Differentiation of skeletal myoblasts is a tightly orchestra

Differentiation of skeletal myoblasts is a tightly orchestrated process that involves myoblast proliferation, np e withdrawal, expression of muscle-specific genes, and fusion into multinucleated myofibers (Horsley and Pavlath, 2004; Krauss, 2010). The maintenance of muscle mass is important for staving off the risk of metabolic syndrome and age-related muscle loss, which affects the quality of life (Sanchez et al., 2013). Myoblast differentiation is also under the control of several signaling pathways, which include Akt and p38MAPK. These signaling pathways play critical roles in early differentiation induction events, such as activation of the MyoD transcription factor through enhancing the heterodimerization of MyoD with its partner E proteins, activation of Mef2 by phosphorylation, changes in chromatin-remodeling at muscle-specific genes, and cell survival (Bae et al., 2009, Bae et al., 2010; Serra et al., 2007; Simone et al., 2004).
Materials and methods
Results
Discussion Endogenous estrogens are well recognized as playing important roles in the regulation and maintenance of sex-specific myogenic differentiation. However, BPA or its metabolites are well known obesogens that induce adipogenesis (Boucher et al., 2015), and little is known about their myogenic effects, and their interactions with endogenous estrogens. In addition, skeletal muscle is expected to be a target tissue for estrogens, because two ER isoforms are expressed in skeletal muscle (Wiik et al., 2009). BPA has been reported to bind to hormone receptors, and influence multiple endocrine pathways (Deutschmann et al., 2013; Matsushima et al., 2008; Riu et al., 2011). In particular, BPA reduces Akt phosphorylation in skeletal muscle, and alters serum adipocytokine levels, thereby inducing glucose intolerance (Moon et al., 2015). In this study, we investigated the effects and molecular mechanism of BPA on myoblast differentiation, and found BPA has no effects on the proliferation or apoptosis of C2C12 myoblasts; however, it influences the capacity for myoblast differentiation at low concentration (less than 1 μM) (Fig. 1A). In addition, we found that BPA and estradiol inhibit myoblast differentiation with reduced muscle gene expression and myotube formation (Figs. 1A, 4A and 5A). Like the effect of BPA on glucose intolerance (Boucher et al., 2015), in the present study BPA exposure reduced Akt phosphorylation, which plays a critical role in myoblast differentiation. As there were no overt signs that BPA-induced cell death or cytotoxicity was associated with differentiation, BPA-inhibitory effect on Akt activation is specific to the differentiation program. The binding of estrogen (17ß-estradiol) to the ligand-binding domain of ERα exerts genomic effects on DNA sequences called estrogen response elements (EREs), which regulate the transcription of target genes. However, 17ß-estradiol decreases the levels of myogenic regulatory factors (MRFs) during skeletal muscle formation, and acts to repress myogenesis in mouse C2C12 myoblast and mouse satellite cells (Ogawa et al., 2011). Thus, it is conceivable that BPA might also suppress MRFs, which can be observed in higher dose of BPA (Fig. 1C). Efficient myoblast differentiation requires not only Akt but also p38 MAPK activation, and it appears that regulates cell cycle control, MyoD dimerization with E proteins, and Mef2 transcriptional activity in myoblast differentiation (Bae et al., 2009; Krauss, 2010). However, the exposure of BPA in myoblasts did not affect p38MAPK activities on myogenesis (data not shown). Previous studies have shown that APPL1/Akt might form different pools of signaling complexes of JLP/p38MAPK in myoblast differentiation (Bae et al., 2010). The suggestion that BPA inhibits mechanisms for the activation of Akt in differentiating myoblasts might include signaling by APPL1, an interacting partner of Akt (graphic highlight). To compare the toxicity of BPA to that of E2, we used somewhat high concentration, 1 μM. As human daily exposure to BPA is approx. 1/228-1/45 μM (Yang et al., 2014), we can inversely apply uncertainty factor, 100, i.e., 1/2-2.2 μM to mice. for For real exposure issues, we tried to match animal exposure levels, 1 μM of BPA and E2, to human real exposure levels