Myoblast Proliferation Research Articles

The Spatial-Temporal Alternative Splicing Profile Reveals the Functional Diversity of FXR1 Isoforms in Myogenesis.

Alternative splicing (AS) is a fundamental mechanism contributing to proteome diversity, yet its comprehensive landscape and regulatory dynamics during skeletal muscle development remain largely unexplored. Here, the temporal AS profiles are investigated during myogenesis in five vertebrates, conducting comprehensive profiling across 27 developmental stages in skeletal muscle and encompassing ten tissues in adult pigs. The analysis reveals a pervasive and evolutionarily conserved pattern of alternative exon usage throughout myogenic differentiation, with hundreds of skipped exons (SEs) showing developmental regulation, particularly within skeletal muscle. Notably, this study identifies a muscle-specific SE (exon 15) within the Fxr1 gene, whose AS generates two dynamically expressed isoforms with distinct functions: the isoform without exon 15 (Fxr1E15 -) regulates myoblasts proliferation, while the isoform incorporating exon 15 (Fxr1E15+) promotes myogenic differentiation and fusion. Transcriptome analysis suggests that specifically knocking-down Fxr1E15+ isoform in myoblasts modulates differentiation by influencing gene expression and splicing of specific targets. The increased inclusion of exon 15 during differentiation is mediated by the binding of Rbm24 to the intron. Furthermore, in vivo experiments indicate that the Fxr1E15+ isoform facilitates muscle regeneration. Collectively, these findings provide a comprehensive resource for AS studies in skeletal muscle development, underscoring the diverse functions and regulatory mechanisms governing distinct Fxr1 isoforms in myogenesis.

Novel sol-gel derived layered double oxides and layered double oxide/γ-Al2O3 with tunable selectivity as promising adsorbents for L-lactate uptake from the myoblast culture medium

Cultured meat production, a biotechnological solution for global protein demand and sustainability, faces economic challenges due to the high demand for cell culture medium. Contamination with cell metabolites, particularly L-lactate, impedes myoblast proliferation in high-density suspension culture. Eliminating L-lactate is crucial for reusing the medium and overcoming economic barriers in mass myoblast culture. Retaining essential nutrients like D-glucose, amino acids, and vitamins after L-lactate removal is imperative for sustained myoblast growth. This study proposes a sustainable and economically feasible adsorption technique utilizing novel sol-gel derived Mg-Al layered double oxides and Mg-Al layered double oxide/γ-Al2O3 with tunable selectivity as a promising tool for L-lactate uptake from the myoblast medium. In an aqueous solution, L-lactate uptake and selectivity over D-glucose were enhanced with the decrease in total Mg/Al ratio and crystallinity, increase in Al2O3 dispersion in adsorbent, specific surface area, and porosity. The latter was achieved successfully by metal alkoxide sol-gel method employing an organic structure-directing agent, where LDO/amorphous alumina nanohybrid with Mg/Al = 1.23 was successfully prepared in mixed methanol-ethanol solution in a relatively short time. A simple mixture of Mg-Al LDO with Mg/Al = 2 and nano-γ-Al2O3 prepared by a sustainable and economically feasible co-precipitation method was found to have comparable efficiency and selectivity to a costly sol-gel derived material. In myoblast medium, lower total Mg/Al was associated with higher cytocompatibility, pH-stability, and selectivity, while surface properties of materials did not play a significant role. Nano-γ-Al2O3 was identified as the most promising material for selective L-lactate uptake from the myoblast culture medium. Thus, further optimization of the medium treatment process with nano-γ-Al2O3 can realize cost-effective high-density myoblast culture with circular use of the medium – a novel sustainable process in the biotechnology field.

Interferon-β-Induced Injury During Pediatric Muscle Differentiation: Insight Into Juvenile Dermatomyositis Pathogenesis.

