Myosin Heavy Chain Protein Expression Research Articles

Bidirectional effect of uric acid on C2C12 myotubes and its partial mechanism.

To explore the effects and mechanisms of different concentrations of uric acid on skeletal muscle cells. C2C12 myoblasts were differentiated into myotubes and then exposed to a medium containing uric acid (0 μM, 200 μM, 400 μM, 600 μM, 800 μM, 1000 μM, 1200 μM, 1400 μM). The myotube diameters were observed under light microscopy; the expressions of myosin heavy chain (MyHC), autophagy-related proteins (LC3BII/LC3BI, P62), cGAS, and p-Sting/Sting proteins were analyzed using Western blotting or immunoprecipitation; and oxidative stress and mitochondrial damage were evaluated using ROS, mtDNA and JC-1 assays. Cell viability was measured via CCK8 assay, and 1000-μM uric acid was selected for follow-up experiments. Furthermore, C2C12 myotubes were divided into a blank control group (Ctrl), a high-uric-acid group (HUA), and an HUA plus cGASn inhibitor group (HUA + RU.521). Then, the myotube diameter was observed, oxidative stress and mitochondrial damage were evaluated, and MyHC and autophagy-related protein expressions were analysed. C2C12 myotubes cultured in 400-μM uric acid medium had the greatest myotube diameter and the highest MyHC protein expression. At 1000-μM uric acid, the diameter and MyHC protein expression were significantly decreased, LCB3II/LCB3I expression was notably increased, and the level of p62 protein expression was considerably decreased. RU.521 partially alleviated the HUA-induced C2C12 myotubes changes. Uric acid bidirectionally affected C2C12 myotubes: 400-μΜ uric acid promoted myotube growth, while 1000-μΜ uric acid triggered myotube atrophy with increased autophagy. Inhibiting cGAS-Sting signaling attenuated HUA-induced C2C12 myotube autophagy and atrophy. Geriatr Gerontol Int 2024; 24: 430-439.

Effect of resveratrol on skeletal slow-twitch muscle fiber expression via AMPK/PGC-1α signaling pathway in bovine myotubes

The purpose of this study was to evaluate the impact of resveratrol on slow-twitch muscle fiber expression in bovine myotubes. The results revealed that resveratrol enhanced slow myosin heavy chain (MyHC) and suppressed fast MyHC protein expression, accompanied by increased MyHC I/IIa and decreased MyHC IIx/IIb mRNA levels in bovine myotubes (P < 0.05). Resveratrol also enhanced the activities of succinic dehydrogenase (SDH), malate dehydrogenase (MDH) and the mitochondrial DNA (mtDNA) content, but reduced lactate dehydrogenase (LDH) activity (P < 0.05). Meanwhile, the protein and gene expression of AMPK, SIRT1 and PGC-1α were upregulated by resveratrol (P < 0.05). Furthermore, PGC-1α inhibitor SR-18292 could attenuate resveratrol-induced muscle fiber conversion from fast-twitch to slow-twitch. These results suggest that resveratrol might promote muscle fiber type transition from fast-twitch to slow-twitch through the AMPK/PGC-1α signaling pathway and mitochondrial biogenesis in bovine myotubes.

TESTOSTERONE INDUCES POSTMENOPAUSAL HYPERTENSIVE HYPERTROPHY THROUGH INHIBITING THE ACTIVATION OF AMPK- FOXO1/MURF1 SIGNALING PATHWAY

