iPSCs ameliorate hypoxia-induced autophagy and atrophy in C2C12 myotubes via the AMPK/ULK1 pathway

BACKGROUND: Duchenne muscular dystrophy (DMD) is an X-linked lethal genetic disorder for which there is no effective treatment. Previous studies have shown that stem cell transplantation into mdx mice can promote muscle regeneration and improve muscle function, however, the specific molecular mechanisms remain unclear. DMD suffers varying degrees of hypoxic damage during disease progression. This study aimed to investigate whether induced pluripotent stem cells (iPSCs) have protective effects against hypoxia-induced skeletal muscle injury. RESULTS: In this study, we co-cultured iPSCs with C2C12 myoblasts using a Transwell nested system and placed them in a DG250 anaerobic workstation for oxygen deprivation for 24 h. We found that iPSCs reduced the levels of lactate dehydrogenase and reactive oxygen species and downregulated the mRNA and protein levels of BAX/BCL2 and LC3II/ LC3I in hypoxia-induced C2C12 myoblasts. Meanwhile, iPSCs decreased the mRNA and protein levels of atrogin-1 and MuRF-1 and increased myotube width. Furthermore, iPSCs downregulated the phosphorylation of AMPKA and ULK1 in C2C12 myotubes exposed to hypoxic damage. CONCLUSIONS: Our study showed that iPSCs enhanced the resistance of C2C12 myoblasts to hypoxia and inhibited apoptosis and autophagy in the presence of oxidative stress. Further, iPSCs improved hypoxia-induced autophagy and atrophy of C2C12 myotubes through the AMPK/ULK1 pathway. This study may provide a new theoretical basis for the treatment of muscular dystrophy in stem cells.

Cholic and deoxycholic acids induce mitochondrial dysfunction, impaired biogenesis and autophagic flux in skeletal muscle cells

BACKGROUND: Skeletal muscle is sensitive to bile acids (BA) because it expresses the TGR5 receptor for BA. Cholic (CA) and deoxycholic (DCA) acids induce a sarcopenia-like phenotype through TGR5-dependent mechanisms. Besides, a mouse model of cholestasis-induced sarcopenia was characterised by increased levels of serum BA and muscle weakness, alterations that are dependent on TGR5 expression. Mitochondrial alterations, such as decreased mitochondrial potential and oxygen consumption rate (OCR), increased mitochondrial reactive oxygen species (mtROS) and unbalanced biogenesis and mitophagy, have not been studied in BA-induced sarcopenia.METHODS: We evaluated the effects of DCA and CA on mitochondrial alterations in C2C12 myotubes and a mouse model of cholestasis-induced sarcopenia. We measured mitochondrial mass by TOM20 levels and mitochondrial DNA; ultrastructural alterations by transmission electronic microscopy; mitochondrial biogenesis by PGC-1α plasmid reporter activity and protein levels by western blot analysis; mitophagy by the co-localisation of the MitoTracker and LysoTracker fluorescent probes; mitochondrial potential by detecting the TMRE probe signal; protein levels of OXPHOS complexes and LC3B by western blot analysis; OCR by Seahorse measures; and mtROS by MitoSOX probe signals. RESULTS: DCA and CA caused a reduction in mitochondrial mass and decreased mitochondrial biogenesis. Interestingly, DCA and CA increased LC3II/LC3I ratio and decreased autophagic flux concordant with raised mitophagosome-like structures. In addition, DCA and CA decreased mitochondrial potential and reduced protein levels in OXPHOS complexes I and II. The results also demonstrated that DCA and CA decreased basal, ATP-linked, FCCP-induced maximal respiration and spare OCR. DCA and CA also reduced the number of cristae. In addition, DCA and CA increased the mtROS. In mice with cholestasis-induced sarcopenia, TOM20, OXPHOS complexes I, II and III, and OCR were diminished. Interestingly, the OCR and OXPHOS complexes were correlated with muscle strength and bile acid levels. CONCLUSION: Our results showed that DCA and CA decreased mitochondrial mass, possibly by reducing mitochondrial biogenesis, which affects mitochondrial function, thereby altering potential OCR and mtROS generation. Some mitochondrial alterations were also observed in a mouse model of cholestasis-induced sarcopenia characterised by increased levels of BA, such as DCA and CA.

Muscle architecture of the vastus lateralis and rectus femoris in the production of knee extensor torque in physically inactive women

Abstract Aim: This study aimed to verify the data reliability of muscle architecture (MA) variables, and the relationship between MA and the isometric peak torque (PT) of the monoarticular and biarticular knee extensor (KE) muscle in physically inactive women. Methods: Ten physically inactive women (24.0 ± 1.64 years; 162.9 ± 5.34 cm; 63.5 ± 11.90 kg) participated in the study. An ultrasound device assessed the MA variables (muscle thickness, fascicle length, and pennation angle) of the Vastus Lateralis (VL) and Rectus Femoris (RF), and an isokinetic dynamometer assessed the PT. Pearson correlation evaluated the relationship between PT and MA variables, with a significance level of 5%. Additionally, the intraclass correlation coefficient, coefficient of variation, and standard error of measurement. Results: Excellent reliability between images was observed, and no significant relationships were observed between the PT and MA variables of the VL and RF. Conclusion: Isolated variables of the MA of a monoarticular or a biarticular muscle do not influence the production of the isometric PT of the KE.

