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Várias condições podem estar associadas com a atrofia muscular, tais como inatividade, envelhecimento, septicemia, diabetes, câncer e uso de glicocorticoides. Todas estas condições levam a atrofia muscular através de mecanismos que incluem aumento da degradação proteica e/ou redução na síntese proteica, envolvendo pelo menos cinco sistemas: lisossomal, da calpaína, das caspases, metaloproteinases e o sistema ubiquitina-proteasoma (SUP). Glicocorticoides, tais como a dexametasona, acarretam atrofia muscular atuando em quase todos esses sistemas, com significante ativação do SUP e lisossomal, afetando uma importante via de trofismo muscular, a via do IGF-1/PI-3K/Akt/mTOR. Ácidos graxos poli-insaturados, como o Ômega-3 (ômega-3), têm sido utilizados de forma benéfica na atenuação da atrofia muscular que ocorre na septicemia e na caquexia associada ao câncer, no entanto, sua atuação sobre a atrofia muscular induzida por glicocorticoides ainda não foi avaliada. Objetivo: Avaliar se a suplementação do ácido graxo ômega-3 influenciaria o desenvolvimento da atrofia muscular induzida pela dexametasona em ratos. Metodologia: Vinte e quatro ratos Wistar suplementados e não suplementados com ômega-3 (40 dias) foram submetidos à administração de dexametasona subcutânea (5mg/Kg/dia) nos últimos 10 dias, formando assim quatro grupos: Controle (CT), dexametasona (DX), ômega3 e dexametasona+ômega3 (DX+ômega3). Através de estudo de comportamento motor, histológico, PCR em tempo real e Western Blotting foram avaliados respectivamente, o número de grandes e pequenos movimentos em campo aberto; a área de secção transversa das fibras musculares (fibras I, IIA e IIB); a expressão dos genes MyoD, Miogenina, MuRF-1, Atrogina-1 e Miostatina; e a expressão de proteínas relacionadas com a via do IGF-1/PI-3K/Akt/mTOR: Akt, GSK3beta, FOXO3a e mTOR, totais e fosforiladas. Resultados: A dexametasona produziu diminuição na quantidade de pequenos movimentos, atrofia muscular em fibras do tipo IIB e diminuição na expressão de P-Akt, P-GSK3ômega e P-FOXO3a/FOXO3a total. A suplementação com Ômega-3 não se mostrou eficaz na atenuação de tais alterações. Por outro lado, o Ômega-3 associado à dexametasona (grupo DX+3) induziu a maior expressão de atrogenes (MuRF-1 e atrogina-1) causando, adicionalmente, maior atrofia muscular em fibras do tipo I e IIA, além de menor expressão gênica de Miogenina. O Ômega-3 de forma isolada conduziu de forma significativa a maior expressão de Miostatina e MyoD, e de forma não significante elevou a expressão proteica de mTOR total e induziu menor ganho de peso corporal dos animais ao fim do estudo. Conclusão: A suplementação de Ômega-3 não foi capaz de atenuar as alterações comportamentais, atrofia muscular e perda de peso corporal causadas pela administração de dexametasona, levando por outro lado a maior atrofia das fibras musculares e aumento na expressão de atrogenes. Desta forma, este estudo sugere que suplementos alimentares usualmente considerados benéficos para saúde, tal como o ácido graxo Ômega-3, podem agir em interação com alguns medicamentos, como os glicocorticoides, potencializando seus efeitos colaterais / Many conditions can be related to muscle atrophy, such as inactivity, aging, sepsis, diabetes, cancer, as well as, glucocorticoid treatment. All these conditions lead to muscle atrophy through mechanisms that include increase of protein degradation and/or decrease of protein synthesis involving at least five systems: lysossomal, calpain, caspases, metaloproteinases and ubiquitin proteasome system (UPS). Glucocorticoids, such as dexamethasone cause muscle atrophy acting in almost all of these systems, with a significant UPS activation and affecting an important pathway related to muscular trophism, IGF-1/PI-3k/Akt/mTOR pathway. Poly-unsaturated fatty acids, such as Omega-3 (omega-3), have been used beneficially to attenuation of muscle atrophy that occur in sepsis and cachexia related to cancer, however, its action in the glucocorticoid-induced muscle atrophy, has never been evaluated. Objective: Assess whether the omega-3 supplementation would influence the development of dexamethasone-induced muscle atrophy in rats. Methods: Twenty four Wistar rats supplemented and non-supplemented with omega-3 (40 days) were submitted to dexamethasone administration (5mg/kg/day) during the last 10 days, thus establishing 4 groups: control (CT), dexamethasone (DX), omega-3 and dexamethasone+omega-3 (DX+ omega-3). The amount of large and small movements in open field; muscle fiber cross sectional areas (I, IIA and IIB); MyoD, Myogenin, MuRF-1, Atrogin-1 and Myostatin gene expression; and protein expression of Akt, GSK3omega, FOXO3a and mTOR, total and phosphorylated forms were assessed, respectively, by: motor behavior testing, histological reactions, Real-time PCR and Western Blotting analysis. Results: Dexamethasone administration induced significant decrease of small motor movements, atrophy in type IIB muscle fibers and decrease of P-Akt, P-GSK3omega and P-FOXO3a/total FOXO3a expression. Omega-3 