Objective: It has been found that relative elevated level of testosterone could exacerbates postmenopausal hypertensive left ventricular hypertrophy(PM-HTN-LVH), but the mechanisms responsible are unknown. Moreover, MuRF1, a muscle-specific E3 ubiquitin ligase in the ubiquitination process, was newly identified as a negative regulatory factor of myocardial and muscle tissue in exerting a protective functions on the cardiomyocyte hypertrophy. There are little study to certify whether MuRF1 took part in testosterone-induced PM-HTN-LVH. Design and method: Femal spontaneously hypertensive rats (SHR) were ovariectomized (OVX group) or sham-operated (Sham group) at 12 weeks and then the OVX group were intramuscularly injected the testosterone (T 2.85 mg/kg/ quaque omni die im) (OVX+T group) or vehicle (OVX+ vehicle group) for 4 weeks. Futher, Cultured cardiomyocytes were respectively stimulated with different concentration gradients of testosterone for 24h, then measured the expression of myosin heavy chain (MHC) and MuRF1, as well as the phosphorylation levels of AMPK(p-AMPK) and Forkhead box O 1 (p-FOXO1), were separately measured using western blot. Results: After 4 weeks of testosterone supplementation, SHR heart exhibited significant increments in the heart weight/tibial length and the levels of IVST, LVPWT, LVM and the expression of MHC, and have a lower expression of the MuRF1, p-AMPK and p-FOXO1 in the OVX+T group, compared with OVX group and Sham group. Moreover, the expression level of p-AMPK and p-FOXO1 in the heart were at least twice as many as gastrocnemius muscle among the three group, but the MuRF1 was equivalent between heart and gastrocnemius muscle. In vivo, after the treatment of different gradients of testosterone for 24h, the protein expression of MHC showed an S-shaped growth trend and MuRF1, p-AMPK and p-FOXO1 presented with reverse S-shaped reduction along with increasing of testosterone concentration, peaking at 10-6M testosterone. Conclusions: The present study demonstrates that testosterone induces PM-HTN-LVH by modulating the atrophy related AMPK-FOXO1/MuRF1 signaling pathway in vitro and vivo. Increased the heart expression of MuRF1 by activation of AMPK may reverse testosterone-mediated PM-HTN-LVH, which might cause little change of MuRF1 expression in the skeletal muscle resulting in the muscle atrophy.

Palmitic Acid Inhibits Myogenic Activity and Expression of Myosin Heavy Chain MHC IIb in Muscle Cells through Phosphorylation-Dependent MyoD Inactivation.

Sarcopenia associated with aging and obesity is characterized by the atrophy of fast-twitch muscle fibers and an increase in intramuscular fat deposits. However, the mechanism of fast-twitch fiber-specific atrophy remains unclear. In this study, we aimed to assess the effect of palmitic acid (PA), the most common fatty acid component of human fat, on muscle fiber type, focusing on the expression of fiber-type-specific myosin heavy chain (MHC). Myotubes differentiated from C2C12 myoblasts were treated with PA. The PA treatment inhibited myotube formation and hypertrophy while reducing the gene expression of MHC IIb and IIx, specific isoforms of fast-twitch fibers. Consistent with this, a significant suppression of MHC IIb protein expression in PA-treated cells was observed. A reporter assay using plasmids containing the MHC IIb gene promoter revealed that the PA-induced reduction in MHC IIb gene expression was caused by the suppression of MyoD transcriptional activity through its phosphorylation. Treatment with a specific protein kinase C (PKC) inhibitor recovered the reduction in MHC IIb gene expression levels in PA-treated cells, suggesting the involvement of the PA-induced activation of PKC. Thus, PA selectively suppresses the mRNA and protein expression of fast-twitch MHC by modulating MyoD activity. This finding provides a potential pathogenic mechanism for age-related sarcopenia.

Effects of dietary l-theanine supplementation on pork quality and muscle fiber type transformation in finishing pigs.

This experiment aimed to investigate effects of dietary l-theanine supplementation on pork quality and muscle fiber type transformation in finishing pigs. In a 30-day experiment, 18 healthy Duroc × Landrace × Yorkshire (DLY) pigs with an average body weight of 86.03 ± 0.83 kg were randomly divided into three groups (a basal diet or a basal diet supplemented with 500 and 1000 ppml-theanine, respectively), with six duplicates and one pig per replicate. The results showed that dietary 1000 ppml-theanine supplementation significantly reduced (P < 0.05) b*24 h and drip loss. Dietary 1000 ppml-theanine supplementation significantly increased (P < 0.05) slow myosin heavy chain (MyHC) protein expression and the percentage of slow-twitch fibers, as well as significantly decreased (P < 0.05) fast MyHC protein expression and the percentage of fast-twitch fibers, accompanied by an increase in succinate dehydrogenase (SDH) and malate dehydrogenase (MDH) activities and a decrease in lactate dehydrogenase (LDH) activity. In addition, the adenosine monophosphate (AMP)-activated protein kinase (AMPK) signaling pathway was activated by l-theanine. Together, this study demonstrated for the first time that dietary supplementation of 1000 ppml-theanine can improve pork color and drip loss and promote muscle fiber type transformation from fast-twitch to slow-twitch in finishing pigs. © 2022 Society of Chemical Industry.

Ellagic Acid Alters Muscle Fiber-Type Composition and Promotes Mitochondrial Biogenesis through the AMPK Signaling Pathway in Healthy Pigs.

