RT Journal Article SR Electronic T1 An immortalised mesenchymal stem cell line maintains mechano-responsive behaviour and can be used as a reporter of substrate stiffness JF bioRxiv FD Cold Spring Harbor Laboratory SP 269225 DO 10.1101/269225 A1 Asier Galarza Torre A1 Joshua E. Shaw A1 Amber Wood A1 Hamish T. J. Gilbert A1 Oana Dobre A1 Paul Genever A1 Keith Brennan A1 Stephen M. Richardson A1 Joe Swift YR 2018 UL http://biorxiv.org/content/early/2018/02/21/269225.abstract AB The mechanical environment can influence cell behaviour, including changes to transcriptional and proteomic regulation, morphology and, in the case of stem cells, commitment to lineage. However, current tools for characterizing substrates’ mechanical properties, such as atomic force microscopy (AFM), often do not fully recapitulate the length and time scales over which cells ‘feel’ substrates. Here, we show that an immortalised, clonal line of human mesenchymal stem cells (MSCs) maintained responsiveness to substrate mechanics observed in primary cells, and could be used as a reporter of stiffness. MSCs were cultured on soft and stiff polyacrylamide hydrogels. In both primary and immortalised MSCs, stiffer substrates promoted increased cell spreading, expression of lamin-A/C and translocation of mechano-sensitive proteins YAP1 and MKL1 to the nucleus. Stiffness was also found to regulate transcriptional markers of lineage. A GFP-YAP / RFP-H2B reporter construct was designed and virally delivered to the immortalised MSCs for in situ detection of substrate stiffness. MSCs with stable expression of the reporter showed GFP-YAP to be increasingly co-localized with nuclear RFP-H2B on stiffer substrates, enabling development of a cellular reporter of substrate stiffness. This will facilitate mechanical characterisation of new materials developed for applications in tissue engineering and regenerative medicine.