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1.
FASEB J ; 36(8): e22458, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35867073

RESUMO

Mechanosensors control muscle integrity as demonstrated in mice. However, no information is available in human muscle about the distribution of mechanosensors and their adaptations to mechanical loading and environmental conditions (hypoxia). Here, we hypothesized that mechanosensors show fiber-type-specific distributions and that loading and environmental conditions specifically regulate mechanosensors. We randomly subjected 28 healthy males to one of the following groups (n = 7 each) consisting of nine loading sessions within 3 weeks: normoxia moderate (NM), normoxia intensive (NI), hypoxia moderate (HM), and hypoxia intensive (HI). We took six biopsies: pre (T0), 4 h (T1), and 24 h (T2) after the third as well as 4 h (T3), 24 h (T4), and 72 h (T5) after the ninth training session. We analyzed subjects' maximal oxygen consumption (V̇O2 max), maximal power output (Pmax), muscle fiber types and cross-sectional areas (CSA), fiber-type-specific integrin-linked kinase (ILK) localizations as well as ILK, vinculin and talin protein and gene expressions in dependence on loading and environmental conditions. V̇O2 max increased upon NM and HM, Pmax upon all interventions. Fiber types did not change, whereas CSA increased upon NI and HI, but decreased upon HM. ILK showed a type 2-specific fiber type localization. ILK, vinculin, and talin protein and gene expressions differed depending on loading and environmental conditions. Our data demonstrate that mechanosensors show fiber type-specific distributions and that exercise intensities rather than environmental variables influence their profiles in human muscles. These data are the first of their kind in human muscle and indicate that mechanosensors manage the mechanosensing at a fiber-type-specific resolution and that the intensity of mechanical stimulation has a major impact.


Assuntos
Fibras Musculares Esqueléticas , Talina , Humanos , Hipóxia/metabolismo , Masculino , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/metabolismo , Proteínas Serina-Treonina Quinases , Talina/metabolismo , Vinculina
2.
J Exp Biol ; 226(20)2023 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-37721047

RESUMO

In response to a mechanical stimulus, tendons have a slower tissue renewal rate compared with muscles. This could, over time, lead to a higher mechanical demand (experienced strain) for the tendon, especially when a high strain magnitude exercise is repeated without sufficient recovery. The current study investigated the adaptive responses of the human triceps surae (TS) muscle-tendon unit (MTU) and extracellular matrix turnover-related biomarkers to repetitive high tendon strain cyclic loading. Eleven young adult males performed a progressive resistance exercise over 12 consecutive days, consisting of high Achilles tendon (AT) strain cyclic loading (90% MVC) with one leg once a day (LegT1) and the alternate leg three times a day (LegT3). Exercise-related changes in TS MTU mechanical properties and serum concentrations of extracellular matrix turnover-related biomarkers were analysed over the intervention period. Both legs demonstrated similar increases in maximal AT force (∼10%) over the 12 day period of exercise. A ∼20% increase in maximal AT strain was found for LegT3 (P<0.05) after 8 consecutive exercise days, along with a corresponding decrease in AT stiffness. These effects were maintained even after a 48 h rest period. The AT mechanical properties for LegT1 were unaltered. Biomarker analysis revealed no sign of inflammation but there was altered collagen turnover and a delay in collagen type I synthesis. Accordingly, we suggest that tendon is vulnerable to frequent high magnitude cyclic mechanical loading as accumulation of micro-damage can potentially exceed the rate of biological repair, leading to increased maximal tendon strain.

3.
Cell Mol Life Sci ; 79(6): 321, 2022 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-35622133

RESUMO

BACKGROUND: Skeletal muscles (SkM) are mechanosensitive, with mechanical unloading resulting in muscle-devastating conditions and altered metabolic properties. However, it remains unexplored whether these atrophic conditions affect SkM mechanosensors and molecular clocks, both crucial for their homeostasis and consequent physiological metabolism. METHODS: We induced SkM atrophy through 14 days of hindlimb suspension (HS) in 10 male C57BL/6J mice and 10 controls (CTR). SkM histology, gene expressions and protein levels of mechanosensors, molecular clocks and metabolism-related players were examined in the m. Gastrocnemius and m. Soleus. Furthermore, we genetically reduced the expression of mechanosensors integrin-linked kinase (Ilk1) and kindlin-2 (Fermt2) in myogenic C2C12 cells and analyzed the gene expression of mechanosensors, clock components and metabolism-controlling genes. RESULTS: Upon hindlimb suspension, gene expression levels of both core molecular clocks and mechanosensors were moderately upregulated in m. Gastrocnemius but strongly downregulated in m. Soleus. Upon unloading, metabolism- and protein biosynthesis-related genes were moderately upregulated in m. Gastrocnemius but downregulated in m. Soleus. Furthermore, we identified very strong correlations between mechanosensors, metabolism- and circadian clock-regulating genes. Finally, genetically induced downregulations of mechanosensors Ilk1 and Fermt2 caused a downregulated mechanosensor, molecular clock and metabolism-related gene expression in the C2C12 model. CONCLUSIONS: Collectively, these data shed new lights on mechanisms that control muscle loss. Mechanosensors are identified to crucially control these processes, specifically through commanding molecular clock components and metabolism.


