Embryonic Muscle Research Articles

Three-dimensional visualization and quantitative analysis of embryonic and fetal thigh muscles using magnetic resonance and phase-contrast X-ray imaging.

The musculoskeletal system around the human hip joint has acquired a suitable structure for erect bipedal walking. However, little is known about the process of separation and maturation of individual muscles during the prenatal period, when muscle composition is acquired. Understanding the maturation process of the normal musculoskeletal system contributes to elucidating the acquisition of bipedal walking in humans and to predicting normal growth and detecting congenital muscle disorders and anomalies. In this study, we clarify the process of thigh muscle maturation from the embryonic stage to the mid-fetal stage using serial sections, phase-contrast X-ray computed tomography, and magnetic resonance imaging. We also provide a 4D atlas of human thigh muscles between 8 and 23 weeks of gestation. As a result, we first show that muscle separation in the lower thigh tends to progress from the superficial to the deep layers and that all musculoskeletal components are formed by Carnegie Stage 22. Next, we show that femur and muscle volume grow in correlation with crown-rump length. Finally, we show that the anterior, abductor, and posterior muscle groups in the thigh contain a high percentage of monoarticular muscle volume by the end of the embryonic period. This ratio approaches that of adult muscle composition during normal early fetal development and is typical of bipedal walking. This study of fetal muscle composition suggests that preparation for postnatal walking may begin in early fetal period.

Temporal Expression of Myogenic Regulatory Genes in Different Chicken Breeds during Embryonic Development

The basic units of skeletal muscle in all vertebrates are multinucleate myofibers, which are formed from the fusion of mononuclear myoblasts during the embryonic period. In order to understand the regulation of embryonic muscle development, we selected four chicken breeds, namely, Cornish (CN), White Plymouth Rock (WPR), White Leghorn (WL), and Beijing-You Chicken (BYC), for evaluation of their temporal expression patterns of known key regulatory genes (Myomaker, MYOD, and MSTN) during pectoral muscle (PM) and thigh muscle (TM) development. The highest expression level of Myomaker occurred from embryonic days E13 to E15 for all breeds, indicating that it was the crucial stage of myoblast fusion. Interestingly, the fast-growing CN showed the highest gene expression level of Myomaker during the crucial stage. The MYOD gene expression at D1 was much higher, implying that MYOD might have an important role after hatching. Histomorphology of PM and TM suggested that the myofibers was largely complete at E17, which was speculated to have occurred because of the expression increase in MSTN and the expression decrease in Myomaker. Our research contributes to lay a foundation for the study of myofiber development during the embryonic period in different chicken breeds.

N-CAM Expression: The Study of Muscle Disease in a Tertiary Center of Thailand

Background: Neural cell adhesion molecule (N-CAM), or CD56, is a cell-surface glycoprotein that plays a critical role in mediating intercellular adhesion in the central nervous system. It is also found to be expressed on embryonic muscle but disappears in healthy adult muscle. However, denervated or regenerating muscles can express N-CAM. Objective: To evaluate the value of N-CAM expressions in diagnosing muscle diseases in the Thai population. Materials and Methods: The present study used immunohistochemistry to interpret N-CAM expression in 75 muscle biopsy specimens diagnosed with myopathies and non-myopathies from a 3-year retrospective cohort. Results: Using the chi-squared test, there was statistically significant associativity between N-CAM expression and the diagnosis of inflammatory myopathy, but not in muscular dystrophy, neurogenic change, or non-specific myopathy even though they also showed N-CAM expression. The inflammatory myopathy showed positive N-CAM in 15 out of 17 cases with statistical significance (p<0.001, OR 14.250, 95% CI 2.960 to 68.606). The authors did not find any N-CAM expression in the present study mitochondrial myopathy or non-myopathy cases, whose biopsies were defined as a control group due to lack of myopathic change and non-specific clinical symptoms. Conclusion: The authors suggest the evaluation of N-CAM expression as a complementary tool in the diagnosis of muscle diseases that have denervated or regenerating muscle fibers, especially in inflammatory myopathy. Keywords: Neural cell adhesion molecule; N-CAM; CD56; Muscle disease; Myopathy

