Myogenic Regulatory Factors Research Articles

Protective effects of chlorogenic acid on growth, intestinal inflammation, hepatic antioxidant capacity, muscle development and skin color in channel catfish Ictalurus punctatus fed an oxidized fish oil diet

Under oxidative stress condition, the protective effects of dietary chlorogenic acid (CGA) supplementation on liver antioxidant capacity, intestinal inflammation and barrier function, muscle development and skin coloration in channel catfish Ictalurus punctatus were explored in the current study. With that purpose, I. punctatus were fed five experimental diets containing 2% fresh fish oil (FFO, 9.2 meqO2/kg) or 2% oxidized fish oil (OFO, 897.4 meqO2/kg) without or with CGA supplementation (0.02%, 0.04% and 0.08%) for 8 weeks. Upon comparative analysis, the oxidized fish oil consumption significantly lowered weight gain rate, decreased intestinal villi length and muscular thickness values and the tight junction proteins mRNA abundance, augmented the intestinal proinflammatory factors, attenuated hepatic antioxidant enzymes activities and related genes mRNA expression levels, influenced the myogenic regulatory factors expression profile and impacted the myocyte density, myocyte area values as well as the skin pigments contents compared to the FFO treatment. Collectively, long-term feeding of the oxidized fish oil diet suppressed the growth performance, destroyed intestinal structural integrity, caused intestinal inflammation and hepatic oxidative stress, impacted the skeletal development and skin color of I. punctatus. Whereas CGA supplementation in oxidized fish oil diets partially counteracted the negative effects of the oxidized fish oil on I. punctatus in terms of increasing the growth performance, improving the intestinal mucosal structure, alleviating hepatic oxidative stress and intestinal inflammation, recompiling the myogenic regulatory factors expression and improving skin color. In conclusion, CGA has great potential to be an aquatic feed additive.

Monochromatic Green Light Stimulation during Incubation Alters Hepatic Glucose Metabolism That Improves Embryonic Development in Yangzhou Goose Eggs

The influence of monochromatic green light stimulation on hatching performance and embryo development has been studied in chickens, but not geese. The liver has crucial functions in the regulation of energy metabolism during embryogenesis, but its involvement in green light transduction is still unidentified. We aimed to determine the influence of monochromatic green light on Yangzhou goose hatching performance and embryo development. We also investigated the metabolomics and transcriptomic responses of the embryonic liver to green light to determine the underlying molecular mechanisms. Eggs were incubated under either 12 h of monochromatic green light/dark (12 L:12D) cycles or 24 h of darkness (0G:24D). Green light promoted embryonic development and hatching performance, also affected the expression of myogenic regulatory factors associated with muscle development. It also shortened hatching time and elevated plasma levels of growth hormone and insulin-like growth factor-1. Metabolomics and transcriptomic results revealed differentially expressed genes and metabolites with enhanced gluconeogenesis/glycolysis and increased plasma glucose and pyruvate levels under green light. Hence, the growth-promoting effect possibly through regulating energy metabolism in the liver and myogenic regulatory factors in muscle. Our findings provide important and novel insights into the mechanisms underlying the beneficial effects of green light on goose embryos.

Mealworm Ethanol Extract Enhances Myogenic Differentiation and Alleviates Dexamethasone-Induced Muscle Atrophy in C2C12 Cells.

Aging, and other disease-related muscle disorders are serious health problems. Dexamethasone (DEX), a synthetic glucocorticoid, can trigger skeletal muscle atrophy. This study examined the effects of mealworm (Tenebrio molitor larva) ethanol extract (TME) on C2C12 myoblast differentiation and DEX-induced myotube atrophy. TME induced myotube formation compared to the differentiation medium (DM) group. TME also significantly increased the mRNA expression of muscle creatine kinase (CKm) and myogenic regulatory factors (MRFs), such as myogenin (MyoG), myogenic factor (Myf)5, and MRF4 (Myf6). TME dramatically increased the muscle-specific protein, MyoG, compared to the control, whereas the expression of myogenic differentiation 1 (MyoD) remained unchanged. It also activated the mammalian target of rapamycin (mTOR) signaling pathway. In the DEX-induced muscle atrophy C2C12 model, TME reduced the gene expression of atrogin-1, muscle RING finger protein-1 (MuRF-1), and myostatin, which are involved in protein degradation in skeletal muscles. Furthermore, TME elevated the phosphorylation of forkhead box O3 (FoxO3α) and protein kinase B (Akt). These findings suggest that TME can enhance myotube hypertrophy by regulating the mTOR signaling pathway, and can rescue DEX-induced muscle atrophy by alleviating atrophic muscle markers mediated by Akt activation. Thus, TME can be a potential therapeutic agent for treating muscle weakness and atrophy.

