Fabrication of nanofiber coated with l-arginine via electrospinning technique: a novel nanomatrix to counter oxidative stress under crosstalk of co-cultured fibroblasts and satellite cells.

Myofiber size is dynamically regulated, increasing and decreasing depending on muscle use. Hypertrophy is defined by increases in myofiber cross-sectional area and mass as well as myofibrillar protein content. Myofibers contain many hundreds of nuclei, each of which has a nuclear domain. A nuclear

Novel roles of Xin-repeat protein in skeletal muscle: a new insight into monogenetic myopathies.

Autologous Transplantation of SM/C-2.6+ Satellite Cells Transduced with Micro-dystrophin CS1 cDNA by Lentiviral Vector into mdx Mice

Objeetive To study the effect of implantation of muscle satellite cells (MSCs) transtected with myogenic differentiation factor D (MyoD) high expression plasmids on denervated gastrocnemius muscle atrophy in rats.Methods Plasmid of high expression of MyoD was constructed and used to transfect MSCs derived from Sprague-Dawley rats.These transfected MSCs were transplanted into the denervated gastrocnemius muscles in a rat sciatic nerve transection model.At 2 and 4 weeks postoperatively,gastrocnemius muscles were harvested for measurement of wet muscle weight and muscle fiber cross sectional area from H&E stained tissue sections.The expression of MyoD was quantified with real-time PCR.Results Confocal microscopy detected MyoD expression in the transfected MSCs with a transfection efficiency of 43 ％.Gastrocnemius muscle wet weight of the MyoD transfected MSC group was higher than those of the other groups (P ＜ 0.05).The level of MyoD gene expression was higher in the MyoD transfected MSC group than in the other groups (P ＜ 0.05).Muscle cell degeneration and cell death was noted in the control group at 2 and 4 weeks postoperatively.Vacuole formation and myofiber rupture were also observed.Myofiber regeneration was present in the MSC group and MyoD transfected MSC group.Muscle fiber cross sectional area in the MyoD transfected MSC group was higher than the other groups at both 2 and 4 week time points.The differences were statistically significant (P ＜ 0.05).Conclusion Transplantation of high MyoD expression MSCs can delay denervation atrophy of gastrocnemius muscle in rats. Key words: Muscular atrophy; Skeletal muscle satellite cells; Myogenic differentiation factor D; Denervated muscle atrophy; Gene therapy

The process of myogenesis, which involves the growth and differentiation of muscle cells, is a crucial determinant of meat yield and quality in beef cattle. Essential nutrients, such as vitamins D and A, play vital roles in the development and maintenance of various tissues, including muscle. However, limited knowledge exists regarding the specific effects of vitamins A and D in bovine muscle. Therefore, the aim of this study was to investigate the impact of vitamins A and D treatment on myogenic fusion and differentiation in bovine satellite cells (BSC). BSC were isolated from Korean native beef cattle, specifically from four female cows approximately 30 mo old. These individual cows were used as biological replicates (n = 3 or 4), and we examined the effects of varying concentrations of vitamins A (All-trans retinoic acid; 100 nM) and D (1,25-dihydroxy-vitamin D3; 1 nM, 10 nM, and 100 nM), both individually and in combination, on myoblast fusion and myogenic differentiation during the growth phase (48 h) or differentiation phase (6 d). The results were statistically analyzed using GLM procedure of SAS with Tukey's test and t-tests or one-way ANOVA where appropriate. The findings revealed that vitamin A enhanced the myoblast fusion index, while vitamin D treatment decreased the myoblast fusion index during the growth phase. Furthermore, vitamin A treatment during the differentiation phase promoted terminal differentiation by regulating the expression of myogenic regulatory factors (Myf5, MyoD, MyoG, and Myf6) and inducing myotube hypertrophy compared to the control satellite cells (P < 0.01). In contrast, vitamin D treatment during the differentiation phase enhanced myogenic differentiation by increasing the mRNA expression of MyoG and Myf6 (P < 0.01). Moreover, the combined treatment of vitamins A and D during the growth phase increased myoblast fusion and further promoted myogenic differentiation and hypertrophy of myotubes during the differentiation phase (P < 0.01). These results suggest that vitamin A and D supplementation may have differential effects on muscle development in Korean native beef cattle during the feeding process.

