Differentiating Human Pluripotent Stem Cells to Cardiomyocytes Using Purified Extracellular Matrix Proteins

Abstract

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) can be differentiated into cardiomyocytes (hESC-CMs and iPSC-CMs, respectively), which hold great promise for cardiac regenerative medicine and disease modeling efforts. However, the most widely employed differentiation protocols require undefined substrates that are derived from xenogeneic (animal) products, contaminating resultant hESC- and iPSC-CM cultures with xenogeneic proteins and limiting their clinical applicability. Additionally, typical hESC- and iPSC-CM protocols produce CMs that are significantly contaminated by non-CMs and that are immature, requiring lengthy maturation procedures. In this review, we will summarize recent studies that have investigated the ability of purified extracellular matrix (ECM) proteins to support hESC- and iPSC-CM differentiation, with a focus on commercially available ECM proteins and coatings to make such protocols widely available to researchers. The most promising of the substrates reviewed here include laminin-521 with laminin-221 together or Synthemax (a synthetic vitronectin-based peptide coating), which both resulted in highly pure CM cultures. Future efforts are needed to determine whether combinations of specific purified ECM proteins or derived peptides could further improve CM maturation and culture times, and significantly improve hESC- and iPSC-CM differentiation protocols.

Adult Human Heart ECM Improves Human iPSC-CM Function via Mitochondrial and Metabolic Maturation

“…Regarding what causes these observed effects, there are extensive studies investigating and comparing ECM substrates (i.e., Laminin-521, -221, Collagen-I, Collagen-IV, Fibronectin, Vitronectin) alone or in combination, for their ability to support iCM differentiation [50]. Certain ECM proteins, such as fibronectin, have been reported to promote the formation of precardiac mesoderm, while others, like laminins (Laminin-111, -521) and collagen-IV, have been found to inhibit cardiogenesis [51].…”

Heart-on-a-chip systems: disease modeling and drug screening applications

“…However, because Matrigel is derived from animals (Engelbreth-Holm-Swarm mouse sarcoma cells), it lacks chemical definition, which can lead to varying concentration of proteins and a lack of reproducibility between studies. 133 These discrepancies have motivated investigations into whether particular protein isoforms can support hESC and hiPSC growth and differentiation. Laminin-111, -511, and -521 have had the most success in supporting the growth and efficient differentiation of pluripotent hiPSCs and hESCs, especially laminin-521 combined with laminin-211 or -221.…”

Uncovering the Genetic Basis of Congenital Heart Disease: Recent Advancements and Implications for Clinical Management

“…However, multiple approaches to enhance cardiomyocyte maturation or to make use of 3-dimensional culture models have been developed, such as cardiac organoids, microtissues, engineered heart tissues, and biomimetic culture systems, which may help to improve the suitability of hiPSCs to model CHD in a dish. 106 , 107 , 108 , 109 , 110 , 111 , 112 …”