Juvenile dermatomyositis (JDM) involves up-regulated type I interferons (IFNs), including IFNβ, yet pathologic mechanisms remain poorly understood. We aimed to characterize the functional and structural effects of IFNβ on in vitro human pediatric myoblast growth and differentiation in a three-dimensional skeletal muscle model (myobundles). Myobundles fabricated from myoblasts of a healthy pediatric donor were exposed to IFNβ at 0 to 5,600 IU/mL during growth (days 1-4), differentiation (days 4-11), and/or mature (days 11-18) periods. To assess myobundle structure and function, contractile force, kinetics, and fatigue were measured at day 18 with subsequent immunohistochemistry. Myobundles were not functionally affected by IFNβ exposure during growth period alone. However, when IFNβ exposure continued through differentiation, myobundles became dysfunctional (P < 0.0001). IFNβ during differentiation or mature periods alone resulted in dose-dependent decreases in contractility, with greater decrease in the differentiation alone group (P < 0.0001). Twitch kinetics and fatigue remained largely unchanged when myobundles were exposed to IFNβ only during growth, yet twitch time slowed (P < 0.005) and fatigue decreased (P < 0.002) when myobundles were exposed during differentiation or mature stages alone. Nuclei density and myofiber size and organization also decreased when IFNβ was added during differentiation period alone. IFNβ decreases pediatric myobundle contractile function most significantly during differentiation of myoblasts to myotubes. Function is not affected when IFNβ exposure is limited to myoblast proliferation alone. These findings implicate a pathologic role for IFNβ in JDM by impairing myoblast differentiation, leading to subsequent loss of function and ongoing need for muscle regeneration and repair.

CLIC5 promotes myoblast differentiation and skeletal muscle regeneration via the BGN-mediated canonical Wnt/β-catenin signaling pathway

Myoblast differentiation plays a vital role in skeletal muscle regeneration. However, the protein-coding genes controlling this process remain incompletely understood. Here, we showed that chloride intracellular channel 5 (CLIC5) exerts a critical role in mediating myogenesis and skeletal muscle regeneration. Deletion of CLIC5 in skeletal muscle leads to reduced muscle weight and decreases the number and differentiation potential of satellite cells. In vitro, CLIC5 consistently inhibits myoblast proliferation while promoting myotube formation. CLIC5 promotes myogenic differentiation by activating the canonical Wnt/β-catenin signaling pathway in a biglycan (BGN)–dependent manner. CLIC5 deletion impairs muscle regeneration. Paired box gene 7 (Pax7) expression and the activity of BGN-mediated canonical Wnt/β-catenin signaling are reduced in CLIC5-deficient mice. Conversely, increasing CLIC5 levels in skeletal muscles enhances muscle regeneration capacity. In conclusion, our findings underscore CLIC5 as a pivotal regulator of myogenesis and skeletal muscle regeneration, functioning through interaction with BGN to activate the canonical Wnt/β-catenin signaling pathway.

METTL16 inhibits differentiation and promotes proliferation and slow myofibers formation in chicken myoblasts

N6-methyladenosine (m6A) plays a crucial regulatory role in muscle growth and development. In our previous studies, we identified a m6A methyltransferase, Methyltransferase like 16 (METTL16), which is associated with chicken muscle development and muscle fiber type conversion. To further understand the regulatory role of METTL16 in chicken muscle function, we analyzed its expression in muscle tissues with different myofiber type compositions and in chicken primary myoblasts (CPMs) at various stages. We also manipulated METTL16 expression in CPMs to examine its effects on cell proliferation, differentiation, muscle fiber type formation, and global m6A RNA methylation status. Our results showed that METTL16 expression increased during myoblast proliferation and gradually decreased in the late differentiation stage. Furthermore, METTL16 exhibited specific expression in slow-twitch muscles. Cell Counting Kit-8 assays, 5-Ethynyl-2'-deoxyuridine staining, RT-qPCR, Western blot, and immunofluorescence analyses showed that METTL16 promotes myoblast proliferation while inhibiting myoblast differentiation. We also observed that METTL16 induces the upregulation of slow-twitch myosin heavy chain (MyHC) and slow-twitch-specific genes in myotubes, while downregulating fast-twitch MyHC and fast-twitch-specific genes. Furthermore, both interference and overexpression of METTL16 led to changes in overall cellular m6A modification levels and Methyltransferase like 3 (METTL3) expression levels. These findings confirm that METTL16 plays a key role in myoblast proliferation, differentiation, and muscle fiber type formation in chickens. Considering the role of myoblasts in chicken muscle growth and meat quality regulation, METTL16 may serve as a key target for molecular selection in chicken meat traits.