Dietary Leucine Supplementation Improves Muscle Fiber Growth and Development by Activating AMPK/Sirt1 Pathway in Blunt Snout Bream (Megalobrama amblycephala).

This research is aimed at evaluating the effects of leucine supplementation on muscle fibers growth and development of blunt snout bream through a feeding trial and a primary muscle cells treatment. An 8-week trial with diets containing 1.61% leucine (LL) or 2.15% leucine (HL) was conducted in blunt snout bream (mean initial weight = 56.56 ± 0.83 g). Results demonstrated that the specific gain rate and the condition factor of fish in the HL group were the highest. The essential amino acids content of fish fed HL diets was significantly higher than that fed LL diets. The texture (hardness, springiness, resilience, and chewiness), the small-sized fiber ratio, fibers density, and sarcomere lengths in fish all obtained the highest in the HL group. Additionally, the proteins expression related with the activation of the AMPK pathway (p-Ampk, Ampk, p-Ampk/Ampk, and Sirt1) and the expression of genes (myogenin (myog), myogenic regulatory factor 4 (mrf4) and myoblast determination protein (myod), and protein (Pax7) related to muscle fiber formation were significantly upregulated with increasing level of dietary leucine. In vitro, the muscle cells were treated with 0, 40 and 160 mg/L leucine for 24 h. The results showed that treated with 40 mg/L leucine significantly raised the protein expressions of BCKDHA, Ampk, p-Ampk, p-Ampk/Ampk, Sirt1, and Pax7 and the gene expressions of myog, mrf4, and myogenic factor 5 (myf5) in muscle cells. In summary, leucine supplementation promoted muscle fibers growth and development, which may be related to the activation of BCKDH and AMPK.

An Origanum majorana Leaf Diet Influences Myogenin Gene Expression, Performance, and Carcass Characteristics in Lambs.

Myogenin is a protein in the myogenic regulatory factor family that plays an important role in determining carcass and meat traits and is vital for the growth and development of livestock muscles. The objective of this study was to determine the impact of Origanum majorana leaf (MOL) on the myogenin gene expression of lambs. Twenty-four male Kermani lambs were used in a completely randomized design using two experimental groups (0% Origanum majorana L. = MOL0 and 4% Origanum majorana L. = MOL4). Final weight, average daily gain, hot and cold carcass weight, feed conversion ratio, empty body weight, hot and cold dressing percentage, the weight of the shoulder, loin, leg, and lean meat, and the lean/bone ratio were measured. A standard kit was used for extracting total RNA from the loin, leg, and shoulder muscles of the lambs’ tissues. The cDNA was synthesized, a real-time PCR was performed, and the obtained data were analyzed. The results of this study showed that the effect of MOL4 on dry matter intake is not significant. The MOL4 diet increased final weight by 8.22%, average daily gain by 28.57%, hot carcass weight by 11.38%, cold carcass weight by 13.43%, feed conversion ratio by 31.03%, empty body weight by 9.38%, hot dressing percentage by 2.92%, cold dressing percentage by 3.75%, shoulder weight by 56.70%, loin weight by 8.98%, leg weight by 10.90%, lean meat weight by 14.62%, and the lean/bone ratio by 2.85% (p < 0.05) compared to the MOL0. Along with adding MOL4 in the lambs’ diets, in comparison with MOL0, there was higher expression of myogenin in the loin (3.5 times), leg (3.9 times), and shoulder (3.6 times) muscles of the lambs. Due to the fact that adding Origanum majorana to the diet of the lambs enhanced the expression of the myogenin gene in the loin, leg, and shoulder muscles and increased parameters related to growth, it can be used to improve the parameters related to growth and to increase myogenin gene expression in the muscle of lambs.