Ellagic acid (EA), because of its remarkable health-promoting ability, has aroused widespread interest in the fields of nutrition and medicine. However, no reports showed that EA regulates mitochondrial biogenesis as well as muscle fiber-type composition in pigs. Our study found that dietary 75 and 150 mg/kg EA obviously augmented the slow myosin heavy chain (MyHC) protein level, the number of slow-twitch muscle fibers, and the activity of malate dehydrogenase (MDH) in the longissimus thoracis (LT) muscle of growing-finishing pigs. In contrast, dietary 75 and 150 mg/kg EA decreased the fast MyHC level, the number of fast-twitch muscle fibers, and the activity of lactate dehydrogenase (LDH) in the LT muscle. In addition, our further study found that dietary 75 and 150 mg/kg EA promoted the mitochondrial DNA (mtDNA) content, the mRNA expressions of ATP synthase (ATP5G), mtDNA transcription factor A (TFAM), AMP-activated protein kinase α1 (AMPKα1), peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) and sirtuin 1 (Sirt1), and the level of phospho-LKB1 (P-LKB1), phospho-AMPK (P-AMPK), Sirt1, and PGC-1α in the LT muscle. In vitro, 5, 10, and 20 μmol/L EA treatment upregulated the level of slow MyHC, but only 10 μmol/L EA treatment decreased fast MyHC protein expression in porcine skeletal muscle satellite cells (PSCs). In addition, our data again found that 10 μmol/L EA treatment promoted the mtDNA content, the mRNA levels of ATP5G, mitochondrial transcription factor b1 (TFB1M), citrate synthase (Cs), AMPKα1, PGC-1α, and Sirt1, and the protein expressions of P-AMPK, P-LKB1, PGC-1α, and Sirt1 in PSCs. What is more, inhibition of the AMPK signaling pathway by AMPKα1 siRNA significantly eliminated the improvement of EA on muscle fiber-type composition as well as the mtDNA content in PSCs. In conclusion, EA altered muscle fiber-type composition and promoted mitochondrial biogenesis through the AMPK signaling pathway.

2,6-Dimethoxy-1,4-benzoquinone increases skeletal muscle mass and performance by regulating AKT/mTOR signaling and mitochondrial function

Background2,6-Dimethoxy-1,4-benzoquinone (DMBQ), a natural phytochemical present in fermented wheat germ, has been reported to exert anti-cancer, anti-inflammatory, and anti-adipogenic effects. However, the effect of DMBQ on muscle hypertrophy and myoblast differentiation has not been elucidated. PurposeWe investigated the effect of DMBQ on skeletal muscle mass and muscle function and then determined the possible mechanism of DMBQ. MethodsTo examine myogenic differentiation and hypertrophy, confluent C2C12 cells were incubated in differentiation medium with or without various concentrations of DMBQ for 4 days. In animal experiments, C57BL/6 mice were fed DMBQ-containing AIN-93 diet for 7 weeks. Grip strength, treadmill, microscopic evaluation of muscle tissue, western blotting, and quantitative real-time PCR were performed. ResultsDMBQ significantly increased fusion index, myotube size, and the protein expression of myosin heavy chain (MHC). DMBQ increased the phosphorylation of protein kinase B (AKT) and p70 ribosomal protein S6 kinase (S6K), whereas the phosphorylation of these proteins was abolished by the phosphoinositide 3-kinase inhibitor LY294002 in C2C12 cells. In addition, DMBQ treatment increased peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1α), which programs mitochondrial biogenesis, protein levels compared with control C2C12 cells. DMBQ significantly increased maximal respiration and spare respiratory capacity in C2C12 cells. In animal experiments, DMBQ increased skeletal muscle weights and skeletal muscle fiber size compared with the control group values. In addition, the DMBQ group showed increased grip strength and running distance on an accelerating treadmill. The protein expression of total MHC, MHC1, MHC2A, and MHC2B in skeletal muscle was upregulated by DMBQ supplementation. We found that DMBQ increased the phosphorylation of AKT and mammalian target of rapamycin (mTOR), as well as downstream S6K and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) in skeletal muscle. DMBQ also stimulated mRNA expression of PGC1α, accompanied by an increase in mitochondrial DNA content, oxidative phosphorylation (OXPHOS) proteins, and oxidative enzyme activity. ConclusionCollectively, DMBQ was shown to increase skeletal muscle mass and performance by regulating the AKT/mTOR signaling pathway and enhancing mitochondrial function, which might be useful for the treatment and prevention of skeletal muscle atrophy.

