Differentiation Of Myoblast Cells Research Articles

Alpha Mangostin promotes myogenic differentiation of C2C12 mouse myoblast cells

Mangosteen, a fruit mainly produced in Southeast Asia, has been used as food and as an antipyretic and for treating skin diseases. The xanthones contained in mangosteen have many physiological activities including melanin suppression and anticancer activities, but little is known about the physiological effects of the most abundant xanthone, α-mangostin (α-MG) on myoblasts. In this study, we applied α-MG to C2C12 cells that had been induced to differentiate using 2% HS, and analyzed the physiological action of α-MS and the underlying mechanism in the context of myogenic differentiation. α-MG increased the survival rate of C2C12 cells in a concentration-dependent manner. Analysis of the morphological changes in the cells showed that α-MG significantly enhanced the myogenic differentiation of C2C12 myoblasts, whereas the mitochondrial number was only slightly affected. Expression analysis of differentiation-related proteins in C2C12 cells revealed that α-MG promoted the expression of MyoD and Myogenin. Thus, the present study revealed that α-MG improves the survival and myogenic differentiation of C2C12 myoblasts.

3D myotube guidance on hierarchically organized anisotropic and conductive fibers for skeletal muscle tissue engineering.

Tissue engineering represents a promising approach for the functional restoration of large volumetric skeletal muscle loss. However, design and fabrication of 3D hierarchically organized scaffolds that closely mimic the natural micro-/nanostructures of skeletal muscle extracellular matrix (ECM) is still an ongoing challenge. Here, we constructed a hierarchically organized, anisotropic and conductive scaffold with microscale melt electrowriting (MEW) grooves parallel aligned on the top of unidirectionally oriented nanofibrous mesh to guide myoblast cell alignment, elongation and differentiation into myotubes. A 7-days study of H9c2 myoblast cells cultured on the scaffolds indicated that the combination of nanoscale and microscale anisotropic surface topography enhanced myogenesis. Specifically, the parallel patterned scaffold effectively enhanced both the elongation and maturation of myotubes, as indicated by the increased myotube length (>600μm), higher heavy myosin chain (MHC) surface coverage and maturation index on the parallel patterned scaffold compared to their pure (2.4-fold MHC surface coverage, 1.6-fold maturation index) and perpendicular patterned counterparts (1.7-fold MHC surface coverage, 1.5-fold maturation index). Furthermore, a gold nanolayer coating on the aligned nanofibrous mesh surface provided electroactive cues to enhance the formation of myotubes. With the increase of Au coating thickness, improved myoblasts alignment (74.6%, 80.5%, 85.6%, 94.9% for Au 0, 10, 40, 70nm respectively) and higher MHC-positive expression (57.5% for Au 0nm, 90.3%, 92.5%, 95.0% for Au10, 40, 70nm respectively) were observed. Finally, the formation and morphology of myotubes were also found dependent on the spacing between microgrooves (100, 200, 300μm), where myotubes formed on 200μm spacing scaffold showed the highest alignment (within 10° along nanofibers) and elongation (>850μm). The hierarchically organized, anisotropic and conductive fibers hold great potential for regeneration of skeletal muscle tissue.

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

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

Low Levels of Serum Tryptophan Underlie Skeletal Muscle Atrophy.

Sarcopenia is a poor prognosis factor in some cancer patients, but little is known about the mechanisms by which malignant tumors cause skeletal muscle atrophy. Tryptophan metabolism mediated by indoleamine 2,3-dioxygenase is one of the most important amino acid changes associated with cancer progression. Herein, we demonstrate the relationship between skeletal muscles and low levels of tryptophan. A positive correlation was observed between the volume of skeletal muscles and serum tryptophan levels in patients with diffuse large B-cell lymphoma. Low levels of tryptophan reduced C2C12 myoblast cell proliferation and differentiation. Fiber diameters in the tibialis anterior of C57BL/6 mice fed a tryptophan-deficient diet were smaller than those in mice fed a standard diet. Metabolomics analysis revealed that tryptophan-deficient diet downregulated glycolysis in the gastrocnemius and upregulated the concentrations of amino acids associated with the tricarboxylic acid cycle. The weights and muscle fiber diameters of mice fed the tryptophan-deficient diet recovered after switching to the standard diet. Our data showed a critical role for tryptophan in regulating skeletal muscle mass. Thus, the tryptophan metabolism pathway may be a promising target for preventing or treating skeletal muscle atrophies.