MiRNA sequencing of Embryonic Myogenesis in Chengkou Mountain Chicken

BackgroundSkeletal muscle tissue is among the largest organ systems in mammals, essential for survival and movement. Embryonic muscle development determines the quantity and quality of muscles after the birth of an individual. MicroRNAs (miRNAs) are a significant class of non-coding RNAs that bind to the 3’UTR region of mRNA to regulate gene function. Total RNA was extracted from the leg muscles of chicken embryos in different developmental stages of Chengkou Mountain Chicken and used to generate 171,407,341 clean small RNA reads. Target prediction, GO, and KEGG enrichment analyses determined the significantly enriched genes and pathways. Differential analysis determined the significantly different miRNAs between chicken embryo leg muscles at different developmental stages. Meanwhile, the weighted correlation network analysis (WGCNA) identified key modules in different developmental stages, and the hub miRNAs were screened following the KME value.ResultsThe clean reads contained 2047 miRNAs, including 721 existing miRNAs, 1059 known miRNAs, and 267 novel miRNAs. Many genes and pathways related to muscle development were identified, including ERBB4, MEF2C, FZD4, the Wnt, Notch, and MAPK signaling pathways. The WGCNA established the greenyellow module and gga-miR-130b-5p for E12, magenta module and gga-miR-1643-5p for E16, purple module and gga-miR-12218-5p for E19, cyan module and gga-miR-132b-5p for E21.ConclusionThese results lay a foundation for further research on the molecular regulatory mechanism of embryonic muscle development in Chengkou mountain chicken and provide a reference for other poultry and livestock muscle development studies.

Laminin α5_CD239_Spectrin is a candidate association that compensates the linkage between the basement membrane and cytoskeleton in skeletal muscle fibers

The linkage between the basement membrane (BM) and cytoskeleton is crucial for muscle fiber stability and signal transduction. Mutations in the linkage molecules can cause various types of muscular dystrophies. The different severities and times of onset suggest that compensatory linkages occur at the sarcolemma. Cluster of differentiation 239 (CD239) binds to the α5 subunit of laminin-511 extracellularly and is connected to spectrin intracellularly, resulting in a linkage between the BM and cytoskeleton. In this study, we explored the linkage of laminin α5_CD239_spectrin in skeletal muscles. Although laminin α5, CD239, and spectrin were present in embryonic skeletal muscles, they disappeared in adult skeletal muscle tissues, except for the soleus and diaphragm. Laminin α5_CD239_spectrin was localized in the skeletal muscle tissues of Duchenne muscular dystrophy and congenital muscular dystrophy mouse models. The experimental regeneration of skeletal muscle increased the CD239-mediated linkage, indicating that it responds to regeneration, but not to genetic influence. Furthermore, in silico analysis showed that laminin α5_CD239_spectrin was upregulated by steroid therapy for muscular dystrophy. Therefore, CD239-mediated linkage may serve as a therapeutic target to prevent the progression of muscular dystrophy.

Transcriptome-wide study revealed m6A and miRNA regulation of embryonic breast muscle development in Wenchang chickens.

N6-Methyladenosine (m6A) modification has been shown to play important role in skeletal muscle development. Wenchang chickens are commonly used as a high-quality animal model in researching meat quality. However, there have been no previous reports regarding the profile of m6A and its function in the embryonic breast muscle development of Wenchang chickens. In this paper, we identified different developmental stages of breast muscle in Wenchang chickens and performed m6A sequencing and miRNA sequencing in the breast muscle of embryos. Embryo breast muscles were weighed and stained with hematoxylin–eosin after hatching. We found that myofibers grew fast on the 10th day after hatching (E10) and seldom proliferated beyond the 19th day after hatching (E19). A total of 6,774 differentially methylated genes (DMGs) were identified between E10 and E19. For RNA-seq data, we found 5,586 differentially expressed genes (DEGs). After overlapping DEGs and DMGs, we recorded 651 shared genes (DEMGs). Subsequently, we performed miRNA-seq analysis and obtained 495 differentially expressed miRNAs (DEMs). Then, we overlapped DEMGs and the target genes of DEMs and obtained 72 overlapped genes (called miRNA-m6A-genes in this study). GO and KEGG results showed DEMGs enriched in many muscle development-related pathways. Furthermore, we chose WNT7B, a key regulator of skeletal muscle development, to perform IGV visualization analysis and found that the m6A levels on the WNT7B gene between E10 and E19 were significantly different. In conclusion, we found that miRNAs, in conjunction with m6A modification, played a key role in the embryonic breast muscle development of Wenchang chickens. The results of this paper offer a theoretical basis for the study of m6A function in muscle development and fat deposition of Wenchang chickens.

