Biochemical and Biophysical Research Communications
Modulation of satellite cell adhesion and motility following BMP2-induced differentiation to osteoblast lineage
Section snippets
Materials and methods
Cell culture and differentiation protocol. C2C12 and MM14 mouse myoblast cell lines were maintained as previously described [15], [16]. For the current studies, cells were plated onto 6-well tissue culture plates at a density of 1.5 × 105 cells/cm2. Cells were cultured for 7 days with or without BMP2 (300 ng/ml, Peprotech Inc.). To examine alkaline phosphatase activity, cultures were processed as described previously [19].
Flow cytometry and immunofluorescence staining. Standard procedures for flow
Induction of osteogenic differentiation
Both the MM14 and C2C12 myoblasts are well characterized for their potential for myogenic differentiation and capability for forming mature myotubes in culture. These two cell lines were originally derived from adult muscle satellite cells. They both express muscle-specific markers and are competent for differentiating into myotubes and incorporate into functional muscle fibers after transplantation in vivo. In addition, the MM14 cells have been reported to exhibit different migratory
Conclusion
The current study has identified that α7 integrin may be a specific marker that defines myoblasts with pluripotent stem cell activity. Also, modulation of integrin receptor expression occurs in parallel with the conversion of myoblasts cells to osteogenic lineage. These alterations in adhesion are potentially important for the optimization of cell functions such as adhesion, motility, remodeling, and tissue repair. Similar changes in adhesion receptors must occur in embryonic and other adult
Acknowledgments
The skilled technical assistance by Baomei Liu and Eric Gschweng is gratefully acknowledged. These studies were supported by a grant from the National Institutes of Health (R01 DE15404).
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2018, Developmental BiologyCitation Excerpt :BMPs and their inhibitors define where and when muscle formation occurs (Re'em-Kalma et al., 1995; Hirsinger et al., 1997; Reshef et al., 1998). During skeletal muscle regeneration, low concentrations of BMP2, 4, and 7 maintain proliferation of satellite cells and myoblasts (Amthor et al., 1998; Ozeki et al., 2007; Ono et al., 2010; Wang et al., 2010; Friedrichs et al., 2011; Sartori et al., 2013;), whereas, high levels of BMP2, 4, 5, or 7 inhibit myogenesis, induce chondrogenesis, and ultimately, osteogenesis of satellite cells, C2C12 myoblasts, C3H10T1/2 MSCs, and other MSCs, with continued exposure in culture (Katagiri et al., 1994; Schmitt et al., 2003; Shea et al., 2003; Bandyopadhyay et al., 2006; Knippenberg et al., 2006; Ozeki et al., 2007; Friedrichs et al., 2011; Takács et al., 2013; Liao et al., 2014; Zhou et al., 2016). Proliferating lizard satellite cells expressed bmp2, 5, and 7 at high levels, unlike their murine counterparts (Fig. 2).
Differentiation of human skeletal muscle stem cells into odontoblasts is dependent on induction of α1 integrin expression
2014, Journal of Biological ChemistryCitation Excerpt :Extracellular matrix (ECM) proteins at a suitable concentration were added as soluble proteins to the growth medium for the indicated time, and then cell proliferation was evaluated using a BrdU cell proliferation enzyme-linked immunosorbent assay (ELISA) (Roche Applied Science) as described previously (16, 17). The protocol for RT-PCR has been described previously (19). The PCR within the exponential phase of the amplification curve was performed for 25 cycles for MYOD (muscle-specific marker); FOXD3 and SOX10 (neural crest markers); MEOX1 (presomitic mesoderm marker); RUNX2 (osteogenic transcription factor); DSPP, dentin matrix protein 1 (DMP-1), and MMP-20 (odontoblast markers); osteopontin and osteocalcin (osteoblast markers); and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in a thermal cycler (GeneAmp PCR System 9700, Applied Biosystems).
Effect of RGD functionalization and stiffness modulation of polyelectrolyte multilayer films on muscle cell differentiation
2013, Acta BiomaterialiaCitation Excerpt :In order to investigate the possible role of β-chain integrins in cell adhesion knockdown of β1 or β3 integrin or both using the siRNA approach was studied. These integrins are known to be involved in cell mechano-sensing and both are present in C2C12 myoblasts [50,51]. The cell area of the transfected cells was quantified after 4 h adhesion (Fig. 3D).