-
Poultry Science May 2024Long-term intensive genetic selection has led to significant differences between broiler and layer chickens, which are evident during the embryonic period. Despite this,...
Long-term intensive genetic selection has led to significant differences between broiler and layer chickens, which are evident during the embryonic period. Despite this, there is a paucity of research on the genetic regulation of the initial formation of muscle fiber morphology in chick embryos. Embryonic d 17 (E17) is the key time point for myoblast fusion completion and muscle fiber morphology formation in chickens. This study aimed to explore the genetic regulatory mechanisms underlying the early muscle fiber morphology establishment in broiler chickens of Cornish (CC) and White Plymouth Rock (RR) and layer chickens of White Leghorn (WW) at E17 using the transcriptomic and chromatin accessibility sequencing of pectoral major muscles. The results showed that broiler chickens exhibited significant higher embryo weight and pectoral major muscle weight at E17 compared to layer chickens (P = 0.000). A total of 1,278, 1,248, and 892 differentially expressed genes (DEGs) of RNA-seq data were identified between CC vs. WW, RR vs. WW, and CC vs. RR, separately. All DEGs were combined for cluster analysis and they were divided into 6 clusters, including cluster 1 with higher expression in broilers and cluster 6 with higher expression in layers. DEGs in cluster 1 were enriched in terms related to macrophage activation (P = 0.002) and defense response to bacteria (P = 0.002), while DEGs in cluster 6 showed enrichment in protein-DNA complex (P = 0.003) and monooxygenase activity (P = 0.000). ATAC-seq data analysis identified a total of 38,603 peaks, with 13,051 peaks for CC, 18,780 peaks for RR, and 6,772 peaks for WW. Integrative analysis of transcriptomic and chromatin accessibility data revealed GOLM1, ISLR2, and TOPAZ1 were commonly upregulated genes in CC and RR. Furthermore, screening of all upregulated DEGs in cluster 1 from CC and RR identified GOLM1, ISLR2, and HNMT genes associated with neuroimmune functions and MYOM3 linked to muscle morphology development, showing significantly elevated expression in broiler chickens compared to layer chickens. These findings suggest active neural system connectivity during the initial formation of muscle fiber morphology in embryonic period, highlighting the early interaction between muscle fiber formation morphology and the nervous system. This study provides novel insights into late chick embryo development and lays a deeper foundation for further research.
PubMed: 38833745
DOI: 10.1016/j.psj.2024.103882 -
Tissue Engineering. Part A Jun 2024The fusion index is a key indicator for quantifying the differentiation of a myoblast population, which is often calculated manually. In addition to being...
The fusion index is a key indicator for quantifying the differentiation of a myoblast population, which is often calculated manually. In addition to being time-consuming, manual quantification is also error prone and subjective. Several software tools have been proposed for addressing these limitations but suffer from various drawbacks, including unintuitive interfaces and limited performance. In this study, we describe MyoFInDer, a Python-based program for the automated computation of the fusion index of skeletal muscle. At the core of MyoFInDer is a powerful artificial intelligence-based image segmentation model. MyoFInDer also determines the total nuclei count and the percentage of stained area and allows for manual verification and correction. MyoFInDer can reliably determine the fusion index, with a high correlation to manual counting. Compared with other tools, MyoFInDer stands out as it minimizes the interoperator variability, minimizes process time and displays the best correlation to manual counting. Therefore, it is a suitable choice for calculating fusion index in an automated way, and gives researchers access to the high performance and flexibility of a modern artificial intelligence model. As a free and open-source project, MyoFInDer can be modified or extended to meet specific needs.
PubMed: 38832871
DOI: 10.1089/ten.TEA.2024.0049 -
ELife Jun 2024Styxl2, a poorly characterized pseudophosphatase, was identified as a transcriptional target of the Jak1-Stat1 pathway during myoblast differentiation in culture. Styxl2...