Assuntos
Relógios Biológicos , Mecanorreceptores , Músculo Esquelético , Atrofia Muscular , Animais , Relógios Biológicos/genética , Relógios Biológicos/fisiologia , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Expressão Gênica , Elevação dos Membros Posteriores , Masculino , Mecanorreceptores/metabolismo , Mecanotransdução Celular/genética , Mecanotransdução Celular/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/genética , Atrofia Muscular/metabolismo , Doenças Musculares/genética , Doenças Musculares/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo
4.
Int J Mol Sci ; 24(9)2023 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-37175428

RESUMO

Mechanosensing plays an essential role in maintaining tissue functions. Across the human body, several tissues (i.e., striated muscles, bones, tendons, ligaments, as well as cartilage) require mechanical loading to exert their physiological functions. Contrary, mechanical unloading triggers pathological remodeling of these tissues and, consequently, human body dysfunctions. At the cellular level, both mechanical loading and unloading regulate a wide spectrum of cellular pathways. Among those, pathways regulated by oxidants such as reactive oxygen species (ROS) represent an essential node critically controlling tissue organization and function. Hence, a sensitive balance between the generation and elimination of oxidants keeps them within a physiological range. Here, the Nuclear Factor-E2-related factor 2/Antioxidant response element (Nrf2/ARE) system plays an essential role as it constitutes the major cellular regulation against exogenous and endogenous oxidative stresses. Dysregulations of this system advance, i.a., liver, neurodegenerative, and cancer diseases. Herein, we extend our comprehension of the Nrf2 system to the aforementioned mechanically sensitive tissues to explore its role in their physiology and pathology. We demonstrate the relevance of it for the tissues' functionality and highlight the imperative to further explore the Nrf2 system to understand the physiology and pathology of mechanically sensitive tissues in the context of redox biology.


Assuntos
Elementos de Resposta Antioxidante , Fator 2 Relacionado a NF-E2 , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Mecanotransdução Celular , Fator 2 Relacionado a NF-E2/metabolismo , Oxidantes , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo
5.
Nat Mater ; 20(6): 892-903, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33495631

RESUMO

The basement membrane (BM) is a special type of extracellular matrix and presents the major barrier cancer cells have to overcome multiple times to form metastases. Here we show that BM stiffness is a major determinant of metastases formation in several tissues and identify netrin-4 (Net4) as a key regulator of BM stiffness. Mechanistically, our biophysical and functional analyses in combination with mathematical simulations show that Net4 softens the mechanical properties of native BMs by opening laminin node complexes, decreasing cancer cell potential to transmigrate this barrier despite creating bigger pores. Our results therefore reveal that BM stiffness is dominant over pore size, and that the mechanical properties of 'normal' BMs determine metastases formation and patient survival independent of cancer-mediated alterations. Thus, identifying individual Net4 protein levels within native BMs in major metastatic organs may have the potential to define patient survival even before tumour formation. The ratio of Net4 to laminin molecules determines BM stiffness, such that the more Net4, the softer the BM, thereby decreasing cancer cell invasion activity.


Assuntos
Membrana Basal/metabolismo , Fenômenos Mecânicos , Metástase Neoplásica , Fenômenos Biomecânicos , Linhagem Celular Tumoral , Humanos , Netrinas/metabolismo
6.
Medicina (Kaunas) ; 57(7)2021 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-34357009

RESUMO

Background and Objectives: Angiogenesis describes the outgrowth of new capillaries from already existing ones. Different biomarkers regulate this process. Physical exercise and hypoxia are key stimuli for the activation of different angiogenic molecules, such as the vascular endothelial growth factor (VEGF). matrix metalloproteases (MMPs)-2 and -9 or the extracellular matrix cleavage fragment endostatin. The present study aimed to investigate influences of short-term, intensive cycling exercise under both normoxic and normobaric hypoxic conditions on the mentioned parameters. Materials and Methods: Twelve male subjects (age: 23.3 ± 2.0 years) participated in the study. All subjects conducted four intensive cycling tests until individual exhaustion in a randomized order under the following conditions: normoxia, 2000 m, 3000 m and 4000 m above sea level. Blood samples were taken before (pre) and 10 min, 30 min, 60 min and 240 min post exercise and were analyzed by ELISA. Results: VEGF showed a significantly reduced concentration compared to the pre-value solely under 4000 m at 10 min post exercise. MMP-2 showed significantly reduced concentrations at 240 min post exercise under 4000 m. MMP-9 increased at 240 min post exercise under both 2000 m and 4000 m conditions. Endostatin was significantly increased at 10 min post exercise independently of the applied stimulus. Conclusions: The presented data show that intensive short-term exercise bouts facilitate the bioavailability of angiogenic, ECM (extracellular matrix)-related biomarkers. This finding is interesting for both health- and performance-related research as it demonstrates the positive effects of intensive short exercise interventions.


