Mechanism Of Egr1 In The Proliferation Of Mouse Myoblasts

Skeletal muscles are an important component of the human body and essential organs for vital activities such as exercise and metabolism.In clinical practice,there are still many insurmountable problems in the prevention and treatment of muscle injury and muscle degenerative diseases.The key to solving these problems is to study the molecular mechanisms of skeletal muscle growth and development.Egr1 is widely expressed in various types of cells and participates in important physiological processes such as cell proliferation,differentiation,invasion,and apoptosis.Previous laboratory research has shown that Egr1 can promote the differentiation of C2C12 cells,and further studies have shown that Egr1 also plays a role in the proliferation of mouse myoblasts.Therefore,we chose to study the effect and mechanism of Egr1 on mouse myoblast proliferation through C2C12 cells and mouse muscle injury repair models,in order to provide theoretical basis for the treatment of muscle injury and muscle related diseases.The main research content includes:(1)Western blotting technology was used to detect the protein expression pattern of Egr1 during the proliferation of C2C12 cells.(2)Construct an overexpression and interference vector of Egr1,and detect the effect of Egr1 on the proliferation of C2C12 cells using Western blotting,Ed U,and flow cytometry.(3)Chromatin immunoprecipitation and double Luciferase reporter gene technology were used to verify the binding of Egr1 and Pax7 promoter region.(4)Overexpression and interference of Egr1 during the proliferation process of C2C12 cells,and Western blotting was used to detect the regulatory effect of Egr1 on Pax7.(5)Western blotting and immunohistochemical staining were used to detect the expression and localization of Egr1 and Pax7 during muscle injury repair in mice,as well as the regulation of Egr1 on Pax7.The results indicate that:(1)During the proliferation process of C2C12 cells,the expression level of Egr1 protein gradually increases,reaching its highest value after 24 hours of proliferation culture.This is consistent with the expression pattern of proliferation related marker proteins CCNB1 and PCNA,indicating that Egr1 may be involved in the regulation of C2C12 cell proliferation process.(2)After overexpression of Egr1,the protein levels of CCNB1 and PCNA significantly increased.Ed U test results showed an increase in cell proliferation rate,and flow cytometry test results showed a significant increase in the proportion of cells in the S phase;After inhibiting Egr1,the protein levels of CCNB1 and PCNA significantly decreased.Ed U test results showed a decrease in cell proliferation rate,and flow cytometry test results showed a significant decrease in the proportion of cells in the S phase.This indicates that Egr1 can regulate the proliferation of C2C12 cells.(3)The results of chromatin immuno coprecipitation showed that the two binding sites predicted by Egr1 in the Pax7 promoter region were correct;The double Luciferase reporter gene detection showed that the activity of Luciferase was significantly reduced after two Egr1 binding sites on the Pax7 promoter were mutated at the same time or separately,which proved that Egr1 could bind to two binding sites in the Pax7 promoter.(4)After overexpression and interference with Egr1,the protein expression level of Pax7 increases and decreases,indicating that Egr1 can regulate the expression of Pax7.(5)The Western blotting test results showed that during the process of muscle injury repair,the expression patterns of Egr1 and Pax7 proteins were consistent,gradually increasing from day 0to day 3 of the injury,and reaching a peak on day 3.After upregulating Egr1 expression levels by injecting IGF1 into the injury site,the expression level of Pax7 protein significantly increased.The immunohistochemical staining results showed that on the third day of muscle injury,the positive signals of Egr1 and Pax7 at the injury site were the strongest in muscle satellite cells.Prove that Egr1 can regulate the proliferation of muscle satellite cells by regulating the expression of Pax7 during the repair process of muscle injury in mice.In vivo and in vitro studies have shown that Egr1 can regulate its expression by binding to the Pax7 gene promoter region,promote the proliferation of mouse skeletal muscle satellite cells,and accelerate the process of skeletal muscle damage repair.