Integrin stimulation by collagen I at the progenitor stage accelerates maturation of human iPSC-derived cardiomyocytes

Abstract

Cell manufacturing challenges have hampered effective preclinical evaluations of mature cardiac cells derived from human-induced pluripotent stem cells (hiPSCs). These challenges mainly stem from standard differentiation methods yielding cardiac cells of an immature phenotype, low cell yields and the need for extended culture for enhanced maturation. Although the intricate relationship between extracellular matrix (ECM) components and integrin expression levels plays a pivotal role during heart development, the impact of differentiation and maturation of cardiac cells on integrin behavior has not been thoroughly studied. This study postulates that cardiac cell maturation is significantly influenced by the timing of integrin stimulation via cell-matrix interactions. We profiled integrin expression levels throughout the differentiation process of cardiac cells and assessed the effects of utilizing defined ECM components as culture substrates on cell adhesion, proliferation, differentiation, and maturation. Our findings reveal that integrins facilitate hiPSC adhesion to ECM coated culture surfaces and underscores dynamic alterations in integrin expression during cardiac cell differentiation. Remarkably, we observed significant enrichments in α2 and β1 collagen integrin levels at the progenitor and differentiated stages. These shifts in collagen integrin levels were associated with enhanced cell seeding efficiency on collagen-type I surfaces and altered population doubling times. The stimulation of collagen integrins at the progenitor stage markedly boosted cardiac cell maturation, demonstrated by a significant (∼3-fold) increase in cardiac troponin I expression compared to the standard method after 15 days of culture. Enhanced maturation levels were further supported by significant increases in sarcomere development, maturation gene expression, morphological features, improved beating potency, and fatty acid metabolism dependency. Cardiac maturation driven by collagen was abrogated upon inhibition of collagen integrins targeted with selective pharmacological blockers, affirming their indispensable role in maturation without affecting cardiac differentiation levels. Our work confirms that stimulating collagen integrins at the progenitor stage is a potential strategy to achieve rapid maturation of hiPSC-derived cardiac cells.

Statement of significance

This study offers a novel strategy guided by integrin expression levels for generating hiPSC-CMs with improved maturation features in a short culture period (<16 days). The improvements in cardiac cell maturation were achieved by stimulating collagen type 1 integrin at the progenitor stage. The potential benefits of this method for regenerative cardiac repair will pave the way for the preclinical examination of mature cardiac cells in tissues to advance cell manufacturing and cardiac toxicity studies.