Gut Microbiota-Derived Indole-3-Carboxylate Influences Mucosal Integrity and Immunity Through the Activation of the Aryl Hydrocarbon Receptors and Nutrient Transporters in Broiler Chickens Challenged With Eimeria maxima.

Two studies were conducted to evaluate the effects of indole-3-carboxylate (ICOOH) as a postbiotic on maintaining intestinal homeostasis against avian coccidiosis. In the first study, an in vitro culture system was used to investigate the effects of ICOOH on the proinflammatory cytokine response of chicken macrophage cells (CMCs), gut integrity of chicken intestinal epithelial cells (IECs), differentiation of quail muscle cells (QMCs), and primary chicken embryonic muscle cells (PMCs) and anti-parasitic effect against Eimeria maxima. Cells to be tested were seeded in the 24-well plates and treated with ICOOH at concentrations of 0.1, 1.0, and 10.0 µg. CMCs were first stimulated by lipopolysaccharide (LPS) to induce an innate immune response, and QMCs and PMCs were treated with 0.5% and 2% fetal bovine serum, respectively, before they were treated with ICOOH. After 18 h of incubation, cells were harvested, and RT-PCR was performed to measure gene expression of proinflammatory cytokines of CMCs, tight junction (TJ) proteins of IECs, and muscle cell growth markers of QMCs and PMCs. In the second study, in vivo trials were carried out to study the effect of dietary ICOOH on disease parameters in broiler chickens infected with E. maxima. One hundred twenty male broiler chickens (0-day-old) were allocated into the following four treatment groups: 1) basal diet without infection (CON), 2) basal diet with E. maxima (NC), 3) ICOOH at 10.0 mg/kg feed with E. maxima (HI), and 4) ICOOH at 1.0 mg/kg feed with E. maxima (LO). Body weights (BWs) were measured on 0, 7, 14, 20, and 22 days. All groups except the CON chickens were orally infected with E. maxima on day 14. Jejunal samples were collected for lesion score and the transcriptomic analysis of cytokines and TJ proteins. In vitro, ICOOH increased the expression of TJ proteins in IECs and decreased IL-1β and IL-8 transcripts in the LPS-stimulated CMCs. In vivo, chickens on the HI diet showed reduced jejunal IL-1β, IFN-γ, and IL-10 expression and increased expression of genes activated by aryl hydrocarbon receptors and nutrient transporters in E. maxima-infected chickens. In conclusion, these results demonstrate the beneficial effects of dietary ICOOH on intestinal immune responses and barrier integrity in broiler chickens challenged with E. maxima. Furthermore, the present finding supports the notion to use microbial metabolites as novel feed additives to enhance resilience in animal agriculture.

TMBIM5 loss of function alters mitochondrial matrix ion homeostasis and causes a skeletal myopathy.

Ion fluxes across the inner mitochondrial membrane control mitochondrial volume, energy production, and apoptosis. TMBIM5, a highly conserved protein with homology to putative pH-dependent ion channels, is involved in the maintenance of mitochondrial cristae architecture, ATP production, and apoptosis. Here, we demonstrate that overexpressed TMBIM5 can mediate mitochondrial calcium uptake. Under steady-state conditions, loss of TMBIM5 results in increased potassium and reduced proton levels in the mitochondrial matrix caused by attenuated exchange of these ions. To identify the in vivo consequences of TMBIM5 dysfunction, we generated mice carrying a mutation in the channel pore. These mutant mice display increased embryonic or perinatal lethality and a skeletal myopathy which strongly correlates with tissue-specific disruption of cristae architecture, early opening of the mitochondrial permeability transition pore, reduced calcium uptake capability, and mitochondrial swelling. Our results demonstrate that TMBIM5 is an essential and important part of the mitochondrial ion transport system machinery with particular importance for embryonic development and muscle function.

Genome-Wide Identification and Characterization of Long Non-Coding RNAs in Embryo Muscle of Chicken.

