Fig. 3: Myogenic differentiation under mechanical stretching.
From: Shaping 3D minimal model tissues with mechanical constraints to orchestrate muscle differentiation
a Myogenic marker expression levels, determined by quantitative reverse transcription polymerase chain reaction (RT-qPCR). Relative mRNA expression levels are measured for CTL spheroids at D1 and D3, and for stretched aggregates at D3. RPLP0 was used as a housekeeping gene. At least N = 6 for CTL spheroids with 4 independent experiments, N = 4 to 8 in 4 to 8 independent experiments for stretched aggregates at D3 for each gene. Mean ± standard error on the mean (SEM) is indicated and each point indicates an independent experiment. Myod1, myogenic differentiation 1, Myog, myogenin, Myf6, myogenic factor 6, Tnnt, troponin T, Ckm, creatin kinase M-type, Myh, myosin heavy chain; mRNA, messenger RNA; D, day. b Western Blot of CTL spheroids at D1 and D3 and stretched aggregates at D3 for myosin heavy chain, MHC and myogenic factor 6, Myf6. α-tubulin is used as a loading protein control. c Representative confocal image of a stretched aggregate at D3. Immunostaining of MHC (red) is superimposed with nuclei (blue). Scale bar = 20 μm. d Representative confocal image of a stretched aggregate at D2. Both F-actin (green) and nuclei (blue) are visible. Scale bar = 20 μm. One can notice long actin filaments and the presence of fused (multinucleated) cells. e Representative confocal images of a spheroid and stretched aggregates at D3. Immunostaining of troponin T protein (magenta) is superimposed with nuclei (cyan) in a transverse cryosection of a CTL spheroid (left), of a stretched aggregate (middle) and in a longitudinal cryosection of a stretched aggregate. Scale bar = 200 μm.
