TGFβ-mediated signaling is intact in cultured VICs
(A) Relative expression of TGFβ ligands in rat VICs and thoracic aorta compared with heart muscle RNA.
(B) Relative expression of TGFβ receptors in rat VICs and thoracic aorta compared with heart muscle RNA.
(C) Exogenous hTGFβ stimulation in VICs causes concentration-dependent increases in SMAD3 phosphorylation with peak response reached at 1 h and sustained for at least 48 h.
(D) After 48-h treatment with 10 ng/mL hTGFβ, myofibroblast targets, ACTA2 and Tagln, were upregulated, indicative of enhanced activation of VICs.
(E) Following 1-h treatment with TGFβR inhibitors that included 10 ng/mL hTGFβ stimulation during the last 30 min of compound treatment, the inhibitors caused concentration-dependent decreases in SMAD3 phosphorylation.
(F) Decreases in SMAD3 phosphorylation without hTGFβ stimulation.
All transcriptional data in (A and B) were normalized with housekeeping gene, Hrpt1, and calculated relative to rat heart muscle RNA. Represented data presented in (A–F) are the average of three experiments ± SEM. Statistical comparisons employed one-way or two-way ANOVA followed by Dunnett's test. ∗p ≤ 0.05; ∗∗p ≤ 0.01; ∗∗∗p ≤ 0.005; ∗∗∗∗p ≤ 0.0001.