SCSMRD: A database for single-cell skeletal muscle regeneration

Skeletal muscle regeneration is a complex process where various cell types and cytokines are involved.Single-cell RNA-sequencing (scRNA-seq) provides the opportunity to deconvolute heterogeneous tissue into individual cells based on their transcriptomic profiles.Recent scRNA-seq studies on mouse muscle regeneration have provided insights to understand the transcriptional dynamics that underpin muscle regeneration.However,a database to investigate gene expression profiling during skeletal muscle regeneration at the single-cell level is lacking.Here,we collected over 105 000 cells at 7 key regenerative time-points and non-injured muscles and developed a database,the Singlecell Skeletal Muscle Regeneration Database (SCSMRD).SCSMRD allows users to search the dynamic expression profiles of genes of interest across different cell types during the skeletal muscle regeneration process.It also provides a network to show the activity of regulons in different cell types at different time points.Pesudotime analysis showed the state changes trajectory of muscle stem cells (MuSCs) during skeletal muscle regeneration.This database is freely available at https://scsmrd.fengs-lab.com.

Factors influencing myogenic differentiation of adipose-derived stem cells and their application in muscle regeneration

Skeletal muscle regeneration mainly depends on muscle satellite cells;however,these cells are not sufficient for supporting repair and regeneration in volumetric muscle loss(VML),Duchenne muscular dystrophy,and other muscle injuries or muscle diseases.As such,much work has been conducted in recent years to search for myogenic stem cells.Adipose-derived stem cells(ADSCs)have a wide range of sources,rapid growth,and multi-directional differentiation potential,and have become vital candidates for muscle regeneration.Multiple factors influence the myogenic differentiation capacity of ADSCs.This paper reviews the regulatory aspects and possible factors that have been identified in recent years to affect myogenic differentiation of ADSCs.Based on these factors,gene editing,and perfusion concepts,a method was proposed to achieve maximal differentiation efficiency of ADSCs.This study focused on the application of ADSCs in muscle regeneration and disease.Based on the importance of myogenic differentiation of ADSCs for the repair and regeneration of muscle damage,this study provides a basis for future research surrounding the efficient induction of myogenic differentiation of ADSCs in vitro.

3D-printed tissue repair patch combining mechanical support and magnetism for controlled skeletal muscle regeneration

Physical forces,such as magnetic and mechanical stimulation,are known to play a significant role in the regulation of cell response.In the present study,a biomimetic regeneration patch was fabricated using E-jet 3 D printing,which integrates mechanical and magnetic stimulation in a biocompatible"one-pot reaction"strategy when combined with a static magnetic field(SMF).The magneto-based therapeutic regeneration patch induced myoblasts to form aligned and multinucleated myotubes,regulated the expression of myogenic-related genes,and activated the p38αmitogen-activated protein kinase pathway via the initiation of myogenic differentiation.To validate the efficiency of the proposed strategy,the regeneration patch was implanted into mice and exposed to a suitable SMF,which resulted in significantly enhanced in vivo skeletal muscle regeneration.The findings demonstrated that appropriate external physical stimulation provides a suitable biophysical microenvironment that is conducive to tissue regeneration.The method used in the present study represents a promising technique to induce the regeneration of damaged skeletal muscle tis sue.

Research Progress of Vitamins on Muscle Regeneration

Muscle satellite cells,as muscle stem cells,play a critical role in the process of muscle regeneration,and effective muscle regeneration helps to restore muscle function and maintain the homeostasis of muscle tissues.In damaged muscle,muscle satellite cells are activated to form new myofibers through the process of cell proliferation,migration,differentiation and fusion to complete muscle tissue regeneration.Meanwhile,this process is mainly affected by endogenous gene expression and many exogenous factors.Researches in recent years have shown that vitamins,as important nutrients,play an extremely important role in the process of muscle regeneration.Therefore,this article reviewed the roles of vitamins in the regeneration of muscle satellite cells,according to the latest research progress.It would provide more theoretical and data support for the regeneration and repair of muscle damage,muscle atrophy and other muscle diseases,so that it could be better applied in the field of muscle regeneration researches and serve human health.