Alterações miogênicas em células musculares humanas infectadas pelo vírus zika: susceptibilidade de mioblastos e resistência de miotubos / Myogenic alterations in human muscle cells infected by zika virus: myoblast susceptibility and myotube resistance

O músculo esquelético é responsável pelo movimento e manutenção da postura, e é um órgão produtor de miocinas e altamente metabólico, onde alterações em sua fisiologia podem ter consequências sistêmicas. Esse tecido é alvo para diferentes arboviroses, e mialgia é um sintoma frequentemente relatado. O músculo esquelético é composto majoritariamente por fibras musculares, e uma pequena população de células progenitoras denominadas células satélites (SC), que em caso de lesão podem ser ativadas, proliferam e se diferenciam, sendo capazes de regenerar o tecido muscular. Recentemente nosso grupo demonstrou que SC em proliferação (mioblastos) são infectadas pelo vírus ZIKA (ZIKV), enquanto células diferenciadas e fusionadas (miotubos) não apresentam proteínas virais. O presente trabalho avaliou alterações miogênicas e o perfil transcricional de mioblastos e miotubos humanos após tratamento com ZIKV, com o objetivo de identificar fatores e mecanismos envolvidos na susceptibilidade e resistência destas células à infecção. Confirmamos infecção produtiva do ZIKV nos mioblastos, que apresentaram uma redução no número de células expressando a molécula KI67 em altas concentrações (característico de células em mitose). A análise de sequenciamento mostrou perturbação das vias do ciclo celular em mioblastos infectados, que ainda apresentaram enriquecimento de vias relacionadas à morte celular. Também confirmamos a ausência de infecção produtiva nos miotubos. Interessantemente, verificamos que o ZIKV entra nas células diferenciadas, mas não consegue replicar o RNA viral, e a análise do transcriptoma identificou um enriquecimento de vias e modulação de genes antivirais maior ou exclusivamente nas células diferenciadas em comparação aos mioblastos infectados. Além disso, miotubos expostos ao ZIKV aparentam ter aumento de fusão/hipertrofia. Ao contrário dos mioblastos, miotubos apresentaram enriquecimento de vias relacionadas a organização da matriz extracelular. Dados preliminares do nosso grupo mostraram que o cultivo de mioblastos sobre a isoforma de laminina 511 levou à redução da infecção pelo ZIKV. Contudo, em nossos ensaios, a infecção pelo ZIKV não modulou a expressão de receptores para LM e o bloqueio do receptor de LM, a integrina α6, não reduziu a infecção pelo ZIKV em mioblastos. Nosso trabalho mostrou que a infecção pelo ZIKV induz resposta imune e antiviral, que foram enriquecidos em mioblastos e miotubos, sendo que estes apresentaram uma assinatura única de vias e genes antivirais, que poderiam explicar a resistência frente ao ZIKV.

Study of muscle fibers of the extensor digitorium longus and soleus muscles of C57BL/6 females exposed to glyphosate during pregnancy and lactation / Estudo das fibras musculares dos músculos extensor longo dos dedos e sóleo de fêmeas C57BL/6 expostas ao glifosato durante a prenhez e a lactação

ABSTRACT Objective To evaluate the morphology and morphometry of the muscles extensor digitorium longus and soleus of C57BL/6 females, who were exposed to glyphosate during pregnancy and lactation. Methods Twelve female mice from the C57BL/6 lineage were used. After detection of pregnancy, they were divided into a Control Group, which received only water, and a Glyphosate Group, which received water with 0.5% glyphosate during pregnancy and lactation. Both groups received ad libitum standard diet. After weaning, the females were euthanized and weighed; naso-anal length was measured, and fats were collected and weighed. The muscles extensor digitorium longus and soleus were collected, and their length and weight were measured. Then, the muscles were fixed in Methacarn to perform the histological study of muscle fibers. Results Glyphosate Group presented lower weight gain during pregnancy and also lower final body weight and naso-anal length; however, the other body parameters evaluated did not present a significant difference in relation to the Control Group. Significant differences were also not observed in the analysis of muscle fibers and connective tissue. Conclusion Exposure to 0.5% glyphosate during pregnancy and lactation resulted in lower weight gain during pregnancy, final weight, and naso-anal length. Despite not directly altering the morphology of muscle tissue, these results may indicate enough exposure to interfere with animal metabolism.

RESUMO Objetivo Avaliar a morfologia e a morfometria dos músculos extensor longo dos dedos e sóleo de fêmeas C57BL/6 expostas ao glifosato durante a prenhez e lactação. Métodos Foram utilizados 12 camundongos fêmeas da linhagem C57BL/6. Após detecção da prenhez, foram separadas em Grupo Controle, que recebeu somente água, e Grupo Glifosato, que recebeu água com 0,5% de glifosato durante a prenhez e lactação. Ambos os grupos receberam dieta padrão ad libitum. Após o desmame, as fêmeas foram eutanasiadas e pesadas; o comprimento nasoanal foi mensurado, e as gorduras foram coletadas e pesadas. Os músculos extensor longo dos dedos e sóleo foram coletados, e seu comprimento e peso foram mensurados. Em seguida, os músculos foram fixados em Methacarn para a realização do estudo histológico das fibras musculares. Resultados O Grupo Glifosato apresentou menor ganho de peso durante a prenhez e também menor peso corporal final e comprimento nasoanal, entretanto os demais parâmetros corporais avaliados não apresentaram diferença significativa em relação ao Grupo Controle. Na análise das fibras musculares e do tecido conjuntivo, também não foram observadas diferenças significativas. Conclusão A exposição a 0,5% de glifosato durante a prenhez e lactação resultou em menor ganho de peso na gestação, peso final e comprimento nasoanal, o que pode indicar que, apesar de não alterar a morfologia do tecido muscular diretamente, a exposição foi suficiente para interferir no metabolismo dos animais.