supplementation was not able to attenuate these changes. Instead, omega-3 associated to dexamethasone (DX+ omega-3 group) additionally induced higher muscle atrophy in type I, IIA muscle fibers, and reduced expression of Myogenin. The isolated use of Omega-3 led to a significant higher expression of Myostatin and MyoD, and a non-significant increase of total mTOR protein expression and less body weight gain at end of study. Conclusion: Supplementation of omega-3 was not able to attenuate motor behavioral changes, muscle atrophy and loss of body weight caused by dexamethasone administration, leading on the other hand to higher muscle fibers atrophy and increase in atrogenes expression. Therefore, this study suggests that food supplements, usually considered benefic to the health, such as Omega-3 fatty acid, may interact with some medications, such as glucocorticoids, potentiating its side effects

Várias condições podem estar associadas com a atrofia muscular, tais como inatividade, envelhecimento, septicemia, diabetes, câncer e uso de glicocorticoides. Todas estas condições levam a atrofia muscular através de mecanismos que incluem aumento da degradação proteica e/ou redução na síntese proteica, envolvendo pelo menos cinco sistemas: lisossomal, da calpaína, das caspases, metaloproteinases e o sistema ubiquitina-proteasoma (SUP). Glicocorticoides, tais como a dexametasona, acarretam atrofia muscular atuando em quase todos esses sistemas, com significante ativação do SUP e lisossomal, afetando uma importante via de trofismo muscular, a via do IGF-1/PI-3K/Akt/mTOR. Ácidos graxos poli-insaturados, como o Ômega-3 (ômega-3), têm sido utilizados de forma benéfica na atenuação da atrofia muscular que ocorre na septicemia e na caquexia associada ao câncer, no entanto, sua atuação sobre a atrofia muscular induzida por glicocorticoides ainda não foi avaliada. Objetivo: Avaliar se a suplementação do ácido graxo ômega-3 influenciaria o desenvolvimento da atrofia muscular induzida pela dexametasona em ratos. Metodologia: Vinte e quatro ratos Wistar suplementados e não suplementados com ômega-3 (40 dias) foram submetidos à administração de dexametasona subcutânea (5mg/Kg/dia) nos últimos 10 dias, formando assim quatro grupos: Controle (CT), dexametasona (DX), ômega3 e dexametasona+ômega3 (DX+ômega3). Através de estudo de comportamento motor, histológico, PCR em tempo real e Western Blotting foram avaliados respectivamente, o número de grandes e pequenos movimentos em campo aberto; a área de secção transversa das fibras musculares (fibras I, IIA e IIB); a expressão dos genes MyoD, Miogenina, MuRF-1, Atrogina-1 e Miostatina; e a expressão de proteínas relacionadas com a via do IGF-1/PI-3K/Akt/mTOR: Akt, GSK3beta, FOXO3a e mTOR, totais e fosforiladas. Resultados: A dexametasona produziu diminuição na quantidade de pequenos movimentos, atrofia muscular em fibras do tipo IIB e diminuição na expressão de P-Akt, P-GSK3ômega e P-FOXO3a/FOXO3a total. A suplementação com Ômega-3 não se mostrou eficaz na atenuação de tais alterações. Por outro lado, o Ômega-3 associado à dexametasona (grupo DX+3) induziu a maior expressão de atrogenes (MuRF-1 e atrogina-1) causando, adicionalmente, maior atrofia muscular em fibras do tipo I e IIA, além de menor expressão gênica de Miogenina. O Ômega-3 de forma isolada conduziu de forma significativa a maior expressão de Miostatina e MyoD, e de forma não significante elevou a expressão proteica de mTOR total e induziu menor ganho de peso corporal dos animais ao fim do estudo. Conclusão: A suplementação de Ômega-3 não foi capaz de atenuar as alterações comportamentais, atrofia muscular e perda de peso corporal causadas pela administração de dexametasona, levando por outro lado a maior atrofia das fibras musculares e aumento na expressão de atrogenes. Desta forma, este estudo sugere que suplementos alimentares usualmente considerados benéficos para saúde, tal como o ácido graxo Ômega-3, podem agir em interação com alguns medicamentos, como os glicocorticoides, potencializando seus efeitos colaterais / Many conditions can be related to muscle atrophy, such as inactivity, aging, sepsis, diabetes, cancer, as well as, glucocorticoid treatment. All these conditions lead to muscle atrophy through mechanisms that include increase of protein degradation and/or decrease of protein synthesis involving at least five systems: lysossomal, calpain, caspases, metaloproteinases and ubiquitin proteasome system (UPS). Glucocorticoids, such as dexamethasone cause muscle atrophy acting in almost all of these systems, with a significant UPS activation and affecting an important pathway related to muscular trophism, IGF-1/PI-3k/Akt/mTOR pathway. Poly-unsaturated fatty acids, such as Omega-3 (omega-3), have been used beneficially to attenuation of muscle atrophy that occur in sepsis and cachexia related to cancer, however, its action in the glucocorticoid-induced muscle atrophy, has never been evaluated. Objective: Assess whether the omega-3 supplementation would influence the development of dexamethasone-induced muscle atrophy in rats. Methods: Twenty