Highly Aligned Ternary Nanofiber Matrices Loaded with MXene Expedite Regeneration of Volumetric Muscle Loss

Current therapeutic approaches for volumetric muscle loss(VML)face challenges due to limited graft availability and insufficient bioactivities.To overcome these limitations,tissue-engineered scaffolds have emerged as a promising alternative.In this study,we developed aligned ternary nanofibrous matrices comprised of poly(lactide-co-ε-caprolactone)integrated with collagen and Ti_(3)C_(2)T_(x)MXene nanoparticles(NPs)(PCM matrices),and explored their myogenic potential for skeletal muscle tissue regeneration.The PCM matrices demonstrated favorable physicochemical properties,including structural uniformity,alignment,microporosity,and hydrophilicity.In vitro assays revealed that the PCM matrices promoted cellular behaviors and myogenic differentiation of C2C12 myoblasts.Moreover,in vivo experiments demonstrated enhanced muscle remodeling and recovery in mice treated with PCM matrices following VML injury.Mechanistic insights from next-generation sequencing revealed that MXene NPs facilitated protein and ion availability within PCM matrices,leading to elevated intracellular Ca^(2+)levels in myoblasts through the activation of inducible nitric oxide synthase(i NOS)and serum/glucocorticoid regulated kinase 1(SGK1),ultimately promoting myogenic differentiation via the m TOR-AKT pathway.Additionally,upregulated i NOS and increased NO–contributed to myoblast proliferation and fiber fusion,thereby facilitating overall myoblast maturation.These findings underscore the potential of MXene NPs loaded within highly aligned matrices as therapeutic agents to promote skeletal muscle tissue recovery.

Bioactive biodegradable polycitrate nanoclusters enhances the myoblast differentiation and in vivo skeletal muscle regeneration via p38 MAPK signaling pathway

Complete skeletal muscle repair and regeneration due to severe large injury or disease is still a challenge.Biochemical cues are critical to control myoblast cell function and can be utilized to develop smart biomaterials for skeletal muscle engineering.Citric acid-based biodegradable polymers have received much attention on tissue engineering,however,their regulation on myoblast cell differentiation and mechanism was few investigated.Here,we find that citrate-based polycitrate-polyethylene glycol-polyethylenimine(POCG-PEI600)nanoclusters can significantly enhance the in vitro myoblast proliferation by probably reinforcing the mitochondrial number,promote the myotube formation and full-thickness skeletal muscle regeneration in vivo by activating the myogenic biomarker genes expression of Myod and Mhc.POCG-PEI600 nanoclusters could also promote the phosphorylation of p38 in MAP kinases(MAPK)signaling pathway,which led to the promotion of the myoblast differentiation.The in vivo skeletal muscle loss rat model also confirmed that POCG-PEI600 nanoclusters could significantly improve the angiogenesis,myofibers formation and complete skeletal muscle regeneration.POCG-PEI600 nanocluster could be also biodegraded into small molecules and eliminated in vivo,suggesting their high biocompatibility and biosafety.This study could provide a bioactive biomaterial-based strategy to repair and regenerate skeletal muscle tissue.

Procyanidins-crosslinked small intestine submucosa: A bladder patch promotes smooth muscle regeneration and bladder function restoration in a rabbit model

Currently the standard surgical treatment for bladder defects is augmentation cystoplasty with autologous tissues,which has many side effects.Biomaterials such as small intestine submucosa(SIS)can provide an alternative scaffold for the repair as bladder patches.Previous studies have shown that SIS could enhance the capacity and compliance of the bladder,but its application is hindered by issues like limited smooth muscle regeneration and stone formation since the fast degradation and poor mechanical properties of the SIS.Procyanidins(PC),a natural bio-crosslinking agent,has shown anti-calcification,anti-inflammatory and anti-oxidation properties.More importantly,PC and SIS can crosslink through hydrogen bonds,which may endow the material with enhanced mechanical property and stabilized functionalities.In this study,various concentrations of PC-crosslinked SIS(PC-SIS)were prepared to repair the full-thickness bladder defects,with an aim to reduce complications and enhance bladder functions.In vitro assays showed that the crosslinking has conferred the biomaterial with superior mechanical property and anti-calcification property,ability to promote smooth muscle cell adhesion and upregulate functional genes expression.Using a rabbit model with bladder defects,we demonstrated that the PC-SIS scaffold can rapidly promote in situ tissue regrowth and regeneration,in particular smooth muscle remodeling and improvement of urinary functions.The PC-SIS scaffold has therefore provided a promising material for the reconstruction of a functional bladder.