Tipo de fibra muscular y su relación con el abordaje fonoaudiológico en los trastornos de la deglución / Type of muscle fibreand its relationship to the Speech-Language Pathology approach in swallowing disorders

Los actuales protocolos de intervención en deglución no especifican parámetros de los ejercicios tales como la cantidad de series, duración, tiempos de repetición, entre otros. Lo anterior, es un desafío para la intervención fonoaudiológica en los trastornos de la deglución. El objetivo de este trabajo es acercar al profesional fonoaudiólogo al conocimiento de las fibras musculares, ya que es un conocimiento que se debe considerar antes de indicar ejercicios musculares cráneo-cérvico-orales. Por ello, se describen las principales fibras musculares con sus respectivas características fundamentales, como lo son: la resistencia a la fatiga y la velocidad de contracción. Se revisa la literatura sobre la dispersión de las fibras musculares, de algunos de los principales músculos que participan en el proceso deglutorio. También, se analizan diversas particularidades de los músculos de la zona cráneo-cérvico-oral. Se describen además las diferentes dificultades para evaluar esta musculatura. Finalmente, se expone la relevancia práctica de conocer estos tipos de fibras musculares y las perspectivas futuras de este enfoque basado en parámetros del ejercicio y la medición de variables objetivas.

Swallowing intervention protocols in dysphagia do not specify the parameters of the exercises, namely the duration, frequencyand number of series, all of which present a challenge in speech therapy intervention in swallowing disorders. The purpose of this review is to provide Speech and Language Pathologists with background information concerning crucial muscle fibres used in swallowing therapy and to subsequently indicate appropriate skull-cervical-oral muscle exercises. We describe the primary muscle and its characteristics such as fatigue resistance and contraction speed, review the types of fibres of some of the main muscles that participate in the swallowing process, and present the particularities of the muscles of the cranio-cervical-oral area in comparison with the rest of the skeletal muscles. We also provide information regarding the difficulties in evaluating the musculature involvedin the swallowing process before finally highlighting the relevance of understanding the roles and characteristics of these muscles for clinical practice

Plasticidad muscular y fibras híbridas en la musculatura masticatoria: revisión de literatura / Muscle plasticity and hybrid fibers in the masticatory musculature: literature review

La musculatura masticatoria se caracteriza por presentar fibras híbridas que en los últimos años se han relacionado con el fenómeno de plasticidad muscular. El objetivo del estudio fue describir la relación entre la plasticidad muscular y las fibras musculares híbridas presentes en la musculatura masticatoria, mediante una revisión narrativa de literatura. Para esto, se realizó una búsqueda electrónica en PUBMED, ScienceDirect y BIREME, utilizando las palabras claves: "Muscle Plasticity", "Hybrid Muscle Fibers" y "Hybrid Fibers". Fueron seleccionados documentos que reportan las isoformas de cadena pesada de miosina (MHC) presentes en los músculos masticatorios de humanos y otros mamíferos, junto a los cambios vinculados a demandas funcionales. Se describe la presencia de fibras puras tipo I y tipo II, además de otras isoformas como la MHC-la, MHC-IIM, MHC-fetal y MHC-cardíaca. Sin embargo, un porcentaje considerable de fibras en la musculatura masticatoria son híbridas, es decir, expresan a más de una isoforma de MHC, las cuales también son diferentes a nivel intermuscular e intramuscular. Las influencias locales pueden contribuir a la variación de la expresión del tipo de fibra. En la musculatura masticatoria, el destete, la dureza de los alimentos, el bruxismo, la morfología craneofacial y el uso de prótesis dentales genera cambios a nivel de los músculos masticatorios, donde es común la presencia de fibras híbridas. Se concluye la presencia importante de fibras híbridas en la musculatura masticatoria y su relación con la plasticidad muscular a lo largo del ciclo vital, debido a cambios funcionales y patológicos. Es importante que los terapeutas de habla y motricidad orofacial profundicen en el conocimiento de la fisiología del comportamiento oromiofuncional.

The masticatory musculature is characterized by presenting hybrid fibers that in recent years have been related to the phenomenon of muscle plasticity. The objective of the study was to describe the relationship between muscle plasticity and the hybrid muscle fibers present in the masticatory muscles, through a narrative review. For this, an electronic search was conducted in PUBMED, ScienceDirect and BIREME, using the keywords: "Muscle Plasticity", "Hybrid Muscle Fibers" and "Hybrid Fibers". Documents that report the myosin heavy chain (MHC) isoforms present in the masticatory muscles of humans and other mammals were selected, along with the changes linked to functional demands. The presence of type I and type II pure fibers were described, in addition to other isoforms such as MHC-la, MHC-IIM, MHC-fetal and MHC-cardiac. However, a significant percentage of fibers in the masticatory muscles are hybrids, that is, they express more than one MHC isoform, which are also different at the intermuscular and intramuscular level. Local influences can contribute to the variation of fiber type expression. In the chewing muscles, weaning, the hardness of food, bruxism, craniofacial morphology and the use of dental prostheses generate changes at the level of the chewing muscles, where the presence of hybrid fibers is common. The important presence of hybrid fibers in the masticatory muscles and their relationship with muscle plasticity throughout the life cycle, due to functional and pathological changes, is concluded. It is important for Speech Therapy andMyofunctional Therapy to deepen their understanding of the physiology of oromyofunctional behavior.