Substitution of soybean meal with Clostridium autoethanogenum protein in grass carp (Ctenopharygodon idella) diets: Effects on growth performance, feed utilization, muscle nutritional value and sensory characteristics

Iso-nitrogenous and iso-lipidic diets replace 0 %, 30 %, and 100 % of soybean meal (SBM) with Clostridium autoethanogenum protein (CAP) (namely, CAP0, CAP30 and CAP100) were prepared to evaluate their effects on the growth performance, feed utilization, muscle nutritional value and sensory characteristics of grass carp. The results indicated that the growth performance and feed utilization of grass carp fed a diet containing CAP groups were comparable to those of the control group (P > 0.05). However, CAP-containing diet enhanced pH, cohesiveness, springiness, gumminess and chewiness of muscle (P < 0.05), thereby improving the muscle physical properties. The CAP100 group did not affect the muscle content of crude protein and crude fat (P > 0.05), nevertheless it increased the content of essential and non-essential amino acids (P < 0.05), which improved the nutritional value of grass carp muscle. The CAP30 group promoted the synthesis of proteins in grass carp by regulating rapamycin signaling pathway which increased the crude protein content in muscle. Meanwhile, the CAP30-containing diet upregulated the expression of genes myogenic factor 5 (Myf5), myogenic regulatory factor 4 (Mrf4), myogenin (MyoG), myogenic determining factor (MyoD) and myosin heavy chain (MyHC) proteins which participate in muscle growth and development of the muscle. Compared with the control group, the tenderness and the content of inosinic acid in the CAP100 group muscle were increased (P < 0.05), and the content of volatile compounds which regulate fishy smell was decreased in muscles in the CAP100 group (P < 0.05). Collectively, our results indicated that CAP improved the nutritional value and sensory properties of the muscle without affecting the growth performance and feed utilization of grass carp.

Effect of Mechanical Loading of Senescent Myoblasts on Their Myogenic Lineage Progression and Survival

During aging, muscle cell apoptosis increases and myogenesis gradually declines. The impaired myogenic and survival potential of the aged skeletal muscle can be ameliorated by its mechanical loading. However, the molecular responses of aged muscle cells to mechanical loading remain unclear. This study examined the effect of mechanical loading of aged, proliferating, and differentiated myoblasts on the gene expression and signaling responses associated with their myogenic lineage progression and survival. Control and aged C2C12 cells were cultured on elastic membranes and underwent passive stretching for 12 h at a low frequency (0.25 Hz) and different elongations, varying the strain on days 0 and 10 of myoblast differentiation. Activation of ERK1/2 and Akt, and the expression of focal adhesion kinase (FAK) and key myogenic regulatory factors (MRFs), MyoD and Myogenin, were determined by immunoblotting of the cell lysates derived from stretched and non-stretched myoblasts. Changes in the expression levels of the MRFs, muscle growth, atrophy, and pro-apoptotic factors in response to mechanical loading of the aged and control cells were quantified by real-time qRT-PCR. Mechanical stretching applied on myoblasts resulted in the upregulation of FAK both in proliferating (day 0) and differentiated (day 10) cells, as well as in increased phosphorylation of ERK1/2 in both control and aged cells. Moreover, Akt activation and the expression of early differentiation factor MyoD increased significantly after stretching only in the control myoblasts, while the late differentiation factor Myogenin was upregulated in both the control and aged myoblasts. At the transcriptional level, mechanical loading of the proliferating myoblasts led to an increased expression of IGF-1 isoforms and MRFs, and to downregulation of muscle atrophy factors mainly in control cells, as well as in the upregulation of pro-apoptotic factors both in control and aged cells. In differentiated cells, mechanical loading resulted in an increased expression of the IGF-1Ea isoform and Myogenin, and in the downregulation of atrophy and pro-apoptotic factors in both the control and aged cells. This study revealed a diminished beneficial effect of mechanical loading on the myogenic and survival ability of the senescent muscle cells compared with the controls, with a low strain (2%) loading being most effective in upregulating myogenic/anabolic factors and downregulating atrophy and pro-apoptotic genes mainly in the aged myotubes.