four Wistar rats supplemented and non-supplemented with omega-3 (40 days) were submitted to dexamethasone administration (5mg/kg/day) during the last 10 days, thus establishing 4 groups: control (CT), dexamethasone (DX), omega-3 and dexamethasone+omega-3 (DX+ omega-3). The amount of large and small movements in open field; muscle fiber cross sectional areas (I, IIA and IIB); MyoD, Myogenin, MuRF-1, Atrogin-1 and Myostatin gene expression; and protein expression of Akt, GSK3omega, FOXO3a and mTOR, total and phosphorylated forms were assessed, respectively, by: motor behavior testing, histological reactions, Real-time PCR and Western Blotting analysis. Results: Dexamethasone administration induced significant decrease of small motor movements, atrophy in type IIB muscle fibers and decrease of P-Akt, P-GSK3omega and P-FOXO3a/total FOXO3a expression. Omega-3 supplementation was not able to attenuate these changes. Instead, omega-3 associated to dexamethasone (DX+ omega-3 group) additionally induced higher muscle atrophy in type I, IIA muscle fibers, and reduced expression of Myogenin. The isolated use of Omega-3 led to a significant higher expression of Myostatin and MyoD, and a non-significant increase of total mTOR protein expression and less body weight gain at end of study. Conclusion: Supplementation of omega-3 was not able to attenuate motor behavioral changes, muscle atrophy and loss of body weight caused by dexamethasone administration, leading on the other hand to higher muscle fibers atrophy and increase in atrogenes expression. Therefore, this study suggests that food supplements, usually considered benefic to the health, such as Omega-3 fatty acid, may interact with some medications, such as glucocorticoids, potentiating its side effects

Thesis (MSc (Physiological Sciences))--Stellenbosch University, 2008. / Introduction: Skeletal muscle atrophy is a mitigating complication that is characterized by a reduction in muscle fibre cross-sectional area as well as protein content, reduced force, elevated fatigability and insulin resistance. It seems to be a highly ordered and regulated process and signs of this condition are often seen in inflammatory conditions such as cancer, AIDS, diabetes and chronic heart failure (CHF). It has long been understood that an imbalance between protein degradation (increase) and protein synthesis (decrease) both contribute to the overall loss of muscle protein. Although the triggers that cause atrophy are different, the loss of muscle mass in each case involves a common phenomenon that induces muscle proteolysis. It is becoming evident that interactions among known proteolytic systems (ubiquitin-proteosome) are actively involved in muscle proteolysis during atrophy. Factors such as TNF-α and ROS are elevated in a wide variety of chronic inflammatory diseases in which skeletal muscle proteolysis presents a lethal threat. There is an increasing body of evidence that implies TNF-α may play a critical role in skeletal muscle atrophy in a number of clinical settings but the mechanisms mediating its effects are not completely understood. It is also now apparent that the transcription factor NF-κB is a key intracellular signal transducer in muscle catabolic conditions. This study investigated the various proposed signalling pathways that are modulated by increasing levels of TNF-α in a skeletal muscle cell line, in order to synthesize our current understanding of the molecular regulation of muscle atrophy. Materials and Methods: L6 (rat skeletal muscle) cells were cultured under standard conditions where after reaching ± 60-65% confluency levels, differentiation was induced for a maximum of 8 days. During the last 2 days, myotubes were incubated with increasing concentrations of recombinant TNF-α (1, 3, 6 and 10 ng/ml) for a period of 40 minutes, 24 and 48 hours. The effects of TNF-α on proliferation and cell viability were measured by MTT assay and Trypan Blue exclusion technique. Morphological assessment of cell death was conducted using the Hoechst 33342 and Propidium Iodide staining method. Detection of apoptosis was assessed by DNA isolation and fragmentation assay. The HE stain was used for the measurement of cell size. In order to determine the source and amount of ROS production, MitoTracker Red CM-H2 X ROS was utilised. Ubiquitin expression was assessed by immunohistochemistry. PI3-K activity was calculated by using an ELISA assay and the expression of signalling proteins was analysed by Western Blotting using phospho-specific and total antibodies. Additionally, the antioxidant Oxiprovin was used to investigate the quantity of ROS production in TNF-α-induced muscle atrophy. Results and Discussion: Incubation of L6 myotubes with increasing concentrations of recombinant TNF-α revealed that the lower concentrations of TNF-α used were not toxic to the cells but data analysis of cell death showed that 10 ng/ml TNF-α induced apoptosis and necrosis. Long-term treatment with TNF-α resulted in an increase in the upregulation of TNF- α receptors, specifically TNF-R1. The transcription factors NF-κB and FKHR were