Late-stage iCMCs show better cardiac function measured by PIV method.

(A) Differentiated and beating cardiomyocytes visualized by high frame rate (1/10 sec) video microscopy as in S1 Movie. (B) Velocity field snapshot of the area marked by a white rectangle in panel A. Red lines represent PIV-calculated displacements, their end point is marked by black dots. Locations without red lines were stationary during the 0.1 sec long time interval. (C) The beat pattern (PIV-derived displacements, measured relative to a stationary reference state and averaged over the entire field of view) indicates that CMC contractility is periodic with a steady waveform. (D) Time development of a tissue culture area that initially consisted three aggregates. Microscopic fields (D1, D3, D5, and the video microscopy is given in S2, S3 and S4 Movies, respectively) and characteristic beat patterns (D2, D4, D6) are shown for three consecutive days after the onset of beating at day 7. Red, green and blue curves correspond to the areas marked as 1, 2 and 3 in the microscopic fields, respectively. During the time course of three days, the aperiodic and asynchronous beat patterns consolidate into a periodic and synchronous one. (E) Average profiles of contractile peaks are shown for beat patterns characterizing area 3 in panel D from day 8 [blue], day 9 [green] and day 10 [red]. Blue, green and red colors indicate progressively older cultures. As cardiomyocytes mature, the contractile periods become shorter. (F) Average period lengths obtained from 8 different cultures at various days in vitro. As cultures mature, the beating frequency tends to increase up to 1.4 Hz, at 37 days after the onset of beating. Representative images are from three repeated experiments.