Naringin induces skeletal muscle fiber type transformation via AMPK/PGC-1α signaling pathway in mice and C2C12 myotubes

A large number of studies have shown that polyphenols can regulate skeletal muscle fiber type transformation through AMPK signal. However, the effects and mechanism of naringin (a natural polyphenol) on muscle fiber type transformation still remains unclear. Thus, we hypothesized that naringin would induce the transformation of skeletal muscle fibers from type II to type I by AMPK signaling. C2C12 myotubes and BALB/c mice models were used to test this hypothesis. We found that naringin significantly increased the protein expression of slow myosin heavy chain (MyHC), myoglobin and troponin I type I slow skeletal (Troponin I-SS) and the activities of succinate dehydrogenase (SDH) and malate dehydrogenase (MDH), and significantly decreased fast MyHC protein expression and lactate dehydrogenase (LDH) activity, accompanied by the activation of AMPK and the activity of peroxisome proliferator activated receptor-γ coactivator-1α (PGC-1α) in mice and C2C12 myotubes. Further inhibition of AMPK activity by compound C showed that the above effects were significantly inhibited in C2C12 myotubes. In conclusion, naringin promotes the transformation of skeletal muscle fibers from type II to type I through AMPK/PGC-1α signaling pathway, which not only enriches the nutritional and physiological functions of naringin, but also provides a theoretical basis for the regulation of muscle fiber type transformation by nutritional approaches.

Ablation of VDR Induced Atrophy of Skeletal Muscle via Locally Affecting the Renin‐Angiotensin System

Objective Our previous studies demonstrated that vitamin D is a vital protector against tissue injuries by locally suppressing renin-angiotensin system (RAS). It is known that vitamin D deficiency is associated with a range of muscle disorders, such as muscle atrophy. This study aimed to investigate the biological role of vitamin D receptor (VDR) in skeletal muscle and the underlying mechanism using a mice model with deletion of VDR associated with dexamethasone-induced muscle atrophy. Materials and Methods The wild type and VDR(-/-) mice were treated with vehicle and dexamethasone (i.p., once daily) for 10 consecutive days. The murine myoblast cell line C2C12 and the skeletal muscle satellite cells were applied as in vitro models in this study. Results Treatment with the angiotensin II (Ang II) type 1 receptor blocker olmesartan eliminated Ang II-induced atrophy of myotube cells, reflected by alleviating myotube cell atrophy, suggesting that Ang II could aggravate atrophy of skeletal muscle. The ablation of VDR in muscle satellite cells worsened the phenotype of muscle atrophy induced by Ang II. After challenging to dexamethasone, VDR-null mice exhibited more severe amyotrophy compared with wild-type counterparts, manifested by significant decrease in muscle wet weight coefficient of the tibialis anterior muscle and gastrocnemius as well as the reduction of limb grip and weight-loading swimming time. Similarly, the dystrophin fluorescence staining and the haematoxylin eosin (H&E) staining showed much more decrease in cross-sectional area of skeletal muscle in the VDR-null mice in a comparison with the counterparts. MAFbx and MuRF1 are two E3 ubiquitin ligases that are important regulators for ubiquitin-mediated protein degradation in skeletal muscle, while myogenic differentiation-1 (MyoD) and myosin heavy chain (MHC) are necessary to maintain myocytes proliferation and differentiation. VDR ablation significantly accelerated the up-regulation of MAFbx and MuRF1 proteins and down-regulated protein expression of MyoD and MHC in skeletal muscle tissue of mice with muscle atrophy. Conclusion These results provided evidences that VDR might attenuate skeletal muscle atrophy at least in part by suppressing the renin-angiotensin system.

Smyd1 is essential for myosin expression and sarcomere organization in craniofacial, extraocular, and cardiac muscles