rapidly activated thus resulting in the induction of the ubiquitin-proteosome pathway. Activation of this pathway produced significant increases in the expression of E3 ubiquitin ligases MuRF-1 and MAFbx. Muscle fibre diameter appeared to have decreased with increasing TNF-α concentrations in part due to the suppressed activity of the PI3-K/Akt pathway as well as significant reductions in differentiation markers. Western blot analysis also showed that certain MAPKs are activated in response to TNF-α. No profound changes were observed with ROS production. Finally, the use Oxiprovin significantly lowered cell viability and ROS production. These findings suggest that TNF-α may elicit strong catabolic effects on L6 myotubes in a dose and time dependent manner. Conclusion: These observations suggest that TNF-α might have beneficial effects in skeletal muscle in certain circumstances. This beneficial effect however is limited by several aspects which include the concentration of TNF-α, cell type, time of exposure, culture conditions, state of the cell (disturbed or normal) and the cells stage of differentiation. The effect of TNF-α can be positive or negative depending on the concentration and time points analysed. This action is mediated by various signal transduction pathways that are thought to cooperate with each other. More understanding of these pathways as well as their subsequent upstream and downstream constituents is obligatory to clarify the central mechanism/s that control physiological and pathophysiological effects of TNF-α in skeletal muscle.

INTRODUÇÃO: Existem poucos estudos demonstrando que anormalidades musculares esqueléticas em pacientes com insuficiência cardíaca crônica persistem meses após o transplante cardíaco. No presente estudo, objetivamos avaliar massa muscular, e força muscular periférica e respiratória em pacientes no pré-transplante cardíaco, e no seguimento precoce (6 meses) e tardio (1,5 e 3 anos) pós-transplante cardíaco. Objetivamos verificar ainda a correlação entre força muscular periférica e respiratória em pacientes no pré e pós-transplante cardíaco. Comparamos, por fim, os dados de pacientes do pré-transplante cardíaco com um grupo controle de indivíduos saudáveis sem doença cardíaca. MÉTODOS: Tratou-se de estudo prospectivo do tipo coorte. Foram selecionados todos os pacientes em lista de espera para transplante cardíaco do Hospital de Messejana, do período de agosto de 2011 a março de 2013. Avaliamos idade, gênero, causas da insuficiência cardíaca, hipertensão, diabetes, tempo de espera na lista, tempo de internamento pós-transplante, tempo de ventilação mecânica, medida da força muscular respiratória, da força muscular periférica, da espessura do adutor do polegar, média bilateral da área de secção transversal do músculo psoas maior, índice de massa corporal e creatinina em todos os pacientes do estudo e no grupo controle. As variáveis de massa e força muscular foram medidas por meio de tomografia computadorizada, paquimetria, manovacuometria e dinamometria. RESULTADOS: Foram encontrados 25 pacientes elegíveis e 23 foram incluídos. Ocorreram 8 óbitos no seguimento precoce, 4 no seguimento tardio e, ao final de 3 anos de seguimento, 11 pacientes sobreviveram com enxerto funcionando. Foram selecionados 23 indivíduos saudáveis para o grupo controle, pareados para gênero, idade, peso e altura. Quando comparamos as variáveis de massa e força muscular dos pacientes do grupo pré-transplante cardíaco com o grupo controle de indivíduos saudáveis foram encontradas diminuição da força muscular periférica (27,0 kg/f vs. 38,2 kg/f), da área de secção transversal do músculo psoas ( 1.238,9 mm2 vs. 1.533,1 mm2) da espessura do músculo adutor do polegar (16,5 mm vs. 23,9 mm), da força muscular inspiratória (60,2 cmH2O vs. 94,8 cmH2O) e da força muscular expiratória (75,2 cmH2O vs. 102,17 cmH2O) nos pacientes do grupo pré-transplante cardíaco. Na comparação entre os períodos pré-transplante cardíaco, seguimento precoce e seguimento tardio pós-transplante cardíaco houve aumento estatisticamente significante (p < 0,001) das seguintes variáveis: força muscular periférica (27,3 kg/f vs. 34,7 kg/f), da área de secção transversal do músculo psoas ( 1.305,4 mm2 vs. 1.431,3 mm2) da espessura do músculo adutor do polegar (15,9 mm vs. 20,2 mm), da força muscular inspiratória (59,5 cmH2O vs. 90,9 cmH2O) e da força muscular expiratória (79,5 cmH2O vs. 101,8 cmH2O) nos 11 pacientes sobreviventes. Ao final do seguimento tardio pós-transplante cardíaco todas as variáveis de massa e força muscular atingiram níveis semelhantes àqueles do grupo controle, exceto a espessura do músculo adutor do polegar. CONCLUSÃO: Os achados demonstraram haver sarcopenia em pacientes no pré-transplante cardíaco, visto que houve diminuição da massa muscular e da força muscular periférica e respiratória confirmando a presença de dois critérios, requisito para fazer o diagnóstico de sarcopenia. O transplante cardíaco proporcionou aumento da força muscular respiratória, da força muscular periférica, da espessura do músculo adutor do polegar e aumento da