In vivo Fluorescence Imaging of Muscle Cell Regeneration by Transplanted EGFP-labeled Myoblasts

Numerical densities of myonuclei and satellite cells in muscle fiber types in the aging human thyroarytenoid muscle: An immunohistochemical and stereological study using confocal laser scanning microscopy

Composed of a diverse variety of cells, the skeletal muscle is one of the body's tissues with the remarkable ability to regenerate after injury. One of the key players in the regeneration process is the muscle satellite cell (MuSC), a stem cell population for skeletal muscle, as it is the source of new myofibers. Maintaining MuSC quiescence during homeostasis involves complex interactions between MuSCs and other cells in their corresponding niche in adult skeletal muscle. After the injury, MuSCs are activated to enter the cell cycle for cell proliferation and differentiate into myotubes, followed by mature myofibers to regenerate muscle. Despite decades of research, the exact mechanisms underlying MuSC maintenance and activation remain elusive. Traditional methods of analyzing MuSCs, including cell cultures, animal models, and gene expression analyses, provide some insight into MuSC biology but lack the ability to replicate the 3-dimensional (3-D) in vivo muscle environment and capture dynamic processes comprehensively. Recent advancements in imaging technology, including confocal, intra-vital, and multi-photon microscopies, provide promising avenues for dynamic MuSC morphology and behavior to be observed and characterized. This chapter aims to review 3-D and live-imaging methods that have contributed to uncovering insights into MuSC behavior, morphology changes, interactions within the muscle niche, and internal signaling pathways during the quiescence to activation (Q-A) transition. Integrating advanced imaging modalities and computational tools provides a new avenue for studying complex biological processes in skeletal muscle regeneration and muscle degenerative diseases such as sarcopenia and Duchenne muscular dystrophy (DMD).

ObjectiveRecovery of the quadriceps femoris muscle after anterior ligament reconstruction is impaired. The aim of this study was to investigate satellite cell content and function of the vastus lateralis muscle after anterior ligament reconstruction.MethodsBiopsies were obtained from the vastus lateralis muscle of 16 recreational athletes immediately before and again 12 weeks after anterior ligament reconstruction. Total satellite cell number (Pax7+), activated (Pax7+/MyoD+), differentiating (Pax7–/MyoD+), and apoptotic (Pax7+/TUNEL+) satellite cells, myofibers expressing myosin heavy chain (MHC) I and II, and neonatal MHC (MHCneo) were determined immunohistochemically.ResultsAfter anterior ligament reconstruction, the number of apoptotic satellite cells was significantly (p = 0.019) increased, concomitant with a significant (p < 0.001) decrease in total satellite cell number, with no change in activated and differentiating satellite cell number. MHCneo+ myofibers tended towards an increase.CONCLUSIONSatellite cell apoptosis and the reduction in the satellite cell pool might provide an explanation for prolonged quadriceps muscle atrophy after anterior ligament reconstruction.LAY ABSTRACTProtracted muscle atrophy is common after anterior ligament reconstruction, even if athletes adhere to a structured rehabilitation programme. Satellite cells, the stem cells of skeletal muscle, play an important role in recovery of an atrophied muscle. Exercise can activate satellite cells, induce their proliferation, and probably also differentiation of these stem cells. The current study evaluated satellite cell content and function in biopsies from the vastus lateralis muscle of 16 recreational athletes immediately before and 12 weeks after anterior ligament reconstruction. After anterior ligament reconstruction, an increased number of satellite cells showed signs of apoptosis (cell death). Furthermore, total satellite cell number was decreased, with no change in the numbers of activated and differentiating satellite cells. The number of regenerating myofibers expressing neonatal myosin tended to increase. In conclusion, satellite cell apoptosis and the reduced satellite cell number might provide an explanation for the impaired muscle recovery after anterior ligament reconstruction.

