Abstract
Satellite cells, as stem cells of adult skeletal muscle, are tightly associated with the differentiated muscle fibers and remain quiescent in the absence of muscle damage. In response to an injury, the quiescent satellite cell is activated by soluble factors, including FGFs released from injured myofibers. Using immunostaining, we here first show that TRPC1 channels are highly expressed in satellite cells attached to muscle fibers. Since CD34, a traditional stem cell marker, was recently found to be expressed in skeletal muscle satellite cells we labeled living satellite cells in their physiological niche associated with host FDB fibers using anti-CD34-FITC antibody. We then monitored intra-cellular calcium in anti-CD34-FITC labeled satellite cells attached to muscle fibers using the calcium sensitive dye X rhod-1 which has little fluorescence cross talk with FITC. FGF2 increased intracellular calcium in satellite cells, which was antagonized by the TRPC channel blocker SKF 96365. Immunostaining showed that NFATc3 is highly expressed in satellite cells, but not in host FDB fibers. Elevation of intracellular calcium by FGF2 is accompanied by nuclear translocation of NFATc3 and NFATc2 and by an increase in the number of MyoD positive cells per muscle fiber, both of which were attenuated by TRPC blocker SKF 96365. Our results suggest a novel pathway of satellite cell activation where FGF2 enhances calcium influx through a TRPC channel, and the increased cytosolic calcium leads to both NFATc3 and NFATc2 nuclear translocation and enhanced number of MyoD positive satellite cells per muscle fiber.
Highlights
Muscle satellite cells, the muscle resident stem cells of adult skeletal muscle, are located between the basal lamina and the multinucleated mature muscle fibers, and remain quiescent in the absence of muscle damage
Our interest in transient receptor potential canonical (TRPC) in satellite cells was sparked by the report that in neural stem cells FGF2 evokes calcium transients mediated by TRPC1 (Fiorio Pla et al, 2005)
To examine whether TRPC1 is differentially expressed in satellite cells compared to the host flexor digitorum brevis (FDB) fibers, we carried out immunofluorescent staining experiments on FDB cultures with satellite cells attached
Summary
The muscle resident stem cells of adult skeletal muscle, are located between the basal lamina and the multinucleated mature muscle fibers, and remain quiescent in the absence of muscle damage. In response to an injury, the quiescent satellite cell is activated by soluble factors, including fibroblast growth factors (FGFs) released from injured myofibers, as well as by possible cessation of release of quiescence maintaining factors from the previously undamaged host or other fibers (Montarras et al, 2013). FGF2 is a strong activator of skeletal muscle satellite cells in response to muscle injury. The number of satellite cells entering proliferation and expressing MyoD, as well as the overall number of satellite cells subsequently transiting into the myogenic state, is enhanced in the presence of FGF2 (Yablonka-Reuveni et al, 1999; Shefer et al, 2006). It was proposed that the enhanced number of satellite cells undergoing myogenesis in the presence of FGF2 reflects a specific effect of the growth factor in recruiting satellite cells into active myogenesis (Yablonka-Reuveni and Rivera, 1994)
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