Branched-chain amino acids promotes the repair of exercise-induced muscle damage via enhancing macrophage polarization.

The repair of exercise-induced muscle damage (EIMD) is closely related with inflammation. Branched-chain amino acids (BCAAs), as a nutritional supplement, promote EIMD repair; however, the underlying mechanism remains unclear. In vivo, Sprague-Dawley rats were subjected to Armstrong's eccentric exercise (a 120-min downhill run with a slope of -16° and a speed of 16mmin-1) to induce EIMD and BCAA supplement was administered by oral gavage. Protein expression of macrophages (CD68 and CD163) and myogenic regulatory factors (MYOD and MYOG) in gastrocnemius was analyzed. Inflammatory cytokines and creatine kinase (CK) levels in serum was also measured. In vitro, peritoneal macrophages from mice were incubated with lipopolysaccharide (LPS) or IL-4 with or without BCAAs in culture medium. For co-culture experiment, C2C12 cells were cultured with the conditioned medium from macrophages prestimulated with LPS or IL-4 in the presence or absence of BCAAs. The current study indicated BCAA supplementation enhanced the M1/M2 polarization of macrophages in skeletal muscle during EIMD repair, and BCAAs promoted M1 polarization through enhancing mTORC1-HIF1α-glycolysis pathway, and promoted M2 polarization independently of mTORC1. In addition, BCAA-promoted M1 macrophages further stimulated the proliferation of muscle satellite cells, whereas BCAA-promoted M2 macrophages stimulated their differentiation. Together, these results show macrophages mediate the BCAAs' beneficial impacts on EIMD repair via stimulating the proliferation and differentiation of muscle satellite cells, shedding light on the critical role of inflammation in EIMD repair and the potential nutritional strategies to ameliorate muscle damage.

Myomaker and Myomixer Characterization in Gilthead Sea Bream under Different Myogenesis Conditions

Skeletal muscle is formed by multinucleated myofibers originated by waves of hyperplasia and hypertrophy during myogenesis. Tissue damage triggers a regeneration process including new myogenesis and muscular remodeling. During myogenesis, the fusion of myoblasts is a key step that requires different genes' expression, including the fusogens myomaker and myomixer. The present work aimed to characterize these proteins in gilthead sea bream and their possible role in in vitro myogenesis, at different fish ages and during muscle regeneration after induced tissue injury. Myomaker is a transmembrane protein highly conserved among vertebrates, whereas Myomixer is a micropeptide that is moderately conserved. myomaker expression is restricted to skeletal muscle, while the expression of myomixer is more ubiquitous. In primary myocytes culture, myomaker and myomixer expression peaked at day 6 and day 8, respectively. During regeneration, the expression of both fusogens and all the myogenic regulatory factors showed a peak after 16 days post-injury. Moreover, myomaker and myomixer were present at different ages, but in fingerlings there were significantly higher transcript levels than in juveniles or adult fish. Overall, Myomaker and Myomixer are valuable markers of muscle growth that together with other regulatory molecules can provide a deeper understanding of myogenesis regulation in fish.

Dietary DHA Enhanced the Textural Firmness of Common Carp (Cyprinus carpio L.) Fed Plant-Derived Diets through Restraining FoxO1 Pathways.