Electrospun "Hard-Soft" Interpenetrating Nanofibrous Tissue Scaffolds Facilitating Enhanced Mechanical Strength and Cell Proliferation.

"Soft" hydrogel-based macroporous scaffolds have been widely used in tissue engineering and drug delivery applications due to their hydrated interfaces and macroporous structures, but have drawbacks related to their weak mechanics and often weak adhesion to cells. In contrast, "hard" poly(caprolactone) (PCL) electrospun fibrous networks have desirable mechanical strength and ductility but offer minimal interfacial hydration and thus limited capacity for cell proliferation. Herein, we demonstrate the fabrication of interpenetrating nanofibrous networks based on coelectrospun PCL and poly(oligoethylene glycol methacrylate) (POEGMA) nanofibers that exhibit the mechanical benefits of PCL but the interfacial hydration benefits of hydrogels. The electrospinning process results in partially aligned but interpenetrating fiber network with minimal internal phase separation, leading to anisotropic but strong mechanical properties even in the hydrated state; apparent ultimate tensile strengths of the swollen scaffolds ranged from 429 ± 39 kPa in the direction of fiber alignment (longitudinal) to 86 ± 25 kPa perpendicular to fiber alignment (cross-longitudinal), typical of PCL-based scaffolds and enabling efficient suture retention in different directions. However, contact angle measurements indicate hydrogel-like interfacial properties due to the presence of the interpenetrating POEGMA network. C2C12 myoblast proliferation in the PCL-POEGMA scaffolds was 50% higher than that observed on PCL-only scaffolds, a result attributed to the presence of the more hydrophilic POEGMA interpenetrating nanofiber network. Overall, this method is demonstrated to represent a facile single-step strategy to fabricate strong macroporous but still interfacially hydrophilic scaffolds for tissue engineering applications.

Skeletal muscle-derived exosomes selectively coated miRNAs and participate in myoblast proliferation and differentiation mediated by miR-4331-3p

The phenotypic characteristics and meat quality of skeletal muscles are collectively determined by muscle cells and their intricate interactions with the extracellular microenvironment. In this study, we evaluated muscle fiber phenotypes in the longissimus dorsi (HC-L) and psoas major (HC-P) of Hechuan black pigs. The results revealed significant differences in muscle fiber diameter, density, and type (P < 0.05). Subsequently, co-culture experiments with myoblasts demonstrated that skeletal muscle-derived exosomes (SKM-Exos) promoted myoblast proliferation and differentiation with P-Exo exhibiting superior efficacy in promoting the augmentation of MyHCIIa fiber. Furthermore, SKM-Exos are inherently heterogeneous, and the microRNAs (miRNAs) present in SKM-Exos are selectively coated. Notably, the expression of miR-4331-3p was significantly higher in SKM-Exos than in the corresponding skeletal muscles. The expression of miR-4331-3p was significantly elevated in the SKM-Exos of HC-L compared to that of HC-P, and it interacted with differentially expressed genes between HC-L and HC-P. Moreover, miR-4331-3p enhanced myoblast proliferation and inhibited differentiation. Our findings offer valuable insights into the molecular processes that contribute to meat formation, including intricate cellular interactions.

Androgen receptor regulates the differentiation of myoblasts under cyclic mechanical stretch and its upstream and downstream signals

