Figure 6.

TWIST1 enhanced Wnt/β-catenin signaling and regulated expression of vascularization-related genes

A: Luciferase analysis showed that Wnt/β-catenin signaling activity was increased in HEK293STF cells transfected with TWIST1 plasmids. B, C: Western blotting showed that cyclin D1 and phosphorylation of GSK3β proteins increased when TWIST1 was overexpressed in HRECs. Western blotting with anti-Flag and TWIST1 showed successful transfection of TWIST1 plasmids. GAPDH was used as the loading control. Data are presented as mean±standard error of the mean (SEM). Sample size n=4. Data are from one typical experiment of three independent experiments. ^**: P<0.01; *: P<0.05, two-tailed t-test. D: RNA-seq showed that vascularization-related genes were up-regulated when TWIST1 was overexpressed in HRECs. E: RT-qPCR verified RNA-seq results in HRECs for indicated genes. ^***: P<0.001. Sample size n=4. F: RT-qPCR verified that expression of most of TWIST1-targeting genes identified by RNA-seq were increased in the lungs of Tg-Twist1^iEC+ mice. ^***: P<0.001;^**: P<0.01; ^*: P<0.05; ns, no significant difference. Sample size n=4.