Assuntos
Exercício Físico , Fator A de Crescimento do Endotélio Vascular , Adulto , Biomarcadores , Humanos , Hipóxia , Masculino , Metaloproteinase 2 da Matriz , Adulto Jovem
7.
Cell Mol Life Sci ; 76(15): 2987-3004, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30701284

RESUMO

Mechanosensors govern muscle tissue integrity and constitute a subcellular structure known as costameres. Costameres physically link the muscle extracellular matrix to contractile and signaling 'hubs' inside muscle fibers mainly via integrins and are localized beneath sarcolemmas of muscle fibers. Costameres are the main mechanosensors converting mechanical cues into biological events. However, the fiber type-specific costamere architecture in muscles is unexplored. We hypothesized that fiber types differ in the expression of genes coding for costamere components. By coupling laser microdissection to a multiplex tandem qPCR approach, we demonstrate that type 1 and type 2 fibers indeed show substantial differences in their mechanosensor complexes. We confirmed these data by fiber type population-specific protein analysis and confocal microscopy-based localization studies. We further show that knockdown of the costamere gene integrin-linked kinase (Ilk) in muscle precursor cells results in significantly increased slow-myosin-coding Myh7 gene, while the fast-myosin-coding genes Myh1, Myh2, and Myh4 are downregulated. In parallel, protein synthesis-enhancing signaling molecules (p-mTORSer2448, p < 0.05; p-P70S6KThr389, tendency with p < 0.1) were reduced upon Ilk knockdown. However, overexpression of slow type-inducing NFATc1 in muscle precursor cells did not change Ilk or other costamere gene expressions. In addition, we demonstrate fiber type-specific costamere gene regulation upon mechanical loading and unloading conditions. Our data imply that costamere genes, such as Ilk, are involved in the control of muscle fiber characteristics. Further, they identify costameres as muscle fiber type-specific loading management 'hubs' and may explain adaptation differences of muscle fiber types to mechanical (un)loading.


Assuntos
Costâmeros/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Actinina/genética , Actinina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Linhagem Celular , Costâmeros/genética , Regulação da Expressão Gênica , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Cadeias Pesadas de Miosina/genética , Cadeias Pesadas de Miosina/metabolismo , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Estresse Mecânico , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
8.
Cell Physiol Biochem ; 52(3): 633-652, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30907990

RESUMO

BACKGROUND/AIMS: Calcium (Ca²âº) coordinates skeletal muscle functions by controlling contractions as well as signaling pathways and transcriptional properties. The ryanodine receptor 1 (RyR1), its phosphorylation site (pRyR1Ser²84°) and its stabilizers navigate Ca²âº oscillations to command muscle signaling cascades and transcriptional activities. While chronic exercise increases pRyR1Ser²84°, investigations on acute exercise's effects on RyR1 and Ca²âº-dependent modifications of skeletal muscle are rare. The aim of this study was to examine molecular events leading to RyR1 phosphorylation in a physiological model of acute exercise. We hypothesized that exercise-induced RyR1 phosphorylation is associated with altered Ca²âº-dependent physiological phenotypes. METHODS: We analyzed pRyR1Ser²84°, its stabilizers, involved signaling pathways, and Ca²âº-sensitive muscle-determining factors (i.e. NFATc1 and epigenetic histone H3 modifications) in rat muscles upon one single running bout of either concentric or eccentric contractions. RESULTS: Both acute exercises significantly increased pRyRSer²84° levels in muscles, which was accompanied by dissociations of stabilizers from RyR1. Additionally, RyR1 phosphorylation-inducing signaling cascades PTEN/CaMKII/ PKA were significantly activated upon exercise. Further, RyR1 phosphorylations were associated with increased Ca²âº-dependent NFATc1 nuclear abundances as well as increased Ca²âº-dependent epigenetic H3 acetylations pointing to a pRyR1Ser²84°-dependent rapid and novel Ca²âº equilibrium upon exercise. CONCLUSION: Our data report synergistic actions of several distinct pathways to modify RyR1 function to govern physiological phenotypes, here expressed as increased nuclear NFATc1 abundances and epigenetic H3 modifications.