Our previous studies have demonstrated the important roles of androgen receptor (AR) in myoblast proliferation regulated by 15 % (mimic appropriate exercise) and 20 % (mimic excessive exercise) mechanical stretches. Except for myoblast proliferation, differentiation is also an important factor affecting muscle mass and strength. But the role of AR in stretch-regulated myoblast differentiation and AR's upstream and downstream signals remain unknown. In the present study, firstly the differences of myogenic differentiation between C2C12 (with AR expression) and L6 (without AR expression) myoblasts induced by 15 % and 20 % mechanical stretches were compared; secondly, AR antagonist flutamide and AR agonist GTx-007 were used in 15 % and 20 % stretched myoblasts respectively to confirm AR's roles in stretch-regulated myoblast differentiation; thirdly, RNA-seq, molecular dynamic simulation (MD) and co-immunoprecipitation were performed to screen the downstream and upstream molecules of AR during stretches. We found that (1) 15 % stretch increased while 20 % stretch decreased myotube number in differentiating C2C12 and L6 myoblasts, with more significant changes in C2C12 cells than L6 cells; (2) in stretched C2C12 myoblasts, AR antagonist flutamide inhibited 15 % stretch-promoted differentiation while AR agonist GTx-007 reversed 20 % stretch-inhibited differentiation (reflected by changes in myotube number, MHC contents of fast-twitch and slow-twitch fiber, and the levels of myogenic regulatory factors (MRFs) such as MyoD and myogenin); (3) KEGG analysis of RNA-seq showed that the differently expressed genes (DEGs) in C2C12 cells induced by 15 % stretch were enriched in FoxO and JAK-STAT signaling pathways, while DEGs by 20 % stretch were enriched in FoxO and MAPK signaling pathways; (4) MD and co-immunoprecipitation showed that β1 integrin could interact with AR and influence AR's activity in C2C12 cells. In conclusion, AR plays important roles in myoblast differentiation promoted by 15 % stretch while inhibited by 20 % stretch, which was fulfilled through FoxO-MRFs. In addition, α7β1 integrin may be a bridge linking mechanical stretch and AR. This study is beneficial to deeply understand the roles and mechanisms of AR in stretch-regulated muscle mass and strength; and reports firstly that myoblasts sense mechanical stimulus and transmit into intracellular AR via α7β1 integrin.

Macrophages in the Context of Muscle Regeneration and Duchenne Muscular Dystrophy.

Macrophages are essential to muscle regeneration, as they regulate inflammation, carry out phagocytosis, and facilitate tissue repair. These cells exhibit phenotypic switching from pro-inflammatory (M1) to anti-inflammatory (M2) states during muscle repair, influencing myoblast proliferation, differentiation, and myofiber formation. In Duchenne Muscular Dystrophy (DMD), asynchronous muscle injuries disrupt the normal temporal stages of regeneration, leading to fibrosis and failed regeneration. Altered macrophage activity is associated with DMD progression and physiopathology. Gaining insight into the intricate relationship between macrophages and muscle cells is crucial for creating effective therapies aimed at treating this muscle disorder. This review explores the dynamic functions of macrophages in muscle regeneration and their implications in DMD.

IGF2BP1-mediated the stability and protein translation of FGFR1 mRNA regulates myogenesis through the ERK signaling pathway

N6-methyladenosine (m6A) is the most prevalent post-transcriptional modification of RNAs and plays a key regulatory role in various biological processes. As a member of the insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) family, IGF2BP1 has recently demonstrated its ability to specifically bind m6A-modified sites within mRNAs and effectively regulate their mRNA stability. However, the precise roles of IGF2BP1 in mammalian skeletal muscle development, along with its downstream mRNA targets during myogenesis, have yet to be fully elucidated. Here, we observed that IGF2BP1 expression significantly decreased during myogenic differentiation. Knockdown of IGF2BP1 significantly inhibited myoblast proliferation while promoted myogenic differentiation. In contrast, IGF2BP1 overexpression robustly stimulated myoblast proliferation but suppressed their differentiation. Combined analysis of high-throughput sequencing and RNA stability assays revealed that IGF2BP1 can enhance fibroblast growth factor receptor 1 (FGFR1) mRNA stability and promote its translation in an m6A-dependent manner, thereby regulating its expression level and the Extracellular Signal-Regulated Kinase (ERK) pathway. Additionally, knockdown of FGFR1 rescued the phenotypic changes (namely increased cell proliferation and suppressed differentiation) induced by IGF2BP1 overexpression via attenuating ERK signaling. Taken together, our findings suggest that IGF2BP1 maintains the stability and translation of FGFR1 mRNA in an m6A-dependent manner, thereby inhibiting skeletal myogenesis through activation of the ERK signaling pathway. This study further enriches the understanding of the molecular mechanisms by which RNA methylation regulates myogenesis, providing valuable insights into the role of IGF2BP1-mediated post-transcriptional regulation in muscle development.