La rigidez cadavérica, el espasmo cadavérico y tipos de fibras musculares / The cadaveric rigidity, the cadaveric spasm and types of muscle fibers

La rigidez cadavérica (rigor mortis) es un proceso no muy bien comprendido por la mayoría de los médicos. El conocimiento de la intimidad del proceso de la rigidez cadavérica es de vital importancia ya que es una de las variables que junto con las livideces (livor mortis) y la temperatura (algor mortis) del cadáver ayudan a determinar el cronotanatodiagnóstico, tanatocronodiagnóstico o intervalo postmortal del período inmediato de la muerte. Para entender el mecanismo de la rigidez y el espasmo cadavérico es preciso hacer un repaso de la contracción muscular fisiológica en el vivo. Hay que tener presente que el tipo de fibra muscular predominante modificará las características de la contracción muscular fisiológica en el vivo, y también la rigidez y el espasmo cadavérico. (AU)

The cadaveric rigidity (rigor mortis) is a process which is not very well understood by the majority of the doctors. The knowledge of the intimacy of the cadaveric stiffness process is of vital importance since it is one of the variables that, as well as the postmortem lividity (livor mortis) and the body temperature post mortem (algor mortis) help determine the chronotanatodiagnostic, tanatochronodiagnostic or postmortal interval of the immediate period of death. In order to understand the mechanism of stiffness and cadaveric spasm, it is necessary to review the physiological muscle contraction in vivo. We should keep in mind that the predominant type of muscle fiber will modify the characteristics of physiological muscle contraction in vivo, as well as stiffness and cadaveric spasm. (AU)

Análise da ativação muscular durante o movimento de alcance nas condições ativo, ativo-assistido e autoassistido em pacientes pós-AVE / Análisis de la activación muscular durante el movimiento de alcance en las condiciones activo, activo-asistido y auto-asistido en pacientes post-AVE / Analysis of muscle activation during reaching movement in active, active-assisted and self-assisted conditions in post-stroke patients

RESUMO O Acidente Vascular Encefálico (AVE) é uma patologia que frequentemente causa limitações motoras nos Membros Superiores (MMSS) gerando prejuízos funcionais nos movimentos de alcance. O objetivo do estudo foi analisar o recrutamento muscular do membro superior parético durante três condições de alcance: ativo, ativo-assistido e autoassistido, através de dados eletromiográficos das fibras anteriores do Músculo Deltoide (MD), Bíceps Braquial (BB) e Tríceps Braquial (TB). Estudo do tipo transversal que utilizou como testes clínicos o miniexame do estado mental, escala de equilíbrio de Berg, medida de independência funcional, escala modificada de Ashworth e escala de Fugl-Meyer - seção MMSS. A coleta dos dados eletromiográficos de superfície foi realizada utilizando-se o eletromiógrafo e eletrodos de configuração bipolar da EMG System do Brasil com três canais posicionados nos pontos motores do MD (fibras anteriores), BB e TB de ambos os membros superiores. As variáveis clínicas apresentaram resultados de comprometimento motor, cognitivo e funcional leves. Os dados eletromiográficos mostraram que o MD e TB durante o alcance ativo-assistido contraíram mais que no alcance autoassistido (p<0.05). Os MD e TB apresentaram diferenças significativas durante os movimentos de alcance, enquanto que o músculo BB não mostrou alterações. Entre os diversos tipos de alcance, o ativo-assistido foi o que proporcionou maior ativação muscular. Sugere-se que sejam feitos ensaios clínicos para verificar a eficácia dos treinamentos.

RESUMEN El Accidente Vascular Encefálico (AVE) es una patología que frecuentemente causa limitaciones motoras en los Miembros Superiores (MMSS) generando perjuicios funcionales en los movimientos de alcance. El objetivo del estudio fue analizar el reclutamiento muscular del miembro superior parético durante tres condiciones de alcance: activo, activo-asistido y auto-asistido, a través de datos electromiográficos de las fibras anteriores del Músculo Deltóide (MD), Bíceps Braquial (BB) y Tríceps Braquial (TB). Estudio del tipo transversal que utilizó como pruebas clínicas el mini-examen del estado mental, escala de equilibrio de Berg, medida de independencia funcional, escala modificada de Ashworth y escala de Fugl-Meyer - sección MMSS. La recolección de los datos electromiográficos de superficie fue realizada utilizando el electromiografo y electrodos de configuración bipolar de la EMG System de Brasil con tres canales colocados en los puntos motores del MD (fibras anteriores), BB y TB de ambos miembros superiores. Las variables clínicas presentaron resultados de compromiso motor, cognitivo y funcional leves. Los datos electromiográficos mostraron que el MD y el TB durante el alcance activo-asistido contrajeron más que en el alcance auto-asistido (p<0.05). Los MD y TB presentaron diferencias significativas durante los movimientos de alcance, mientras que el músculo BB no mostró alteraciones. Entre los diversos tipos de alcance, el activo asistido fue el que proporcionó mayor activación muscular. Se sugiere que se realicen ensayos clínicos para verificar la eficacia de los entrenamientos.