Skeletal and cardiac muscles are striated myofibers that contain highly organized sarcomeres for muscle contraction. Recent studies revealed that Smyd1, a lysine methyltransferase, plays a key role in sarcomere assembly in heart and trunk skeletal muscles. However, Smyd1 expression and function in craniofacial muscles are not known. Here, we analyze the developmental expression and function of two smyd1 paralogous genes, smyd1a and smyd1b, in craniofacial and cardiac muscles of zebrafish embryos. Our data show that loss of smyd1a (smyd1amb5) or smyd1b (smyd1bsa15678) has no visible effects on myogenic commitment and expression of myod and myosin heavy-chain mRNA transcripts in craniofacial muscles. However, myosin heavy-chain protein accumulation and sarcomere organization are dramatically reduced in smyd1bsa15678 single mutant, and almost completely diminish in smyd1amb5; smyd1bsa15678 double mutant, but not in smyd1amb5 mutant. Similar defects are also observed in cardiac muscles of smyd1bsa15678 mutant. Defective craniofacial and cardiac muscle formation is associated with an upregulation of hsp90α1 and unc45b mRNA expression in smyd1bsa15678 and smyd1amb5; smyd1bsa15678 mutants. Together, our studies indicate that Smyd1b, but not Smyd1a, plays a key role in myosin heavy-chain protein expression and sarcomere organization in craniofacial and cardiac muscles. Loss of smyd1b results in muscle-specific stress response.

Arginine promotes skeletal muscle fiber type transformation from fast-twitch to slow-twitch via Sirt1/AMPK pathway

This study investigated the effect of arginine on skeletal muscle fiber type transformation in mice and in C2C12 myotubes. Our data showed that dietary supplementation of arginine in mice significantly up-regulated the slow myosin heavy chain (MyHC), troponin I-SS, sirtuin1 (Sirt1) and peroxisome proliferator activated receptor-γ coactivator-1α (PGC-1α) protein expressions, as well as significantly down-regulated the fast MyHC protein expression. In C2C12 myotubes, arginine significantly increased the protein level of slow MyHC and the number of slow MyHC-positive cells, as well as significantly decreased the protein level of fast MyHC and the number of fast MyHC-positive cells. We also showed that arginine increased the activities of succinic dehydrogenase and malate dehydrogenase and decreased the activity of lactate dehydrogenase in mice and in C2C12 myotubes. Here we found that AMP-activated protein kinase (AMPK) was activated by arginine in mice and in C2C12 myotubes. However, inhibition of AMPK activity by compound C significantly attenuated the effects of arginine on slow MyHC and fast MyHC expressions in C2C12 myotubes. Finally, we showed that inhibition of Sirt1 expression by EX527 attenuated arginine-induced increase in the protein levels of phospho-AMPK and slow MyHC, the mRNA level of nitric oxide synthase (NOS) and the contents of NOS and NO, as well as decrease in fast MyHC protein level. Together, our findings indicated that arginine promotes skeletal muscle fiber type switching from fast-twitch to slow-twitch via Sirt1/AMPK pathway.

The double‐edged sword of endoplasmic reticulum stress in uremic sarcopenia through myogenesis perturbation

BackgroundSarcopenia is the age‐related degeneration characterized with the decline of skeletal muscle mass, strength, and function. The imbalance of protein synthesis and degradation which jeopardizes immune, hormone regulation, and muscle‐motor neuron connection is the main cause of sarcopenia. There is limited knowledge regarding molecular mechanism of sarcopenia. As the endoplasmic reticulum is the control centre of the protein syntheses and degradation, we hypothesized that endoplasmic reticulum stress and unfolded protein response (UPR) play an important in the development of sarcopenia. Understanding the sarcopenia molecular mechanisms may benefit the therapeutic diagnosis and treatment in the future.MethodsMouse myoblast C2C12 cells are exposed to designated time and concentration of indoxyl sulfate (IS), a uremic toxin of chronic kidney disease. The proliferation, differentiation, and the expression of atrogin 1 are examined. The protein and mRNA expression of IS treated‐C2C12 cells are inspected to distinguish the role of ER stress and oxidative stress underlying the sarcopenia.ResultsIndoxyl sulfate inhibits myoblast differentiation. We demonstrate that as the number of multi‐nuclei myotube decreased, the differentiation markers including myoD, myoG, and myosin heavy chain are also suppressed. Indoxyl sulfate inhibits myoblast proliferation and induces the myotubular atrophy marker atrogin‐1 protein expression. Indoxyl sulfate stimulates eIF2α phosphorylation and XBP1 mRNA splicing in UPR. Interestingly, the oxidative stress is related to eIF2α phosphorylation but not XBP1 mRNA splicing. The eIF2α phosphorylation triggered by IS reduces myoD, myoG, and myosin heavy chain protein expression, which represents the anti‐myogenic modulation on the early differentiation event. The XBP1 mRNA splicing induced by IS, however, is considered the adaptive response to restore the myogenic differentiation.ConclusionsOur studies indicated that the ER stress and UPR modulation are critical in the chronic kidney disease uremic toxin‐accumulated sarcopenia model. We believe that UPR‐related signals showed great potential in clinical application.