Simple SummaryThis study preliminarily determined the expression pattern of lncRNA during leg muscle development in the middle and late stages of embryo in chicken, and revealed regulatory lncRNAs and pathways associated with skeletal muscle development, providing a theoretical basis for further research on the molecular mechanism of muscle development in embryo.Embryonic muscle development determines the state of muscle development and muscle morphological structure size. Recent studies have found that long non-coding RNAs (lncRNAs) could influence numerous cellular processes and regulated growth and development of flora and fauna. A total of 1056 differentially expressed lncRNAs were identified by comparing the different time points during embryonic muscle development, which included 874 new lncRNAs. Here, we found that there were different gene expression patterns on the 12th day of embryo development (E12). Herein, WGCNA and correlation analyses were used to predict lncRNA function on E12 through the screening and identification of lncRNAs related to muscle development in the embryo leg muscles of Chengkou mountain chickens at different times. GO and KEGG functional enrichment analysis was performed on target genes involved in cis-regulation and trans-regulation. An interaction network diagram was constructed based on the muscle development pathways, such as Wnt, FoxO, and PI3K-AKT signaling pathways, to determine the interaction between mRNAs and lncRNAs. This study preliminarily determined the lncRNA expression pattern of muscle development during the middle and late embryonic stages of Chengkou mountain chickens, and provided a basis to analyze the molecular mechanism of muscle development.

Med12 is Essential for Skeletal Muscle Development

Tight regulation of tissue‐ and cell‐specific gene expression is essential for healthy physiological development. Mediator is a multisubunit protein complex that links basal transcriptional machinery, transcription factors, and epigenetic modifiers to gene targets to coordinate transcriptional regulation during development. One subunit, MED12, controls tissue‐ and cell‐specific gene expression. Some mutations in MED12 cause neurological developmental disorders, while others are associated with uterine leiomyomas. In addition to these diseases, human MED12 mutations have been linked to muscle disorders such as skeletal muscle hypotonia and congenital heart defects, but the function of MED12 in muscle and muscle cell development has not been elucidated. The objective in this study was to discover a molecular role of MED12 in transcriptional control of muscle development. We hypothesized that MED12 is required for healthy muscle development, and that MED12 helps recruit myogenic transcription factors (TFs) to Mediator at the Pre‐Initiation Complex (PIC) to achieve transcriptional control. Our lab generated a conditional skeletal muscle‐specific Med12 knockout mouse model (Med12mKO) driven by a Myogenin‐Cre transgene that is expressed during embryonic muscle development. Med12mKOmice have a runted phenotype, smaller muscles, disorganized muscle architecture, and suffer premature death, compared to littermate controls. Gene expression analysis by RNA‐sequencing (RNA‐seq) of muscle from 1‐day old Med12mKO and control mice revealed that Med12 deletion dysregulates metabolic and structural genes, including several myosin genes. MED12 is a transcriptional regulator, but does not directly bind DNA. To identify putative candidate TFs that regulate muscle gene expression in a MED12‐dependent manner we performed Upstream Regulator Analysis (URA) of dysregulated genes in Med12mKO muscle. Multiple TFs that regulate muscle gene expression were predicted to do so via MED12, including SRF, which is known to regulate expression of many myosin genes. Using differentiating C2C12 mouse myoblasts as a cell model of myogenesis, knockdown of Med12by shRNA resulted in diminished myotube size and reduced expression of myosin genes, recapitulating the gene expression and muscle phenotype observed in the Med12mKOmodel. However, no interaction was detected between SRF and MED12 in C2C12 myoblasts by Co‐Immunoprecipitation (Co‐IP), which did not support our hypothesis of muscle gene expression regulation through a MED12‐SRF mechanism. To further investigate how loss of MED12 dysregulates muscle gene expression, we used an unbiased approach to identify TFs that interact with MED12. We immunoprecipitated MED12 from C2C12 myoblasts and identified interacting TFs using mass spectrometry. We are currently investigating other TFs through which MED12 could be regulating gene expression during muscle development. We conclude that Med12 is required for normal skeletal muscle development, including normal gene expression of structural and metabolic genes during myogenesis.

Morphological association between muscle attachments and ossification sites in the late cartilaginous skull of tuatara embryos.

During development, the embryonic cartilaginous skull in most vertebrates is partially replaced by bones with endochondral and perichondral ossifications. Muscle attachments are thought to influence the patterns of ossification and, hence, the differentiation of the skull. To investigate the association between muscle attachments and early ossifications of reptilian embryos, we conducted digital 3D reconstructions of the cranium, the head, and the neck musculature from a histological section series of a late term embryonic tuatara, Sphenodon punctatus, with a total body length of 52 mm. As the sole living rhynchocephalian species, it is an important outgroup in comparative studies of squamate evolution. We found that head and neck muscles are largely associated with early ossification of the basal plate and the palatoquadrate, and with three other ossifications in an older specimen with a total body length of 72 mm. These results suggest that tensile forces resulting from embryonic muscle contraction are largely, but not exclusively, correlated with the area of endochondral ossification in the chondrocranium and palatoquadrate in tuatara. Beyond little-known genetic factors, the complexity of chondrocranial architecture, the progress of its development, and the effect of multiple muscle transmitting forces in the chondrocranium must be considered to provide a more comprehensive discussion of the mechanical properties of the embryonic skull.

