Synergistic short-term and long-term effects of TGF-ß1 and 3 on collagen production in differentiating myoblasts.

ABSTRACT

Skeletal muscle fibrosis and regeneration are modulated by transforming growth factor ß (TGF-ß) superfamily. Amongst them, TGF-ß1 is a highly potent pro-fibrotic factor, while TGF-ß3 has been implicated to reduce scar formation and collagen production in skin and vocal mucosa. However, little is known about the individual and combined short- and long-term effects of TGF-ß1 and TGF-ß3 on collagen expression in myoblasts and myotubes. Here we show that in C2C12 myoblasts TGF-ß1 and/or TGF-ß3 increased mRNA expression of Ctgf and Fgf-2 persistently after 3 h and of Col1A1 after 24 h, while TGF-ß1+TGF-ß3 mitigated these effects after 48 h incubation. Gene expression of Tgf-ß1 was enhanced by TGF-ß1 and/or TGF-ß3 after 24 h and 48 h. However, Tgfbr1 mRNA expression was reduced at 48 h. After 48 h incubation with TGF-ß1 and/or TGF-ß3, Col3A1 and Col4A1 mRNA expression levels were decreased. Myoblasts produced collagen after three days incubation with TGF-ß1 and/or TGF-ß3 in a dose independent manner. Collagen deposition was doubled when myoblasts differentiated into myotubes and TGF-ß1 and/or TGF-ß3 did not stimulate collagen production any further. TGF-ß type I receptor (TGFBR1) inhibitor, LY364947, suppressed TGF-ßs-induced collagen production. Collagen I expression was higher in myotubes than in myoblasts. TGF-ß1 and/or TGF-ß3 inhibited myotube differentiation which was antagonized by LY364947. These results indicate that both C2C12 myoblasts and myotubes produce collagen. Whereas TGF-ß1 and TGF-ß3 individually and simultaneously stimulate collagen production in C2C12 differentiating myoblasts, in myotubes these effects are less prominent. In muscle cells, TGF-ß3 is ineffective to antagonize TGF-ß1-induced collagen production.