Signaling pathways of adipose stem cell-derived exosomes promoting muscle regeneration

Severe muscle injury is still a challenging clinical problem. Exosomes derived from adipose stem cells (ASC-exos) may be a potential therapeutic tool, but their mechanism is not completely clear. This review aims to elaborate the possible mechanism of ASC-exos in muscle regeneration from the perspective of signal pathways and provide guidance for further study. Literature cited in this review was acquired through PubMed using keywords or medical subject headings, including adipose stem cells, exosomes, muscle regeneration, myogenic differentiation, myogenesis, wingless/integrated (Wnt), mitogen-activated protein kinases, phosphatidylinositol-4,5-bisphosphate 3-kinase/protein kinase B (PI3K/Akt), Janus kinase/signal transducers and activators of transcription, and their combinations. We obtained the related signal pathways from proteomics analysis of ASC-exos in the literature, and identified that ASC-exos make different contributions to multiple stages of skeletal muscle regeneration by those signal pathways.

Injectable muscle-adhesive antioxidant conductive photothermal bioactive nanomatrix for efficiently promoting full-thickness skeletal muscle regeneration

The completed skeletal muscle regeneration resulted from severe injury and muscle-related disease is still a challenge.Here,we developed an injectable muscle-adhesive antioxidant conductive bioactive photothermo-responsive nanomatrix for regulating the myogenic differentiation and promoting the skeletal muscle regeneration in vivo.The multifunctional nanomatrix was composed of polypyrrole@polydopamine(PPy@PDA,342±5.6 nm)nanoparticles-crosslinked Pluronic F-127(F127)-polycitrate matrix(FPCP).The FPCP nanomatrix demonstrated inherent multifunctional properties including excellent photothermo-responsive and shear-thinning behavior,muscle-adhesive feature,injectable ability,electronic conductivity(0.48±0.03 S/m)and antioxidant activity and photothermal function.The FPCP nanomatrix displayed better photothermal performance with near-infrared irradiation,which could provide the photo-controlled release of protein(91%±2.6%of BSA was released after irradiated 3 times).Additionally,FPCP nanomatrix could significantly enhance the cell proliferation and myogenic differentiation of mouse myoblast cells(C2C12)by promoting the expressions of myogenic genes(MyoD and MyoG)and myosin heavy chain(MHC)protein with negligible cytotoxicity.Based on the multifunctional properties,FPCP nanomatrix efficiently promoted the full-thickness skeletal muscle repair and regeneration in vivo,through stimulating the angiogenesis and myotube formation.This study firstly indicated the vital role of multifunctional PPy@PDA nanoparticles in regulating myogenic differentiation and skeletal muscle regeneration.This work also suggests that rational design of bioactive matrix with multifunctional feature would greatly enhance the development of regenerative medicine.

Effects of PGC-1αoverexpression on the myogenic response during skeletal muscle regeneration

The ability of skeletal muscle to regenerate from injury is crucial for locomotion,metabolic health,and quality of life.Peroxisome proliferator-activated receptor-γcoactivator-1α(PGC1A)is a transcriptional coactivator required for mitochondrial biogenesis.Increased mitochondrial biogenesis is associated with improved muscle cell differentiation,however PGC1A's role in skeletal muscle regeneration following damage requires further investigation.The purpose of this study was to investigate the role of skeletal muscle-specific PGC1A overexpression during regeneration following damage.22 C57BL/6J(WT)and 26 PGC1A muscle transgenic(A1)mice were injected with either phosphate-buffered saline(PBS,uninjured control)or Bupivacaine(MAR,injured)into their tibialis anterior(TA)muscle to induce skeletal muscle damage.TA muscles were extracted 3-or 28-days postinjury and analyzed for markers of regenerative myogenesis and protein turnover.Pgc1a mRNA was~10–20 fold greater in A1 mice.Markers of protein synthesis,AKT and 4EBP1,displayed decreases in A1 mice compared to WT at both timepoints indicating a decreased protein synthetic response.Myod mRNA was~75%lower compared to WT 3 days post-injection.WT mice exhibited decreased cross-sectional area of the TA muscle at 28 days post-injection with bupivacaine compared to all other groups.PGC1A overexpression modifies the myogenic response during regeneration.