LncRNA 2310043L19Rik inhibits differentiation and promotes proliferation of myoblast by sponging miR-125a-5p.

Although many long non-coding RNAs (lncRNAs) have been identified in muscle, some of their physiological functions and regulatory mechanisms remain elusive. Here we report the functional identification and characterization of a novel lncRNA 2310043L19Rik (lnc-231), which is highly expressed in muscle. The expression level of lnc-231 in skeletal muscle of young mice is higher than that in aged mice. Functional analysis showed that overexpression of lnc-231 restrained differentiation and promoted proliferation of myoblast, while inhibition of lnc-231 revealed completely opposite effects in vitro. RNA molecules of lnc-231 acted mechanistically as competing endogenous RNAs (ceRNA) to target miR-125a-5p, whereas miR-125a-5p binds to the 3’-UTR of E2F3 mRNA to inhibit its function. Collectively, lncRNA 2310043L19Rik promotes proliferation and inhibits differentiation of myoblast cells by attenuating the function of miR-125a-5p.

Effect of aqueous extract of ajwa dates on C2C12 myoblast cell proliferation, migration and differentiation

Chemotherapeutic agents generate side effects or cytotoxicity to other cells that include muscle cells. Doxorubicin, one of the mostly used chemotherapeutic agents, has been known to cause a loss of skeletal muscle mass which known as cachexia. On the other hand, chemopreventive agents, which mostly derived from phytochemicals, have been known to show less cytotoxity to cancer cells than chemotherapeutic agents. However, their effects on the muscle cells remain largely unknown. In this study, we investigated the effect of ajwa dates extract in C2C12 myoblast cells. C2C12 cells are satellite cells or adult muscle stem cells which can differentiate to from multinucleated myotube. MTT assays had been performed to determine the effect of ajwa dates extract on cell viability. In addition, effects on cell migration were examined by wound healing method. Proliferation assay was done to investigate the effect of ajwa dates extract to cell growth. Furthermore, C2C12 differentiation into myotubes was also investigated. The results of the cell viability assay against C2C12 cells showed that low concentration of ajwa dates was not toxic for C2C12 cells, whereas higher concentration of ajwa dates (10 µg/µl) decreased cell viability. In addition, the effect of ajwa dates extract (1 µg/µl) on cell migration depended on cell seeding density. At a lower cell density, the extract might promote myoblasts migration, whereas at a higher cell density it inhibited the cell migration that may be related to the induction of myocytes differentiation. Overall, we found that ajwa dates extract exhibited low or no cytotoxicity on C2C12 myoblast cells.

Uniaxially crumpled graphene as a platform for guided myotube formation

Graphene, owing to its inherent chemical inertness, biocompatibility, and mechanical flexibility, has great potential in guiding cell behaviors such as adhesion and differentiation. However, due to the two-dimensional (2D) nature of graphene, the microfabrication of graphene into micro/nanoscale patterns has been widely adopted for guiding cellular assembly. In this study, we report crumpled graphene, i.e., monolithically defined graphene with a nanoscale wavy surface texture, as a tissue engineering platform that can efficiently promote aligned C2C12 mouse myoblast cell differentiation. We imparted out-of-plane, nanoscale crumpled morphologies to flat graphene via compressive strain-induced deformation. When C2C12 mouse myoblast cells were seeded on the uniaxially crumpled graphene, not only were the alignment and elongation promoted at a single-cell level but also the differentiation and maturation of myotubes were enhanced compared to that on flat graphene. These results demonstrate the utility of the crumpled graphene platform for tissue engineering and regenerative medicine for skeletal muscle tissues.