Omega-3 fatty acids have a positive effect on the muscle textural firmness of fish, while the intrinsic mechanism is poorly understood. To investigate the potential mechanism of textural modification caused by dietary docosahexaenoic acid ( DHA) in common carp (Cyprinus carpio L.), three plant-derived diets with varying DHA levels (0%, 0.5%, 1%, D1-D3) were prepared to feed juveniles (initial weight 15.27 ± 0.77 g) for 8 weeks, and the muscular texture, fibers density, and transcriptome were analyzed. The results showed that the growth performance, muscular DHA content, fibers density, and texture of the fish fed diets D2 and D3 were significantly ameliorated compared with the fish fed diet D1. The muscular transcriptome profiles indicated that the up-regulated genes of fish fed dietary DHA mainly in response to muscle proliferation, as well as the FoxO pathway, were significantly enriched in the D2 and D3 groups. Consistent with this, the Quantitative Real-Time PCR (qRT-PCR ) assays indicated that the expression of myogenic regulatory factors (myog, myod, mrf4, mrf5) was up-regulated in the high-DHA groups. Additionally, the expression of foxo1 (inhibitor of myofiber development) mRNA was down-regulated, while its negative regulatory pathway (MAPK and PI3K) was activated in the D2 and D3 groups. The results suggested that the DHA supplementation is beneficial to modifying the muscular textural firmness of common carp fed plant-derived diets, which could be attributed to the inhibition of FoxO1 pathways.

Abstract 14259: Novel Long Noncoding RNA, Ppp1r1b-LncRNA , Epigenetically Promotes Myogenic Differentiation Program in Cardiac and Skeletal Myocytes

Introduction: Emerged as critical regulators of the transcriptome, the lncRNAs play important roles in cardiac development and might be targeted to treat human cardiomyocyte dysfunction in congenital heart disease. In our work, we identified a novel lncRNA, Ppp1r1b-lncRNA , as an essential, functionally conserved, positive regulator of myogenic differentiation. Hypothesis: Ppp1r1b-lncRNA promotes myogenic differentiation program of cardiac and skeletal myocytes via epigenic mechanisms. Methods: Antisense oligonucleotides (GapmeR) were used to suppress Ppp1r1b-lncRNA . Chromatin immunoprecipitation (CHIP) and chromatin isolation by RNA purification (CHIRP) were used to analyze gene specific histone modification level and lncRNA: chromatin interaction, respectively. RNA pull-down and RNA immunoprecipitation (RIP) were used to identify Ppp1r1b-lncRNA : protein interactome. Results: By silencing Ppp1r1b-lncRNA, skeletal myoblasts and human-induced pluripotent stem cell derived cardiomyocytes (hiPSCs-CMs) failed to develop fully differentiated myotubes. Analysis of GapmeR injected neonatal mouse heart further confirmed the role of Ppp1r1b-lncRNA in normal myogenesis. Members of muscle specific transcription factors, including MyoD1 and TBX5, were not induced during myogenic differentiation in the Ppp1r1b-lncRNA depleted cardiac and skeletal muscle precursor cells. Histone modification analysis revealed enrichment of H3K27 tri-methylation at MyoD1 and TBX5 promoters in GapmeR treated skeletal myoblasts. Furthermore, Ppp1r1b-lncRNA was found to bind to Ezh2, and ChIRP analysis revealed enriched interaction of Ppp1r1b-lncRNA with Myod1 and Tbx5 promoters, suggesting that Ppp1r1b-lncRNA induces transcription of myogenic regulatory factors by interacting with the polycomb repressive complex 2 (PRC2) at the chromatin interface. Correspondingly, the silencing of Ppp1r1b-lncRNA increased EZH2 binding at promoter regions of myogenic transcription factors. Conclusions: Ppp1r1b-lncRNA promotes myogenic differentiation by inhibiting histone methylation on the promotors of myogenic master regulators via competing for PRC2 binding with chromatin during heart and skeletal muscle development.

Slow twitch paraspinal muscle dysregulation in adolescent idiopathic scoliosis exhibiting HIF-2α misexpression.