The Effect of Increasing mTOR Signaling on the Speed of Regeneration in D. dorotocephala planaria

Muscle strains are a common injury that can occur during physical activity or exercise, and they can range from mild to severe depending on the extent of the damage. The mTOR pathway is a highly conserved signaling pathway that has been implicated in various cellular processes, including tissue regeneration. Previous studies that have investigated protein synthesis in mice have concluded that the mTOR pathway can improve muscle regeneration. However, the specific effects of mTOR pathway activation on muscle regeneration in planarians have yet to be fully explored. Therefore, this study aimed to investigate the use of Arginine and Leucine to stimulate the mTOR pathway for muscle strains in planarians. During the experiment, planarians were amputated, and different dosages of Arginine and Leucine were used to stimulate the mTOR pathway in Dugesia dorotocephala. The speed of muscle regeneration was measured over 14 days in five groups, with thirty planarians in each group. The results showed that the rate of regeneration from 0.134 mM Leucine solution showed a significant increase compared to the control group, while the groups exposed to 0.1, 0.3 mM Arginine, and 0.0134 Leucine did not show significant changes in muscle regeneration. These findings suggest that mTOR pathway activation may enhance muscle regeneration in planarians and that the effects may be dose-dependent. These findings have important implications for developing new treatments for tissue damage. Further studies are needed to fully understand the mechanisms of mTOR pathway activation on muscle regeneration in planarians and its potential use in tissue engineering and regenerative medicine.

IGF-1, bFGF EXPRESSION AND VASCULAR REGENERATION IN ACUTE INFARCTED CANINE MYOCARDIUM AFTER AUTOLOGUS SKELETAL MUSCLE SATELLITE CELL IMPLANTATION

Objective To study the cell growth factor secretion and vascular regeneration in acute in-farcted myocardium after autologous skeletal muscle satellite cell implantation. Methods Autologous skeletal muscle satellite cells from adult mongrel canine were implanted into the acute myocardial infarct site via the ligated left anterior descending (LAD) artery. Specimens were harvested at 2, 4 , 8 weeks after implantation for the expression of insulin-like growth factor-1 (IGF-1), basic fibroblast growth factor ( bFGF) and the vascular density. Results The expression of IGF-1, bFGF and the vascular density in skeletal muscle satellite cell implant group were higher than that in the control group. Conclusion The skeletal muscle satellite cells, after being implanted into the acute myocardial infarction, not only showed myocardial regeneration, but also showed the ability to secrete the cell factors, hence representing a positive effect on the regeneration of the infarcted myocardium.

Membrane and Subcellular Muscle Injury Are Induced by Single or Multiple Exposure to Cigarette Smoke

Cigarette smoking is the main cause of chronic obstructive pulmonary disease (COPD). Diaphragm injury is observed in patients with COPD. However, the potential role of smoking in triggering or perpetuating muscle injury is unknown. The present study was aimed at evaluating the potential role of commercial tobacco smoke as a direct cause of skeletal muscle injury in experimental conditions. Seventy Wistar rats (170 - 250 g) were assigned to smoking (n = 49) or non-smoking (n = 21) groups. The smoking groups were submitted to a single or multiple (i.e., five or thirty) daily sessions of cigarette smoking in an inhalatory chamber (time length: 2 h each session). The level of exposure was constant and assessed by CO concentrations (50 ppm) and serum cotinine analysis. Animals submitted to a single smoke exposure and the corresponding controls were euthanized in groups at 0 h, 2 h, 4 h, 24 h or 48 h after completing the exposure. Animals submitted to multiple exposures were euthanized at 0 h after smoking. Samples from vastus lateralis muscle were obtained and processed for assessing cell injury and selected protein expression. Monoclonal anti-albumin antibodies were used to identify muscle fibers with sarcolemmal (membrane) injury. Subcellular muscle injury was assessed using transmission electron microscopy (EM). MyoD, myogenin and α-tubulin were immunodetected using western blot techniques. Exposure to cigarette smoke associated with significant membrane damage (mean relative difference (MRD) with controls: +181%, p = 0.004) and sarcomere disruptions (MRD: +226%, p = 0.001). Expression of MyoD and myogenin (normalized to α-tubulin) were significantly increased at 4 h and remained increased at 48 h post-exposure. We conclude that not only a single but also consecutive exposure to tobacco smoke have acute deleterious effects on peripheral muscle structure. A rapid induction of subrogate markers of skeletal muscle stress and repair processes associates to sarcolemmal and sarcomere damage.

