Definitive endoderm differentiation of human-induced pluripotent stem cells using signaling molecules and IDE1 in three-dimensional polymer scaffold

Human-induced pluripotent stem cells (hiPSCs) are considered to be potentially able to differentiate into all human cell lineages and thus hold promise as an unlimited source for cell replacement therapies in clinical applications. Definitive endoderm (DE) formation is the first and crucial step in the development of visceral organs such as liver, lung, pancreas and so forth. Therefore, efficient generation of DE cells ensures the efficient generation of eventual target cells used in cell therapy. In the present study, Matrigel-coated poly(lactic acid)/gelatin (PLA/gelatin) nanofibrous scaffolds were utilized to investigate the proliferation and differentiation of hiPSCs into DE cells. Analyses of DE-specific markers including Sox17, FoxA2, and Gooscoid (Gsc) genes revealed higher levels of mRNA and protein expression in the differentiated hiPSCs cells cultured on PLA/gelatin scaffolds than cells differentiated in two-dimensional (2D) culture. Our results showed that three-dimensional (3D) cultures could significantly promote DE differentiation in comparison with 2D culture. Also using small molecules such as inducer of definitive endoderm 1 (IDE1) and signaling molecules such as Activin A and Wnt3a could enhance the DE differentiation of hiPSCs with Activin A/Wnt3a being significantly more potent in both 2D and 3D cultures compared to IDE1. The results of this study may have impact in tissue engineering and cell replacement therapy of visceral organs-related diseases.