Dissolution, bioactivity and osteogenic properties of composites based on polymer and silicate or borosilicate bioactive glass.

Bioactive glass (BAG)/Poly (Lactic Acid) (PLA) composites have great potential for bone tissue engineering. The interest in these materials is to obtain a scaffold with tailorable properties bringing together the advantages of the composites' constituents such as the biodegradability, bioactivity and osteoinduction. The materials studied are PLA/13-93 and PLA/13-93B20 (20% of SiO2 is replaced with B2O3 in the 13-93 composition). To characterize them, they were dissolved in TRIS buffer and Simulated Body Fluid (SBF) in vitro. Over the 10 weeks of immersion in TRIS, the ion release from the composites was constant. Following immersion in SBF for 2 weeks, the hydroxyapatite (HA) layer was found to precipitate at the composites surface. By adding Boron, both these reactions were accelerated, as the borosilicate glass dissolves faster than pure silicate glass alone. Polymer degradation was studied and showed that during immersion, the pure PLA rods maintained their molecular weight whereby the composites decreased with time, but despite this the mechanical properties remained stable for at least 10 weeks. Their ability to induce osteogenic differentiation of myoblastic cells was also demonstrated with cell experiments showing that C2C12 cells were able to proliferate and spread on the composites. The Myosin Heavy Chain and Osteopontin were tracked by immunostaining the cells and showed a suppression of the myosin signal and the presence of osteopontin, when seeded onto the composites. This proves osteoinduction occurred. In studying the mineralization of the cells, it was found that BAG presence conditions the synthesizing of mineral matter in the cells. The results show that these composites have a potential for bone tissue engineering.

Circular RNA circ-FoxO3 Inhibits Myoblast Cells Differentiation.

CircRNA is a type of closed circular non-coding RNA formed by reverse splicing and plays an important role in regulating the growth and development of plants and animals. To investigate the function of circ-FoxO3 in mouse myoblast cells’ (C2C12) differentiation and proliferation, we used RT-qPCR to detect the expression level of circ-FoxO3 in mouse myoblast cells at different densities and different differentiation stages, and the specific interference fragment was used to inhibit the expression level of circ-FoxO3 in myoblast cells to observe its effect on myoblast cells proliferation and differentiation. We found that the expression level of circ-FoxO3 in myoblast cells increased with the prolongation of myoblast cells differentiation time, and its expression level decreased with the proliferation of myoblast cells. At the same time, we found that the differentiation ability of the cells was significantly increased (p < 0.05), but the cell proliferation was unchanged (p > 0.05) after inhibiting the expression of circ-FoxO3 in myoblast cells. Combining the results of bioinformatics analysis and the dual luciferase reporter experiment, we found that circ-FoxO3 is a sponge of miR-138-5p, which regulates muscle differentiation. Our study shows that circ-FoxO3 can inhibit the differentiation of C2C12 myoblast cells and lay a scientific foundation for further study of skeletal muscle development at circRNA levels.

Iron accumulation causes impaired myogenesis correlated with MAPK signaling pathway inhibition by oxidative stress.