ABSTRACT A cerebrovascular accident (CVA) is a disease that often causes upper limb motor limitations and functional losses in reaching movements. The objective of this study was to analyze the muscle recruitment of the paretic upper limb during three reaching conditions: active, active-assisted and self-assisted, through electromyographic data of anterior fibers of Deltoid Muscle (DM), Biceps Brachii (BB) and Triceps Brachii (TB). Cross-sectional study that used as clinical trials the Mini-Mental State Examination, Berg balance scale, functional independence measure, the modified Ashworth scale, and the Fugl-Meyer assessment - upper limbs section. Surface electromyographic data were collected using the electromyograph and bipolar electrode configuration of the EMG System do Brasil with three channels positioned in the motor points of DM (anterior fibers), BB and TB of both upper limbs. Clinical variables showed mild motor, cognitive, and functional impairment. Electromyographic data showed that DM and TB contracted more during active-assisted than during self-assisted exercise (p<0.05). DM and TB presented significant differences during reaching movements, while the BB muscle showed no changes. Among the different reaching exercises, the active-assisted was the one that provided greater muscle activation. Clinical trials are suggested to verify the effectiveness of the training.

Muscle changes with high-intensity aerobic training in an animal model of renal disease

Abstract Purpose: To analyze the muscle changes with high-intensity aerobic training (HIAT) in an animal model of renal disease (RD). Methods: Twenty one adult Wistar rats were divided into 3 groups: healthy sedentary (HS), RD sedentary (RDS), RD aerobic training (RDAT). RDS and RDAT were subjected to unilateral renal ischemia-reperfusion (10 min) and 21days after that, RDAT was subjected to 6 weeks HIAT (swimming). Serum creatinine (Cr) and muscle morphometry (cross-sectional area = CSA) of gastrocnemius were analyzed. Results: Cr was higher (p = 0.0053) in RDS (0.82 ± 0.04) than in the others (RDAT 0.55 ± 0.04; HS 0.55 ± 0.04). Morphometric analysis (class interval of CSA in μm2/absolute frequency of muscle fibers in each class) indicated that 50th percentile occurred in: HS 7th class (3000.00-3499.00/515), RDS, 8th class (3500.00-3999.00/484), RDAT 5th class (2000.00-2499.00/856). CSA of largest fibers in RDS, RDAT, HS was 9953.00 μm2, 9969.00 μm2,11228.00 μm2, respectively. High frequency of fibers with lower CSA occurred in 4th, 5th, 6th and 7th class in RDA, absence of fibers into 22nd, 23rd classes (RDS and RDAT). Conclusion: HIAT in an animal model of RD resulted in increased the number of muscle fibers with smaller CSA.

Morphological analysis of articularis genus and involvement in muscle synchronicity with vastus intermedius / Análisis morfológico del músculo articular de la rodilla y su participación en la sincronicidad muscular con el músculo vasto intermedio

The articularis genus, which lies under the vastus intermedius, has been regarded as part of the quadriceps femoris. However, they are well known to have different function because their respective origins and insertions are mutually distinct. These muscles are considered to work almost simultaneously when the knee is extended. The electromyogram has been used to demonstrate muscle co-contraction. However, the articularis genus is deeper than other muscles. Moreover, it is difficult to analyze it by surface electromyogram. The relative proportions of muscle fiber types and the characteristics of these fiber types are important determinants of the surface electromyogram. Furthermore, biomechanical analysis of AG has remained unclear. This study investigated the ratio of muscle fiber types in these muscles. Muscle samples from seven human cadaveric specimens were used with application of immunofluorescence double staining. Results show that in the vastus intermedius and articularis genus, the percentage of Type I fibers was significantly higher than that of Type II fibers. No significant difference was found in the mean percentages of Type I and Type II fiber types. The percentages of Type I and Type II fibers in articularis genus muscle were correlated positively to the percentage in the vastus intermedius. These results suggest that similar muscle fiber compositions of these muscles might reflect their contraction during the same active phase of knee extension, despite their different functions.

El músculo articular de la rodilla, que se encuentra cubierto por el músculo vasto intermedio, se ha considerado como parte del músculo cuádriceps femoral. Sin embargo, es sabido que tienen diferentes funciones debido a que sus respectivos orígenes e inserciones son mutuamente distintas. Se considera que estos músculos trabajan de forma casi simultánea cuando la rodilla está extendida. El electromiograma se ha usado para demostrar la contracción muscular. Sin embargo, el músculo articular de la rodillas es más profundo que otros músculos. Además, es difícil analizarlo por electromiograma de superficie. Las proporciones relativas de los tipos de fibras musculares y las características de estos tipos de fibras son importantes determinantes del electromiograma de superficie. Además, el análisis biomecánico de músculo articular de la rodilla no ha sido claro. Este estudio investigó la proporción de tipos de fibras musculares en estos músculos. Se usaron muestras musculares de siete cadáveres humanos con la aplicación de doble tinción de inmunofluorescencia. Los resultados muestran que en los músculos articular de la rodilla y vasto intermedio, el porcentaje de fibras de Tipo I fue significativamente mayor que el de las fibras de Tipo II. No se encontraron diferencias significativas en los porcentajes medios de los Tipo I y Tipo II. Los porcentajes de fibras Tipo I y Tipo II en el músculo articular de la rodilla se correlacionaron positivamente con el porcentaje en el músculo vasto intermediario. Estos resultados sugieren que las composiciones de las fibras musculares similares de estos músculos podrían reflejar su contracción durante la misma fase activa de la extensión de la rodilla, a pesar de sus diferentes funciones.

