Submitted by Ronildo Prado (ronisp@ufscar.br) on 2016-09-21T19:00:11Z No. of bitstreams: 1 DissGHRC.pdf: 1218495 bytes, checksum: fb017af87bda85cd4fb78f46ec458e37 (MD5) / Approved for entry into archive by Ronildo Prado (ronisp@ufscar.br) on 2016-09-21T19:00:52Z (GMT) No. of bitstreams: 1 DissGHRC.pdf: 1218495 bytes, checksum: fb017af87bda85cd4fb78f46ec458e37 (MD5) / Approved for entry into archive by Ronildo Prado (ronisp@ufscar.br) on 2016-09-21T19:01:01Z (GMT) No. of bitstreams: 1 DissGHRC.pdf: 1218495 bytes, checksum: fb017af87bda85cd4fb78f46ec458e37 (MD5) / Made available in DSpace on 2016-09-21T19:01:08Z (GMT). No. of bitstreams: 1 DissGHRC.pdf: 1218495 bytes, checksum: fb017af87bda85cd4fb78f46ec458e37 (MD5) Previous issue date: 2014-09-04 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Menopause is highly associated with the increase of several metabolic disorders. In addition menopause promotes accelerated loss of muscle strength and mass, process known as sarcopenia, generating a deleterious role in the performance of physical functions. In this sense, resistance training has been recommended for the treatment of numerous diseases associated with menopause, as well as sarcopenia, as a nonpharmacological intervention. The aim of this study was to investigate the effects of the resistance training using the vertical ladder as a model on the muscle cross-sectional area and the percentage of muscle fiber types in the plantaris muscle of intact or ovariectomized rats and/or under hormone reposition therapy. Holztman adult female rats were distributed in 7 groups: Sham operated sedentary (Sham-Sed), Sham operated resistance trained (Sham-Tr), Ovariectomized sedentary (Ovx-Sed), Ovariectomized resistance trained and hormone reposition (Ovx-Tr-TRH), Ovariectomized sedentary and hormone reposition (Ovx-TRH), Ovariectomized resistance trained (Ovx-Tr), Ovariectomized and placebo reposition (Ovx-TRH-Pl). During a 12-week resistance training the animals climbed the vertical ladder of 1.1m with weights attached to their tails. Each session was carried out every 72 hours with 4-9 climbs. The ovariectomy promoted a reduction in cross-sectional area of the type I and II fiber when compared to Sham-Sed group, whereas the resistance training in the ovariectomy (Ovx-Tr group) was able to increase the cross-sectional area of the two fiber types (p<0.05). Moreover, TRH did not prevent the reduction of cross-sectional area of the type II fiber however it attenuated the reduction in cross-sectional area of type I fiber. The ovariectomy, hormone reposition and resistance training had no effect into promote alterations in the absolute number of muscle fiber types of the plantaris muscle and there was no correlation in the percentage of fiber muscle types. Our results indicate that resistance training can be a useful tool to prevent skeletal muscle mass reduction induced by ovariectomy. / A menopausa está associada ao risco aumentado de várias alterações metabólicas. Além disso, promove perda acelerada de massa e força muscular, em um processo de sarcopenia, gerando um papel deletério no desempenho de funções físicas. Nesse sentido, o treinamento resistido, como intervenção não farmacológica, vem sendo recomendado para o tratamento de várias doenças associadas à menopausa bem como a sarcopenia. O objetivo deste estudo foi verificar os efeitos do treinamento resistido em escada sobre a área de secção transversa e porcentagem dos tipos de fibras do músculo plantar de ratas intactas ou ovariectomizadas, e/ou submetidas à reposição hormonal. Foram utilizadas 77 ratas Holztman divididas em 7 grupos: Sham operado sedentário (Sham-Sed); Sham operado treinado (Sham-Tr); Ovariectomizado sedentário (Ovx- Sed); Ovariectomizado treinado e com terapia de reposição hormonal (Ovx-Tr-TRH); Ovariectomizado sedentário com terapia de reposição hormonal (Ovx-TRH); Ovariectomizado treinado (Ovx-Tr); e Ovariectomizado com reposição placebo (Ovx- TRH-Pl). Um período de 12 semanas de treinamento resistido foi realizado, durante o qual os animais subiram uma escada vertical de 1,1 m com pesos atados na cauda. As sessões foram realizadas a cada 72 horas, com 4-9 subidas. Foi possível observar que a Ovx promoveu uma redução da área de secção transversa das fibras tipo I e tipo II em relação ao grupo Sham-Sed, enquanto que o TR na condição de Ovx (grupo Ovx-TR) foi capaz de aumentar a área de secção transversa dos dois tipos de fibras (p<0,05). Além disso, a TRH adotada não preveniu a redução da área de secção transversa das fibras tipo II, mas atenuou a redução da área de secção transversa das fibras tipo I. A Ovx, a TRH e o Tr não promoveram alterações com relação ao número absoluto dos tipos de fibras encontradas no músculo plantar e não ocorreu nenhuma correlação relevante na porcentagem dos tipos de fibras musculares. Nossos achados indicaram que o Tr pode ser uma ferramenta importante na prevenção da redução da massa do músculo esquelético induzida pela Ovx.

