Combining genome and tissue engineering for next-generation human biomimetics.

Tissue engineering for cardiovascular implants has largely utilized primary human cells to generate human tissue-engineered matrices (hTEMs). However, due to donor-to-donor variability and limited passage numbers, a more robust alternative to primary cells would be beneficial. To overcome these limitations, we have defined a new differentiation protocol for human-induced pluripotent stem cells (hiPSCs) into isogeneic cardiac fibroblast-like cells (iCFs) using animal sera-free and chemically defined methods. Morphology, extracellular matrix (ECM) deposition, and global transcriptomics revealed similarity between iCFs and primary human cardiac fibroblasts. Additionally, by overexpressing specific ECM and ECM-related proteins through gene-editing approaches, the ECM composition can be modulated as a building block to create "designer" next-generation hTEMs. Proteomics of gene-edited iCF-derived hTEMs demonstrated an increase in proteins involved in collagen and elastic fiber assembly. Furthermore, analysis of gene-edited iCF-derived hTEM mechanical functionality through biaxial mechanical testing exhibited increased collagen function, attributed to increased crosslinking and maturation. In sum, we have combined hiPSC technology with genome engineering to lay the foundation for next-generation tissue engineering applications by generating a novel cell source, gene-edited iCFs, that are able to modulate the composition as well as the functional mechanics of hTEMs.