Skeletal muscle atrophy is caused by disruption in the homeostatic balance of muscle degeneration and regeneration under various pathophysiological conditions. We have previously reported that iron accumulation induces skeletal muscle atrophy via a ubiquitin ligase-dependent pathway. However, the potential effect of iron accumulation on muscle regeneration remains unclear. To examine the effect of iron accumulation on myogenesis, we used a mouse model with cardiotoxin (CTX)-induced muscle regeneration in vivo and C2C12 mouse myoblast cells in vitro. In mice with iron overload, the skeletal muscles exhibited increased oxidative stress and decreased expression of satellite cell markers. Following CTX-induced muscle injury, these mice also displayed delayed muscle regeneration with a decrease in the size of regenerating myofibers, reduced expression of myoblast differentiation markers, and decreased phosphorylation of MAPK signaling pathways. In vitro, iron overload also suppressed the differentiation of C2C12 myoblast cells but the suppression could be reversed by superoxide scavenging using tempol. Excess iron inhibits myogenesis via oxidative stress, leading to an imbalance in skeletal muscle homeostasis.-Ikeda, Y., Satoh, A., Horinouchi, Y., Hamano, H., Watanabe, H., Imao, M., Imanishi, M., Zamami, Y., Takechi, K., Izawa-Ishizawa, Y., Miyamoto, L., Hirayama, T., Nagasawa, H., Ishizawa, K., Aihara, K.-I., Tsuchiya, K., Tamaki, T. Iron accumulation causes impaired myogenesis correlated with MAPK signaling pathway inhibition by oxidative stress.

Cytocompatible, Injectable, and Electroconductive Soft Adhesives with Hybrid Covalent/Noncovalent Dynamic Network.

Synthetic conductive biopolymers have gained increasing interest in tissue engineering, as they can provide a chemically defined electroconductive and biomimetic microenvironment for cells. In addition to low cytotoxicity and high biocompatibility, injectability and adhesiveness are important for many biomedical applications but have proven to be very challenging. Recent results show that fascinating material properties can be realized with a bioinspired hybrid network, especially through the synergy between irreversible covalent crosslinking and reversible noncovalent self‐assembly. Herein, a polysaccharide‐based conductive hydrogel crosslinked through noncovalent and reversible covalent reactions is reported. The hybrid material exhibits rheological properties associated with dynamic networks such as self‐healing and stress relaxation. Moreover, through fine‐tuning the network dynamics by varying covalent/noncovalent crosslinking content and incorporating electroconductive polymers, the resulting materials exhibit electroconductivity and reliable adhesive strength, at a similar range to that of clinically used fibrin glue. The conductive soft adhesives exhibit high cytocompatibility in 2D/3D cell cultures and can promote myogenic differentiation of myoblast cells. The heparin‐containing electroconductive adhesive shows high biocompatibility in immunocompetent mice, both for topical application and as injectable materials. The materials could have utilities in many biomedical applications, especially in the area of cardiovascular diseases and wound dressing.

Age-Dependent Oxidative Stress Elevates Arginase 1 and Uncoupled Nitric Oxide Synthesis in Skeletal Muscle of Aged Mice.

Aging is associated with reduced muscle mass (sarcopenia) and poor bone quality (osteoporosis), which together increase the incidence of falls and bone fractures. It is widely appreciated that aging triggers systemic oxidative stress, which can impair myoblast cell survival and differentiation. We previously reported that arginase plays an important role in oxidative stress-dependent bone loss. We hypothesized that arginase activity is dysregulated with aging in muscles and may be involved in muscle pathophysiology. To investigate this, we analyzed arginase activity and its expression in skeletal muscles of young and aged mice. We found that arginase activity and arginase 1 expression were significantly elevated in aged muscles. We also demonstrated that SOD2, GPx1, and NOX2 increased with age in skeletal muscle. Most importantly, we also demonstrated elevated levels of peroxynitrite formation and uncoupling of eNOS in aged muscles. Our in vitro studies using C2C12 myoblasts showed that the oxidative stress treatment increased arginase activity, decreased cell survival, and increased apoptotic markers. These effects were reversed by treatment with an arginase inhibitor, 2(S)-amino-6-boronohexanoic acid (ABH). Our study provides strong evidence that L-arginine metabolism is altered in aged muscle and that arginase inhibition could be used as a novel therapeutic target for age-related muscle complications.