Styxl2, a poorly characterized pseudophosphatase, was identified as a transcriptional target of the Jak1-Stat1 pathway during myoblast differentiation in culture. Styxl2 is specifically expressed in vertebrate striated muscles. By gene knockdown in zebrafish or genetic knockout in mice, we found that Styxl2 plays an essential role in maintaining sarcomere integrity in developing muscles. To further reveal the functions of Styxl2 in adult muscles, we generated two inducible knockout mouse models: one with being deleted in mature myofibers to assess its role in sarcomere maintenance, and the other in adult muscle satellite cells (MuSCs) to assess its role in de novo sarcomere assembly. We find that Styxl2 is not required for sarcomere maintenance but functions in de novo sarcomere assembly during injury-induced muscle regeneration. Mechanistically, Styxl2 interacts with non-muscle myosin IIs, enhances their ubiquitination, and targets them for autophagy-dependent degradation. Without Styxl2, the degradation of non-muscle myosin IIs is delayed, which leads to defective sarcomere assembly and force generation. Thus, Styxl2 promotes de novo sarcomere assembly by interacting with non-muscle myosin IIs and facilitating their autophagic degradation.
Topics: Animals; Mice; Mice, Knockout; Proteolysis; Sarcomeres; Zebrafish; Phosphoprotein Phosphatases
PubMed: 38829202
DOI: 10.7554/eLife.87434 -
British Poultry Science Jun 20241. The development of chicken skeletal muscle is directly relevant to poultry husbandry production. Numerous studies have suggested that circular RNA play pivotal roles...
1. The development of chicken skeletal muscle is directly relevant to poultry husbandry production. Numerous studies have suggested that circular RNA play pivotal roles in muscle development. However, the functions and mechanisms of most circRNA in chicken myogenesis remain largely unknown.2. This study identified a novel circSESN1 based on existing sequencing data and examined its authenticity and subcellular localisation by enzyme digestion and RNA fluorescence in situ hybridisation. Additionally, there was a positive correlation between the expression levels of circSESN1 and the developmental stage of chicken muscle.3. Mechanistically, knockdown or overexpression of circSESN1 was performed in primary myoblasts to validate its function. The interactions between circSESN1, miR-16-5p, and the target gene 1 (1) were investigated using bioinformatics analysis and a dual fluorescein reporter system. Real-time qPCR, a cell proliferation assay, and immunofluorescence staining techniques were used to investigate the promotion effect of circSESN1 on myoblast proliferation and differentiation by miR-16-5p/1 pathway.4. The results demonstrated that the newly identified chicken circSESN1 directly sponges gga-miR-16-5p to regulate 1 gene expression, promoting myoblast proliferation and differentiation.
PubMed: 38828863
DOI: 10.1080/00071668.2024.2360004 -
BioRxiv : the Preprint Server For... May 2024Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of motoneurons (MNs), and despite progress, there is no effective...
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of motoneurons (MNs), and despite progress, there is no effective treatment. A large body of evidence shows that astrocytes expressing ALS-linked mutant proteins cause non-cell autonomous toxicity of MNs. Although MNs innervate muscle fibers and ALS is characterized by the early disruption of the neuromuscular junction (NMJ) and axon degeneration, there are controversies about whether muscle contributes to non-cell-autonomous toxicity to MNs. In this study, we generated primary skeletal myotubes from myoblasts derived from ALS mice expressing human mutant SOD1 (termed hereafter mutSOD1). Characterization revealed that mutSOD1 skeletal myotubes display intrinsic phenotypic and functional differences compared to control myotubes generated from non-transgenic (NTg) littermates. Next, we analyzed whether ALS myotubes exert non-cell-autonomous toxicity to MNs. We report that conditioned media from mutSOD1 myotubes (mutSOD1-MCM), but not from control myotubes (NTg-MCM), induced robust death of primary MNs in mixed spinal cord cultures and compartmentalized microfluidic chambers. Our study further revealed that applying mutSOD1-MCM to the MN axonal side in microfluidic devices rapidly reduces mitochondrial axonal transport while increasing Ca2+ transients and reactive oxygen species (i.e., H O ). These results indicate that soluble factor(s) released by mutSOD1 myotubes cause MN axonopathy that leads to lethal pathogenic changes.
PubMed: 38826246
DOI: 10.1101/2024.05.24.595817 -
International Journal of Biological... Jun 2024The mass proliferation of seed cells and imitation of meat structures remain challenging for cell-cultured meat production. With excellent biocompatibility, high water...