Assuntos
Núcleo Celular/metabolismo , Histonas/metabolismo , Músculo Esquelético/metabolismo , Fatores de Transcrição/metabolismo , Acetilação , Actinina/metabolismo , Animais , Cálcio/metabolismo , Domínio Catalítico , Proteínas Quinases Dependentes de AMP Cíclico/química , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Feminino , Contração Muscular , Fosforilação , Condicionamento Físico Animal , Ratos , Ratos Sprague-Dawley , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Sarcômeros/ultraestrutura
9.
Cell Physiol Biochem ; 52(6): 1309-1324, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31050280

RESUMO

BACKGROUND/AIMS: Different approaches have been considered to improve heart reconstructive medicine and direct delivery of pluripotent stem cell-derived cardiomyocytes (PSC-CMs) appears to be highly promising in this context. However, low cell persistence post-transplantation remains a bottleneck hindering the approach. Here, we present a novel strategy to overcome the low engraftment of PSC-CMs during the early post-transplantation phase into the myocardium of both healthy and cryoinjured syngeneic mice. METHODS: Adult murine bone marrow mesenchymal stem cells (MSCs) and PSC-CMs were co-cultured on thermo-responsive polymers and later detached through temperature reduction, resulting in the protease-free generation of cell clusters (micro-tissues) composed of both cells types. Micro-tissues were transplanted into healthy and cryo-injured murine hearts. Short term cell retention was quantified by real-time-PCR. Longitudinal cell tracking was performed by bioluminescence imaging for four weeks. Transplanted cells were further detected by immunofluorescence staining of tissue sections. RESULTS: We demonstrated that in vitro grown micro-tissues consisting of PSC-CMs and MSCs can increase cardiomyocyte retention by >10fold one day post-transplantation, but could not fully rescue a further cell loss between day 1 and day 2. Neutrophil infiltration into the transplanted area was detected in healthy hearts and could be attributed to the cellular implantation rather than tissue damage exerted by the transplantation cannula. Injected PSC-CMs were tracked and successfully detected for up to four weeks by bioluminescence imaging. CONCLUSION: This approach demonstrated that in vitro grown micro-tissues might contribute to the development of cardiac cell replacement therapies.


Assuntos
Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Miocárdio/patologia , Miócitos Cardíacos/transplante , Animais , Células da Medula Óssea/citologia , Linhagem Celular , Rastreamento de Células , Técnicas de Cocultura , Imunidade Inata , Masculino , Células-Tronco Mesenquimais/metabolismo , Camundongos , Microscopia de Fluorescência , Infarto do Miocárdio/patologia , Infarto do Miocárdio/terapia , Miocárdio/imunologia , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Infiltração de Neutrófilos , Imagem Óptica , Células-Tronco Pluripotentes/citologia , Polímeros/química
10.
J Muscle Res Cell Motil ; 38(3-4): 317-330, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-29190010

RESUMO

Skeletal muscle tissue is mechanosensitive, as it is able to sense mechanical impacts and to translate these into biochemical signals making the tissue adapt. Among its mechanosensitive nature, skeletal muscle tissue is the largest metabolic organ of the human body. Disturbances in skeletal muscle mechanosensing and metabolism cause and contribute to many diseases, i.e. muscular dystrophies/myopathies, cardiovascular diseases, COPD or diabetes mellitus type 2. A less commonly focused muscle-related disorder is clinically known as muscle contractures that derive from cerebral palsy (CP) conditions in young and adults. Muscle contractures are characterized by gradually increasing passive muscle stiffness resulting in complete fixation of joints. Different mechanisms have been identified in CP-related contractures, i.e. altered calcium handling, altered metabolism or altered titin regulation. The muscle-related extracellular matrix (ECM), specifically collagens, plays a role in CP-related contractures. Herein, we focus on mechanically sensitive complexes, known as costameres (Cstms), and discuss their potential role in CP-related contractures. We extend our discussion to the ECM due to the limited knowledge of its role in CP-related contractures. The aims of this review are (1) to summarize CP-related contracture mechanisms, (2) to raise novel hypotheses on the genesis of contractures with a focus on Cstms, and (3) to stimulate novel approaches to study CP-related contractures.


