Abstract 17896: Activation of Innate Immunity is Required for Efficient Nuclear Reprogramming and Endothelial Differentiation
Introduction: Stem cell therapy for vascular regeneration has been investigated using embryonic stem cells. We recently generated endothelial cells (ECs) from human induced pluripotent stem cells (hiPSCs) and investigated their potential to promote the perfusion of ischemic tissue in a murine model of peripheral arterial disease (PAD). However, to utilize iPSCs therapeutically, the cells should be generated via non-integrating approaches to avoid integration of foreign DNA.
Objective: The present study highlights underlying mechanisms of reprogramming and investigates the role of novel pathways in enhancing nuclear reprogramming for potential clinical application.
Results: In seeking to develop novel non-integrating approaches using cell-permeant proteins (CPPs), we discovered a striking difference in the pattern of gene expression induced by viral versus protein-based delivery of the reprogramming factors. This suggested that a signaling pathway required for efficient nuclear reprogramming was activated by the retroviral, but not CPP approach. In both gain- and loss-of function studies, we find that activation of toll-like receptor 3 (TLR3) plays a role in the efficient reprogramming of human cells using viral approaches. Stimulation of TLR3 causes rapid changes in the expression of epigenetic modifiers, with chromatin remodeling and changes in gene expression that favors induction of pluripotency. Importantly, knowing that this pathway is critical, we were able to generate hiPSCs using CPPs by adding a TLR3 agonist (Poly IC) to the reprogramming protocol. Furthermore, the insights obtained from the TLR pathway made it feasible to transdifferentiate fibroblasts to EC directly (rather than reprogramming to pluripotentiality, followed by differentiation to EC) when provided with the essential growth factors.
Conclusion: Recognition of the role of innate immunity signaling in reprogramming may advance the therapeutic application of iPSCs. We intend to develop an efficient protein-based system to generate EC and determine their therapeutic potential in animal models of PAD. Furthermore, we have discovered an important signaling pathway in reprogramming, which may have implications in cancer biology and regenerative medicine.
- © 2012 by American Heart Association, Inc.