Morphological Characterization of Diaphragm in Common Squirrel Monkey (Saimiri sciureus)

ABSTRACT The wall of the diaphragm can be affected by congenital or acquired alterations which allow the passage of viscera between the abdominal and chest cavities, allowing the formation of a diaphragmatic hernia. We characterized morphology and performed biometrics of the diaphragm in the common squirrel monkey Saimiri sciureus. After fixation, muscle fragments were collected and processed for optical microscopy. In this species the diaphragm muscle is attached to the lung by phrenopericardial ligament. It is also connected to the liver via the coronary and falciform ligaments. The muscle is composed of three segments in total: 1) sternal; 2) costal, and 3) a segment consisting of right and left diaphragmatic pillars. The anatomical structures analyzed were similar to those reported for other mammals. Histological analysis revealed stable, organized muscle fibers with alternation of light and dark streaks, indicating transverse striation.

Morphological changes in rat rectus abdominis muscle induced by diabetes and pregnancy

The urethral muscle of diabetic pregnant rats is affected by long-term mild diabetes and short-term severe diabetes, which plays a crucial role in the pathogenesis of pelvic floor disorders. We hypothesized that muscles outside the pelvis are subject to similar changes. The current study aimed at analyzing the effects of long-term mild and short-term severe diabetes on the structure and ultrastructure of fiber muscles and collagen in rats' rectus abdominis (RA) muscle. Therefore, the RA muscle of virgin, pregnant, long-term mild diabetic, short-term severe diabetic, long-term mild diabetic pregnant and short-term severe diabetic pregnant 3-month-old Wistar rats were collected. The structure was analyzed by picrosirius red staining, immunohistochemistry for fast and slow muscle fibers and transmission electron microscopy. We investigated two levels of STZ- induced diabetes: long-term mild diabetes (blood glucose level: 120-200 mg/dL) and short-term severe diabetes (blood glucose level >300 mg/dL). Long-term mild diabetic pregnant and short-term severe diabetic pregnant rats had decreased fast fibers and increased slow fibers, disrupted areas of sarcomere, intermyofibrillar mitochondria and myelin figures in the RA muscle. Both groups enabled us to analyze the specific influence of pregnancy, separately from diabetes. The current study demonstrated that diabetes and pregnancy induced intramuscular transformation and reorganization of RA muscle with a switch of fiber type adjusting their architecture according to intensity and duration of hyperglycemic insult within pregnancy.

Action of vanillin (Vanilla planifolia) on the morphology of tibialis anterior and soleus muscles after nerve injury / Ação da vanilina (Vanilla planifolia) sobre a morfologia dos músculos tibial anterior e sóleo após lesão nervosa

ABSTRACT Objective To evaluate the action of vanillin (Vanilla planifolia) on the morphology of tibialis anterior and soleus muscles after peripheral nerve injury. Methods Wistar rats were divided into four groups, with seven animals each: Control Group, Vanillin Group, Injury Group, and Injury + Vanillin Group. The Injury Group and the Injury + Vanillin Group animals were submitted to nerve injury by compression of the sciatic nerve; the Vanillin Group and Injury + Vanillin Group, were treated daily with oral doses of vanillin (150mg/kg) from the 3rd to the 21st day after induction of nerve injury. At the end of the experiment, the tibialis anterior and soleus muscles were dissected and processed for light microscopy and submitted to morphological analysis. Results The nerve compression promoted morphological changes, typical of denervation, and the treatment with vanillin was responsible for different responses in the studied muscles. For the tibialis anterior, there was an increase in the number of satellite cells, central nuclei and fiber atrophy, as well as fascicular disorganization. In the soleus, only increased vascularization was observed, with no exacerbation of the morphological alterations in the fibers. Conclusion The treatment with vanillin promoted increase in intramuscular vascularization for the muscles studied, with pro-inflammatory potential for tibialis anterior, but not for soleus muscle.

RESUMO Objetivo Avaliar a ação da vanilina (Vanilla planifolia) sobre a morfologia dos músculos tibial anterior e sóleo após lesão nervosa periférica. Métodos Ratos Wistar foram divididos em quatro grupos, com sete animais cada, sendo Grupo Controle, Grupo Vanilina, Grupo Lesão e Grupo Lesão + Vanilina. Os animais dos Grupos Lesão e Grupo Lesão + Vanilina foram submetidos à lesão nervosa por meio da compressão do nervo isquiático, e os Grupos Vanilina e Grupo Lesão + Vanilina foram tratados diariamente com doses orais de vanilina (150mg/kg) do 3o ao 21o dia após a indução da lesão nervosa. Ao término do experimento, os músculos tibial anterior e sóleo foram dissecados e seguiram o processamento de rotina em microscopia de luz, para posterior análise morfológica. Resultados A compressão nervosa promoveu alterações morfológicas características de denervação, sendo que o tratamento com vanilina foi responsável por respostas distintas nos músculos estudados. Para o tibial anterior, houve aumento do número de células satélites, núcleos centrais e atrofia das fibras, bem como desorganização fascicular. Já no sóleo, houve apenas aumento da vascularização, sem exacerbação das alterações morfológicas nas fibras. Conclusão O tratamento com vanilina promoveu o aumento da vascularização intramuscular para os músculos estudados, com potencial pró-inflamatório para o tibial anterior, o que não ocorreu no músculo sóleo.