Com o envelhecimento observa-se redução da massa muscular, aumento da resistência à insulina e queda nos níveis séricos de alguns hormônios, entre eles o DHEA. Ratos com 12-14 meses de idade apresentam regulação músculo-específica em vias da insulina e IGF 1 em músculos com populações distintas de tipos de fibras e como indicador de sarcopenia apresentam redução da proporção e tamanho de fibras tipo 2b. Dose única de DHEA não modula as vias da insulina e IGF 1 no músculo esquelético e reduz tamanho de fibras tipo 1 no EDL. / Reductions in skeletal muscle mass, increased insulin resistance, and decrements in several hormones serum levels, including DHEA, are features observed in aging. Twelve to fourteen months old rats present specific regulation in the insulin and the IGF-1 intracellular pathway in distinct skeletal muscle fiber composition and show reductions in size and proportion of muscle composed predominantly by fiber type 2. DHEA treatment had no effect in these intracellular pathways but induces reduction in the size of type 1 fiber in the EDL muscle.

Metabolic Syndrome Insulin Resistance Is Associated with Discordant Distribution of GLUT4 and the Insulin Receptor in Fast-Twitch and Slow-Twitch Muscle Fiber Types We have previously shown that We have previously shown that strength training alone improved insulin responsiveness in sedentary controls but not in metabolic syndrome subjects. Immunoblots of metabolic syndrome subjects[apos] muscle homogenates showed training-related increases in GLUT4 and mitochondrial enzymes was half that seen in the controls. To determine if this was due to changes primarily in fast-twitch fibers (strength fibers), we performed immunohistochemical (IHC) studies on muscle sections from these subjects to quantify fiber-specific changes in GLUT4, phospho-AMPK, phospho-mTOR, ATP synthase, and the insulin receptor. Signal intensity in confocal microscopic images was digitally quantified and the amount in each fiber type was adjusted by the fiber composition and the average size of each fiber type. Fiber type was classified using monoclonal antibodies against slow-twitch (type 1 fibers) and fast-twitch (type 2a and 2b fibers) myosin heavy chains. At baseline, both groups had slightly more insulin receptor in slow-twitch fibers, and most of the ATP synthase (mitochondrial marker) was in fast-twitch fibers. In controls, 55% of GLUT4 was in slow-twitch fibers, whereas metabolic syndrome subjects had only 33% of their GLUT4 in slow-twitch fibers. The IHC data showed modest increases in GLUT4 (9-25%), and substantial increases of ATP synthase (55-95%), and insulin receptors (44-104%) in both fiber types in both groups. Training-related increases were seen in phospho-AMPK (25% in slow-twitch, 15% in fast-twitch) only in the control subjects but no change in phospho-mTOR in either subject group. At baseline, metabolic syndrome subjects[apos] muscle had 56% of insulin receptors expressed in slow-twitch fibers, but only 33% of the GLUT4 was in these fibers. Thus, the untrained muscle composition of the metabolic syndrome subjects exhibited a mismatch between insulin receptors and GLUT4 in their fiber-specific distributions. This mismatch may contribute to the insulin resistance seen in the metabolic syndrome and may be involved in the diminished insulin sensitivity response to strength training in these subjects.