Ontogeny of myosin isoform expression and prehensile function in the tail of the gray short-tailed opossum (Monodelphis domestica).

Terrestrial opossums use their semiprehensile tail for grasping nesting materials as opposed to arboreal maneuvering. We relate the development of this adaptive behavior with ontogenetic changes in myosin heavy chain (MHC) isoform expression from 21 days to adulthood. Monodelphis domestica is expected to demonstrate a progressive ability to flex the distal tail up to age 7 mo, when it should exhibit routine nest construction. We hypothesize that juvenile stages (3-7 mo) will be characterized by retention of the neonatal isoform (MHC-Neo), along with predominant expression of fast MHC-2X and -2B, which will transition into greater MHC-1β and -2A isoform content as development progresses. This hypothesis was tested using Q-PCR to quantify and compare gene expression of each isoform with its protein content determined by gel electrophoresis and densitometry. These data were correlated with nesting activity in an age-matched sample of each age group studied. Shifts in regulation of MHC gene transcripts matched well with isoform expression. Notably, mRNA for MHC-Neo and -2B decrease, resulting in little-to-no isoform translation after age 7 mo, whereas mRNA for MHC-1β and -2A increase, and this corresponds with subtle increases in content for these isoforms into late adulthood. Despite the tail remaining intrinsically fast-contracting, a critical growth period for isoform transition is observed between 7 and 13 mo, correlating primarily with use of the tail during nesting activities. Functional transitions in MHC isoforms and fiber type properties may be associated with muscle "tuning" repetitive nest remodeling tasks requiring sustained contractions of the caudal flexors.NEW & NOTEWORTHY Little is understood about skeletal muscle development as it pertains to tail prehensility in mammals. This study uses an integrative approach of relating both MHC gene and protein expression with behavioral and morphometric changes to reveal a predominant fast MHC expression with subtle isoform transitions in caudal muscle across ontogeny. The functional shifts observed are most notably correlated with increased tail grasping for nesting activities.

A Rapid Automated Protocol for Muscle Fiber Population Analysis in Rat Muscle Cross Sections Using Myosin Heavy Chain Immunohistochemistry.

Quantification of muscle fiber populations provides a deeper insight into the effects of disease, trauma, and various other influences on skeletal muscle composition. Various time-consuming methods have traditionally been used to study fiber populations in many fields of research. However, recently developed immunohistochemical methods based on myosin heavy chain protein expression provide a quick alternative to identify multiple fiber types in a single section. Here, we present a rapid, reliable and reproducible protocol for improved staining quality, allowing automatic acquisition of whole cross sections and automatic quantification of fiber populations with ImageJ. For this purpose, embedded skeletal muscles are cut in cross sections, stained using myosin heavy chains antibodies with secondary fluorescent antibodies and DAPI for cell nuclei staining. Whole cross sections are then scanned automatically using a slide scanner to obtain high-resolution composite pictures of the entire specimen. Fiber population analyses are subsequently performed to quantify slow, intermediate and fast fibers using an automated macro for ImageJ. We have previously shown that this method can identify fiber populations reliably to a degree of ±4%. In addition, this method reduces inter-user variability and time per analyses significantly using the open source platform ImageJ.

Moderate-intensity resistance exercise alters skeletal muscle molecular and cellular structure and function in inactive older adults with knee osteoarthritis.

High-intensity resistance exercise (REX) training increases physical capacity, in part, by improving muscle cell size and function. Moderate-intensity REX, which is more feasible for many older adults with disease and/or disability, also increases physical function, but the mechanisms underlying such improvements are not understood. Therefore, we measured skeletal muscle structure and function from the molecular to the tissue level in response to 14 wk of moderate-intensity REX in physically inactive older adults with knee osteoarthritis (n = 17; 70 ± 1 yr). Although REX training increased quadriceps muscle cross-sectional area (CSA), average single-fiber CSA was unchanged because of reciprocal changes in myosin heavy chain (MHC) I and IIA fibers. Intermyofibrillar mitochondrial content increased with training because of increases in mitochondrial size in men, but not women, with no changes in subsarcolemmal mitochondria in either sex. REX increased whole muscle contractile performance similarly in men and women. In contrast, adaptations in single-muscle fiber force production per CSA (i.e., tension) and contractile velocity varied between men and women in a fiber type-dependent manner, with adaptations being explained at the molecular level by differential changes in myosin-actin cross-bridge kinetics and mechanics and single-fiber MHC protein expression. Our results are notable compared with studies of high-intensity REX because they show that the effects of moderate-intensity REX in older adults on muscle fiber size/structure and myofilament function are absent or modest. Moreover, our data highlight unique sex-specific adaptations due to differential cellular and subcellular structural and functional changes.NEW & NOTEWORTHY Moderate-intensity resistance training causes sex-specific adaptations in skeletal muscle structure and function at the cellular and molecular levels in inactive older adult men and women with knee osteoarthritis. However, these responses were minimal compared with high-intensity resistance training. Thus adjuncts to moderate-intensity training need to be developed to correct underlying cellular and molecular structural and functional deficits that are at the root of impaired physical function in this mobility-limited population.