Pleiotropic Gene HMGA2 Regulates Myoblast Proliferation and Affects Body Size of Sheep.

Uncovering genes associated with muscle growth and body size will benefit the molecular breeding of meat Hu sheep. HMGA2 has proven to be an important gene in mouse muscle growth and is associated with the body size of various species. However, its roles in sheep are still limited. Using sheep myoblast as a cell model, the overexpression of HMGA2 significantly promoted sheep myoblast proliferation, while interference with HMGA2 expression inhibited proliferation, indicated by qPCR, EdU, and CCK-8 assays. Furthermore, the dual-luciferase reporter system indicated that the region NC_056056.1: 154134300-154134882 (-618 to -1200 bp upstream of the HMGA2 transcription start site) was one of the habitats of the HMGA2 core promoter, given the observation that this fragment led to a ~3-fold increase in luciferase activity. Interestingly, SNP rs428001129 (NC_056056.1:g.154134315 C>A) was detected in this fragment by Sanger sequencing of the PCR product of pooled DNA from 458 crossbred sheep. This SNP was found to affect the promoter activity and was significantly associated with chest width at birth and two months old, as well as chest depth at two and six months old. The data obtained in this study demonstrated the phenotypic regulatory role of the HMGA2 gene in sheep production traits and the potential of rs428001129 in marker-assisted selection for sheep breeding in terms of chest width and chest depth.

438 DNA methylation patterns and transcriptional regulation during pig fetal skeletal muscle development

Abstract Fetal development is controlled by a complex cascade of highly regulated and coordinated gene expression patterns. Epigenetic mechanisms have important roles in regulating development and differentiation. Among such mechanisms, DNA methylation exhibits context-specific associations with gene expression and has been shown to be highly dynamic during developmental processes. We performed whole-genome bisulfite sequencing (WGBS) to assess DNA methylation in pig longissimus dorsi muscle at 41- and 70-d gestation (dg), as well as RNA- and small RNA-sequencing to identify coordinated changes in methylation and expression between myogenic stages. We identified 45,739 differentially methylated regions (DMRs) between stages, and the majority (n = 34,232) were hypomethylated at 70 vs. 41 dg. Developmental DMRs exhibited feature-specific enrichment in gene regulatory regions, as well as in regions proximal to micro-RNAs (miRNAs) that have known roles in myogenesis. Integration of methylation and transcriptomic data revealed strong associations between differential gene methylation and transcript abundance. We surveyed myogenic regulatory factor (MRF) genes to determine if differential methylation was present in expected genomic regions. Within the MYF5 and MYF6 locus, MYF5 was significantly promoter-hypermethylated at 70 dg, whereas MYF6 was significantly hypomethylated upstream of its transcription start site. MYF5 is the earliest MRF to be expressed and primarily functions in myoblast proliferation and determination, while MYF6 functions in muscle cell differentiation. Thus, these patterns were consistent with expected downregulation of MYF5 and upregulation of MYF6 as muscle development progresses and demonstrate that differential methylation is evident at myogenic transcription factors. Differential miRNA methylation was significantly negatively correlated with abundance, and dynamic expression of assayed miRNAs persisted postnatally. Motif analysis revealed significant enrichment of myogenic regulatory factor motifs among hypomethylated regions, suggesting that DNA hypomethylation may function to increase accessibility of muscle-specific transcription factors. We also show that developmental DMRs are enriched for GWAS SNPs associated with muscle physiology and meat quality traits, demonstrating the potential for epigenetic processes to influence phenotypic diversity. Our results enhance understanding of DNA methylation dynamics in pig fetal skeletal muscle and reveal putative cis-regulatory elements governed by epigenetic processes during porcine myogenesis.