massa muscular do psoas / INTRODUCTION: There are few studies demonstrating that skeletal muscle abnormalities in patients with chronic heart failure persist for months after heart transplantation. In this study, we aimed to evaluate muscle mass, and peripheral and respiratory muscle strength in patients in pre-heart transplantation, and in the early (6 months) and late (1.5 to 3 years) follow-up after heart transplantation. We also aimed to verify the correlation between peripheral and respiratory muscle strength in patients before and after heart transplantation. Finally, we compared the pre-heart transplantation patients\' data with a control group of healthy individuals without heart disease. METHODS: It was a prospective cohort study. We selected all patients on the waiting list for heart transplantation of Messejana\'s Hospital from August 2011 to March 2013. Age, gender, cause of heart failure, hypertension, diabetes, period on the waiting list, post-transplantation hospitalization time, mechanical ventilation time, measurements of respiratory muscle strength (maximum inspiratory muscle strength and maximum expiratory muscle strength), peripheral muscle strength (hand grip strength), adductor pollicis muscle thickness, the bilateral average of the major psoas muscle cross-sectional area, body mass index and serum creatinine were assessed in all the patients in the study and in control groups. Mass and muscle strength variables were measured using computed tomography, pachymetry, manometry and dynamometry. RESULTS: We found 25 eligible patients and 23 were included. There were 8 deaths in the early follow-up period; by the end of 3-year follow-up there were 11 surviving patients with functioning graft. We selected 23 healthy subjects for the control group, matched for gender, age, weight and height. When we compared the variables mass and muscle strength of the pre heart transplant patients with healthy control subjects were found decreased peripheral muscle strength (hand grip strength) (27.0 kg/f vs. 38.2 kg/f), of the psoas muscle\'s cross-section area (1238.9 mm2 vs. 1533.1 mm2), the adductor pollicis muscle thickness (16.5 mm vs. 23.9 mm), maximum inspiratory muscle strength (60.2 cmH2O vs. 94 8 cmH2O) and maximum expiratory muscle strength (75.2 cm H2O vs. 102.17 cmH2O) in patients in the pre heart transplant group. Comparing the pre-heart transplant periods, the early and late heart transplantation follow-up there was a statistically significant increase (p < 0.001) the following variables: peripheral muscle strength (hand grip strength) (27.3 kg / f vs. 34.7 kg / f ), the psoas muscle\'s cross-sectional area (1305.4 vs. 1431.3 mm 2 mm 2) the adductor pollicis muscle thickness (15.9 mm vs. 20.2 mm), maximum inspiratory muscle strength (59.5 cmH2O vs. 90.9 cm H2O) and maximum expiratory muscle strength (79.5 cm H2O vs. 101.8 cm H2O) in the 11 surviving patients. At the end of post-heart transplant late follow-up all variables mass and muscle strength reached similar levels to those of the control group, except the adductor pollicis muscle thickness. CONCLUSION: The findings showed that there was sarcopenia in patients in pre-heart transplantation period, since there was a decrease in muscle mass and a decrease in muscle strength of peripheral and respiratory muscles confirming the presence of at least two criteria, a requirement to make the diagnosis of sarcopenia. Heart transplantation has provided increased respiratory muscle strength, increased peripheral muscle strength, increased the adductor pollicis muscle thickness and increased psoas muscle mass

Les particules ribonucléoprotéiques (RNP) sont impliquées dans divers mécanismes cellulaires : les UsnRNP, la SRP, les RNP à boîtes C/D et H/ACA dans la modification des ARN et la maturation des ARN ribosomiques, et la télomérase dans le maintien des extrémités chromosomiques. L'assemblage de ces RNP est un processus complexe faisant intervenir de nombreux facteurs, dont le complexe SMN. Un déficit de l’une des protéines de ce complexe conduit à l’amyotrophie spinale. Il est essentiel à la survie cellulaire et est nécessaire à l’assemblage des UsnRNP et de la SRP. Il est suggèré que le complexe SMN joue un rôle dans la biogenèse des RNP à boîtes C/D et H/ACA. Nous avons montré des interactions in vitro et des associations in cellulo entre le complexe SMN et la protéine NUFIP (un facteur d’assemblage de ces RNP). Ces résultats suggèrent l’existence d’un lien fonctionnel entre le complexe SMN et NUFIP dans l'assemblage des RNP à boîtes C/D et H/ACA et de la snRNP U4. Des interactions in vitro entre le complexe SMN et la protéine NAF1 (un facteur d’assemblage des RNP à boîtes H/ACA) ont révélés, que le complexe SMN est capable de s’associer avec la RNP à boîtes H/ACA en formation. Si le complexe SMN intervient dans l’assemblage des RNP, on peut supposer que cet assemblage soit défectueux dans la SMA. Nous avons montré que certains ARN sont accumulés dans la moelle épinière et le cerveau de souris SMA. La télomérase est réactivée dans les cellules cancéreuses. En collaboration avec