THE IMPROVEMENT OF INFARCTED MYOCARDIAL CONTRACTILE FORCE AFTER AUTOLOGOUS SKELETAL MUSCLE SATELLITE CELL IMPLANTATION

Objective To study the improvement of infarcted myocardial contractile force after autologous skeletal muscle satellite cell implantation via intracoronary arterial perfusion. Methods Skeletal muscle cells were harvested from gluteus max of adult mongrel dogs and the cells were cultured and expanded before being labeled with DAPI (4’, 6-diamidino-2-phenylindone). The labeled cells were then implanted into the acute myocardial infarct site via the ligated left anterior descending (LAD) coronary artery. Specimens were taken at 2nd, 4th, 8th week after myoblast implantation for histologic and contractile force evaluation, respectively. Results The satellite cells with fluorescence had been observed in the infarct site and also in papi- llary muscle with consistent oriented direction of host myocardium. A portion of the implanted cells had differen- tiated into muscle fibers. Two weeks after implantation, the myocardial contractile force showed no significant difference between the cell implant group and control group. At 4 and 8 week, the contractile force in the cell implant group was better than that in control group. Conclusion The skeletal muscle satellite cells, implanted into infarct myocardium by intracoronary arterial perfusion, could disseminate through the entire infarcted zone with myocardial regeneration and improve the contractile function of the infarcted myocardium.

Immunomodulatory extracellular matrix hydrogel induces tissue regeneration in a model of partial glossectomy

While oropharyngeal cancer treatment regimens,including surgical resection,irradiation,and chemotherapy,are effective at removing tumors,they lead to muscle atrophy,denervation,and fibrosis,contributing to the pathogenesis of oropharyngeal dysphagia-difficulty swallowing.Current standard of care of rehabilitative tongue strengthening and swallowing exercises is ineffective.Here,we evaluate an alternative approach utilizing an acellular and injectable biomaterial to preserve muscle content and reduce fibrosis of the tongue after injury.Skeletal muscle extracellular matrix(SKM)hydrogel is fabricated from decellularized porcine skeletal muscle tissue.A partial glossectomy injury in the rat is used to induce tongue fibrosis,and SKM hydrogels along with saline controls are injected into the site of scarring two weeks after injury.Tissues are harvested at 3 and 7 days post-injection for gene expression and immunohistochemical analyses,and at 4 weeks post-injection to evaluate histomorphological properties.SKM hydrogel reduces scar formation and improves muscle regeneration at the site of injury compared to saline.SKM additionally modulates the immune response towards an anti-inflammatory phenotype.This study demonstrates the immunomodulatory and tissue-regenerative capacity of an acellular and minimally invasive ECM hydrogel in a rodent model of tongue injury.

Accelerated regeneration of the skeletal muscle in RNF13-knockout mice is mediated by macrophage-secreted IL-4/IL-6

RING finger protein 13 （RNF13） is a newly identified E3 ligase reported to be functionally significant in the reg- ulation of cancer development, muscle cell growth, and neuronal development. In this study, the function of RNF13 in cardiotoxin-induced skeletal muscle regener- ation was investigated using RNF13-knockout mice. RNF13-/- mice exhibited enhanced muscle regeneration --characterized by accelerated satellite cell proliferation --compared with wild-type mice. The expression of RNF13 was remarkably induced in macrophages rather than in the satellite cells of wild-type mice at the very early stage of muscle damage. This result indicated that inflammatory cells are important in RNF13-mediated satellite cell functions. The cytokine levels in skeletal muscles were further analyzed and showed that RNF13＂i＂ mice produced greater amounts of various cytokines than wild-type mice. Among these, IL-4 and IL-6 levels significantly increased in RNF13-/＂ mice. The accelerated muscle regeneration phenotype was abrogated by inhibiting IL-4/IL-6 action in RNF13＂/- mice with blocking antibodies. These results indicate that RNF13 deficiency promotes skeletal muscle regeneration via the effects on satellite cell niche mediated by IL-4 and IL-6.

