Stiffness of Hydrogels Regulates Cellular Reprogramming Efficiency Through Mesenchymal‐to‐Epithelial Transition and Stemness Markers

Abstract

The stiffness of hydrogels has been reported to direct cell fate. Here, we found that the stiffness of hydrogels promotes the reprogramming of mouse embryonic fibroblasts into induced pluripotent stem cells (iPSCs). We prepared cell culture substrates of various stiffnesses (0.1, 1, 4, 10, and 20 kPa) using a polyacrylamide hydrogel. We found that culture on a soft hydrogel plays an important role in inducing cellular reprogramming into iPSCs via activation of mesenchymal-to-epithelial transition and enhancement of stemness marker expression. These results suggest that physical signals at the interface between cell and substrate can be used as a potent regulator to promote cell fate changes associated with reprogramming into iPSCs, which may lead to effective and reproducible iPSC-production.

On the role of mechanics in driving mesenchymal-to-epithelial transitions

“…Knock-down of these kinases that disrupt the F-actin network, including TESK1 and LIMK2 that phosphorylate the actin-binding protein cofilin [102], induce dramatic cellular phenotypic changes, turns fibroblasts into epithelial cells, and enhances iPSC efficiency. Similarly, MEFs cultured on a soft hydrogels increased MET and iPSC formation [103]. Together these data suggest a key role for cellular mechanics and tension maintained by cytoskeleton in regulating MET during iPSC formation.…”

Substrate-mediated reprogramming of human fibroblasts into neural crest stem-like cells and their applications in neural repair

“…Topographical cues may affect the direct reprogramming of fibroblasts into induced neurons [18]. The stiffness of hydrogels could regulate the cellular reprogramming efficiency [19]. However, the influence of flat culture substrates on cellular reprogramming has received little attention so far.…”

Biophysical regulation of cell reprogramming

“…0.1 kPa polyacrylamide gels induced MET and increased expression of Oct4 and Sox2 on adhered cells during reprogramming relative to the 20 kPa gels [27]. Similarly in an earlier study, softening of substrate stiffness has been shown to achieve reprogramming in HEK-293 cells simply through changes in actin force without any transcription factors [28].…”