l’équipe de J-M Vignaud (CHU central, Nancy), nous avons montré une augmentation du taux des protéines cœur des RNP à boîtes H/ACA et de NUFIP dans les cellules tumorales / Ribonucleoprotein particles (RNPs) are involved in various cellular mechanisms in eukaryotic cells: UsnRNP, SRP, C/D and H/ACA box RNPs in RNA modifications and rRNA maturation and telomerase in the synthesis of the chromosome extremities. RNP assembly is a very complex process, which involves numerous factors. One of these factors is the SMN complex. Decreased level of one of its components leads to spinal muscular atrophy. It is essential for cell survival and necessary for UsnRNP and SRP assembly. It is suggested that the SMN complex plays a role in C/D and H/ACA RNP biogenesis. We showed in vitro interactions and in cellulo associations between the SMN complex and the protein NUFIP (an assembly factor of these RNP). These results suggest the existence of a functional link between the SMN complex and NUFIP in the assembly of the C/D and H/ACA box RNPs and the U4 snRNP. In vitro interactions between the SMN complex and the protein NAF1 (an assembly factor of the H/ACA boxes RNPs) revealed, that the SMN complex is capable of joining with the H/ACA boxes RNPs in formation. If the SMN complex intervenes in the RNPs assembly, we can suppose that this assembly is defective in the SMA. We showed that any ARN is accumulated in the spinal cord and the brain of SMA mouse. The telomerase is reactivated in cancer cells. In association with the team of J-M Vignaud (CHU central, Nancy), we showed an increase of H/ACA box RNP proteins and NUFIP in these cancer cells

Les protéines SREBP-1 sont des facteurs de transcription connus pour leur rôle dans la régulation du métabolisme lipidique. Plus récemment des études faites in vitro (myotubes humains en culture primaire) et in vivo (muscle tibial de souris) ont montré que la surexpression de SREBP-1a ou SREBP-1c induit une atrophie musculaire et bloque la différenciation musculaire, en inhibant notamment l’expression des protéines structurales du muscle squelettique et des facteurs de la différenciation musculaire (MRFs). Les travaux de thèse présentés dans ce manuscrit ont eu pour but de décrypter le mécanisme de l’atrophie induite par SREBP-1 et de déterminer comment les protéines SIRT1 pourraient réguler ce facteur de transcription. L’atrophie musculaire résulte d’un déséquilibre entre la quantité de protéines synthétisées et dégradées. Dans nos études, nous montrons que SREBP-1 régule la synthèse protéique et la dégradation protéique, respectivement via le contrôle négatif de l’expression des MRFs et via le contrôle de l’expression des atrogènes, MuRF1 et Atrogin-1. Dans le muscle squelettique, nous démontrons que la désacétylase SIRT1 régule l’activité transcriptionnelle de SREBP-1. Les protéines SREBP-1 et SIRT1 étant toutes deux impliquées dans la régulation du métabolisme lipidique, nous mettons en évidence une nouvelle voie de signalisation reliant le métabolisme énergétique et nutritionnel avec l’activité transcriptionnelle de SREBP-1 dans le muscle. Étant donné le rôle de SIRT1 et SREBP-1 dans le métabolisme lipidique et musculaire, nous nous sommes intéressés au rôle des phospholipides et plus particulièrement des céramides dans la régulation de la masse musculaire.Nos études montrent que la régulation de la quantité de céramides par la cytokine TNFα régule la masse musculaire. Ainsi, nos travaux mettent en évidence de nouveaux liens entre le métabolisme lipidique et la régulation de la masse et du métabolisme musculaire. / SREBP-1 transcription factors are involved in lipid metabolism regulation. Recently, in vitro and in vivo studies have shown that SREBP-1a or SREBP-1c overexpression induce muscular atrophy and block muscular differentiation, notably by inhibiting structural proteins and Myogenics Regulatory Factors (MRFs) expression. The aims of this work are the mecanism determination of the muscular atrophy induced by SREBP-1 overexpression and the elucidation of the role of SIRT1 proteins on SREBP-1 regulation.The muscular atrophy results from an imbalance between the amount of synthesized and degraded proteins. In our studies, we shown that SREBP-1 regulates protein synthesis and protein degradation, respectively via a negative control of MRFs expression and via a control of atrogenes expression, MuRF1 and Atrogin-1. In skeletal muscle, we shown that SIRT1 desacetylase enzyme regulates SREBP-1 transcription activity. Because of SREBP-1 and SIRT1 proteins involvement in lipid metabolism regulation, our results suggest a new signalisation pathway linking energetic metabolism and SREBP-1 transcriptionnal activity in muscle. As SIRT1 and SREBP-1 have a role on lipid and muscular metabolism, we took an interest in phospholipids involvement and more specifically in ceramides involvement in muscle mass regulation. Our studies shown that the regulation of the amount of ceramids by the TNFα regulates muscle mass. Thus, our work allows to identify new links between lipid metabolism and muscle mass and metabolism regulation.

