Fig 1: Characterization of nuclear indentation on micropatterned surfaces. a) Representative fluorescence microscopy images of MSC nuclear phenotypes on 5 × 5 micropatterned substrates, depicting actin (green) and nuclei (blue). Scale bar, 20 µm. b) Confocal z-stacks demonstrating nuclear “hole” extending through the nuclear compartment. c) Schematic representation of two potential types of nuclear deformation, either nuclear indentation (in which the micropillar deforms the nuclear envelope, thus displacing genetic material), or nuclear perforation (in which a new nuclear envelope is formed around the micropillar to produce a doughnut shaped nucleus). Cytosol is shown in red, genetic material in blue, and nuclear envelope in green. d). Representative images of lamin A/C (green), DNA (blue), and actin (red). Asterisks demonstrate regions of lamin A/C accumulation on the surface of micropillars and corresponding regions of DNA displacement respectively. Arrows denote regions of peripheral nuclear envelope accumulation of lamin A/C. Scale bar, 5 µm. e) Quantification of lamin A/C staining on flat and the 5 × 5 substrates, respectively. f) Representative false-coloured FIB/SEM image of MSCs cultured on 5 × 5 substrates. Micropillars are colored red, with nuclei colored blue. Scale bar, 5 µm g) RT-PCR and h) Western blotting analysis of LMNA and lamin A/C expression profiles respectively across flat, 1 × 5, 5 × 5 and 10 × 5 micropillars. Data was collected from three independent replicates across three MSC donors. RT-PCR data is presented as mean ± SD relative to TCP control samples.