Effect of Beta‐hydroxybutyrate on Myoblast Proliferation and Differentiation

Ketogenic diets (KDs) and fasting have been promoted for weight loss and health. KDs cause an increase in circulating levels of ketone bodies, including beta‐hydroxybutyrate (βHB). The effects of amplified levels of βHB on skeletal muscle proliferation and differentiation are largely unknown. Therefore, we tested whether βHB has an effect on proliferation and differentiation of C2C12 myoblast cells in vitro. Experiment 1: to assess proliferation, myoblasts were plated sparsely and incubated in growth media for 6 hours. They were then incubated overnight (~12 hours) with 0 – 10 mM bHB followed by the addition of BrDU to the media for an additional 2 hours to label proliferating cells. The cells were fixed and stained for BrDU and DAPI and proliferation index was calculated as the percentage of BrDU+ nuclei/total nuclei. We found that myoblast proliferation did not increase significantly with βHB. Experiment 2: to assess differentiation, myoblasts were grown to 60% confluence in growth media, followed by differentiation media for 4 days with 0–10 mM bHB. The myotubes were then fixed and stained with DAPI and antibodies against myosin heavy chain. Differentiation index was calculated as the percentage of nuclei associated with MHC+ myotubes. We found that myoblast differentiation increased 22% with 0.6125 mM βHB, and tended to increase (p = 0.12) at 1.25 mM bHB. This suggests that moderate levels of bHB (similar to those achieved in those on KDs or fasting) may be beneficial to muscle repair and/or regeneration.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Circular RNA circHIPK3 Promotes the Proliferation and Differentiation of Chicken Myoblast Cells by Sponging miR-30a-3p.

Circular RNAs and microRNAs widely exist in various species and play crucial roles in multiple biological processes. It is essential to study their roles in myogenesis. In our previous sequencing data, both miR-30a-3p and circular HIPK3 (circHIPK3) RNA, which are produced by the third exon of the HIPK3 gene, were differentially expressed among chicken skeletal muscles at 11 embryo age (E11), 16 embryo age (E16), and 1-day post-hatch (P1). Here, we investigated their potential roles in myogenesis. Proliferation experiment showed that miR-30a-3p could inhibit the proliferation of myoblast. Through dual-luciferase assay and Myosin heavy chain (MYHC) immunofluorescence, we found that miR-30a-3p could inhibit the differentiation of myoblast by binding to Myocyte Enhancer Factor 2 C (MEF2C), which could promote the differentiation of myoblast. Then, we found that circHIPK3 could act as a sponge of miR-30a-3p and exerted a counteractive effect of miR-30a-3p by promoting the proliferation and differentiation of myoblasts. Taking together, our data suggested that circHIPK3 could promote the chicken embryonic skeletal muscle development by sponging miR-30a-3p.

Enhanced growth and differentiation of myoblast cells grown on E-jet 3D printed platforms.

BackgroundSkeletal muscle tissue engineering often involves the prefabrication of muscle tissues in vitro by differentiation and maturation of muscle precursor cells on a platform which provides an environment that facilitates the myogenic differentiation of the seeded cells.MethodsPoly lactic-co-glycolic acid (PLGA) 3D printed scaffolds, which simulate the highly complex structure of extracellular matrix (ECM), were fabricated by E-jet 3D printing in this study. The scaffolds were used as platforms, providing environment that aids in growth, differentiation and other properties of C2C12 myoblast cells.ResultsThe C2C12 myoblast cells grown on the PLGA 3D printed platforms had enhanced cell adhesion and proliferation. Moreover, the platforms were able to induce myogenic differentiation of the myoblast cells by promoting the formation of myotubes and up-regulating the expressions of myogenic genes (MyHC and MyOG).ConclusionThe fabricated 3D printed platforms have excellent biocompatibility, thereby can potentially be used as functional cell culture platforms in skeletal tissue engineering and regeneration.

