Expressão de MMP-2, MMP-9 E MT1-MMP durante o desenvolvimento muscular de peito em embriões de galinhas

Abstract

In vertebrates, striated skeletal muscles originate from the epithelial borders of the dermomyotome, a region of the somites, through the differentiation of mononucleated myoblasts. The myoblasts fuse to form multinucleated myotubes that begin to express muscle-specific proteins. Although it is known that metalloproteinases are involved in the morphogenesis of different organs and tissues, there are few in vivo studies demonstrating their expression during the formation of striated skeletal muscle. Thus, the aim of this study was to verify whether there is expression of metalloproteinases 2, 9 and MT1-MMP during the development of the pectoral muscle in chickens at E11, E15 and E19 days of incubation. For this purpose, fertilized eggs donated by Granja Econômica Avícola LTDA (Carambeí-PR) were used and kept in an incubator for 11, 15 and 19 days. At the analyzed times, the embryos were dissected and part of the removed pectoral muscle was kept in culture for 24h in conditioned DMEM medium (without fetal bovine serum) and part was fixed. The samples were subjected to Masson's Trichome staining, immunofluorescence for beta-actin, zymography, including inhibition tests and immunohistochemistry for MMPs. Masson's Trichome staining evidenced the morphological phases of differentiation, from myoblasts to the formation of muscle fibers. Immunofluorescence demonstrated changes in the actin distribution pattern in the cytoplasm of the cells, throughout the analyzed stages. Zymography showed bands of proteolytic activity above 55 kDa suggestive of MMP-2 and confirmed by protease inhibition tests that, in addition to MMP-2, showed expression of serine proteases in the samples. Immunohistochemistry detected the expression of MMP-2 and MT1-MMP at all time points, but did not show the presence of MMP-9. It is therefore concluded that the proteases MMP-2, MT1-MMP and serine proteases are expressed during the muscle differentiation phases in chicken embryos.