Assuntos
Paralisia Cerebral , Contratura , Matriz Extracelular , Mecanotransdução Celular/genética , Proteínas Musculares , Músculo Esquelético , Adulto , Paralisia Cerebral/genética , Paralisia Cerebral/metabolismo , Paralisia Cerebral/patologia , Paralisia Cerebral/fisiopatologia , Contratura/genética , Contratura/metabolismo , Contratura/patologia , Contratura/fisiopatologia , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Humanos , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia
11.
Eur J Appl Physiol ; 116(11-12): 2281-2292, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27695979

RESUMO

PURPOSE: We aimed to determine whether there are different changes in Achilles tendon (AT) mechanical properties in middle-aged, compared to younger runners that might indicate that tendon fatigue, induced by long-distance running, is age-dependent. METHODS: 27 middle-aged (50-67 years) and 22 younger (21-29 years) participants ran a 21 km route at their own pace (mean and SD: old: 3.1 ± 0.3 m s-1; young: 3.6 ± 0.5 m s-1). We tested for changes in the AT force-elongation relationship using dynamometry and ultrasonography during isometric voluntary ankle plantarflexion ramp contractions, conducted 20-28 h pre-run, immediately pre-run, immediately post-run and 20-28 h post-run. Stride frequency and number were examined to estimate cyclic tensile loading characteristics of the tendon during running. RESULTS: Muscle strength decreased significantly (P < 0.05) in both groups immediately post-run (old: 17 %; young: 11 %) and recovered to baseline within 20-28 h post-run. AT stiffness did not change for the younger adults, whereas the middle-aged adults showed a significant (P < 0.05) decrease in AT stiffness (22 %). However, tendon stiffness recovered to baseline 20-28 h post-run. Middle-aged, compared to young adults, demonstrated significantly (P < 0.05) greater stride frequency and number, but no correlations with tendon fatigue changes were determined (R 2 ≤ 0.038). CONCLUSIONS: The results suggest that the plasticity of the AT in response to short-term mechanical loading may be age dependent and that the AT length-tension properties of middle-aged runners may be more vulnerable to change following running compared to younger athletes. However, the observed AT changes in the middle-aged runners dissipated within 20-28 h post-run, suggesting that a tendon viscoelastic recovery mechanism may occur in vivo.


Assuntos
Tendão do Calcâneo/fisiologia , Envelhecimento/fisiologia , Contração Isométrica/fisiologia , Modelos Biológicos , Resistência Física/fisiologia , Corrida/fisiologia , Tendão do Calcâneo/anatomia & histologia , Adulto , Idoso , Envelhecimento/patologia , Simulação por Computador , Módulo de Elasticidade/fisiologia , Humanos , Masculino , Pessoa de Meia-Idade , Estresse Mecânico , Resistência à Tração/fisiologia , Adulto Jovem
12.
Int J Mol Sci ; 17(5)2016 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-27136539

RESUMO

Protein sumoylation is a posttranslational modification triggered by cellular stress. Because general information concerning the role of small ubiquitin-related modifier (SUMO) proteins in adult skeletal muscle is sparse, we investigated whether SUMO-1 proteins will be subjected to time-dependent changes in their subcellular localization in sarcoplasmic and nuclear compartments of human type I and II skeletal muscle fibers in response to acute stimulation by resistance exercise (RE). Skeletal muscle biopsies were taken at baseline (PRE), 15, 30, 60, 240 min and 24 h post RE from 6 male subjects subjected to a single bout of one-legged knee extensions. SUMO-1 localization was determined via immunohistochemistry and confocal laser microscopy. At baseline SUMO-1 was localized in perinuclear regions of myonuclei. Within 15 and up to 60 min post exercise, nuclear SUMO-1 localization was significantly increased (p < 0.01), declining towards baseline levels within 240 min post exercise. Sarcoplasmic SUMO-1 localization was increased at 15 min post exercise in type I and up to 30 min post RE in type II myofibres. The changing localization of SUMO-1 proteins acutely after intense muscle contractions points to a role for SUMO proteins in the acute regulation of the skeletal muscle proteome after exercise.


Assuntos
Exercício Físico , Fibras Musculares Esqueléticas/metabolismo , Proteína SUMO-1/metabolismo , Adulto , Núcleo Celular/metabolismo , Humanos , Imuno-Histoquímica , Lamina Tipo A/metabolismo , Masculino , Microscopia Confocal , Fibras Musculares Esqueléticas/patologia , Retículo Sarcoplasmático/metabolismo , Adulto Jovem
13.
Pflugers Arch ; 467(6): 1343-56, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25070178