Effects of human umbilical mesenchymal stem cell on Wistar rats models of stress urinary incontinence and its possible mechanism

Objective To investigate the effects of human umbilical mesenchymal stem cells (hUCMSCs) on Wistar rat models of stress urinary incontinence and its possible mechanism. Methods hUCMSCs cells were separated and extracted from human umbilical cord of cesarean delivery of full-term pregnancy women. The models of stress urinary incontinence (SUI) of Wistar rats were established by imitating delivery damage and ovary removing surgery, and then randomly divided into three groups: model group, transforming growth factor-β1 (TGF-β1) group, hUCMSCs group, and normal rats as normal group; and every group included ten rats. Maximal bladder capacity (MBC), leak point pressure (LPP), abdominal leak point pressure (ALPP), and sneezing experiment of rats including normal rats, model rats and model rats accepted TGF-β1 or hUCMSCs treatment were examined. When the treatment accomplished, the rats were killed and urethral sphincter were separated and examined by hematoxylin eosin (HE) staining and the proteins of troponin I (TnI), troponin C (TnC), troponin T (TnT), tropomyosin (Tm), actin (AT), myosin heavy chain (MHC) and myosin light chain (MLC) of urethral sphincter was detected with Western blot method. Results The cells of hUCMSCs were extracted and authenticated, SUI models of Wistar rats were successfully established and authenticated. Compared to model group and TGF-β1 group, the MBC, LPP and ALPP of hUCMSCs group dramatically improved (P<0.05), while the positive rate of sneezing experiment significantly dropped (P<0.05). HE staining showed urethral sphincter became attenuation, fracture, sparse and disorder, simultaneously the proteins of TnI, TnC, TnT, Tm, MHC and MLC of urethral sphincter reduced (P<0.05). While the damaged urethral sphincter basically restored the normal structure and the proteins of TnI, TnC, TnT, Tm, MHC and MLC of urethral sphincter up-regulated through hUCMSCs treatment (P<0.05). Conclusions The treatment of hUCMSCs translation could observably improve the clinic symptoms of SUI Wistar rat models, and its mechanism may be related to enhancement of the protein expression of TnI, TnC, TnT, Tm, MHC and MLC of urethral sphincter, and were involved in the rehabilitation and reconstruction of urethral sphincter. Key words: Cord blood stem cell transplantation; Urinary incontinence, stress/SU/PP; Urethra/PP

Cardioprotective effects of early and late aerobic exercise training in experimental pulmonary arterial hypertension

Clinical studies suggest that aerobic exercise can exert beneficial effects in pulmonary arterial hypertension (PAH), but the underlying mechanisms are largely unknown. We compared the impact of early or late aerobic exercise training on right ventricular function, remodeling and survival in experimental PAH. Male Wistar rats were submitted to normal cage activity (SED), exercise training in early (EarlyEX) and in late stage (LateEX) of PAH induced by monocrotaline (MCT, 60mg/kg). Both exercise interventions resulted in improved cardiac function despite persistent right pressure-overload, increased exercise tolerance and survival, with greater benefits in EarlyEX+MCT. This was accompanied by improvements in the markers of cardiac remodeling (SERCA2a), neurohumoral activation (lower endothelin-1, brain natriuretic peptide and preserved vascular endothelial growth factor mRNA), metabolism and mitochondrial oxidative stress in both exercise interventions. EarlyEX+MCT provided additional improvements in fibrosis, tumor necrosis factor-alpha/interleukin-10 and brain natriuretic peptide mRNA, and beta/alpha myosin heavy chain protein expression. The present study demonstrates important cardioprotective effects of aerobic exercise in experimental PAH, with greater benefits obtained when exercise training is initiated at an early stage of the disease.

