Proteomic Approach Using DIA-MS Identifies Morphogenesis-Associated Proteins during Cardiac Differentiation of Human iPS Cells

Human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes have potential applications in regenerative medicine. The quality by design (QbD) approach enables the efficiency and quality assurance in the manufacturing of hiPSC-derived products. It requires a molecular understanding of hiPSC differentiation throughout the differentiation process; however, information on cardiac differentiation remains limited. Proteins associated with the early stages of cardiac differentiation would be useful in the cardiomyocyte quality assessment. Here, we performed quantitative proteomics of hiPSC intermediate cells in the early phase of cardiac differentiation to better understand their molecular characteristics. Proteomic profiles suggested that day 5-7 cells were in the morphogenetic stage of cardiac differentiation. Trophoblast glycoprotein (TPBG) was the most up-regulated protein in the morphogenetic stage; it was previously shown to be up-regulated during differentiation into neural stem cells. Proteomics of TPBG-knockdown cells revealed that TPBG is involved in cell proliferation and is related to the cardiomyocyte yield, suggesting that it could be used as a marker in QbD development. Our approach helps us understand the molecular basis of hiPSC differentiation and could be a powerful tool in QbD-based manufacturing.