| The growth and development process of Vertebrate skeletal muscle tissue includes the embryonic myotome formation (myoblast progenitor cells), muscle stem cells (stem cell), myoblasts (myoblast), myotubes (myotube) formation and fusion and mature. During the ontogeny, muscle development performed in the increase of muscle cells number and the expansion of muscle fiber volume. Skeletal muscle is regulated by many transcription factors, such as Pax7, Myf5, MyoD and Myogenin. In recent years, many muscle development and differentiation related genes was found to be regulated by miRNAs.this not only make the muscle development and differentiation of the adjustment process more complex, but also tell the importance role of miRNAs in skeletal muscle development.Myosin heavy chain (myosin Heavy Chain MHC) is an important structural and functional component of the skeletal muscle. Our previous study found that miRNA could regulate skeletal muscle differentiation by regulating skeletal muscle-specific MHC (MYH1, MYH2, MYH4coding). Undifferentiated skeletal muscle precursor cells mainly express Non-muscle-specific NMHC. With the differentiation and maturation of skeletal muscle,the skeletal muscle-specific MHCs are primarily expressed. Whether the miRNAs play a regulatory role in the process of reduction of NMHC (myogenic differentiation)?There are three NM Ⅱ isoforms NMHC ⅡA, ⅡB and IIC(MYH9, MYH10, MYH14coding) in mammals. Myoblasts can’t complete myogenic differentiation in vitro without NMHC IIA. No appropriate role was found in ⅡB and ⅡC. It showed that NMHC IIA has a regulatory function for muscle differentiation. This paper demonstrates that MYH9expression levels are different in laying hens and broilers. Here, using bioinformatics detection analysis,we found that miR-199could combined with the MYH9mRNA3’-UTR. Recent reports indicated that the miR-199expression level was significantly higher in the broiler compared with hens.Thus, we hypothesized that miR-199may regulate skeletal muscle differentiation by regulating the expression of MYH9.Chicken embryonic development experience21days, in which embryonic skeletal muscle development starting at6days to16days to develop basic mature. We first compared the expression level of MYH9between laying hens and broiler. As a result, we found that both the MYH9and miR-199expression levels in the embryonic muscle with a significant difference, showing a reverse correlation. The3’-UTR cDNA sequence of MYH9was inserted into the PGL-3vector to construct a luciferase reporter gene expression plasmid. We found that miR-199could significantly inhibite transfected luciferase expression, indicating that the MYH9gene is target of miR-199. Layers and Broilers embryonic myoblasts were isolated and cultured and induced to differentiate in vitro. In consequence,we found that the myogenic differentiation of myoblasts of Layers and Broilers is significantly different. Layers myoblasts had a better differentiation than Broilers, and a larger myotube diameter. During differentiation, Layers had a lower expression of miR-199Compared with Brolier. MYH9expression Was negatively correlated with this.To investigate the role of miR-199for the regulation of muscle differentiation both over-expression and knockdown of miR-199a were performed in cultured myoblast. Layer myoblasts transfected with microRNA analogues(mimics) for inhibiting the expression of mir-199could not complete myogenic differentiation. To transfect MicroRNA inhibitors in Broiler myoblasts for suppress the expression of miR-199could significantly promote myogenic differentiation.Taken together, our results demonstrated that miR-199could regulate skeletal muscle differentiation by regulating MYH9expression. our data suggests a novel mechanism under the regulatory effects on myogenesis by miRNAs, and provides a new idea for poultry and livestock meat quality improvement.
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