Figure 3

Cultured vascular smooth muscle cells from transgenic mice have a TGF-β- activated phenotype. (a) Reporter gene assay for β-galactosidase shows equal chemiluminescence in vSMCs from wild-type and transgenic mice. Transgenic fibroblasts from the same animals were used as a positive control. Data are expressed as mean ± SEM; *P < 0.05 from WT (n = 3) and TG (n = 3) animals. (b) β-Galactosidase immunostaining results comparing vSMCs and fibroblasts from wild-type and transgenic animals show negative staining in all vSMCs and positive staining only in transgenic fibroblasts. Images shown are representative of three independent experiments. (c) vSMCs from transgenic mice show increased expression of smoothelin gene by qPCR. Recombinant TGF-β1 induced smoothelin mRNA in wild-type vSMCs, but the response in cells from transgenic mice was attenuated. Immunostaining confirms constitutive overexpression of smoothelin in vSMCs from transgenic mice. (d) Data for transgelin, a second gene important for vSMC cytoskeletal function, show the same trends. Data are expressed as mean ± SEM; *P < 0.05, **P < 0.001; and are representative of three independent experiments examining four littermates for each condition. Original magnification, ×40.