RESUMO

How force development and time under tension (TUT) during resistance exercise (RE) influence anabolic signalling of skeletal muscle is incompletely understood. We hypothesized that high force development during RE is more important for post-exercise-induced signalling than submaximal and fatiguing RE with lower force development but similar TUT. Twenty-two male subjects (24 ± 6 years, 181 ± 9 cm, 79 ± 2 kg) performed three distinct RE modes in the fed state with equal TUT but distinct force output: (i) maximal eccentric RE (ECC, n = 7) three sets, eight reps, 100% eccentric dynamic force; (ii) standard RE (STD, n = 7), three sets, 10 reps, 75% dynamic force; and (iii) high fatiguing single-set RE (HIT, n = 8), 20 reps, 100% eccentric-concentric force; vastus lateralis biopsies were collected at baseline, 15, 30, 60, 240 min and 24 h after RE, and the signalling of mechanosensitive and mammalian target of rapamycin (mTOR)-related proteins was determined. The phosphorylation levels of pFAK(Tyr397), pJNK(Thr183/Tyr185), pAKT(Thr308/Ser473), pmTOR(Ser2448), p4E-BP1(Thr37/46), p70s6k(Thr389)/(Ser421/Thr424) and pS6(Ser235/236) were significantly higher in ECC than those in STD and HIT at several time points (P < 0.01). pJNK(Thr183/Tyr185) and pS6(Ser235/236) levels were significantly higher in type II myofibres in ECC compared with STD and HIT. HIT exerted throughout the weakest signalling response. We conclude that high force development during acute RE is superior for anabolic skeletal muscle signalling than fatiguing RE with lower force output but similar TUT. Our results suggest that this response is substantially driven by the higher activation of type II myofibres during RE.


Assuntos
Contração Muscular , Músculo Esquelético/metabolismo , Treinamento Resistido , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Proteínas de Ciclo Celular , Quinase 2 de Adesão Focal/metabolismo , Humanos , MAP Quinase Quinase 4/metabolismo , Masculino , Fadiga Muscular , Força Muscular , Músculo Esquelético/fisiologia , Fosfoproteínas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Serina-Treonina Quinases TOR/metabolismo
14.
Br J Haematol ; 168(5): 728-36, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25316332

RESUMO

Human red blood cells (RBC) express an active and functional endothelial-like nitric oxide (NO) synthase (RBC-NOS). We report studies on RBC-NOS activity in sickle cell anaemia (SCA), a genetic disease characterized by decreased RBC deformability and vascular dysfunction. Total RBC-NOS content was not significantly different in SCA patients compared to healthy controls; however, using phosphorylated RBC-NOS-Ser(1177) as a marker, RBC-NOS activation was higher in SCA patients as a consequence of the greater activation of Akt (phosphorylated Akt-Ser(473) ). The higher RBC-NOS activation in SCA led to higher levels of S-nitrosylated α- and ß-spectrins, and greater RBC nitrite and nitrotyrosine levels compared to healthy controls. Plasma nitrite content was not different between the two groups. Laser Doppler flowmetric experiments demonstrated blunted microcirculatory NO-dependent response under hyperthermia in SCA patients. RBC deformability, measured by ektacytometry, was reduced in SCA in contrast to healthy individuals, and pre-shearing RBC in vitro did not improve deformability despite an increase of RBC-NOS activation. RBC-NOS activation is high in freshly drawn blood from SCA patients, resulting in high amounts of NO produced by RBC. However, this does not result in improved RBC deformability and vascular function: higher RBC-NO is not sufficient to counterbalance the enhanced oxidative stress in SCA.


Assuntos
Anemia Falciforme/enzimologia , Deformação Eritrocítica , Eritrócitos/enzimologia , Óxido Nítrico Sintase/metabolismo , Estresse Oxidativo , Anemia Falciforme/patologia , Ativação Enzimática , Eritrócitos/patologia , Feminino , Humanos , Masculino , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espectrina/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo
15.
Int J Mol Sci ; 16(1): 1066-95, 2015 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-25569087

RESUMO

Calcium (Ca2+) plays a pivotal role in almost all cellular processes and ensures the functionality of an organism. In skeletal muscle fibers, Ca(2+) is critically involved in the innervation of skeletal muscle fibers that results in the exertion of an action potential along the muscle fiber membrane, the prerequisite for skeletal muscle contraction. Furthermore and among others, Ca(2+) regulates also intracellular processes, such as myosin-actin cross bridging, protein synthesis, protein degradation and fiber type shifting by the control of Ca(2+)-sensitive proteases and transcription factors, as well as mitochondrial adaptations, plasticity and respiration. These data highlight the overwhelming significance of Ca(2+) ions for the integrity of skeletal muscle tissue. In this review, we address the major functions of Ca(2+) ions in adult muscle but also highlight recent findings of critical Ca(2+)-dependent mechanisms essential for skeletal muscle-regulation and maintenance.