Disease Prevention and Alleviation by Human Myoblast Transplantation

Myoblast implantation is a unique, patented technology of muscle regeneration being tested in Phase III clinical trials of muscular dystrophy, ischemic cardiomyopathy, Phase II trial of cancer, and Phase I trial of Type II diabetes. Differentiated and committed, myoblasts are not stem cells. Implanted myoblasts fuse spontaneously among themselves, replenishing genetically normal myofibers. They also fuse with genetically abnormal myofibers of muscular dystrophy, cardiomyopathy, or Type II diabetes, transferring their nuclei containing the normal human genome to provide stable, long-term expression of the missing gene products. They develop to become cardiomyocytes in the infracted myocardium. Myoblasts transduced with VEGF165 allow concomitant regeneration of blood capillaries and myofibers. They are potent biologics for treating heart failure, ischemic cardiomyopathy, diabetic ischemia, erectile dysfunction, and baldness. Myoblasts, because of their small size, spindle shape, and resilience, can grow within wrinkles and on skin surfaces, thus enhancing the color, luster and texture of the skin “plated” with them. They can be injected subcutaneously as a cellular filler to reduce wrinkles. Intramuscular injection of myoblasts can augment the size, shape, consistency, tone and strength of muscle groups, improving the lines, contours and vitality to sculpt a youthful appearance. This highly promising technology has great social economic values in treating hereditary, fatal and debilitating disease conditions.

Function and regulation of muscle stem cells in skeletal muscle development and regeneration:a narrative review

Skeletal muscle plays an essential role in generating the mechanical force necessary to support the movement of our body and daily exercise. Compared with cardiac and smooth muscle, in mammals, skeletal muscle exhibits remarkable regenerative capacity in response to damage. Muscle stem cells, also known as satellite cells, directly contribute to regeneration. Here, we review primary and secondary myogenesis processes with a focus on muscle stem cells, as well as the function and regulation of muscle stem cells in adult muscle regeneration in mammals.

Scaffold tissue engineering strategies for volumetric muscle loss

Volumetric muscle loss(VML)refers to a composite,en bloc loss of skeletal muscle mass resulting in functional impairment.These injuries normally heal with excessive fibrosis,minimal skeletal muscle regeneration,and poor functional recovery.Functional muscle transfer is a treatment option for some patients but is limited both by the degree of functional restoration as well as donor site morbidity.As such,new therapeutic options are necessary.De novo regeneration of skeletal muscle,by way of tissue engineering,is an emerging strategy to treat VML.This review evaluates available scaffolds for promoting skeletal muscle regeneration and functional recovery following VML.The use of growth factors and stem cell therapies,which may augment scaffold integration and skeletal muscle reconstitution,are also discussed.Regenerative medicine with the use of scaffolds is a promising area in skeletal muscle reconstruction and VML treatment.

Regulation of muscle stem cell fate

Skeletal muscle plays a critical role in human health.Muscle stem cells(MuSCs)serve as the major cell type contribut-ing to muscle regeneration by directly differentiating to mature muscle cells.MuSCs usually remain quiescent with occasionally self-renewal and are activated to enter cell cycle for proliferation followed by differentiation upon muscle injury or under pathological conditions.The quiescence maintenance,activation,proliferation,and differentiation of MuSCs are tightly regulated.The MuSC cell-intrinsic regulatory network and the microenvironments work coordi-nately to orchestrate the fate transition of MuSCs.The heterogeneity of MuSCs further complicates the regulation of MuSCs.This review briefly summarizes the current progress on the heterogeneity of MuSCs and the microenviron-ments,epigenetic,and transcription regulations of MuSCs.