Figure 5. Critical role of YAP in promoting smooth muscle cell (SMC) proliferation and the expression of extracellular matrix (ECM) genes in response to cyclic stretch.

A, Human aortic SMCs (in low serum) underwent cyclic stretch for 24 h. Flow cytometry analysis showing that cyclic stretch induced cell proliferation (indicated by EdU labeling) in cultured SMCs (n=4 biologic repeats). B, Western blotting results showing that cyclic stretch induced ECM protein production in cultured SMCs and YAP expression and dephosphorylation in cultured SMCs that underwent cyclic stretch (n=8 biologic repeats). C, Western blotting quantification showing that cyclic stretch induced a marked dephosphorylating of YAP in cultured SMCs. D, Representative immunofluorescence staining (images and quantification data) showing the cyclic stretch–induced nuclear translocation of YAP (n=3 biologic repeats and 3 replicates). E-F, SMCs were transfected with wild-type (WT) YAP or constitutively-active YAP (S127A) (n=5 biologic repeats). Overexpression of YAP induced cell proliferation (E) and ECM protein production (F). G, Silencing YAP with siRNA prevented ECM protein production. H, LOX mRNA levels (n=5 biologic repeats) were increased by cyclic stretch, and silencing Yap with siRNA partially prevented LOX expression induced by cyclic stretch. Two-way analysis of variance (ANOVA) with Bonferroni’s post hoc test for pairwise comparisons was used in (H). A paired Student’s t-test was used in (C). Data are shown as box and whisker plots with the first quartile, minimum, median, third quartile, and maximum in (A), (C), (D), (E), and (H).