Bioactive Nb2C MXene-Functionalized Hydrogel with Microenvironment Remodeling and Enhanced Neurogenesis to Promote Skeletal Muscle Regeneration and Functional Restoration.

The complete structure-functional repair of volumetric muscle loss (VML) remains a giant challenge and biomedical hydrogels to remodel microenvironment and enhance neurogenesis have appeared to be a promising direction. However, the current hydrogels for VML repair hardly achieve these two goals simultaneously due to their insufficient functionality and the challenge in high-cost of bioactive factors. In this study, a facile strategy using Nb2C MXene-functionalized hydrogel (OPTN) as a bioactive scaffold is proposed to promote VML repair with skeletal muscle regeneration and functional restoration. In vitro experiments show that OPTN scaffold can effectively scavenge reactive oxygen species (ROS), guide macrophages polarization toward M2 phenotype, and resist bacterial infection, providing a favorable microenvironment for myoblasts proliferation as well as the endothelial cells proliferation, migration, and tube formation. More importantly, OPTN scaffold with electroactive feature remarkably boosts myoblasts differentiation and mesenchymal stem cells neural differentiation. Animal experiments further confirm that OPTN scaffold can achieve a prominent structure-functional VML repair by attenuating ROS levels, alleviating inflammation, reducing fibrosis, and facilitating angiogenesis, newborn myotube formation, and neurogenesis. Collectively, this study provides a highly promising and effective strategy for the structure-functional VML repair through designing bioactive multifunctional hydrogel with microenvironment remodeling and enhanced neurogenesis.

IGFBP7 promotes the proliferation and differentiation of primary myoblasts and intramuscular preadipocytes in chicken

Though it is well known that insulin-like growth factor (IGF) binding protein 7 (IGFBP7) plays an important role in myogenesis and adipogenesis in mammals, its impact on the proliferation, differentiation, and lipid deposition in chicken primary myoblasts (CPM) and intramuscular preadipocytes remains unexplored. In the present study, we firstly examined the correlation between SNPs within the genomic sequence of the IGFBP7 gene and carcass and blood chemical traits in a F2 resource population by genetic association analysis, and found that a significant correlation between the SNP (4_49499525) located in the intron region of IGFBP7 and serum high-density lipoproteins (HDL). We then examined the expression patterns of IGFBP7 across different stages of proliferation and differentiation in CPMs and intramuscular preadipocytes via qPCR, and explored the biological functions of IGFBP7 through gain- and loss-of-function experiments and a range of techniques including qPCR, CCK-8, EdU, flow cytometry, Western blot, immunofluorescence, and Oil Red O staining to detect the proliferation, differentiation, and lipid deposition in CPMs and intramuscular preadipocytes. We ascertained that the expression levels of the IGFBP7 gene increased as cell differentiation progresses in CPMs and intramuscular preadipocytes, and that IGFBP7 promotes the proliferation and differentiation of these cells, as well as facilitates intracellular lipid deposition. Furthermore, we investigated the regulatory mechanism of IGFBP7 expression by using co-transfection strategy and dual-luciferase reporter assay, and discovered that the myogenic transcription factors (MRF), myoblast determination factor (MyoD) and myogenin (MyoG), along with the adipocyte-specific transcription factor (TF) CCAAT/enhancer-binding protein α (C/EBPα), can bind to the core transcription activation region of the IGFBP7 promoter located 500 bp upstream from the transcription start site, thereby promoting IGFBP7 transcription and expression. Taken together, our study underscores the role of IGFBP7 as a positive regulator for myogenesis and adipogenesis, while also elucidating the functional and transcriptional regulatory mechanisms of IGFBP7 in chicken skeletal muscle development and intramuscular adipogenesis.

