Genome-wide impact of substrate stiffness on embryonic stem cell pluripotency and the signalling pathways controlling it

GOAL: The goal of this study was to assess the impact of substrate stiffness on the gene expression of pluripotent embryonic stem cells, in different heterogeneous conditions. Embryonic stem cells cultured on tissue culture plastic have heterogeneous gene expression when cultured in serum+LIF. We investigated whether culturing cells on soft fibronectin-coated StemBond hydrogels has genome-wide effects and in particular improves the homogeneity of naive pluripotent gene expression. To assess the interplay with multiple signalling pathways, we used multiple pluripotent conditions or differentiating conditions, by adding or removing the MEK inhibitor PD03, and the cytokine LIF. Embryonic stem cells were cultured on fibronectin-coated StemBond hydrogels and tissue culture plastic for either 24hrs or 48hrs. Culture conditions were multiple: serum+LIF (24hrs), serum+LIF (48hrs), serum without LIF (48hrs), serum+PD03 (48hrs), serum+LIF+PD03 (48hrs). 3 replicate samples per condition were sequenced.