Combinatorial extracellular matrix microenvironments promote survival and phenotype of human induced pluripotent stem cell-derived endothelial cells in hypoxia

Abstract

Recent developments in cell therapy using human induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) hold great promise for treating ischemic cardiovascular tissues. However, poor post-transplantation viability largely limits the potential of stem cell therapy. Although the extracellular matrix (ECM) has become increasingly recognized as an important cell survival factor, conventional approaches primarily rely on single ECMs for in vivo co-delivery with cells, even though the endothelial basement membrane is comprised of a milieu of different ECMs. To address this limitation, we developed a combinatorial ECM microarray platform to simultaneously interrogate hundreds of micro-scale multi-component chemical compositions of ECMs on iPSC-EC response. After seeding iPSC-ECs onto ECM microarrays, we performed high-throughput analysis of the effects of combinatorial ECMs on iPSC-EC survival, endothelial phenotype, and nitric oxide production under conditions of hypoxia (1% O2) and reduced nutrients (1% fetal bovine serum), as is present in ischemic injury sites. Using automated image acquisition and analysis, we identified combinatorial ECMs such as collagen IV+gelatin+heparan sulfate+laminin and collagen IV+fibronectin+gelatin+heparan sulfate+laminin that significantly improved cell survival, nitric oxide production, and CD31 phenotypic expression, in comparison to single-component ECMs. These results were further validated in conventional cell culture platforms and within three-dimensional scaffolds. Furthermore, this approach revealed complex ECM interactions and non-intuitive cell behavior that otherwise could not be easily determined using conventional cell culture platforms. Together these data suggested that iPSC-EC delivery within optimal combinatorial ECMs may improve their survival and function under the condition of hypoxia with reduced nutrients.

Combinatorial Extracellular Matrix Microenvironments for Probing Endothelial Differentiation of Human Pluripotent Stem Cells

“…1A) 15, 18 . Each ECM microarray slide consisted of 63 unique ECM compositions derived from the 6 ECM proteins at equal mass ratios (1:1, 1:1:1, etc.).…”

Decellularized Extracellular Matrix Materials for Cardiac Repair and Regeneration

“…This method can be applied to cardiac‐specific ECM to identify regenerative components. Combinatorial methods use multiple ECM‐derived components in 2D or 3D to analyze which combination of materials induces functional changes in stem cells . These systems can also be used to analyze components that may be beneficial to supplement dECM scaffolds and improve reparative outcomes.…”

Cardiac Extracellular Matrix Modification as a Therapeutic Approach

“…These simplistic studies of single ECM components led to the study of combinations of cECM proteins in 2D in order to discern how interactions between proteins influence cardiac cell behavior. When human pluripotent stem cells were seeded on combinatorial ECMs composed of Col IV, Lam 111, and heparan sulfate or Col IV, gelatin, and heparan sulfate, a significant increase in CD31 expression was observed over single-factor ECMs [32]. The study implies that cell behavior is guided through a combination of signals from the ECM, several of which are required in order to enhance differentiation.…”