Identification of crucial circRNAs in skeletal muscle during chicken embryonic development

BackgroundChicken provides humans with a large amount of animal protein every year, in which skeletal muscle plays a leading role. The embryonic skeletal muscle development determines the number of muscle fibers and will affect the muscle production of chickens. CircRNAs are involved in a variety of important biological processes, including muscle development. However, studies on circRNAs in the chicken embryo muscle development are still lacking.ResultsIn the study, we collected chicken leg muscles at 14 and 20-day embryo ages both in the fast- and slow-growing groups for RNA-seq. We identified 245 and 440 differentially expressed (DE) circRNAs in the comparison group F14vsF20 and S14vsS20 respectively. GO enrichment analysis for the host genes of DE circRNAs showed that biological process (BP) terms in the top 20 related to growth in F14vsF20 were found such as positive regulation of transcription involved in G1/S phase of mitotic cell cycle, multicellular organismal macromolecule metabolic process, and multicellular organismal metabolic process. In group S14vsS20, we also found some BP terms associated with growth in the top 20 including actomyosin structure organization, actin cytoskeleton organization and myofibril assembly. A total of 7 significantly enriched pathways were obtained, containing Adherens junction and Tight junction. Further analysis of those pathways found three crucial host genes MYH9, YBX3, IGF1R in both fast- and slow-growing groups, three important host genes CTNNA3, AFDN and CREBBP only in the fast-growing group, and six host genes FGFR2, ACTN2, COL1A2, CDC42, DOCK1 and MYL3 only in the slow-growing group. In addition, circRNA-miRNA network also revealed some key regulation pairs such as novel_circ_0007646-miR-1625-5p, novel_circ_0007646-miR-1680-5p, novel_circ_0008913-miR-148b-5p, novel_circ_0008906-miR-148b-5p and novel_circ_0001640-miR-1759-3p.ConclusionsComprehensive analysis of circRNAs and their targets would contribute to a better understanding of the molecular mechanisms in poultry skeletal muscle and it also plays an important guiding role in the next research.

Quantifying maternal transfer of trace elements and stable isotopes in the endangered pelagic thresher shark (Alopias pelagicus)

To quantify maternal provisioning of nutrients in the pelagic thresher shark (Alopias pelagicus) and the potential for negative impacts, the concentrations of trace elements (essential: Co, Cr, Cu, Mn, Ni, Se, and Zn; nonessential: As, Ba, Cd, Hg, and Pb) and fractionation of stable isotopes (13C and 15N) were analyzed in the muscle and liver of 10 pregnant females and 18 associated embryos. Essential trace elements were observed to be offloaded at higher concentrations to embryos, with the exception of Zn and Ni in liver, while nonessential trace elements were unevenly distributed between maternal-embryo tissues. Observed Hg concentrations were at levels considered toxic in A. pelagicus, but the Se: Hg molar ratios in all embryonic tissues were all greater than one. A negative correlation was observed between transfer ratios and concentrations of all elements in maternal tissue, indicating the existence of a regulatory mechanism in maternal ovaries of A. pelagicus. Compared with maternal specimens, associated embryos had higher δ13C and δ15N values in muscle and liver tissue. Negative correlations were observed between δ13C, δ15N, and Δδ13C values and precaudal length in embryonic muscle tissue potentially reflecting either a dietary-habitat shift in pregnant females during the latter period of gestation or a physiological change modifying fractionation. Higher concentrations of essential elements are linked to potential benefits for embryos during early development, levels of Hg suggested a degree of anthropogenic impact with unknown consequences while the directionality of isotopic fractionation could suggest a potential reproductive migration as a protective mechanism for birthing.

Synthesis and Biological Evaluation of Novel Bufalin Derivatives.