Hipotrofía nos músculos que estabilizam a coluna tem sido identificada nos pacientes com lombalgia. Entretanto, não se sabe se a perda muscular é causa ou conseqüência desta disfunção, nem se ela é influenciada pelo tipo de alteração discal que o indivíduo apresenta. Este estudo testou a hipótese de que a hipotrofía dos pacientes com lombalgia seja dependente do tipo de alteração discal. Para avaliar a condição muscular em diferentes tipos de alteração discal, optou-se por um estudo retrospectivo e por uma seleção aleatória de 78 exames de ressonância magnética de indivíduos com lombalgia. Em cada exame foram realizadas medidas quantitativas da porcentagem de gordura na área de secção transversa dos mm. multífidus e psoas, nos três últimos níveis da coluna lombar. A alteração discal foi encontrada em 95% dos exames, sendo o abaulamento o achado de imagem mais freqüente, seguido pela protrusão discal. A porcentagem de gordura variou conforme o tipo de alteração discal. Nos níveis com abaulamento há em ambos os músculos estudados 6% a mais de tecido gorduroso do que nos níveis onde há protrusão e esta diferença foi estatisticamente significante. Músculos nos níveis onde há protrusão sem fissura no anel fibroso apresentaram maior substituição gordurosa do que aqueles onde há protrusão com fissura. A porcentagem de gordura foi influenciada por características anatômicas como músculo estudado e nível da coluna, e por características como idade e sexo dos sujeitos. Estes resultados indicam que a hipotrofía muscular em pacientes com lombalgia não é um processo uniforme e generalizado, mas sim correlacionado a diferentes variáveis, entre elas o tipo de alteração discal que o paciente apresenta. / Low back pain patients present atrophy on muscles responsible for spine stabilization. However, it is not clear if muscle waste is related to the cause or if it is a consequence of this disfunction. Nor it is clear if muscle athophy is affected by the type of disc pathology. This study tested the hypothesis that muscle waste in low back pain patients influenced by the type of disc derangement. Magnetic resonance scans of 78 low back pain patients were randomly analysed. Cross sectional area percentage of fat tissue in multifidi and psoas major muscles was measured on the lower levels of the lumbar spine. Disc pathology was found in 95% of the exams and disc bulge was the most frequent abnormality, followed by disc protrusion. Fat percentage varied according to disc pathology and this difference was statistically significant. Muscles on levels with disc bulge presented 6% more fat deposits than muscles on levels with disc protrusion. Muscles on levels with discs without anular tear present more fat infiltration than muscles on levels with anular tear. Fat percentage was also influenced by anatomic aspects such as evaluated muscle and spine level, and sample characteristics as age and sex. The results indicated that muscle atrophy in low back pain patients is not a uniform and generalized feature. It is correlated to different variables, such as type of disc pathology

A estimulação elétrica neuromuscular (EENM) é um importante recurso utilizado na medicina esportiva para acelerar processos de recuperação. O objetivo deste estudo foi analisar os efeitos da EENM durante a imobilização do músculo gastrocnêmio, em posições de alongamento (LP) e encurtamento (SP). Para tanto, 60 ratas fêmeas jovens WISTAR foram distribuídas em seis grupos e acompanhadas durante 7 dias: controle (C), eletroestimuladas (EE), imobilizadas em encurtamento (ISP), imobilizadas em alongamento (ILP), imobilizadas em encurtamento e eletroestimuladas (ISP+EE) e imobilizadas em alongamento e eletroestimuladas (ILP+EE). Para a imobilização, o membro posterior direito foi envolvido por uma malha tubular e ataduras de algodão juntamente à atadura gessada. A EENM foi utilizada com uma freqüência de 50 Hz, 10 minutos por dia, totalizando 20 contrações em cada sessão. Após 7 dias os animais foram submetidos à eutanásia e os músculos gastrocnêmios foram retirados para a realização do ensaio mecânico de tração em uma máquina universal de ensaios (EMIC®). A partir dos gráficos carga versus alongamento foram calculadas as seguintes propriedades mecânicas: alongamento no limite de proporcionalidade (ALP), carga no limite de proporcionalidade (CLP), alongamento no limite máximo (ALM), carga no limite máximo (CLM) e rigidez. As imobilizações SP e LP promoveram reduções significativas (p<0,05) nas propriedades de ALP, CLP, ALM e CLM sendo mais acentuada, principalmente, no