The mass proliferation of seed cells and imitation of meat structures remain challenging for cell-cultured meat production. With excellent biocompatibility, high water content and porosity, hydrogels are frequently-studied materials for anchorage-dependent cell scaffolds in biotechnology applications. Herein, a scaffold based on gelatin/alginate/ε-Poly-l-lysine (GAL) hydrogel is developed for skeletal muscle cells, which has a great prospect in cell-cultured meat production. In this work, the hydrogel GAL-4:1, composed of gelatin (5 %, w/v), alginate (5 %, w/v) and ε-Poly-l-lysine (molar ratio vs. alginate: 4:1) is selected as cell scaffold based on Young's modulus of 11.29 ± 1.94 kPa, satisfactory shear-thinning property and suitable porous organized structure. The commercially available C2C12 mouse skeletal myoblasts and porcine muscle stem cells (PMuSCs), are cultured in the 3D-printed scaffold. The cells show strong ability of attachment, proliferation and differentiation after induction, showing high biocompatibility. Furthermore, the cellular bioprinting is performed with GAL-4:1 hydrogel and freshly extracted PMuSCs. The extracted PMuSCs exhibit high viability and display early myogenesis (desmin) on the 3D scaffold, suggesting the great potential of GAL hydrogel as 3D cellular constructs scaffolds. Overall, we develop a novel GAL hydrogel as a 3D-printed bioactive platform for cultured meat research.
Topics: Animals; Alginates; Gelatin; Polylysine; Cell Differentiation; Tissue Scaffolds; Swine; Printing, Three-Dimensional; Cell Proliferation; Mice; Hydrogels; Stem Cells; Meat; Muscle Development; Tissue Engineering; Cell Line; Bioprinting; In Vitro Meat
PubMed: 38821790
DOI: 10.1016/j.ijbiomac.2024.131980 -
Science Advances May 2024The ability of cells to organize into tissues with proper structure and function requires the effective coordination of proliferation, migration, polarization, and...
The ability of cells to organize into tissues with proper structure and function requires the effective coordination of proliferation, migration, polarization, and differentiation across length scales. Skeletal muscle is innately anisotropic; however, few biomaterials can emulate mechanical anisotropy to determine its influence on tissue patterning without introducing confounding topography. Here, we demonstrate that substrate stiffness anisotropy coordinates contractility-driven collective cellular dynamics resulting in C2C12 myotube alignment over millimeter-scale distances. When cultured on mechanically anisotropic liquid crystalline polymer networks (LCNs) lacking topography, C2C12 myoblasts collectively polarize in the stiffest direction. Cellular coordination is amplified through reciprocal cell-ECM dynamics that emerge during fusion, driving global myotube-ECM ordering. Conversely, myotube alignment was restricted to small local domains with no directional preference on mechanically isotropic LCNs of the same chemical formulation. These findings provide valuable insights for designing biomaterials that mimic anisotropic microenvironments and underscore the importance of stiffness anisotropy in orchestrating tissue morphogenesis.
Topics: Anisotropy; Animals; Muscle Fibers, Skeletal; Mice; Extracellular Matrix; Cell Line; Cell Differentiation; Muscle Contraction; Myoblasts
PubMed: 38820155
DOI: 10.1126/sciadv.adn0235 -
Cureus Apr 2024The muscle cells myocytes are differentiated for the purpose of contraction function, which plays a major role in body metabolism and energy haemostasis, through...
BACKGROUND
The muscle cells myocytes are differentiated for the purpose of contraction function, which plays a major role in body metabolism and energy haemostasis, through different metabolic pathways, such as glucose and protein metabolic pathways. Alanine aminotransferase (ALT) plays a crucial role by reversibly catalysing transamination between alanine and a-ketoglutarate to form pyruvate and glutamate and by mediating the conversion of these four major intermediate metabolites. ALT plays important roles for energy homeostasis during fasting and prolonged exercise anaerobically, when muscle protein must first be broken down into its constituent amino acids.
METHODS
Mouse skeletal myoblast cell line C2C12 was cultured in Dulbecco's modified eagle medium (DMEM) growth medium, supplied with 2% horse serum supplemented with 1 uM insulin, 2 mM glutamine and penicillin and streptomycin antibiotics for seven days. The differentiation medium is refreshed every 24 hours. Then, C2C12 cells were treated with insulin and dexamethasone to examine their effects on myocytes' ALT activity.