Adolescent idiopathic scoliosis (AIS) refers to a three-dimensional spinal deformity which has a typical onset during adolescence. In most cases, the cause of the deformity cannot be clearly identified. Unbalanced paraspinal muscle activity in AIS patients was reported and hypoxia was implicated to regulate myogenesis. This study aims to investigate the association between myogenesis/muscle toning and HIF-αs activity in the pathogenesis of AIS. HIF-αs expression was examined by enzyme-linked immunosorbent assay and western blot in paraspinal myoblasts isolated from 18 subjects who underwent deformity correction surgery. QPCR was conducted to measure the gene expression levels of perinatal muscle fiber markers MYH3, MYH8; slow twitch muscle fiber markers MHY7; fast twitch muscle fiber markers MYH4; and myogenic regulatory factors MYF5 and MYOG. Slow and fast twitch muscle fiber composition in concave/convex paraspinal musculature of AIS subjects was evaluated by immunostaining of myosin heavy chain type I (MyHC I) and myosin heavy chain type II (MyHC II). Reduced HIF-2α induction under hypoxia was found in paraspinal myoblast culture of 33% AIS subjects. We detected a suppression of perinatal and slow twitch muscle fiber associated genes, but not fast twitch muscle fiber-associated genes and myogenic regulatory factors in HIF-2α misexpressed AIS myoblasts. Distinct reduction of slow twitch muscle fiber was evidenced in convex paraspinal musculature, suggesting an asymmetric expression of slow twitch muscle fiber in HIF-2α misexpressed AIS patients. This study indicates an association of abnormal HIF-2α expression in paraspinal myoblasts and a disproportionate slow twitch muscle fiber content in the convexity of the curvature in a subset of AIS subjects, suggesting HIF-2α dysregulation as a possible risk factor for AIS. The role of HIF-2α in paraspinal muscle function during spinal growth and its relevance in AIS prognosis warrants further investigation.

Interleukin-11 (IL11) inhibits myogenic differentiation of C2C12 cells through activation of extracellular signal-regulated kinase (ERK)

Cancer-associated cachexia (CAC) is a multifactorial wasting syndrome characterized by loss of skeletal muscle. Interleukin-11 (IL11), one of the IL6 family cytokines, is highly expressed in various types of cancer including cancers frequently associated with cachexia. However, the impact of IL11 on muscle metabolism remains to be determined. Since one of the mechanisms of muscle wasting in cachexia is defective muscle regeneration due to impaired myogenic differentiation, we examined the effect of IL11 on the differentiation of C2C12 mouse myoblasts. Treatment of C2C12 cells with recombinant mouse IL11 resulted in decreased myotube formation. In addition, IL11 treatment reduced the protein and mRNA levels of myosin heavy chain (MHC), a marker of myogenic differentiation. Moreover, the levels of myogenic regulatory factors including myogenin and Mrf4 were significantly reduced by IL11 treatment. IL11 treatment increased the number of BrdU-positive cells and the level of phosphorylated retinoblastoma (Rb) protein, while the levels of p21Waf1 and p27Kip1 were reduced by IL11 treatment in differentiating C2C12 cells, suggesting that IL11 interferes with cell cycle exit during the early stages of myogenic differentiation. Consistent with this, IL11 treatment at the late stage of differentiation did not affect myotube formation and MHC expression. IL11 treatment resulted in an activation of ERK, STAT3, and AKT in differentiating C2C12 cells. However, only ERK inhibitors including PD98059 and U0126 were able to ameliorate the suppressive effect of IL11 on the expression of MHC and myogenin. Additionally, pretreatment with PD98059 and U0126 resulted in improved myotube formation and reduced BrdU staining in IL11-treated cells. Together, our results suggest that IL11 inhibits myogenic differentiation through delayed cell cycle exit in an ERK-dependent manner. To our knowledge, this study is the first to demonstrate an inhibitory role of IL11 in myogenic differentiation and identifies the previously unrecognized role of IL11 as a possible mediator of CAC.

NRF2 Regulates Viability, Proliferation, Resistance to Oxidative Stress, and Differentiation of Murine Myoblasts and Muscle Satellite Cells.

