and their interactions might affect antioxidant capacity and oxidative stress in cardiac tissue. Male Wistar rats (180-200 g; n = 10) were divided into four groups: control ad libitum diet (C), 50% restricted diet (DR), fed with fibre-enriched diet (F), and 50% restricted fibre-enriched diet (DR-F). After 35 days of the treatments, F, DR, and DR-F rats showed low cholesterol, LDL-cholesterol, and triacylglycerol, and high HDL-cholesterol in serum. The DR, DR-F, and F groups had decreased myocardial lipoperoxide and lipid hydroperoxide. The DR-F and F treatments increased superoxide dismutase and glutatione peroxidase (GSH-Px). The DR treatment increased GSH-Px and catalase activities. Dietary fibre beneficial effects were related to metabolic alterations. The F and DR-F groups showed high cardiac glycogen and low lactate dehydrogenase/citrate synthase ratios, indicating diminished anaerobic and elevated aerobic myocardial metabolism in these animals. There was no synergistic effect between dietary restriction and dietary fibre addition, since no differences were observed in markers of oxidative stress in the F and DR-F groups. Dietary fibre supplementation, rather than energy intake and dietary restriction, appears to be the main process retarding oxidative stress in cardiac tissue.

Synergistic action of olive oil supplementation and dietary restriction on serum lipids and cardiac antioxidant defences

Caloric intake is higher than recommended in many populations. Therefore, enhancing olive oil intake alone may not be the most effective way to prevent cardiovascular diseases. The purpose of the present study was to analyse the association of olive oil and dietary restriction on lipid profile and myocardial antioxidant defences. Male Wistar rats (180-200 g, n = 6) were divided into 4 groups: control ad libitum diet (C), 50% restricted diet (DR), fed ad libitum and supplemented with olive oil (3 mL/(kg-day)) (OO), and 50% restricted diet and supplemented with olive oil (DROO). After 30 days of treatments, OO, DR, and DROO groups had increased total cholesterol and high-density lipoprotein cholesterol concentrations. DR and DROO animals showed decreased low-density lipoprotein cholesterol. DROO had the lowest low-density lipoprotein cholesterol concentration. Total lipids and triacylglycerols were raised by dietary restriction and diminished by olive oil. OO rats had higher myocardial Superoxide dismutase and lower catalase and glutathione peroxidase activities than C rats. DR and DROO showed enhanced cardiac Superoxide dismutase, catalase, and glutathione peroxidase activities from the control. Olive oil supplementation alone improved the lipid profile but was more effective when coupled with dietary restriction. There was a synergistic beneficial action of dietary restriction and olive oil on serum lipids and myocardial antioxidant defences.

Morphological changes in the fast vs slow fiber profiles of the urethras of diabetic pregnant rats

Background: This study was undertaken to test the hypothesis that diabetes and pregnancy detrimentally affect the normal function of urethral striated muscles in rats, providing a model for additional studies related to urinary incontinence. The aim of this study was to evaluate morphological alterations in the urethral striated muscles of diabetic pregnant rats. Materials and Methods: Twenty female Wistar rats were distributed into four experimental groups of five rats as follows: virgin, pregnant, diabetic virgin, and diabetic pregnant. Diabetes was induced using streptozotocin administration (40 mg/kg i.v.). The rats were lethally anesthetized, and the urethra and vagina were extracted as a unit. Cryostat sections (6 μm thick) were cut and stained with hematoxylin-eosin, and immunohistochemical procedures were performed and subjected to morphological and semi quantitative analysis. Results: The urethral striated muscle from the diabetic pregnant rats presented with the following variations: thinning and atrophy, disorganization and disruption associated with the colocalization of fast and slow fibers and a steady decrease in the proportion of fast vs slow fibers. Conclusion: Diabetes and pregnancy impair the urethral striated muscle and alter its fiber type distribution. © Copyright G. Marini et al., 2011.