Fig. 2: Contractile forces in C2C12 micro-tissues.

a Bright-field image of a C2C12 micro-tissue focused at the top of the PDMS pillars. The orange arrowhead indicates a high-contrast point that is used for particle image velocimetry analysis. Scale bar: 200 µm. b Kymograph of the region indicated in (a) in response to active tissue contraction. Scale bars: 10 µm and 250 ms. c Calibration curves to determine the spring constants of the PDMS pillar. The spring constants were measured from the force-deflection relationship of pillars at different heights and for PDMS with different crosslinker-to-base ratios. Dots represent slopes of individual force-deflection measurements, and lines represent fits to the data points using the Euler–Bernoulli beam equation. d Static and active contractility of the micro-tissue shown in (a). The stimulation frequency was 1 Hz. e Static contractile forces of C2C12 micro-tissues after 7 days in culture. One group of micro-tissues was prepared as usual (i.e., using ITS differentiation medium), and one group was kept in culture medium for the entire 7 days (instead of switching to ITS medium after 24 h) so that the myocytes did not differentiate. We measured the static contractile forces under control conditions (Ctrl; 1 vol% dimethylsulfoxid), after adding either 10 µM cytochalasin-D (cyto-D), or 100 µM blebbistatin (Blebb) to the medium for 1 h, or after adding Triton X-100 (Tx-100) to the medium for 10 min; n.s., P ≥ 0.05; ★★★, P < 0.001. f Same as (e) for active contractile forces. g Contrast-enhanced brightfield images of exemplary micro-tissues before and after treatment with cyto-D. One micro-tissue was cultured in ITS medium and the other in culture medium before drug treatment. Scale bar: 100 µm.