Escherichia coli O157:H7 est le sérotype le plus souvent incriminé lors de toxi-infection alimentaire par les E. coli entérohémorragiques (EHEC). Il peut être associé, dans les cas les plus graves, à des colites hémorragiques mortelles et au syndrome hémolytique et urémique (SHU), touchant essentiellement les jeunes enfants. Le vecteur alimentaire le plus courant lors de ces contaminations est le boeuf haché. L’étape primaire de la contamination bactérienne se situe lors de l'abattage où les bactéries peuvent être transférées de la peau à la carcasse. Une gaine conjonctive entoure les muscles, sa composition protéique, similaire à la matrice extracellulaire (ECM), pourrait jouer un rôle dans l'adhésion bactérienne. Dans un premier temps, l’étude de l'adhésion et de la colonisation des bactéries aux protéines majeures de l’ECM musculaire, a révélé une forte influence des conditions de croissances sur l’adhésion, l'adhésion étant maximale à 25°C et pH7, en particulier aux collagènes I et III. Chez les EHEC, diverses protéines de surfaces peuvent être potentiellement impliquées dans l’adhésion à l’ECM. Le rôle d'un autotransporteur, l'antigène 43 (Ag43), dans l'autoagrégation, l'adhésion et la formation de biofilm, a été établit chez E. coli O157:H7 EDL933. Par la suite, les interactions entre E. coli O157:H7 et la viande ont été étudiées sur deux muscles modèles de types métabolique et contractile opposés (Soleus oxidatif lent et EDL, glycolytique rapide), caractérisés par microspectroscopie de fluorescence UV couplée au rayonnement synchrotron. Les différents types de fibres musculaires ainsi que l’effet d’une anoxie prolongée simulant la maturation des viandes ont été discriminés par leurs réponses spectrales après une excitation à 275 nm. Un tropisme bactérien plus élevé pour le muscle soleus que pour le muscle EDL a été clairement observé. Bien qu'E. coli O157:H7 adhère de manière similaire aux différents types de fibres musculaires, l'adhésion des bactéries se fait essentiellement au niveau de l'ECM, mettant en évidence le rôle clé de l'ECM et du tissu conjonctif musculaire dans l’adhésion des E. coli O157:H7 à la viande. Ces travaux de recherche sur l’adhésion bactérienne aux muscles squelettiques aux niveaux moléculaires, cellulaires et tissulaires fournissent les premières connaissances sur la physiologie des EHEC lors de la contamination de la viande et constituent un pré-requis indispensable au développement de pratiques et de stratégies innovantes afin de réduire le risque de contamination des viandes. / Escherichia coli O157:H7 is the most prevalent serotype involved in foodborne infection by enterohemorrhagic E. coli (EHEC). It is associated with life-threatening hemorrhagic colitis and the hemolyticuremic syndrome (HUS), which essentially affect young children. The major food vector of EHEC contamination is ground beef. The primary bacterial contamination occurs during the slaughter, essentially at dehiding stage where bacteria can be transferred from hides to carcasses. The connective tissue surrounding the muscle, highly similar to extracellular matrix (ECM) could potentially be a support for bacterial adhesion. When investigating the adhesion and colonization to the main muscle fibrous ECM proteins, the great influence of growth conditions on subsequent bacterial attachment was shown. Maximal adhesion to ECM proteins occurred at 25°C and pH 7, especially to collagens I and III. In EHEC, various surface-exposed protein determinants can be expressed and potentially involved in ECM adhesion. Investigating the autoaggregation, bacterial adhesion and biofilm formation, the involvement of Antigen 43 (Ag43), an autotransporter protein, was demonstrated in E. coli O157:H7 EDL933. Then, the attachment of E. coli O157:H7 to the meat was determined on two different model muscles, with different contractile and metabolic characteristic (Soleus oxidative, slow and EDL glycolytic, fast), previously characterized by UV microspectroscopy coupled to synchrotron radiation fluorescence. The different of muscle fiber types and the effect of a prolonged anoxia simulating maturing meat were discriminated by their spectral responses after excitation at 275 nm. It clearly appeared that bacteria displayed differential tropism as function of the muscle types, higher for the Soleus than the EDL muscles. While E. coli O157:H7 adhered similarly to the different types of muscle fibers, bacterial adherence essentially occurred at the ECM, pinpointing the key role of connective tissue for E. coli O157:H7 adhesion to meat. This first comprehensive investigation of bacterial adhesion to skeletal muscles at molecular, cellular and tissue levels provides new insight in the physiology of the colonization of meat by EHEC and constitutes a prerequisite for the development of innovative practices and strategies to minimize the risk of meat contamination.

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