Regulation of muscle plasticity and trophism by fatty acids: A short review / Regulação da plasticidade e do trofismo muscular pelos ácidos graxos: uma breve revisão

Summary The skeletal muscle tissue has a remarkable ability to alter its plastic structural and functional properties after a harmful stimulus, regulating the expression of proteins in complex events such as muscle regeneration. In this context, considering that potential therapeutic agents have been widely studied, nutritional strategies have been investigated in order to improve the regenerative capacity of skeletal muscle. There is evidence of the modulatory action of fatty acids, such that oleic and linoleic acids, that are abundant in Western diets, on muscle function and trophism. Thus, fatty acids appear to be potential candidates to promote or impair the recovery of muscle mass and function during regeneration, since they modulate intracellular pathways that regulate myogenesis. This study is the first to describe and discuss the effect of fatty acids on muscle plasticity and trophism, with emphasis on skeletal muscle regeneration and in vitro differentiation of muscle cells.

Resumo O tecido muscular esquelético possui a notável capacidade plástica de alterar suas propriedades estruturais e funcionais após um estímulo lesivo, regulando a expressão de proteínas durante eventos complexos como a regeneração muscular. Nesse contexto, considerando que possíveis agentes terapêuticos vêm sendo amplamente estudados, estratégias nutricionais têm sido investigadas na perspectiva de melhorar a capacidade regenerativa do músculo esquelético. Há evidências da ação modulatória dos ácidos graxos, como os ácidos oleico e linoleico, que são abundantes nas dietas ocidentais, sobre a função muscular e o trofismo. Nesse sentido, os ácidos graxos parecem ser potenciais candidatos para promover ou prejudicar a recuperação da massa e a função muscular durante a regeneração, uma vez que modulam vias intracelulares reguladoras da miogênese. Este trabalho é o primeiro a descrever e discutir o efeito dos ácidos graxos sobre a plasticidade e o trofismo muscular, com ênfase na regeneração do músculo esquelético e na diferenciação de células musculares in vitro.

Effects of duodenal-jejunal bypass on structure of diaphragm in western diet obese rats

Abstract Purpose: To evaluate the effects of duodenal-jejunal bypass (DJB) on the diaphragm muscle of obese rats fed on a western diet (WD) . Methods: Eighteen male Wistar rats were fed a standard rodent chow diet (CTL group) or WD ad libitum. After 10 weeks, WD rats were submitted to sham (WD SHAM) or duodenal-jejunal bypass (WD DJB). The structure, ultrastructure, collagen content and the morphometry of the neuromuscular junctions (NMJs) were analyzed two months after surgery. Results: WD SHAM rats displayed an increase in body weight, the Lee index and retroperitoneal and peri-epididymal fat pads compared to the CTL group. DJB did not alter these parameters. The muscle fiber structure and NMJs were similar in the WD SHAM and CTL groups. However, the WD SHAM group showed alterations in the fiber ultrastructure, such as loosely arranged myofibrils and Z line disorganization. In addition, WD SHAM animals presented a considerable amount of lipid droplets and a reduction in the percentage of collagen compared to the CTL group. DJB did not affect the structure or ultrastructure of the muscle fibers or the NMJs in the diaphragm of the WD DJB animals. Conclusion: Duodenal-jejunal bypass did not improve the alterations observed in the diaphragm of western diet obese-rats.

Myostatin promotes distinct responses on protein metabolism of skeletal and cardiac muscle fibers of rodents

Myostatin is a novel negative regulator of skeletal muscle mass. Myostatin expression is also found in heart in a much less extent, but it can be upregulated in pathological conditions, such as heart failure. Myostatin may be involved in inhibiting protein synthesis and/or increasing protein degradation in skeletal and cardiac muscles. Herein, we used cell cultures and isolated muscles from rats to determine protein degradation and synthesis. Muscles incubated with myostatin exhibited an increase in proteolysis with an increase of Atrogin-1, MuRF1 and LC3 genes. Extensor digitorum longus muscles and C2C12 myotubes exhibited a reduction in protein turnover. Cardiomyocytes showed an increase in proteolysis by activating autophagy and the ubiquitin proteasome system, and a decrease in protein synthesis by decreasing P70S6K. The effect of myostatin on protein metabolism is related to fiber type composition, which may be associated to the extent of atrophy mediated effect of myostatin on muscle.

Image analysis of oxidative and glycolytic muscle fibers during reperfusion injury by segmentation based on regions / Análisis de imágenes de fibras musculares oxidativas y glucolíticas durante la reperfusión mediante segmentación basada en regiones

Different situations cause ischemia and reperfusion injury, affecting tissues under the level of compression. In this research, abnormal characteristics in distribution of muscle fibers types in soleus and extensor carpi radialis longus, during short periods of ischemia and short and long periods of reperfusion, were determined. Fibers were classified by enzyme histochemistry techniques NADH-TR and myosin-ATPase. Measurements of areas were carried out through semiautomatic image processing by using segmentation based on regions, which evidenced significant changes in distribution during reperfusion followed to one and three hours of ischemia, as well as in comparisons of areas for all periods of reperfusion. This study strengthens the evidence about using practical procedures of image analysis in the diagnosis of tisular abnormalities.

Diferentes situaciones causan isquemia y reperfusión afectando los tejidos que se encuentran por debajo del nivel de lesión. En esta investigación, se determinaron las características anormales en la distribución de los tipos de fibras musculares en los músculos sóleo y extensor radial longo del carpo, durante períodos cortos de isquemia y de reperfusión tanto cortos como prolongados. Las fibras se clasificaron mediante las técnicas de tinción de histoquímica enzimática NADH-TR y miosina-ATPasa. Las mediciones de las áreas realizadas mediante procesamiento de imágenes y el uso de la segmentación basada en regiones, evidenciaron cambios significativos en la distribución durante la reperfusión seguida a una y tres horas de isquemia, así como en las comparaciones para todos los períodos de reperfusión. Este estudio es un aporte en la evidencia sobre el uso de procedimientos prácticos de análisis de imágenes en el diagnóstico de alteraciones tisulares.

