Vitamin C regulates skeletal muscle post-injury regeneration by promoting myoblast proliferation through its direct interaction with the Pax7 protein.

Previous studies have shown that vitamin C (VC), an essential vitamin for the human body, can promote the differentiation of muscle satellite cells (MuSCs) in vitro and play an important role in skeletal muscle post-injury regeneration. However, the molecular mechanism of VC regulating MuSC proliferation has not been elucidated. In this study, the role of VC in promoting MuSC proliferation and its molecular mechanism were explored using cell molecular biology and animal experiments. The results showed that VC accelerates the progress of skeletal muscle post-injury regeneration by promoting MuSC proliferation in vivo. VC can also promote skeletal muscle regeneration in the case of atrophy. Using the C2C12 myoblast murine cell line, we observed that VC also stimulated cell proliferation. In addition, after an in vitro study establishing the occurrence of a physical interaction between VC and Pax7, we observed that VC also upregulated the total and nuclear Pax7 protein levels. This mechanism increased the expression of Myf5 (Myogenic Factor 5), a Pax7 target gene. This study establishes a theoretical foundation for understanding the regulatory mechanisms underlying VC-mediated MuSC proliferation and skeletal muscle regeneration. Moreover, it develops the application of VC in animal muscle nutritional supplements and treatment of skeletal muscle-related diseases.

Myogenic Differentiation of iPS Cells Shows Different Efficiency in Simultaneous Comparison of Protocols.

Induced pluripotent stem (iPS) cells constitute a perfect tool to study human embryo development processes such as myogenesis, thanks to their ability to differentiate into three germ layers. Currently, many protocols to obtain myogenic cells have been described in the literature. They differ in many aspects, such as media components, including signaling modulators, feeder layer constituents, and duration of culture. In our study, we compared three different myogenic differentiation protocols to verify, side by side, their efficiency. Protocol I was based on embryonic bodies differentiation induction, ITS addition, and selection with adhesion to collagen I type. Protocol II was based on strong myogenic induction at the embryonic bodies step with BIO, forskolin, and bFGF, whereas cells in Protocol III were cultured in monolayers in three special media, leading to WNT activation and TGF-ß and BMP signaling inhibition. Myogenic induction was confirmed by the hierarchical expression of myogenic regulatory factors MYF5, MYOD, MYF6 and MYOG, as well as the expression of myotubes markers MYH3 and MYH2, in each protocol. Our results revealed that Protocol III is the most efficient in obtaining myogenic cells. Furthermore, our results indicated that CD56 is not a specific marker for the evaluation of myogenic differentiation.

Effect of constant compressive stress induced by imitating Tuina stimulation with various durations on the cell cycle, cellular secretion, apoptosis, and expression of myogenic differentiation and myogenic factor 5 of rat skeletal muscle cells in vitro.

OBJECTIVE: To investigate the effect of constant compressive stress induced by imitating Tuina stimulation with various durations on the cell cycle, cellular secretion, apoptosis, and expression of myogenic regulatory factors (MRFs), myogenic factor 5(Myf5) and myogenic differentiation (MyoD) of rat skeletal muscle cells (RSkMCs) in vitro. METHODS: Third passage RSkMCs were subjected to constant compressive stresses with various durations at 2000 strain for 15, 30, 60, 90, and 120 min via a four-point bending system. The control group (CG) was cultured in the absence of mechanical loading. Alterations of the cell cycle and apoptosis rate were detected by flow cytometry (FCM). The concentrations of interleukin 6 (IL-6) / prostaglandin E2 (PGE2) and nitric oxide (NO) in supernatants were determined by enzyme-linked immunosorbent assays and the nitrate reductase method, respectively. Expression of Myf5 and MyoD was detected by immunohistochemistry. RESULTS: Compared with the CG, a significant alteration was observed in the synthesis phase fraction (SPF) (P < 0.01). The SPF and proliferation index (PI) were reduced from 15 to 90 min, but reached levels similar to those at 120 min. Apoptosis was increased significantly at 30 min (P < 0.05) and especially at 90 and 120 min (P < 0.01). Expression of MyoD and Myf5 was increased significantly at 15, 30, and 90 min (P < 0.01). Compared with 15 and 30 min, MyoD and Myf5 expression at 60 and 120 min was decreased significantly (P < 0.01). Compared with 60 min, MyoD expression at 90 min was increased significantly (P < 0.05), whereas MyoD and Myf5 expression at 120 min was significantly lower (P < 0.05). The IL-6 concentration was increased at 60 min compared with the CG and 15 min (P < 0.05), whereas the concentrations of PGE2 and NO were the highest at 15 and 30 min, respectively, compared with the CG and other time points (P < 0.05). CONCLUSION: The cell cycle, secretion, apoptosis, and Myf5 and MyoD expression of RSkMCs were regulated by compressive stress in a time-dependent manner. SPF and PI were inhibited at short durations (< 90 min), but NO and PGE2 secretion was the highest at shorter durations (< 30 min). With the prolongation of stimulation time, SPF, PI, and apoptosis were increased, but Myf5 and MyoD expression was decreased gradually at 15-30 min.

Intramuscular Injection of Combined Calf Blood Compound (CFC) and Homeopathic Drug Tr14 Accelerates Muscle Regeneration In Vivo.