The graphene‐based nanocomposites are considered as great candidates for enhancing electrical and mechanical properties of nonconductive scaffolds in cardiac tissue engineering. In this study, reduced graphene oxide‐silver (rGO‐Ag) nanocomposites (1 and 2 wt%) were synthesized and incorporated into polyurethane (PU) nanofibers via electrospinning technique. Next, the human cardiac progenitor cells (hCPCs) were seed on these scaffolds for in vitro studies. The rGO‐Ag nanocomposites were studied by X‐ray diffraction (XRD), Raman spectroscopy, and transmission electron microscope (TEM). After incorporation of rGO‐Ag into PU nanofibers, the related characterizations were carried out including scanning electron microscope (SEM), TEM, water contact angle, and mechanical properties. Furthermore, PU and PU/nanocomposites scaffolds were used for in vitro studies, wherein hCPCs showed good cytocompatibility via 3‐(4, 5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide (MTT) assay and considerable attachment on the scaffold using SEM studies. Real‐time polymerase chain reaction (PCR) and immunostaining studies confirmed the upregulation of cardiac specific genes including GATA‐4, T‐box 18 (TBX 18), cardiac troponin T (cTnT), and alpha‐myosin heavy chain (α‐MHC) in the PU/rGO‐Ag scaffolds in comparison with neat PU ones. Therefore, these nanofibrous rGO‐Ag–reinforced PU scaffolds can be considered as suitable candidates in cardiac tissue engineering.

Adenosine 5'-triphosphate (ATP) which is released from neuronal and non-neuronal tissues interacts with cell surface receptors to produce a broad range of physiological responses. The present study addressed the issue of whether the cells of the superior cervical ganglia (SCG) respond to ATP. To this end, the dynamics of the intracellular calcium ion concentration ([Ca2+]i) of neurons and satellite cells in intact SCG was analyzed by laser scanning confocal microscopy. ATP produced an increase of [Ca2+]i in both neurons and satellite cells; initially, ATP elicited [Ca2+]i increase in satellite cells and, subsequently, a [Ca2+]i change in neurons was observed. P1 purinoceptor agonists had no effect on this process, but P2 purinoceptor agonists induced [Ca2+]i increase and suramin totally inhibited ATP-induced [Ca2+]i dynamics in both neurons and satellite cells. In satellite cells, Ca2+ channel blockers and the removal of extracellular Ca2+, but not thapsigargin pretreatment, abolished ATP-induced [Ca2+]i dynamics. In contrast, thapsigargin pretreatment abolished ATP-induced [Ca2+]i dynamics in neurons. Reactive blue-2 inhibited the ATP-induced reaction on neurons alone. Uridine 5'-triphosphate caused a [Ca2+]i increase in neurons and alpha,beta-methylene ATP caused a [Ca2+]i increase in satellite cells. We concluded that neurons respond to extracellular ATP mainly via P2Y purinoceptors and that satellite cells respond via P2X purinoceptors.

We studied the mechanical properties of polyurethane (PU) nanofiber filaments prepared by electrospinning and dry twisting, and compared them with those of the corresponding nonwoven PU nanofiber webs. The morphologies and mechanical properties of the nonwoven PU nanofiber webs and the corresponding PU nanofiber filaments were investigated by scanning electron microscopy (SEM) and through the use of a universal testing machine (UTM), respectively. The tensile strength and the Young’s modulus of the nanofiber filaments improved dramatically as the number of twists or the width of the nanofiber webs was increased compared with the nonwoven PU nanofibers. Moreover, it was found that applying pre-tension was an effective method of increasing the mechanical properties of PU nanofiber filaments.

During skeletal muscle regeneration caused by injury, muscle satellite cells proliferate and migrate toward the site of muscle injury. This migration is mainly induced by hepatocyte growth factor (HGF) secreted by intact myofibers and also released from injured muscle. However, the intracellular machinery for the satellite cell migration has not been elucidated. To examine the mechanisms of satellite cell migration, we utilized satellite cell-derived mouse C2C12 skeletal muscle cells. HGF induced reorganization of actin cytoskeleton to form lamellipodia in C2C12 myoblasts. HGF treatment facilitated both nondirectional migration of the myoblasts in phagokinetic track assay and directional chemotactic migration toward HGF in a three-dimensional migration chamber assay. Endogenous N-WASP and WAVE2 were concentrated in the lamellipodia at the leading edge of the migrating cells. Moreover, exogenous expression of wild-type N-WASP or WAVE2 promoted lamellipodial formation and migration. By contrast, expression of the dominant-negative mutant of N-WASP or WAVE2 and knockdown of N-WASP or WAVE2 expression by the RNA interference prevented the HGF-induced lamellipodial formation and migration. When the cells were treated with LY294002, an inhibitor of phosphatidylinositol 3-kinase, the HGF-induced lamellipodial formation and migration were abrogated. These results imply that both N-WASP and WAVE2, which are activated downstream of phosphati-dylinositol 3-kinase, are required for the migration through the lamellipodial formation of C2C12 cells induced by HGF.

