Abstract 1693 Inhibition of DNA methylation promotes the emergence of endothelial cell fate during stem cell differentiation

Endothelial cells lining of blood vessles are essential for vascular development and tissue homeostasis. Due to the scarcity and limited capacity of primary endothelial cells (ECs), strategies to promote EC phenotypes from various types of stem cells (SCs) are emerging as promising choices for cell-based therapy and vascular tissue regeneration. However, the process of differentiating stem cells into endothelial cells in vitro remains lengthy and inefficient. Here, we employ a combination of defined biochemical stimuli, fluid shear stress, and DNA methylation modulation to enhance the efficiency of inducing EC phenotypes from mesenchymal stromal cells (MSCs) and embryonic stem cells (ESCs). We find that treatment with the DNA methylation inhibitor 5-AzaC effectively induces EC phenotypes and EC-specific gene expression profiles in differentiating MSCs and ESCs. Furthermore, a sustained, low dose 5-AzaC treatment of ESCs at later stage of ESC differentiation significantly improves EC phenotypes and yield. Genome-wide transcriptome analysis reveals key signaling pathways governing development, morphogenesis and blood vessel patterning that are regulated by DNA methylation during ESC to EC differentiation. These findings highlight a pivotal role of DNA methylation in guiding stem cell differentiation into endothelial cell phenotypes, potentially advancing the development of rapid and efficient EC differentiation protocols for clinical applications. This work is in part supported by NSF STC for Emergent Behaviors of Integrated Cellular Systems (EBICS) (Award #: CBET0939511), Georgia Partners in Regenerative Medicine (to YF), Nelson and Bennie Abell Professorship in Biology (to YF), and Georgia Institute of Technology.