Post-transcriptional regulation of myotube elongation and myogenesis by Hoi Polloi

Striated muscle development requires the coordinated expression of genes involved in sarcomere formation and contractility, as well as genes that determine muscle morphology. However, relatively little is known about the molecular mechanisms that control the early stages of muscle morphogenesis. To explore this facet of myogenesis, we performed a genetic screen for regulators of somatic muscle morphology in Drosophila, and identified the putative RNA-binding protein (RBP) Hoi Polloi (Hoip). Hoip is expressed in striated muscle precursors within the muscle lineage and controls two genetically separable events: myotube elongation and sarcomeric protein expression. Myotubes fail to elongate in hoip mutant embryos, even though the known regulators of somatic muscle elongation, target recognition and muscle attachment are expressed normally. In addition, a majority of sarcomeric proteins, including Myosin Heavy Chain (MHC) and Tropomyosin, require Hoip for their expression. A transgenic MHC construct that contains the endogenous MHC promoter and a spliced open reading frame rescues MHC protein expression in hoip embryos, demonstrating the involvement of Hoip in pre-mRNA splicing, but not in transcription, of muscle structural genes. In addition, the human Hoip ortholog NHP2L1 rescues muscle defects in hoip embryos, and knockdown of endogenous nhp2l1 in zebrafish disrupts skeletal muscle development. We conclude that Hoip is a conserved, post-transcriptional regulator of muscle morphogenesis and structural gene expression.

Lifelong Physical Activity Prevents Aging-Associated Insulin Resistance in Human Skeletal Muscle Myotubes via Increased Glucose Transporter Expression

Both aging and physical inactivity are associated with increased development of insulin resistance whereas physical activity has been shown to promote increased insulin sensitivity. Here we investigated the effects of physical activity level on aging-associated insulin resistance in myotubes derived from human skeletal muscle satellite cells. Satellite cells were obtained from young (22 yrs) normally active or middle-aged (56.6 yrs) individuals who were either lifelong sedentary or lifelong active. Both middle-aged sedentary and middle-aged active myotubes had increased p21 and myosin heavy chain protein expression. Interestingly MHCIIa was increased only in myotubes from middle-aged active individuals. Middle-aged sedentary cells had intact insulin-stimulated Akt phosphorylation however, the same cell showed ablated insulin-stimulated glucose uptake and GLUT4 translocation to the plasma membrane. On the other hand, middle-aged active cells retained both insulin-stimulated increases in glucose uptake and GLUT4 translocation to the plasma membrane. Middle-aged active cells also had significantly higher mRNA expression of GLUT1 and GLUT4 compared to middle-aged sedentary cells, and significantly higher GLUT4 protein. It is likely that physical activity induces a number of stable adaptations, including increased GLUT4 expression that are retained in cells ex vivo and protect, or delay the onset of middle-aged-associated insulin resistance. Additionally, a sedentary lifestyle has an impact on the metabolism of human myotubes during aging and may contribute to aging-associated insulin resistance through impaired GLUT4 localization.

Skeletal Muscle Adaptation in Response to Supervised Exercise Training for Intermittent Claudication

ObjectivesThere is evidence that the improvement following supervised exercise for claudication results from skeletal muscle adaptation. The myosin heavy chain (MHC) determines muscle fibre type and therefore efficiency. Immunohistochemical analysis has failed to take account of hybrid MHC expression within myofibres. This study sought evidence of differential MHC protein expression following supervised exercise for claudication. Design38 claudicants were recruited. Subjects undertook a three-month supervised exercise programme. Controls were patients awaiting angioplasty for claudication. Materials and methodsSubjects underwent paired gastrocnemius biopsy. Relative expression of MHC proteins was determined by SDS-PAGE electrophoresis. Non-parametric data is presented as median with the inter-quartile range and parametric as the mean ± standard deviation. ResultsUpon completion of the exercise programme there was a 94% increase (124 (106–145) to 241 (193–265) metres, p = 0.002) in maximum walking distance, which was not evident in the control group. An 11.1% (p = 0.02) increase in MHC I expression was observed in the exercise but not the control group (34.3% ± 6.8 to 45.4% ± 4.4). There was a positive correlation between the change in MHC I expression and the improvement in claudication distance (r = 0.69, p < 0.05). ConclusionsSupervised exercise training for claudication results in an increase in the proportion of MHC type I expression within the symptomatic gastrocnemius muscle.