Bufalin and other cardiac steroids (CS) have been used for centuries for the treatment of congestive heart failure, arrhythmias, and other maladies. However, toxicity and the small therapeutic window of this family of steroids limit their use. Therefore, attempts to synthesize a potent, but less toxic, CS are of major importance. In the present study, two novel bufalin derivatives were synthesized and some of their pharmacological properties were characterized. The reaction of bufalin with Ishikawa’s reagent resulted in the production of two novel bufalin derivatives: bufalin 2,3-ene and bufalin 3,4-ene. The compounds were purified with TLC and HPLC and their structure was verified with UV, NMR, and MS analyses. The biological activities of these compounds were evaluated by testing their ability to inhibit the Na+, K+-ATPase activity of the brain microsomal fraction to induce cytotoxic activity against the NCI-60 human tumor cell line panel and non-cancer human cells, and to increase the force of contraction of quail embryonic heart muscle cells in culture. The two steroids exhibited biological activities similar to those of other CS in the tested experimental systems, but with reduced cytotoxicity, advocating their development as drugs for the treatment of heart failure and arrhythmias.

Embryonic muscle splitting patterns reveal homologies of amniote forelimb muscles.

Limb muscles are remarkably complex and evolutionarily labile. Although their anatomy is of great interest for studies of the evolution of form and function, their homologies among major amniote clades have remained obscure. Studies of adult musculature are inconclusive owing to the highly derived morphology of modern amniote limbs, but correspondences become increasingly evident earlier in ontogeny. We followed the embryonic development of forelimb musculature in representatives of six major amniote clades and found, contrary to current consensus, that these early splitting patterns are highly conserved across Amniota. Muscle mass cleavage patterns and topology are highly conserved in reptiles including birds, irrespective of their skeletal modifications: the avian flight apparatus results from slight early topological modifications that are exaggerated during ontogeny. Therian mammals, while conservative in their cleavage patterns, depart drastically from the ancestral amniote musculoskeletal organization in terms of topology. These topological changes occur through extension, translocation, and displacement of muscle groups later in development. Overall, the simplicity underlying the apparent complexity of forelimb muscle development allows us to resolve conflicting hypotheses about homology and to trace the history of each individual forelimb muscle throughout the amniote radiations.

AMPUTATION OF CERVIX AND NEOCERVIX CREATION USING STURMDORF TECHNIQUE IN RECURRENT CERVICAL SARCOMA BOTRYOIDES: A CASE REPORT

Abstract: Sarcoma botryoides, or embryonal rhabdomyosarcoma, is soft tissue sarcoma arises from embryonic muscle cells and normally present with discharge, bleeding, or a visible mass at the vaginal introitus in infants and children. Aim of this study is to report a case of cervical sarcoma botryoides. Method used is a Case report. A 16-year-old girl, nulliparous, complained abnormal vaginal bleeding and a protruding vaginal mass. Patient underwent first surgery, mass recurrent five months later. The second surgery was performed by cervical amputation and neocervix creation using Sturmdorf sutures technique. Postoperative week 12 demonstrated a healthy neocervix in appearance, her menses have resumed, indicated patency. She has no evidence of disease 3 months after diagnosis. Cervical sarcoma botryoides is a rare malignancy. Clinicians must thoroughly examine any polyp or mass protrudes from vagina. Early diagnosis and treatment are essential for better prognosis. Sarcoma botryoides has a much improved prognosis with multimodality treatment. To determine the long-term survival and future potential fertility outcome, larger study and longer follow-up are needed. Keywords: sarcoma, botyroides, embryonal, rhabdomyosarcoma, neocervix

A selective luminescent probe to monitor cellular ATP: Potential application for in vivo imaging in zebrafish embryo

Adenosine 5′-triphosphate (ATP) is the major energy currency of living cells and its efficient real-time monitoring in vivo has increased the demand for the development of fluorescent chemosensors. For the prompt detection of ATP in submolar range, we have developed an easily accessible colorimetric and fluorescence probe 4-(Di-p-tolylphosphoryl)-7-hydroxychroman-2-one (DPC). The selective nature of DPC towards ATP has been confirmed through UV–vis, fluorescence titration, NMR titration as well as computational studies. Coupled with high mitochondrial membrane potential and ATP levels, the chemosensor DPC shows intense blue fluorescence signal at intracellular compartments in murine splenocytes and A549 human lung cancer cells. Moreover, using live zebrafish (Danio rerio)embryos with high energetic demands, we have successfully achieved fluorescence imaging of ATP at distinct sites in vivo(neural tube, myotome or embryonic muscle tissue, somite boundaries and tail regions) followed by estimation of endogenous ATP level from embryo extract. Collectively, our designed probe demonstrates excellent cellular permeability, sensitivity and specificity for intracellular ATP, and may provide valuable insights in the research of ATP-regulated biological processes in living cells or organisms.