Increased oxidative stress can slow down the regeneration of skeletal muscle and affect the activity of muscle satellite cells (mSCs). Therefore, we evaluated the role of the NRF2 transcription factor (encoded by the Nfe2l2 gene), the main regulator of the antioxidant response, in muscle cell biology. We used (i) an immortalized murine myoblast cell line (C2C12) with stable overexpression of NRF2 and (ii) primary mSCs isolated from wild-type and Nfe2l2 (transcriptionally)-deficient mice (Nfe2l2tKO). NRF2 promoted myoblast proliferation and viability under oxidative stress conditions and decreased the production of reactive oxygen species. Furthermore, NRF2 overexpression inhibited C2C12 cell differentiation by down-regulating the expression of myogenic regulatory factors (MRFs) and muscle-specific microRNAs. We also showed that NRF2 is indispensable for the viability of mSCs since the lack of its transcriptional activity caused high mortality of cells cultured in vitro under normoxic conditions. Concomitantly, Nfe2l2tKO mSCs grown and differentiated under hypoxic conditions were viable and much more differentiated compared to cells isolated from wild-type mice. Taken together, NRF2 significantly influences the properties of myoblasts and muscle satellite cells. This effect might be modulated by the muscle microenvironment.

"A Porcine Model to Study the Differential Expression of Myogenic Regulatory Factors (MRFs) in Piglets vis-a-vis Adult Pigs of Indigenous and Large White Yorkshire Breeds"

"A Porcine Model to Study the Differential Expression of Myogenic Regulatory Factors (MRFs) in Piglets vis-a-vis Adult Pigs of Indigenous and Large White Yorkshire Breeds"

MyoG-enhanced circGPD2 regulates chicken skeletal muscle development by targeting miR-203a

Circular RNAs (circRNAs) are a subclass of RNA macromolecules that are reported to be involved in the regulation of skeletal muscle development. However, the functions and regulatory mechanisms of circRNAs in chicken myogenesis are still largely unclear. Here, we identified a novel circRNA, circGPD2, an RNA macromolecule with a calculated molecular weight of 215 kDa. We discovered that circGPD2 is a muscle-specific circRNA and is strongly expressed in the breast muscle of broilers by utilizing the comparison model of layers and broilers. Functional analysis revealed circGPD2 has a positive role in the proliferation and differentiation of myoblasts, and circGPD2 performs function through the release of the inhibition effect of miR-203a on c-JUN and MEF2C. Besides, the myogenic regulatory factor MyoG enhanced the expression of circGPD2 by targeting the E-box element on the GPD2 promoter. Importantly, lentivirus-mediated circGPD2 knockdown resulted in the breast muscle mass loss of the chicks. Overall, we revealed the crucial role of circGPD2 in chicken myogenesis in vitro and in vivo, and analyzed the upstream and downstream regulation mechanisms of circGPD2. Our study provides an attractive target for molecular marker-assisted breeding to improve the meat yield in the chicken meat industry.

EFFECT ON MYOBLAST DIFFERENTIATION BY EXTREMELY LOW-FREQUENCY PULSED ELECTROMAGNETIC FIELDS

In the myoblasts differentiation process of C2C12 cells, it is imperative to research the expression and synthesis of the myogenic regulatory factors in extremely low-frequency pulsed electromagnetic fields (ELF-PEMFs) exposure. In the present study, using a cell cultivation method of C2C12 cells, the subcultured cells were divided into control and ELF-PEMF treated: A (0[Formula: see text]mT), B (0.5[Formula: see text]mT), C (1.0[Formula: see text]mT) and D (1.5[Formula: see text]mT) groups for five consecutive days training. Furthermore, cells were analyzed using immunochemical staining to detect the cell phenotypic changes, and RT-PCR and Western blot were used to test the expression levels of the myogenic regulatory factor D (MyoD) and the myogenic regulatory factor 5 (Myf5) genes and proteins synthesis in the process of myblasts differentiation. The results indicated that compared with the control group, different intensities of extremely low-frequency pulse electromagnetic fields could induce myoblast nuclear fusion, amplify MyoD and Mfy5 expression, and particularly the 1.5[Formula: see text]mT group at the 5th day exhibited the highest expression of these genes. Our study can pave a way for the development of a non-invasive and effective clinical prevention and treatment strategy in serious disorders like skeletal muscle atrophy.

PREFACE: A SPECIAL SELECTION ON EMERGING TECHNIQUES FOR BIOMECHANICS–PART II

PREFACE: A SPECIAL SELECTION ON EMERGING TECHNIQUES FOR BIOMECHANICS–PART II

The MyoD1 Promoted Muscle Differentiation and Generation by Activating CCND2 in Guanling Cattle.