Skeletal muscle injuries in competitive sports cause lengthy absences of athletes from tournaments. This is of tremendous competitive and economic relevance for both the athletes and their respective clubs. Therapy for structural muscle lesions aims to promote regeneration and fast-track return-to-play. A common clinical treatment strategy for muscle injuries is the intramuscular injection of calf blood compound and the homeopathic drug, Tr14. Although the combination of these two agents was reported to reduce recovery time, the regulatory mechanism whereby this occurs remains unknown. In this in vivo study, we selected a rat model of mechanical muscle injury to investigate the effect of this combination therapy on muscle regeneration. Gene expression analysis and histological images revealed that this combined intramuscular injection for muscle lesions can enhance the expression of pro-myogenic genes and proteins and accelerate muscle regeneration. These findings are novel and depict the positive effects of calf blood compound and the homeopathic drug, Tr14, which are utilized in the field of Sports medicine.

Calf Blood Compound (CFC) and Homeopathic Drug Induce Differentiation of Primary Human Skeletal Muscle Cells.

The use of injections to treat structural muscle injuries is controversially discussed. In our controlled in vitro study, we investigated the biological impact of Actovegin and Traumeel alone and in combination on primary human skeletal muscle cells. Cells were characterized by immunofluorescence staining for myogenic factor 5 (Myf5) and MyoD, and cultured with or without Actovegin and / or Traumeel. The effects of these agents were assayed by cell viability and gene expression of the specific markers MyoD, Myf5, neural adhesion molecule (NCAM), and CD31. Myotube formation was determined by myosin staining. Neither Actovegin nor Traumeel showed toxic effects or influenced cell viability significantly. High volumes of Actovegin down-regulated gene expression of NCAM after 3 days but had no effect on MyoD, Myf5, and CD31 gene expression. High volumes of Traumeel inhibited MyoD gene expression after 3 days, whereas after 7 days MyoD expression was significantly up-regulated. The combination of both agents did not significantly influence cell viability or gene expression. This is the first study demonstrating that Actovegin and Traumeel potentially modulate human skeletal muscle cells. The relevance of these in vitro findings has to be highlighted in further in vivo studies.

Mild hyperbaric oxygen inhibits the growth-related decline in skeletal muscle oxidative capacity and prevents hyperglycemia in rats with type 2 diabetes mellitus.

BACKGROUND: Humans and animals with type 2 diabetes mellitus (T2DM) exhibit low skeletal muscle oxidative capacity and impaired glucose metabolism. The aim of the present study was to investigate the effects of exposure to mild hyperbaric oxygen on these changes in obese rats with T2DM. METHODS: Five-week-old non-diabetic Long-Evans Tokushima Otsuka (LETO) and diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats were divided into normobaric (LETO-NB and OLETF-NB) and mild hyperbaric oxygen (LETO-MHO and OLETF-MHO) groups. The LETO-MHO and OLETF-MHO groups received 1266 hPa with 36% oxygen for 3 h daily for 22 weeks. RESULTS: Fasting and non-fasting blood glucose, HbA1c, and triglyceride levels were lower in the OLETF-MHO group than in the OLETF-NB group (P < 0.05). In the soleus muscle, peroxisome proliferator-activated receptor δ/ß (Pparδ/ß), Pparγ, and PPARγ coactivator-1α (Pgc-1α) mRNA levels were lower in the OLETF-NB group than in all other groups (P < 0.05), whereas myogenin (Myog) and myogenic factor 5 (Myf5) mRNA levels were higher in the OLETF-MHO group than in the LETO-NB and OLETF-NB groups (P < 0.05). The soleus muscles in the OLETF-NB group contained only low-oxidative Type I fibers, whereas those in all other groups contained high-oxidative Type IIA and Type IIC fibers in addition to Type I fibers. CONCLUSIONS: Exposure to mild hyperbaric oxygen inhibits the decline in skeletal muscle oxidative capacity and prevents the hyperglycemia associated with T2DM. Pgc-1α, Myog, and Myf5 mRNA levels appear to be closely associated with skeletal muscle oxidative capacity in rats with T2DM.

Possible correlation between selenoprotein W and myogenic regulatory factors in chicken embryonic myoblasts.

The biological function of selenium (Se) is mainly elicited through Se-containing proteins. Selenoprotein W (SelW), one member of the selenoprotein family, is essential for the normal function of the skeletal muscle system. To investigate the possible relationship of Se in the process of differentiation in chicken myoblasts and the expression of SelW, the cultured chicken embryonic myoblasts were incubated with sodium selenite at different concentrations for 72 h, and then the mRNA levels of SelW and myogenic regulatory factors (MRFs) in myoblasts were determined at 12, 24, 48, and 72 h, respectively. Furthermore, the correlation between SelW mRNA expression and MRF mRNA expression was assessed. The results showed that the sodium selenite medium enhanced the mRNA expression of SelW, Myf-5, MRF4, and myogenin in chicken myoblasts. The mRNA expression levels of MRFs were significantly correlated with those of SelW at 24, 48, and 72 h. These data demonstrate that Se is involved in the differentiation of chicken embryonic myoblasts, and SelW showed correlation with MRFs.

[Effects of extracts from Patrinia heterophylla on gene expression patterns during morphogenesis of chicken limb buds in vivo].

OBJECTIVE: To study the effects of the extracts from Patrinia heterophylla on gene expression patterns during morphogenesis of chicken limb buds in vivo. METHODS: Implanted a bead into an chicken embryo, which was soaked in the extracts from Patrinia heterophylla. Detected the extracts-induced morphogenesis changes (Myf5, Myod and PCNA). RESULTS: The extracts from Patrinia heterophylla (200 mg/mL) could affect limb bud development, reduce gene expression of MyfS, MyoD and PCNA. CONCLUSION: The extracts from Patrinia heterophylla can inhibit cell differentiation and proliferation.