Maintaining resting cardiac fibroblasts in vitro by disrupting mechanotransduction
Fig 4
Inhibiting ROCK and TGFβRI is most efficient for preventing and reversing the myofibroblast phenotype.
A and B) Schematic illustration of treatment regimen. Red arrows indicate the addition of fresh inhibitor solution every 24h. C and D) Immunostaining for smooth muscle α-actin (SMA) in cardiac fibroblasts treated with blockers of ROCK (Y27), TGFβRI (SB) and calcineurin (CsA). Scale bar 100μm. E and F) Quantification of SMA staining intensity. G and H) Expression of myofibroblast markers actin alpha 2 smooth muscle (Acta2), connective tissue growth factor (Ctgf), collagen (Col) 1a1, Col1a2, Col3a1, lysyl oxidase (Lox) and periostin (Postn), and the marker of resting cardiac fibroblasts, transcription factor 21 (Tcf21). One-way ANOVA as indicated beside gene name and Dunnett’s multiple comparisons test for significant differences compared to the “no treatment” control as indicated in the heat map. N = 3 (E and F), 4 (G) 4–7 (H).