Heat induces myogenic transcription factors of myoblast cells via transient receptor potential vanilloid 1 (Trpv1)

Exercise generates heat, blood flow, and metabolic changes, thereby inducing hypertrophy of skeletal muscle cells. However, the mechanism by which heat incudes hypertrophy in response to heat is not well known. Here, we hypothesized that heat would induce differentiation of myoblast cells. We investigated the underlying mechanism by which myoblast cells respond to heat. When mouse myoblast cells were exposed to 42 °C for over 30 min, the phosphorylation level of protein kinase C (PKC) and heat shock factor 1 (Hsf1) increased, and the mRNA and protein expression level of heat shock protein 70 (Hsp70) increased. Inhibitors of transient receptor potential vanilloid 1 (Trpv1), calmodulin, PKC, and Hsf1, and the small interfering RNA‐mediated knockdown of Trpv1 diminished those heat responses. Heat exposure increased the phosphorylation levels of thymoma viral proto‐oncogene 1 (Akt), mammalian target of rapamycin (mTOR), eukaryotic translation initiation factor 4E binding protein 1 (Eif4ebp1), and ribosomal protein S6 kinase, polypeptide 1 (S6K1). The knockdown of Trpv1 decreased these heat‐induced responses. Antagonists of Hsp70 inhibited the phosphorylation level of Akt. Finally, heat increased the protein expression level of skeletal muscle markers such as myocyte enhancer factor 2D, myogenic factor 5, myogenic factor 6, and myogenic differentiation 1. Heat also increased myotube formation. Knockdown of Trpv1 diminished heat‐induced increases of those proteins and myotube formation. These results indicate that heat induces myogenic transcription factors of myoblast cells through the Trpv1, calmodulin, PKC, Hsf1, Hsp70, Akt, mTOR, Eif4ebp1, and S6K1 pathway. Moreover, heat increases myotube formation through Trpv1.

MiR-483 inhibits bovine myoblast cell proliferation and differentiation via IGF1/PI3K/AKT signal pathway.

MicroRNAs (miRNAs) have been established to regulate skeletal muscle development in mammals. However, few studies have been conducted on the regulation of proliferation and differentiation of bovine myoblast cells by miRNAs. The aim of our study was to explore the function of miR-483 in cell proliferation and differentiation of bovine myoblast. Here, we found that miR-483 declined in both proliferation and differentiation stages of bovine myoblast cells. During the proliferation phase, the overexpression of miR-483 downregulated the cell cycle-associated genes cyclin-dependent kinase 2 (CDK2), proliferating cell nuclear antigen (PCNA) messenger RNA (mRNA), and the protein levels. At the cellular level, cell cycle, cell counting kit-8, and 5-ethynyl-2´-deoxyuridine results indicated that the overexpression of miR-483 block cell proliferation. During differentiation, the overexpression of miR-483 led to a decrease in the levels of the myogenic marker genes MyoD1 and MyoG mRNA and protein. Furthermore, the immunofluorescence analysis results showed that the number of MyHC-positive myotubes was reduced. In contrast, the opposite experimental results were obtained concerning both proliferation and differentiation after the inhibition of miR-483. Mechanistically, we demonstrated that miR-483 targetinsulin-like growth factor 1 (IGF1) and downregulated the expression of key proteins in the PI3K/AKT signaling pathway. Altogether, our findings indicate that miR-483 acts as a negative regulator of bovine myoblast cell proliferation and differentiation.

Synthetic curcuminoid analogues abrogate oxidationinduced cell death and promote myogenic differentiation of C2C12 mouse myoblasts