Skeletal muscle satellite cells (SC) are important for maintenance and repair of myofibers. SC function and responsiveness is regulated, at least in part, through interactions with niche components in which they reside. Recent evidence suggests that structural changes occur in the SC niche as a function of aging. In the present study, we aimed to investigate whether age has an impact on the properties of the SC niche. Muscle biopsies were obtained from the vastus lateralis of previously untrained but otherwise healthy young men (YM: 21±0.4 yrs; n=10) and old men (OM: 66±0.9 yrs; n=11) at rest. In addition, OM (n =10) performed a single bout of resistance exercise and additional muscle biopsies were taken 24 h and 48 h post‐exercise. Skeletal muscle SC niche measurements were assessed using high resolution immunofluorescent and confocal microscopy. We report changes in the muscle SC niche with aging and that many of these differences are primarily associated with type II fibres. Type II SC niche laminin thickness was significantly greater in OM (1.86 ± 0.06 μm) as compared to YM (1.55 ± 0.09 μm, p&lt;0.05). Mean intensity of collagen IV was significantly lower in OM (92 ± 10 a.u) as compared to YM (512 ± 17 a.u, p&lt;0.05). In OM, the fibrotic index as measured by Masson's trichrome staining was greater (3.6 ± 0.5 %) at baseline, as compared to YM (2.0 ± 0.5 %; p&lt;0.05). We also observed the presence of SC fully incarcerated by the basal lamina. At baseline, 100% of OM possessed SC that were incarcerated, as compared to ~40% of YM. The percentage of SC in mixed muscle that were incarcerated by laminin was greater in OM (19.0 ± 1.6%) as compared to YM (7.2 ± 2.1; p&lt;0.05). Although this phenomenon was present in both Type I and Type II fibres, it appeared exaggerated in Type I fibres (OM: 28.4 ± 8.2%, YM: 3.6 ± 2.1%; p&lt;0.05). In all cases the greatest proportion of active SC were found in the non‐incarcerated population, whereas incarcerated cells were less likely to activate (24 h post exercise – Non‐incarcerated:32 ± 6% active, Incarcerated: 8 ± 2% active, p&lt;0.05). Consistent with this, at 24 h, there was a significantly lower number of quiescent SC in the non‐incarcerated condition (62 ± 7 %) as compared to the incarcerated condition (92 ± 4%, p&lt;0.05). Additional evidence of age‐related dysfunction includes larger myonuclear CSA, increased expression of COL4α1 mRNA and Laminin subunits α2 and β1 was significantly greater in OM as compared to YM. Together this data suggests there are age‐related changes in the niche that are potentially detrimental to SC function and skeletal muscle health.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The effects of zerumbone on the proliferation and apoptosis of esophagus cancer cells and on the P53 and Bcl-2 expression levels were studied. The esophagus cancer EC-109 cells were cultured and inoculated. The effect of zerumbone on proliferation of EC-109 cells was detected via the Cell Counting Kit-8 (CCK-8) method. Cell apoptosis was detected via TdT-mediated dUTP nick end-labeling (TUNEL) staining. Moreover, the mRNA expression levels of P53 and Bcl-2 were detected via reverse transcription-polymerase chain reaction (RT-PCR), and the protein expression levels of P53 and Bcl-2 were evaluated via western blotting. CCK-8 detection results showed that compared with control group, zerumbone in different concentrations could inhibit the activity of EC-109, and the proliferation inhibition rate was significantly increased in a concentration-dependent manner with the increase of concentration. TUNEL staining showed that cell apoptosis gradually occurred in administration group, and the number of apoptotic cells was increased in a concentration-dependent manner with the increase of concentration. RT-PCR detection results showed that the mRNA expression level of P53 in administration group was significantly increased compared with that in control group, but that of Bcl-2 was significantly decreased. Western blotting showed that the protein expression level of Bcl-2 in administration group in different concentrations was significantly increased with the increase of zerumbone concentration, but that of Bcl-2 was significantly decreased in a concentration-dependent manner. Zerumbone can inhibit the proliferation and induce apoptosis of esophageal cancer EC-109 cells, and its induction of apoptosis may be realized through upregulating the mRNA expression of P53 and downregulating the mRNA expression of Bcl-2, and upregulating the protein expression of P53 and downregulating the protein expression of Bcl-2.