The Role Of Polycomb PHC1 On Differentiation Of Human Embryonic Stem Cells Into Cardiomyocyte

Due to the pluripotency of embryonic stem cells,it has received continuous attention and development in the fields of regenerative and transplantation medicine,disease model building,drug screening,and human developmental biology.Unlike somatic cell types,pluripotent stem cells only exist briefly in the first few days of embryogenesis,and there is no environment capable of maintaining pluripotency in the body for a long time.Therefore,hPSCs in the laboratory rely on cell culture conditions and require various.Pathway signaling stimulation and appropriate culture methods and models maintain hPSCs stemness and promote their differentiation.The formation of the heart is a complex process that involves the rapid specialization and differentiation of cells and the activation and expression of related genes in the order of time and space.This gene is strictly and orderly activated and expressed in strict accordance with the rules of time and space and is largely determined by the regulation of epigenetic modifications.Polycomb group proteins(PcG)are important epigenetic molecules that maintain stable gene expression and determine cell fate.They play a key role in cell identification and maintain proper differentiation.PHC1 is an important component of PcG protein.This thesis attempts to establishan in vitro differentiation system of embryonic stem cells into cardiomyocytes and to identify the specific protein expression of cardiomyocytes by immunofluorescence,and to identify gene markers of mesoderm,cardiac precursors and mature cardiomyocytes during the differentiation process by Quantitative real-time PCR.Construct shRNA for knockdown of embryonic stem cells.Under the condition that knockdown will not affect the maintenance of cell stemness,perform PHC1 knockdown before myocardial differentiation begins and in the middle and late stages of myocardial differentiation,and then construct a tagged PHC1 overexpression plasmid and transfer to Embryonic stem cells were screened for hygromycin to begin cardiomyocyte differentiation to determine the role of PHC1 in the directional differentiation of hESCs into cardiomyocytes.The experiment found that the system for inducing stem cells to be cardiomyocytes has been established through immunofluorescence and real-time quantitative PCR identification;PHC1 in shRNA-specific knockdown stem cells has little effect on embryonic stem cell proliferation and survival,and NANOG,OCT4,and SOX2 are still at high level of transcription;overexpression of PHC1 does not affect the expression of core transcription factors NANOG,OCT-4,SOX-2;PHC1 regulates the early stage of myocardial differentiation,that is,the period of mesoderm formation,through MESP1 to affect cardiac development,eventually.Set a foundation for further understanding the relationship between myocardial differentiation and PcG,and provide ideas for clinical treatment of heart disease through embryonic stem cells.