Expression and Methylation of FGF2, TGF-β and Their Downstream Mediators during Different Developmental Stages of Leg Muscles in Chicken

A number of growth factors determine the proliferation of myoblasts and therefore the number of ultimate myofibers. The members of transforming growth factor-beta (TGF-β) family and the fibroblast growth factor 2 (FGF2) have profound effects on skeletal myoblasts proliferation in various animal systems. To investigate their involvement in different stages of avian skeletal muscle development in vivo, we detected the mRNA expression and DNA methylation profiles of TGF-β2, TGF-β3, FGF2 and their downstream mediators in leg muscles at embryonic day 10, day of hatch and day 45 posthatch, using both Arbor Acres meat-type and White Leghorn egg-type chickens. By real-time PCR, we found that the expression levels of TGF-β2, TGF-β3, Smad3 and FGF2 were significantly (P≤0.01) higher at embryonic day 10, a developmental window of abundant fetal myoblasts expansion, by comparison to day of hatch and day 45 posthatch. The methylation status of the 5′ end region of these four genes was examined subsequently. A section of a CpG island in the 5′ end region of FGF2 was significantly hypomethylated (P≤0.01) at embryonic day 10, compared with neonatal and postnatal stages in both stocks. Our results suggested that TGF-β2, TGF-β3, Smad3 and FGF2 may play important roles in fetal myoblasts proliferation in chicken hindlimb, and the transcription of FGF2 in this wave of myogenesis could be affected by DNA methylation in 5′ flanking region. These outcomes contribute to our knowledge of the growth factors in avian myogenesis. Further investigation is needed to confirm and fully understand their functions in fetal limb myogenesis in birds.