Simple SummaryMuscle proliferation and differentiation is a complex process, which is mainly regulated by the myogenic regulatory factors (MRFs) gene family. As one of its family members, myogenic differentiation 1 (MyoD1) is an important regulator of myoblast differentiation and fibrogenesis, and its main function is to regulate the proliferation of myoblasts and the directed development of myogenic cell lines. In this study, CRISPR/CAS9 technology was used to construct the MyoD1 gene knockout MDBK cell lines, and then the transcriptome of MDBK cells was analyzed based on high-throughput sequencing technology. The differentially expressed genes in the transcriptome and the functional enrichment of differentially expressed genes were analyzed. The differentially expressed genes in the transcriptome and the functional enrichment were analyzed to provide experimental data for further studies on the mechanism of MyoD1 regulating CCND2 in myocyte differentiation.The purpose of this study was to analyze the transcriptome of MyoD1 gene knockout MDBK cells (bovine kidney cells) using high-throughput sequencing. For the first time, CRISPR/CAS9 technology was used to construct a MyoD1 knockout in MDBK cells and transcriptome sequence analysis was used to examine MyoD1-related target gene expression. Transcriptome sequencing indicated the presence of 723 differentially expressed genes (DEGs) by comparing wild type and MyoD1 knockout MDBK cells and included 178 upregulated and 72 downregulated genes. The DEGs are mainly enriched in Pl-3-kinase and AKT, p53 signaling pathways. Quantitative RT-PCR confirmed that PDE1B, ADAMTS1, DPT, and CCND2 were highly expressed in the leg muscle, longissimus dorsi, and shoulder of Guanling cattle, and CCND2 was inhibited after MyoD1 knockout, suggesting it may be a key downstream gene of MyoD1 and associated with muscle formation and differentiation in Guanling cattle. This provides experimental data for subsequent studies on the regulatory mechanisms of muscle differentiation in Guanling cattle.

Interrelated but Not Time-Aligned Response in Myogenic Regulatory Factors Demethylation and mRNA Expression after Divergent Exercise Bouts.

DNA methylation regulates exercise-induced changes in the skeletal muscle transcriptome. However, the specificity and the time course responses in the myogenic regulatory factors DNA methylation and mRNA expression after divergent exercise modes are unknown. This study aimed to compare the time course changes in DNA methylation and mRNA expression for selected myogenic regulatory factors ( MYOD1 , MYF5 , and MYF6 ) immediately after, 4 h after, and 8 h after a single bout of resistance exercise (RE), high-intensity interval exercise (HIIE), and concurrent exercise (CE). Nine healthy but untrained males (age, 23.9 ± 2.8 yr; body mass, 70.1 ± 14.9 kg; peak oxygen uptake [V̇O 2peak ], 41.4 ± 5.2 mL·kg -1 ·min -1 ; mean ± SD) performed a counterbalanced, randomized order of RE (4 × 8-12 repetition maximum), HIIE (12 × 1 min sprints at V̇O 2peak running velocity), and CE (RE followed by HIIE). Skeletal muscle biopsies (vastus lateralis) were taken before (REST) immediately (0 h), 4 h, and 8 h after each exercise bout. Compared with REST, MYOD1 , MYF5 , and MYF6 , mean methylation across all CpGs analyzed was reduced after 4 and 8 h in response to all exercise protocols ( P < 0.05). Reduced levels of MYOD1 methylation were observed after HIIE and CE compared with RE ( P < 0.05). Compared with REST, all exercise bouts increased mRNA expression over time ( MYOD1 at 4 and 8 h, and MYF6 at 4 h; P < 0.05). MYF5 mRNA expression was lower after 4 h compared with 0 h and higher at 8 h compared with 4 h ( P < 0.05). We observed an interrelated but not time-aligned response between the exercise-induced changes in myogenic regulatory factors demethylation and mRNA expression after divergent exercise modes. Despite divergent contractile stimuli, changes in DNA methylation and mRNA expression in skeletal muscle were largely confined to the late (4-8 h) recovery period and similar between the different exercise challenges.