Assuntos
Cálcio/metabolismo , Acoplamento Excitação-Contração/fisiologia , Músculo Esquelético/metabolismo , Humanos , Mitocôndrias/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Transdução de Sinais , Fatores de Transcrição/metabolismo
16.
Int J Mol Sci ; 14(4): 7109-39, 2013 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-23538841

RESUMO

Skeletal muscle is responsible for altered acute and chronic workload as induced by exercise. Skeletal muscle adaptations range from immediate change of contractility to structural adaptation to adjust the demanded performance capacities. These processes are regulated by mechanically and metabolically induced signaling pathways, which are more or less involved in all of these regulations. Nitric oxide is one of the central signaling molecules involved in functional and structural adaption in different cell types. It is mainly produced by nitric oxide synthases (NOS) and by non-enzymatic pathways also in skeletal muscle. The relevance of a NOS-dependent NO signaling in skeletal muscle is underlined by the differential subcellular expression of NOS1, NOS2, and NOS3, and the alteration of NO production provoked by changes of workload. In skeletal muscle, a variety of highly relevant tasks to maintain skeletal muscle integrity and proper signaling mechanisms during adaptation processes towards mechanical and metabolic stimulations are taken over by NO signaling. The NO signaling can be mediated by cGMP-dependent and -independent signaling, such as S-nitrosylation-dependent modulation of effector molecules involved in contractile and metabolic adaptation to exercise. In this review, we describe the most recent findings of NO signaling in skeletal muscle with a special emphasis on exercise conditions. However, to gain a more detailed understanding of the complex role of NO signaling for functional adaptation of skeletal muscle (during exercise), additional sophisticated studies are needed to provide deeper insights into NO-mediated signaling and the role of non-enzymatic-derived NO in skeletal muscle physiology.


Assuntos
Exercício Físico/fisiologia , Músculo Esquelético/fisiologia , Óxido Nítrico/metabolismo , Animais , Citoesqueleto/metabolismo , Humanos , Músculo Esquelético/enzimologia , Óxido Nítrico Sintase/metabolismo , Transdução de Sinais
17.
Physiol Rep ; 11(2): e15574, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36695670

RESUMO

Sedentary behavior has become ingrained in our society and has been linked to cardiovascular diseases. Physical inactivity is the main characteristic of sedentary behavior. However, its impact on cardiovascular disease is not clear. Therefore, we investigated the effect of physical inactivity in an established mouse model on gene clusters associated with cardiac fibrosis, electrophysiology, cell regeneration, and tissue degradation/turnover. We investigated a sedentary group (CTR, n = 10) versus a tail suspension group (TS, n = 11) that caused hindlimb unloading and consequently physical inactivity. Through histological, protein content, and transcript analysis approaches, we found that cardiac fibrosis-related genes partly change, with significant TS-associated increases in Tgfb1, but without changes in Col1a1 and Fn1. These changes are not translated into fibrosis at tissue level. We further detected TS-mediated increases in protein degradation- (Trim63, p < 0.001; Fbxo32, p = 0.0947 as well as in biosynthesis-related [P70s6kb1, p < 0.01]). Corroborating these results, we found increased expression of autophagy markers such as Atg7 (p < 0.01) and ULK1 (p < 0.05). Two cardiomyocyte regeneration- and sarcomerogenesis-related genes, Yap (p = 0.0535) and Srf (p < 0.001), increased upon TS compared to CTR conditions. Finally, we found significant upregulation of Gja1 (p < 0.05) and a significant downregulation of Aqp1 (p < 0.05). Our data demonstrate that merely 2 weeks of reduced physical activity induce changes in genes associated with cardiac structure and electrophysiology. Hence, these data should find the basis for novel research directed to evaluate the interplay of cardiac functioning and physical inactivity.


Assuntos
Cardiomiopatias , Doenças Cardiovasculares , Camundongos , Animais , Elevação dos Membros Posteriores/fisiologia , Miócitos Cardíacos , Fibrose , Autofagia
18.
J Appl Physiol (1985) ; 135(4): 918-931, 2023 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-37675473

RESUMO

Mechanosensing and subsequent mechanotransduction are indispensable for muscle plasticity. Nevertheless, a scarcity of literature exists regarding an all-encompassing understanding of the muscle mechanosensing machinery's response to prolonged loading, especially in conditions that resemble a natural physiological state of skeletal muscle. This study aimed to comprehensively explore the effects of prolonged mechanical loading on mechanosensitive components, skeletal muscle characteristics, and metabolism-related gene clusters. Twenty male C57BL/6J mice were randomly divided into two groups: control and prolonged mechanical loading. To induce prolonged mechanical loading on the triceps brachii (TRI) and biceps brachii (BIC) muscles, a 14-day period of tail suspension was implemented. In TRI only, prolonged mechanical loading caused a mild fast-to-slow fiber type shift together with increased mechanosensor gene and protein levels. It also increased transcription factors associated with slow muscle fibers while decreasing those related to fast-type muscle gene expression. Succinate dehydrogenase activity, a marker of muscle oxidative capacity, and genes involved in oxidative and mitochondrial turnover increased, whereas glycolytic-related genes decreased. Moreover, prolonged mechanical loading stimulated markers of muscle protein synthesis. Taken together, our data show a collective muscle-specific increase in mechanosensor gene and protein levels upon a period of prolonged mechanical loading in conditions that reflect a more natural physiological state of skeletal muscle in mice. We provide additional proof-of-concept that prolonged tail suspension-induced loading of the forelimbs triggers a muscle-specific fast-to-slow fiber type switch, and this coincides with increased protein synthesis-related signaling.NEW & NOTEWORTHY This study provides a comprehensive overview of the effects of prolonged loading on mechanosensitive components in conditions that better reflect the natural physiological state of skeletal muscle. Although the muscle mechanosensing machinery has been widely acknowledged for its responsiveness to altered loading, an inclusive understanding of its response to prolonged loading remains scarce. Our results show a fast-to-slow fiber type shift and an upregulation of mechanosensor gene and protein levels following prolonged loading.