RESULTS
In our study, we found an impact on ALT activity under different influences, including C2C12 differentiation, dexamethasone and insulin treatments, which shed light on the dynamic interplay between ALT activity, alanine metabolism, and cellular states, like differentiation and stress responses.
CONCLUSION
The study provides valuable insights into the dynamic regulation of ALT activity and alanine metabolism in C2C12 cells across differentiation and drug treatments. Further research is encouraged to explore the underlying mechanisms and their implications for muscle function, differentiation and potential therapeutic interventions in metabolic disorders.
PubMed: 38817503
DOI: 10.7759/cureus.59331 -
Journal of Biosciences 2024Adult muscle tissue largely comprised of differentiated myofibers also harbors quiescent muscle-resident stem cells (MuSCs) that are responsible for its maintenance,... (Comparative Study)
Comparative Study
Adult muscle tissue largely comprised of differentiated myofibers also harbors quiescent muscle-resident stem cells (MuSCs) that are responsible for its maintenance, repair and regeneration. Emerging evidence suggests that quiescent MuSCs exhibit a specific metabolic state, which is regulated during physiological and pathological alterations. However, a detailed understanding of the metabolic state of quiescent MuSCs and its alteration during activation and repair is lacking. Direct profiling of MuSCs is challenging because the cells are rare and dispersed, while isolation and enrichment leads to their activation and loss of quiescence. In this study, we employed 1H-nuclear magnetic resonance (NMR) spectroscopy to profile metabolites in an established culture model of quiescent MuSC-derived myoblasts and compared with activated, proliferative and differentiated muscle cells to determine the state-specific metabolome. We report that the proliferating and differentiated cells are highly enriched in metabolites involved in energy generation, the quiescent state is enriched in metabolites related to phospholipid catabolism (glycerophosphocholine and choline) and depleted for phosphocholine which is enriched in proliferating cells. We propose that the ratio of these metabolites may be useful as a biomarker of MuSC quiescence.
Topics: Metabolomics; Animals; Cell Differentiation; Cell Proliferation; Mice; Magnetic Resonance Spectroscopy; Myoblasts; Metabolome; Muscle, Skeletal
PubMed: 38817158
DOI: No ID Found -
BMC Genomics May 2024Skeletal muscle development and fat deposition have important effects on meat quality. The study of regulating skeletal muscle development and fat deposition is of great...
BACKGROUND
Skeletal muscle development and fat deposition have important effects on meat quality. The study of regulating skeletal muscle development and fat deposition is of great significance in improving the quality of carcass and meat. In the present study, whole transcriptome sequencing (including RNA-Seq and miRNA-Seq) was performed on the longissimus dorsi muscle (LDM) of Jinfen White pigs at 1, 90, and 180 days of age.
RESULTS
The results showed that a total of 245 differentially expressed miRNAs were screened in any two comparisons, which may be involved in the regulation of myogenesis. Among them, compared with 1-day-old group, miR-22-5p was significantly up-regulated in 90-day-old group and 180-day-old group. Functional studies demonstrated that miR-22-5p inhibited the proliferation and differentiation of porcine skeletal muscle satellite cells (PSCs). Pearson correlation coefficient analysis showed that long non-coding RNA (lncRNA) LOC106505926 and CXXC5 gene had strong negative correlations with miR-22-5p. The LOC106505926 and CXXC5 were proven to promote the proliferation and differentiation of PSCs, as opposed to miR-22-5p. In terms of mechanism, LOC106505926 functions as a molecular sponge of miR-22-5p to modulate the expression of CXXC5, thereby inhibits the differentiation of PSCs. In addition, LOC106505926 regulates the differentiation of porcine preadipocytes through direct binding with FASN.
CONCLUSIONS
Collectively, our results highlight the multifaceted regulatory role of LOC106505926 in controlling skeletal muscle and adipose tissue development in pigs and provide new targets for improving the quality of livestock products by regulating skeletal muscle development and fat deposition.
Topics: Animals; RNA, Long Noncoding; Muscle Development; Swine; MicroRNAs; Lipogenesis; Cell Differentiation; Cell Proliferation; Satellite Cells, Skeletal Muscle; Muscle, Skeletal; Cells, Cultured
PubMed: 38816813
DOI: 10.1186/s12864-024-10422-y