The Mechanism Of Differentiation Regulated By Hypoxia And The Regulation Of PGC-1α By IGF2in C2C12Myoblasts

(1) Insulin-like growth factors(IGFs) play an important role in the growth and development of skeletal muscle and remodeling process of adult skeletal muscle. IGF2is an regulatory factor of embryonic myogenesis, and an autocrine initiation factor of myogenic cell differentiation in vitro. IGF2plays an important role in adult skeletal muscle regeneration and hypertrophy, as well as other physiological and pathological processes. PGC-1α is an important regulator and receptor of body energy metabolism, so it is rich in the organs and tissues, in order to maintain the body’s energy demands. PGC-1α not only has a very important effect on energy metabolism of skeletal muscle, but also relates with muscle diseases. However the regulatory mechanism of PGC-1α in skeletal muscle is still not completely clear. This study aims at exploring how IGF2regulates PGC-1α in skeletal muscle, which is conducive to the theoretical of certain muscular dystrophy and muscle metabolic diseases.Based on the above research purpose, we conduct series of experiments. Results show that, in mouse C2C12cells, exogenous IGF2can down-regulate PGC-1α mRNA level and transcriptional activity. PI3K/Akt inhibitor LY294002can rescue PGC-1α mRNA level which is down-regulated by IGF2. For further study on the mechanism, we construct the FoxO1mutant. FoxO1is a transcription factor of PGC-1α, and it can be phosphorylated by Akt in three sites, thus inhibiting its transcription activity. FoxO1-3A mutant is transfected into C2C12cells, and IGF2can not inhibit PGC-1α. This result shows that Akt can phosphorylate the transcription factor of PGC-1α leading to down-regulation of PGC-1α. These results suggest that IGF2can inhibit PGC-1α through PI3K/Akt-FoxO1pathway in mouse C2C12myoblasts.(2) Hypoxia is a case that the whole body or partial body lost oxygen supply, such as anemia, hypoventilation and pulmonary fibroma. Oxygen is very important in maintaining the life activities of human body, when the tissue do not get enough oxygen, or can not make full use of oxygen, the metabolism function, and even the morphology may be abnormal. To response to hypoxia stress, biological organisms have formed a series of mechanism for hypoxia response, resulting in that the cell, tissue and whole organisms are able to meet the energy metabolism, biosynthesis and oxygen concentration in the case of lack of oxygen.Skeletal muscle progenitor cells can respond to hypoxia. Growth and movement of skeletal muscle are often in a hypoxic environment, and many skeletal muscle diseases also exhibit hypoxia characteristics. Hypoxia can inhibit the differentiation of myogenic cells, suggesting that in the process of development of the microenvironment hypoxia or skeletal muscle remodeling, pathways dependent of oxygen can inhibit progenitor cell differentiation, until there is sufficient supply of oxygen. But the exact mechanism is not entirely understood.As an oxidative stress receptor, AMPK can be activated by hypoxia, activated AMPK is involved in hypoxia response. This prompted us AMPK may be involved in the inhibition of skeletal muscle differentiation response to hypoxia, exhibiting its energy regulator function, leading to the adaptation of skeletal muscle under energy stress. The purpose of this study is to restrain the mechanism of the inhibition of C2C12myoblast differentiation by AMPK under hypoxia.The results show that, hypoxia can inhibit C2C12myoblasts differentiating into multinucleated myotubes; at the same time AMPK is activated in the prosess of differentiation under hypoxic conditions, and hypoxia inhibiting the differentiation of myoblasts is dependent of AMPK. Further research shows that, AMPK can inhibit the phosphorylation of Akt in the process of differentiation under hypoxic conditions. Under hypoxic conditions, phosphorylation of IRS1Ser789is up-regulated, but AMPK inhibitor can lower its phosphorylation, indicating that activated AMPK under hypoxia can inhibit the activity of IRS1by phosphorylating its Ser789, thus inhibiting the PI3K/Akt signaling pathway, leading to restrain the differentiation.