Abstract
Differentiation of pluripotent stem cells to cardiomyocytes is influenced by culture conditions including the extracellular matrices or similar synthetic scaffolds on which they are grown. However, the molecular mechanisms that link the scaffold with differentiation outcomes are not fully known. Here, we determined by immunofluorescence staining and mass spectrometry approaches that extracellular matrix (ECM) engagement by mouse pluripotent stem cells activates critical components of canonical wingless/integrated (Wnt) signaling pathways via kinases of the focal adhesion to drive cardiomyogenesis. These kinases were found to be differentially activated depending on type of ECM engaged. These outcomes begin to explain how varied ECM composition of in vivo tissues with development and in vitro model systems gives rise to different mature cell types, having broad practical applicability for the design of engineered tissues.
Highlights
We focused on three candidate kinases, namely, integrin-linked kinase (ILK), the serine/threonine p21-activated kinase (PAK), and the focal adhesion kinase (FAK), which have been proposed as important modulators of extracellular matrix (ECM)-associated signaling pathways
Our findings demonstrate early dynamical changes in the ECM composition during cardiomyocyte differentiation concomitant with mesoderm expression, which are mainly regulated by FAK signaling in 2D culture of mouse induced pluripotent stem cells
In order to more accurately identify the signaling pathways transduced by ECM leading to cardiac differentiation of mouse induced pluripotent stem cells (miPSCs), we adapted the formulation consisting of type I collagen, fibronectin, and laminin-111 (CFL) for a 2D cell culture system
Summary
Regenerative therapies for cardiac tissue are in high demand to treat a number of conditions including congenital heart defects, ischemic injury, and myocarditis [1,2]. Combined, these maladies represent more than a third of annual deaths in the United States. 3D bioprinting structures that mimic the composition of native tissue have been proposed as a therapeutic option, gaining traction due to their accessibility and modularity [3,4,5] While this is an exciting area of research, it comes with substantial challenges, with cardiac tissue. In situ differentiation of stem cells or their progeny requires cues for differentiation, and these cues differ between the cell types found in native heart tissue
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