19.
Curr Biol ; 32(14): 3033-3047.e9, 2022 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-35688156

RESUMO

Physiological and pathological cardiac stress induced by exercise and hypertension, respectively, increase the hemodynamic load for the heart and trigger specific hypertrophic signals in cardiomyocytes leading to adaptive or maladaptive cardiac hypertrophy responses involving a mechanosensitive remodeling of the contractile cytoskeleton. Integrins sense load and have been implicated in cardiac hypertrophy, but how they discriminate between the two types of cardiac stress and translate mechanical loads into specific cytoskeletal signaling pathways is not clear. Here, we report that the focal adhesion protein ß-parvin is highly expressed in cardiomyocytes and facilitates the formation of cell protrusions, the serial assembly of newly synthesized sarcomeres, and the hypertrophic growth of neonatal rat ventricular cardiomyocytes (NRVCs) in vitro. In addition, physiological mechanical loading of NRVCs by either the application of cyclic, uni-axial stretch, or culture on physiologically stiff substrates promotes NRVC elongation in a ß-parvin-dependent manner, which is achieved by binding of ß-parvin to α/ß-PIX, which in turn activates Rac1. Importantly, loss-of-function studies in mice also revealed that ß-parvin is essential for the exercise-induced cardiac hypertrophy response in vivo. Our results identify ß-parvin as a novel mechano-responsive signaling hub in hypertrophic cardiomyocytes that drives cell elongation in response to physiological mechanical loads.


Assuntos
Adesões Focais , Miócitos Cardíacos , Animais , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Células Cultivadas , Integrinas/metabolismo , Camundongos , Miócitos Cardíacos/metabolismo , Ratos , Sarcômeros/patologia
20.
FASEB J ; 24(8): 2739-51, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20371616

RESUMO

Cardiomyocytes generated from embryonic stem cells (ESCs) and induced pluripotent stem (iPS) cells are suggested for repopulation of destroyed myocardium. Because contractile properties are crucial for functional regeneration, we compared cardiomyocytes differentiated from ES cells (ESC-CMs) and iPS cells (iPS-CMs). Native myocardium served as control. Murine ESCs or iPS cells were differentiated 11 d in vitro and cocultured 5-7 d with irreversibly injured myocardial tissue slices. Vital embryonic ventricular tissue slices of similar age served for comparison. Force-frequency relationship (FFR), effects of Ca(2+), Ni(2+), nifedipine, ryanodine, beta-adrenergic, and muscarinic modulation were studied during loaded contractions. FFR was negative for ESC-CMs and iPS-CMs. FFR was positive for embryonic tissue and turned negative after treatment with ryanodine. In all groups, force of contraction and relaxation time increased with the concentration of Ca(2+) and decreased with nifedipine. Force was reduced by Ni(2+). Isoproterenol (1 microM) increased the force most pronounced in embryonic tissue (207+/-31%, n=7; ESC-CMs: 123+/-5%, n=4; iPS-CMs: 120+/-4%, n=8). EC(50) values were similar. Contractile properties of iPS-CMs and ESC-CMs were similar, but they were significantly different from ventricular tissue of comparable age. The results indicate immaturity of the sarcoplasmic reticulum and the beta-adrenergic response of iPS-CMs and ESC-CMs.


Assuntos
Células-Tronco Embrionárias/citologia , Ventrículos do Coração/citologia , Células-Tronco Pluripotentes Induzidas/citologia , Contração Miocárdica , Miócitos Cardíacos/citologia , Agonistas Adrenérgicos beta/farmacologia , Animais , Fenômenos Biomecânicos , Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio , Técnicas de Cultura de Células , Diferenciação Celular , Técnicas de Cocultura , Camundongos , Nifedipino/farmacologia , Retículo Sarcoplasmático
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