Growth factor messenger ribonucleic acid expression during differentiation of porcine embryonic myogenic cells

The growth factors, IGF-I and II, their binding proteins, IGFBP, and members of the transforming growth factor (TGF) superfamily (myostatin and TGFbeta1) are known to regulate proliferation and differentiation of myogenic cells. We hypothesized that changes in the relative expression of members of the IGF and TGFbeta systems play a significant role in regulating myogenesis in porcine embryonic myogenic cell (PEMC) cultures. Therefore, determining the expression patterns of these factors during PEMC myogenesis is important. Consequently, we used real-time PCR to explore the pattern of IGF-I; IGF-II; IGFBP-2, -3, and -5; IGF-type-I receptor; myogenin; myostatin; and TGFbeta1 mRNA expression during PEMC myogenesis. The progression of differentiation was assessed using creatine kinase activity and myogenin mRNA expression. As anticipated, creatine kinase activity was low in PEMC cultures at 48 h and increased 20-fold (P < 0.0001) between 48 h and its peak at 144 h. Similarly, myogenin mRNA was low at 48 h and increased approximately 5-fold (P < 0.0001) as differentiation progressed, peaking at 120 h and decreasing at 144 h. The patterns of IGF-I and IGFBP-2 mRNA expression were similar and were relatively lower in 48-h PEMC cultures, increasing approximately 5-fold (P < 0.0001) to their greatest levels at 120 h. In contrast, IGF-II and IGFBP-5 mRNA levels were relatively high at 48 h, peaking at 72 h, and steadily decreasing by 60 and 80%, respectively (P < 0.001), at 144 h. The level of IGF-type-I receptor mRNA was relatively high until 96 h of culture, after which it decreased 40% (P < 0.01), reaching a low at 144 h. Levels of IGFBP-3 mRNA were relatively high at 48 h, dropped approximately 40% to their lowest level at 72 h (P < 0.001), and then increased approximately 60% (P < 0.001) to their greatest levels at 144 h. Levels of TGFbeta1 mRNA decreased approximately 30% (P < 0.0001) between 48 and 96 h, then quickly rebounded to a peak at 120 h, and by 144 h had dropped to the levels seen at 72 h. Myostatin mRNA was at its greatest level at 48 h and declined rapidly between 72 and 96 h, finally decreasing by approximately 80% at 144 h (P < 0.0001). Our data demonstrate that these factors are differentially regulated during PEMC myogenesis and provide new information about their pattern of mRNA expression in cultured porcine muscle cells.