Glucosamine inhibits myoblast proliferation and differentiation, and stimulates myotube atrophy through distinct signal pathways

Glucosamine (GlcN) is one of the dietary supplements used in the treatment of osteoarthritis. Endogenously, GlcN is synthesized from glucose through the hexosamine pathway. In addition to ameliorating arthritis, several biological functions of GlcN have been reported, including insulin resistance in skeletal muscle. However, the regulatory role of GlcN in skeletal muscle development is not clear. We therefore investigated the effect of GlcN on myoblast proliferation, differentiation, and myotube development and their underlying mechanisms in C2C12 cells. Myoblast proliferation was measured by MTT assay. The expressions of MyoD, myogenin (MyoG), and myosin heavy chain (MyHC) were identified as determinants of myoblast differentiation. Expressions of atrogin-1 and muscle RING-finger protein-1 (MuRF-1) were identified as markers of myotube atrophy. The results show that treatment with GlcN significantly reduced myoblast proliferation and phosphorylation of Stat3 and S6K. These findings suggest that GlcN can inhibit growth of myoblasts through inhibiting phosphorylation of Stat3 and S6K. In addition, GlcN significantly suppressed the expression of MyoD, MyoG, and MyHC, as well as myotube formation. Pretreatment of C2C12 myoblast cells with ER stress inhibitors significantly blocked GlcN-inhibited MyHC expression and myotube formation. It can be concluded that GlcN suppressed myogenic differentiation via a pathway that involved ER stress. Moreover, GlcN decreased myotube diameter and expression of MyHC, as well as increased MuRF-1 in C2C12 myotubes. Meanwhile, GlcN also reduced the expressions of phosphorylated Akt and mTOR were stimulated after GlcN treatment in C2C12 myotubes. Thus, GlcN induced skeletal muscle atrophy by inhibiting the protein synthesis pathway. Chronic GlcN infusion also caused skeletal muscle atrophy in mice. In conclusion, GlcN regulated important stages of skeletal muscle development through different signaling pathways.

Functional study of the ST6GAL2 gene regulating skeletal muscle growth and development

ST6GAL2, a member of the sialoglycosyltransferase family, primarily localizes within the cellular Golgi apparatus. However, the role of the ST6GAL2 gene in skeletal muscle growth and development remains elusive. In this study, the impact of the ST6GAL2 gene on the proliferation, differentiation, and apoptosis of primary chicken myoblasts at the cellular level was investigated. Quantitative fluorescent PCR was used to measure the expression levels of genes. Subsequently, using gene knockout mice, we assessed its effects on skeletal muscle growth and development in vivo. Our findings reveal that the ST6GAL2 gene promotes the expression of cell cycle and proliferation-related genes, including CCNB2 and PCNA, and apoptosis-related genes, such as Fas and Caspase-9. At the individual level, double knockout of ST6GAL2 inhibited the formation of both fast and slow muscle fibers in the quadriceps, extensor digitorum longus, and tibial anterior muscle, while promoting their formation in the gastrocnemius and soleus. These results collectively demonstrate that the ST6GAL2 gene facilitates the proliferation, apoptosis, and fusion processes of primary chicken myoblasts. Additionally, it promotes the enlargement of cross-sectional muscle fiber areas and regulates the formation of fast and slow muscle fibers at the individual level, albeit inhibiting muscle fusion. This study provides valuable insights into the role of the ST6GAL2 gene in promoting proliferation of skeletal muscle.

Role of circPAPD7 in regulating proliferation and differentiation of goat skeletal muscle satellite cells

The circular RNA (circRNA) plays a crucial role in various biological processes, particularly posttranscriptional regulation. However, the role of circRNA in the development of goat skeletal muscle has not been thoroughly explored. Here, we identified circPAPD7, which is a novel circular RNA that is preferentially expressed in the skeletal muscle. Functional assays demonstrated that circPAPD7 promoted proliferation and inhibited differentiation in goat skeletal muscle satellite cells (MuSCs). Mechanistically, it was discovered that circPAPD7 interacts with miR-26a-5p. Moreover, the rescue experiments indicated that the overexpression of circPAPD7 may reverse the inhibitory impact of miR-26a-5p on myoblast proliferation and the accelerated effects on differentiation. Furthermore, we provided evidence that circPAPD7 functions as a sponge for miR-26a-5p, thereby facilitating the upregulation of EZH2 expression in goat MuSCs. Together, the results revealed that circPAPD7 promote proliferation and inhibit differentiation of goat MuSCs via the miR-26a-5p/EZH2 pathway.