Morphological and morphometric study of the muscle fibers and neuromuscular junctions of the extensor digitorum longus in aged rats submitted to maternal protein restriction / Estudio morfológico y morfométrico de las fibras musculares y de las intersecciones neuromusculares del músculo extensor largo de los dedos de ratas de edad avanzada sometidas a la restricción proteica materna

Protein restriction implies the functional involvement of several systems and organs, including the skeletal muscle, because it is a protein reservoir in the body. This study sought to analyze the morphological and morphometric features of the muscle fibers and neuromuscular junctions (NMJs) of the extensor digitorum longus (EDL) muscle in rats at 365 days of age, submitted to maternal protein restriction during the gestation and lactation periods. Wistar rats were divided into two groups: a Control Group - mothers fed a normal-protein diet (17 % protein) during pregnancy and lactation; and a Restricted Group - mothers fed a low-protein diet (6 % protein) during pregnancy and lactation. The pups were kept with the mother throughout the lactation period (21 days), after which the offspring received a normal protein diet until 365 days of age. Histological (HE) and histoenzymological (NADH-TR) studies were conducted on the muscle fibers. The muscle was subjected to Nonspecific Esterase reaction to stain the Neuromuscular Junctions. Regarding the animals from the restricted group: the histologic analysis of the muscle fibers showed the presence of centralized nuclei and a diminished area; the histoenzymological study showed the different types of muscle fibers were randomly distributed in the EDL muscle and the area of the Type IIa muscle fiber was smaller; the ultrastructural study revealed disorganization of the Z line, and the presence of lipid droplets and vacuoles containing myelin figures in subsarcolemmal and intramiofibrilar regions; while the analysis of the NMJs exhibited no significant differences between the groups. Protein restriction in the pregnancy and lactation period may have affected the development of skeletal muscle, producing a permanent muscle-fiber deficit in the EDL muscle of the offspring.

La restricción proteica implica compromiso funcional de diversos sistemas y órganos, entre ellos, el músculo estriado esquelético, por ser una reserva de proteína del organismo. De esa forma, el presente trabajo procuró analizar las características morfológicas y morfométricas de las fibras musculares y de las intersecciones neuromusculares (JNMs) del músculo extensor largo de los dedos (EDL) en ratas de 365 días de edad, sometidas a restricción proteica materna durante los periodos de gestación y lactancia. Las ratas Wistar fueron separadas en dos grupos: El grupo Control - madres alimentadas durante la gestación y lactancia con ración normoproteica (17 % de proteína) y Grupo con restricción ­ madres alimentadas durante la gestación y lactancia con ración hipoproteica (6 % de proteína). Las crías permanecieron con la madre durante todo el periodo de lactancia (21 días) y después de este periodo la prole recibió ración normoproteica hasta los 365 días de edad. Se realizó un estudio histológico (HE) e histoenzimológico (NADH-TR) de las fibras musculares. Para la marcación de las JNMs, el músculo fue sometido a la reacción de Esterasa Inespecífica. El análisis histológico de las fibras musculares de los animales del Grupo con restricción mostró la presencia de núcleos centralizados y una disminución del área en el grupo con restricción. En el estudio histoenzimológico, el músculo EDL presentó una distribución aleatoria de los diferentes tipos de fibras musculares y el área de las fibras musculares del tipo IIa fue menor en el grupo con restricción. En relación al estudio ultraestructural, en los animales del grupo con restricción se observó desorganización de la línea Z, presencia de pequeñas gotas de lípidos y vacuolas que abrigaban figuras de mielina en las regiones subsarcolemal e intramiofibrilar. En el análisis de las JNMs no hubo diferencias significativas. La restricción proteica impuesta en el periodo de gestación y lactancia puede haber afectado el desarrollo del músculo esquelético, produciendo un déficit permanente en las fibras musculares del músculo EDL de la prole.

Three days of intermittent stretching after muscle disuse alters the proteins involved in force transmission in muscle fibers in weanling rats

The aim of this study was to determine the effects of intermittent passive manual stretching on various proteins involved in force transmission in skeletal muscle. Female Wistar weanling rats were randomly assigned to 5 groups: 2 control groups containing 21- and 30-day-old rats that received neither immobilization nor stretching, and 3 test groups that received 1) passive stretching over 3 days, 2) immobilization for 7 days and then passive stretching over 3 days, or 3) immobilization for 7 days. Maximal plantar flexion in the right hind limb was imposed, and the stretching protocol of 10 repetitions of 30 s stretches was applied. The soleus muscles were harvested and processed for HE and picrosirius staining; immunohistochemical analysis of collagen types I, III, IV, desmin, and vimentin; and immunofluorescence labeling of dystrophin and CD68. The numbers of desmin- and vimentin-positive cells were significantly decreased compared with those in the control following immobilization, regardless of whether stretching was applied (P<0.05). In addition, the semi-quantitative analysis showed that collagen type I was increased and type IV was decreased in the immobilized animals, regardless of whether the stretching protocol was applied. In conclusion, the largest changes in response to stretching were observed in muscles that had been previously immobilized, and the stretching protocol applied here did not mitigate the immobilization-induced muscle changes. Muscle disuse adversely affected several proteins involved in the transmission of forces between the intracellular and extracellular compartments. Thus, the 3-day rehabilitation period tested here did not provide sufficient time for the muscles to recover from the disuse maladaptations in animals undergoing postnatal development.