grupo ISP. Quando utilizada a EENM, houve acréscimo significativo (p<0,05) destas propriedades somente no grupo ISP. Já, em relação à rigidez, foi observada redução significativa (p<0,05) somente do grupo C para o grupo ISP. Quando utilizada a EENM, a rigidez do grupo ILP+EE foi significativamente (p<0,05) maior e mais próxima do grupo C que a do grupo ISP+EE. Neste modelo experimental, a imobilização dos músculos em alongamento atrasou a queda das propriedades e a estimulação elétrica, contribuiu para a manutenção das propriedades mecânicas durante o período de imobilização, principalmente no grupo ILP+EE. / The neuromuscular electric stimulation (NMES) is an important tool used in sport medicine to accelerate the recovery process. The objective of this study was to analyze the effects of NMES during the immobilization of the gastrocnemius muscle, in lengthened (LP) and shortened positions (SP). Sixty young female rats WISTAR were distributed into six groups and followed for 7 days: control (C), electric stimulation (ES), immobilized in shortened (ISP), immobilized in lengthened (ILP), immobilized in shortened and electric stimulation (ISP+ES) and immobilized in lengthened and electric stimulation (ILP+ES). For the immobilization, the right hind limb was involved by a tubular mesh and cotton rolls and plaster. NMES was used in a frequency of 50 Hz, 10 minutes a day, totaling 20 contractions in each session. After 7 days the animals were killed and their gastrocnemius muscles of the right side were submitted to a mechanical test in traction in an universal test machine (EMIC®). From the curves load versus elongation the following mechanical properties were obtained: elongation in the yield limit (EPL), load in the yield limit (LPL), elongation in the ultimate load (EUL), ultimate load (UL) and stiffness. The immobilizations SP and LP promoted significant reductions (p<0,05) in the properties of EPL, LPL, EUL and UL being more accentuated mainly in group ISP. When used NMES, there was significant increment (p<0,05) of such properties only in group ISP. As for stiffness, significant reduction was observed (p<0,05) only of the group C for group ISP. When the NMES was used, the stiffness of group ILP+EE was significantly (p<0,05) higher and closer to group C than for group ISP+EE. We conclude that in this experimental model the immobilization of the muscles in the lengthened position delayed the reduction of the properties and the electric stimulation contributed to the maintenance of the mechanical properties during the immobilization period, mainly for group ILP+ES.

Spinal Muscular Atrophy (SMA) is a neurodegenerative, inherited disease caused by an insufficient amount of functional Survival of Motor Neurone protein (SMN), though the exact mechanism underlying this is not fully understood. The primary function of SMN is assembling a ring of Sm proteins around small nuclear RNA (snRNA) in an early, cytoplasmic stage of small nuclear ribonucleoprotein (snRNP) biogenesis, a process essential in eukaryotes. SMN, together with several mRNA binding proteins, has been linked to neural transport of mRNA towards areas of growth in Motor neurons for local translation of transcripts. Previous research in our group has found that this may involve Coatomer protein-containing vesicles transported by Dynein and requiring the Sm family protein, SmB, for maintenance. Little is known, however, about what other proteins are also present and required for correct transport and localisation of these vesicles. To further investigate this, we have produced plasmids expressing each Sm protein tagged to fluorescent proteins to help track their behaviour, in some cases for the first time, and developed a detergent-free fractionation protocol to enrich for SMN containing vesicles, providing tools that can be used to further probe behaviour and interactions in the future. Using these approaches, SmN, a neural specific Sm protein, was identified to also be present in SMN-containing vesicles similarly to SmB. Analysis of the interactomes of different Sm proteins identified a novel interactor of SMN, Neurochondrin (NCDN), that appears to be required for the correct localisation of SMN in neural cells. NCDN was found to not associate with snRNPs, indicating an snRNP-independent interaction with SMN. NCDN and SMN both independently associated and co-enriched with Rab5, indicating a potential endocytic and cell polarity role for the interaction. This interaction has the potential to be key in SMA pathology and may have therapeutic potential.