Purpose: To investigate the ability of two synthetic curcuminoid analogues, 6-(4-hydroxy-3-methoxyphenethyl)-5-(3-(4-hydroxy-3-methoxyphenyl)propanoyl)-4-phenyl-3,4-dihydropyrimidin-2(1H)-one (compound A) and 6-(4-hydroxy-3-methoxyphenethyl)-4-(4-hydroxy-3-methoxyphenyl)-5-(3-(4- hydroxy-3-methoxyphenyl)propanoyl)-3,4-dihydropyrimidin-2(1H)-one (compound B), to protect against oxidation-induced cell death and the potential to enhance proliferation and differentiation of C2C12 myoblast cells.Methods: Antioxidant activity of curcuminoid analogues was evaluated by DPPH assay. The cytotoxic activity of the compounds (0 - 25 mM) on C2C12 myoblasts was determined by MTT assay while the effect on cell proliferation was assessed by BrdU uptake. Myoblast cell differentiation was measured by the formation of myotubes and myosin heavy chain (MHC) protein expression using immunofluorescence staining and Western blotting, respectively.Results: Both curcuminoid analogues exhibited strong anti-oxidant activity of up to 3-fold greater than that of ascorbic acid, and were non-toxic to C2C12 myoblasts at concentrations up to 25 mM. Furthermore, these curcuminoid analogues mitigated myoblast cell death induced by oxidative stress. Notably, both analogues (10 nM) had no effect on cell proliferation. However, only compound A significantly enhanced myoblast differentiation comparable to the effects of dihydrotestosterone (1 μM) and estradiol (10 nM).Conclusion: The results suggest that compound A may serve as a lead compound for the development of suitable therapeutic agents for muscle injuries and diseases.Keywords: Curcuminoid analogues, Antioxidant, Cell proliferation, Cell differentiation, Myoblasts

Effects of non-euphoric plant cannabinoids on muscle quality and performance of dystrophic mdx mice.

Duchenne muscular dystrophy (DMD), caused by dystrophin deficiency, results in chronic inflammation and irreversible skeletal muscle degeneration. Moreover, the associated impairment of autophagy greatly contributes to the aggravation of muscle damage. We explored the possibility of using non-euphoric compounds present in Cannabis sativa, cannabidiol (CBD), cannabidivarin (CBDV) and tetrahydrocannabidivarin (THCV), to reduce inflammation, restore functional autophagy and positively enhance muscle function in vivo. Using quantitative PCR, western blots and [Ca2+ ]i measurements, we explored the effects of CBD and CBDV on the differentiation of both murine and human skeletal muscle cells as well as their potential interaction with TRP channels. Male dystrophic mdx mice were injected i.p. with CBD or CBDV at different stages of the disease. After treatment, locomotor tests and biochemical analyses were used to evaluate their effects on inflammation and autophagy. CBD and CBDV promoted the differentiation of murine C2C12 myoblast cells into myotubes by increasing [Ca2+ ]i mostly via TRPV1 activation, an effect that undergoes rapid desensitization. In primary satellite cells and myoblasts isolated from healthy and/or DMD donors, not only CBD and CBDV but also THCV promoted myotube formation, in this case, mostly via TRPA1 activation. In mdx mice, CBD (60mg·kg-1 ) and CBDV (60mg·kg-1 ) prevented the loss of locomotor activity, reduced inflammation and restored autophagy. We provide new insights into plant cannabinoid interactions with TRP channels in skeletal muscle, highlighting a potential opportunity for novel co-adjuvant therapies to prevent muscle degeneration in DMD patients. This article is part of a themed section on 8th European Workshop on Cannabinoid Research. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.10/issuetoc.

The DEAD-box RNA helicase Ddx39ab is essential for myocyte and lens development in zebrafish.

RNA helicases from the DEAD-box family are found in almost all organisms and have important roles in RNA metabolism, including RNA synthesis, processing and degradation. The function and mechanism of action of most of these helicases in animal development and human disease remain largely unexplored. In a zebrafish mutagenesis screen to identify genes essential for heart development we identified a mutant that disrupts the gene encoding the RNA helicase DEAD-box39ab (ddx39ab). Homozygous ddx39ab mutant embryos exhibit profound cardiac and trunk muscle dystrophy, along with lens abnormalities, caused by abrupt terminal differentiation of cardiomyocyte, myoblast and lens fiber cells. Loss of ddx39ab hindered splicing of mRNAs encoding epigenetic regulatory factors, including members of the KMT2 gene family, leading to misregulation of structural gene expression in cardiomyocyte, myoblast and lens fiber cells. Taken together, these results show that Ddx39ab plays an essential role in establishment of the proper epigenetic status during differentiation of multiple cell lineages.