Selenium promotes broiler myoblast proliferation through the ROS/PTEN/PI3K/AKT signaling axis

Selenium (Se), an indispensable trace element in broiler chickens, is closely associated with the growth and development of skeletal muscles. However, the role of Se in the proliferation of broiler myoblasts and its specific biological mechanisms have not been elucidated. In the present study, an in vitro growth model of broiler pectoral myoblasts cultured with Se (Na2SeO3) for 24 h was established. Using light microscopy, Cell Counting Kit-8 (CCK-8) assay, and flow cytometry, we found that compared to the control (Con) group, Se supplemental level obviously promoted myoblast proliferation and prevented cell cycle arrest from the G1 phase to the S + G2 phase. Through intracellular reactive oxygen species (ROS) generation detection, western blotting, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR), the study showed that the reduced ROS production caused by Se supplementation significantly decreased PTEN expression and activated the PI3K/AKT signaling pathway in myoblasts, thereby promoting the P53/P21/CyclinD1-regulated cell cycle progression, as well as the expression of proliferation-related myogenic regulatory factors (MRF). Our findings support the potential of Se to maintain the proliferative capacity of chicken myoblasts and emphasize the importance of Se intake in regulating skeletal muscle growth and development in poultry.

Highly conductive, stretchable, and biocompatible graphene oxide biocomposite hydrogel for advanced tissue engineering

The importance of hydrogels in tissue engineering cannot be overemphasized due to their resemblance to the native extracellular matrix. However, natural hydrogels with satisfactory biocompatibility exhibit poor mechanical behavior, which hampers their application in stress-bearing soft tissue engineering. Here, we describe the fabrication of a double methacrylated gelatin bioink covalently linked to graphene oxide (GO) via a zero-length crosslinker, digitally light-processed (DLP) printable into 3D complex structures with high fidelity. The resultant natural hydrogel (GelGOMA) exhibits a conductivity of 15.0 S m-1as a result of the delocalization of theπ-orbital from the covalently linked GO. Furthermore, the hydrogel shows a compressive strength of 1.6 MPa, and a 2.0 mm thick GelGOMA can withstand a 1.0 kg ms-1momentum. The printability and mechanical strengths of GelGOMAs were demonstrated by printing a fish heart with a functional fluid pumping mechanism and tricuspid valves. Its biocompatibility, electroconductivity, and physiological relevance enhanced the proliferation and differentiation of myoblasts and neuroblasts and the contraction of human-induced pluripotent stem cell-derived cardiomyocytes. GelGOMA demonstrates the potential for the tissue engineering of functional hearts and wearable electronic devices.

A serum-free culture medium production system by co-culture combining growth factor-secreting cells and l-lactate-assimilating cyanobacteria for sustainable cultured meat production

Large-scale production of cultured meat requires bulk culture medium containing growth-promoting proteins from animal serum. However, animal serum for mammalian cell culture is associated with high costs, ethical concerns, and contamination risks. Owing to its growth factor content, conditioned medium from rat liver epithelial RL34 cells can replace animal serum for myoblast proliferation. More seeded cells and longer culture periods are thought to yield higher growth factor levels, resulting in more effective muscle cell proliferation. However, RL34 cells can deplete nutrients and release harmful metabolites into the culture medium over time, potentially causing growth inhibition and apoptosis. This issue highlights the need for waste clearance during condition medium production. To address this issue, we introduced a lactate permease gene (lldP) and an l-lactate-to-pyruvate conversion enzyme gene (lldD) to generate a recombinant l-lactate-assimilating cyanobacterium Synechococcus sp. KC0110 strain. Transwell co-culture of this strain with RL34 cells exhibited a marked reduction in the levels of harmful metabolites, lactate and ammonium, while maintaining higher concentrations of glucose, pyruvate, and pyruvate-derived amino acids than those seen with RL34 cell monocultures. The co-culture medium supported myoblast proliferation without medium dilution or additional nutrients, which was attributed to the waste clearance and nutrient replenishment effects of the KC0110 strain. This culture system holds potential for the production of low-cost, and animal-free cultured meat.