Transcriptome analysis reveals long non-coding natural antisense transcripts involved in muscle development in fetal goat (Capra hircus).

Non-coding RNAs have been shown to play vital roles in muscle development. However, the biological roles of long non-coding natural antisense transcripts, antisense lncRNAs (ASlncRNAs), are largely unknown in embryonic muscle development. Here, we identified a total of 466 ASlncRNAs in the longissimus dorsi muscle. And 48 differentially expressed ASlncRNAs were identified based on pairwise comparisons (P<0.05), sixteen of which were validated by qPCR. Additionally, 466 ASlncRNAs were predicted to target 335 protein coding genes based on complementary base-pairing. Enrichment analysis suggests that ASlncRNAs may be involved in muscle development by negatively regulating the expression of target genes. Furthermore, 170 ASlncRNAs were identified as potential miRNA precursors, suggesting that these ASlncRNAs may be involved in the regulation of muscle development by producing miRNA precursors. Our results provide a catalog of goat muscle-related ASlncRNAs, and will contribute to a fuller understanding of the roles of ASlncRNAs in muscle development.

A Comparative Analysis of Metabolic Profiles of Embryonic Skeletal Muscle from Lantang and Landrace Pigs.

Simple SummaryThe pig is one of the most important domesticated meat animals. Some studies have revealed that pigs with low meat production show more intense myogenesis at the early stage of embryonic muscle development than pigs with high meat production. Here, by gas chromatography–mass spectrometry GC–MS based metabolomics, we concluded that the nucleotide metabolism and energy metabolism of the longissimus lumborum (LL) were increased in Lantang pigs compared with Landrace pigs, indicating rapid synthesis of nucleic acids and ATP to meet the material and energy requirements of rapid cell proliferation and differentiation in Lantang pigs.Elucidation of the complex regulation of porcine muscle development is key to increasing pork output and improving pork quality. However, the molecular mechanisms involved in early porcine embryonic muscle development in different pig breeds remain largely unknown. Here, GC–MS based metabolomics and metabolomic profiling was used to examine the longissimus lumborum (LL) of the Lantang (LT) and the Landrace (LR) pig at embryonic day 35 (E35). Metabolites showed clear separation between LT and LR, with 40 metabolites having higher abundances in LT and 14 metabolites having lower abundances in LT compared with LR. In addition, these metabolic changes were mainly associated with nucleotide metabolism and energy metabolism, such as purine metabolism, pyrimidine metabolism, the pentose phosphate pathway, and the TCA cycle. More interestingly, the contents of DNA, RNA, and ATP per unit mass of LL tissues were higher in LT, indicating rapid synthesis of nucleic acids and ATP, to meet both the material and energy requirements of rapid cell proliferation and differentiation. Furthermore, enzyme activity associated with the TCA cycle and pentose phosphate pathway, including α-ketoglutaric dehydrogenase (KGDH), malate dehydrogenase (MDH), pyruvate dehydrogenase (PDH), succinate dehydrogenase (SDH), and glucose-6-phosphate dehydrogenase (G6PDH), were higher in LT. Based on these results, we conclude that there are significant differences in nucleotide metabolism and energy metabolism of LL between LT and LR, and we speculate that the enhanced nucleic acid metabolism and energy metabolism in LT can meet the material and energy requirements of rapid cell proliferation and differentiation, making myogenesis more intense in LT compared to LR which might be the metabolic mechanism underlying the distinct skeletal muscle development in the two breeds.

Embryonic myosin mutations T187I and R672C result in mechanical and structural dysfunction in hiPSC derived skeletal myotubes

Distal arthrogryposis (DA) is a skeletal muscle disorder characterized by joint contractures at various parts of the body, predominantly in the distal extremities. DA is associated with syndromes like Freeman-Sheldon (FSS), caused by mutations in the sarcomeric protein-encoding gene MYH3 responsible for generating embryonic skeletal muscle myosin during development. We previously reported (Racca et al., 2015) that demembranated myofibrils from affected individuals expressing the R672C mutation in MYH3 exhibited lower specific force and prolonged relaxation.