Abstract 722: Unveiling Ccbe1 Role as a Modulator of Cardiomyocyte Differentiation

Abstract

Chronic heart failure is a major unmet clinical need arising from the loss of viable and functional cardiac muscle, representing a major cause of mortality worldwide. There is a need to identify key molecules and signaling pathways acting on the coronary vasculature system towards regenerative therapies. Human induced pluripotent stem cells (hiPSC) are an attractive cell source to understand the regulatory networks involved in cardiac commitment and cardiomyocyte (CM) differentiation. Particularly, CCBE1 (collagen and calcium-EGF biding domain 1) has been studied as a secreted protein critical for lymphatic/cardiac vascular development and recent reports have proposed that CCBE1 may potentially be used to restore cardiac tissue upon heart injury, through CCBE1-mediated cardiac commitment and/or augmentation of lymphangiogenesis. Therefore, the goal of our work is to understand the molecular pathways underlying CCBE1-based cardiac commitment in loss-of-function studies. By exploring gene editing and “omics” tools we aim to unveil the molecular basis of CCBE1-induced cardiogenesis and provide novel insights towards the development of CCBE1-mediated therapeutic strategies for cardiac regenerative medicine. To selectively knock-down (KD) CCBE1 expression along hiPSC cardiac differentiation, we used modified hiPSC line with CRISPR interference technology (CRISPRi-hiPSC). The CCBE1 KD led to a reduction on the expression of cardiac troponin marker TNNT2 and on the ratios of MYH7:MYH6 and TNNI3:TNNI1. The CCBE1 KD-derived CMs also presented an immature-related ultrastructure, suggesting that CCBE1 may modulate the CM phenotype. On the other hand, the EC differentiation was not impaired by CCBE1 KD. A comprehensive and integrated characterization of the transcriptome and proteome along the differentiation in the presence or absence of CCBE1 is being pursued to identify the key players at cardiac mesoderm and cardiac progenitors stages and their interactions with CCBE1. This work opens new avenues for the identification of CCBE1-modulated proteins/pathways in cardiac commitment, which may contribute for novel cell-based or cell-free approaches towards more efficacious cardiac regenerative therapies.