Screening and Identification of BMP5 as a Key Regulatory Gene for hPSCs Transcardiomyocyte Differentiation

Human pluripotent stem cells (hPSCs)-derived cardiomyocytes have tremendous advantages in generating cell models that are suitable for the study of cardiovascular diseases and their treatment. However, there is a lack of systematic screening and analysis of important genes for inducing hPSCs differentiation into cardiomyocytes.

Gene Expression Omnibus (GEO) databases were used to explore the key genes involved in hPSCs differentiation into cardiomyocytes. Then, the induction of human embryonic stem cells (hESCs) differentiation into cardiomyocytes was verified by alkaline phosphatase (ALP) detection, real-time PCR, western blotting and immunofluorescence experiments. Furthermore, BMP5 overexpression and CRISPER/Cas9-mediated knockout of BMP5 were performed to determine the important role of BMP5 in the differentiation of hESCs into cardiomyocytes.

Through differential expression analysis of high-throughput sequencing datasets from four stages of hPSCs differentiation into cardiomyocytes (undifferentiated hPSCs, mesoderm cells, early cardiomyocytes, and cardiomyocytes), 10 key differentially expressed genes (DEGs) were ultimately identified. Weighted gene co-expression network analysis (WGCNA) and protein-protein interaction (PPI) network analysis subsequently revealed that BMP5 interacted with the transcriptional regulator NKX2-5. During the induction of hESCs differentiation into cardiomyocytes, the expression of BMP5 was significantly increased in the mesoderm, early cardiomyocyte, and cardiomyocyte stages compared with that in the hESCs stage. After overexpression of BMP5 in hESCs and induction of cell differentiation into cardiomyocytes, the number of myocardial beats and the expression levels of cardiomyocyte marker proteins were significantly greater than those in nontransfected hESCs-derived cardiomyocytes. CRISPR/Cas9-mediated knockout of BMP5 in ESCs and then cells were induced to differentiate towards cardiomyocytes, transcriptome sequencing results revealed a significant reduction in the expression levels of genes related to myocardial development.

The role of BMP5 in the differentiation of hESCs into cardiomyocytes was explored, and findings demonstrated that BMP5 promoted the differentiation of hESCs into cardiomyocytes.