Stem cell differentiation: sticky mechanical memory.

S cientists have been fascinated for decades by the ability to drive differentiation of pluripotent or multipotent stem cells down Waddington's epigenetic landscape — a conceptual framework that explains how genes interact with their surroundings to produce a cell's phenotype. Major efforts have identified a number of soluble biochemical factors that control stem cell plasticity and differentiation. More recently, it has become evident that differentiation is also governed by physical cues (such as substrate stiffness 1 , matrix anchoring 2 and cell shape 3) that the cells sense in their culture environment. Writing in Nature Materials, Kristi Anseth and colleagues now show that lineage-commitment decisions of human mesenchymal stem cells (hMSCs) in response to soluble cues are also influenced by past mechanical environments 4. Anseth and co-authors adapted a classical differentiation assay in which primary-bone-marrow-derived hMSCs cultured on substrates with different bulk stiffness were treated in a mixed osteogenic and adipogenic differentiation medium 3. Under these culture conditions, hMSCs adherent to soft substrates (Young's modulus E < 2 kPa) typically commit to the adipogenic lineage, whereas those grown on stiff substrates (E > 10 kPa) prefer to differentiate into osteoblasts. The authors first added hMSCs to tissue culture polystyrene (TCPS, E ~3 GPa) for 10 days, and then transferred the cells to soft substrates before exposing them to mixed (osteogenic and adipogenic) media. Surprisingly, the cells spread on the soft substrates and stained positive for alkaline phosphatase (ALP), an early osteoblast marker. The authors then assessed the nuclear translocation of transcription factors known to be important for osteogenic differentiation 5 , namely yes associated protein (YAP), transcriptional co-activator with PDZ binding domain (TAZ), and Runt-related transcription factor 2 (RUNX2). Even in the absence of differentiation factors, these transcription factors exhibited persistent activation (nuclear localization); indeed, the longer the cells had adhered to stiff surfaces before they were transferred to soft environments, the larger the number of cells with YAP/TAZ and RUNX2 in the nucleus, a situation that was maintained even when the actin cytoskeleton was temporarily disrupted with the small molecule latrunculin A. These experiments suggested that hMSCs retain mechanical memory of the stiffness of the substrates they have been cultured on. Still, the enzymatic transfer of cells from TCPS to compliant substrates can introduce potential artefacts. To circumvent these, Anseth and co-authors took advantage of a previously developed photodegradable poly(ethylene glycol) (PEG) hydrogel 6 because it could be softened …