Fig. 7

Notch signal blockade abrogates the contractile phenotype and promotes the secretory phenotype in vSMCs. a-c vSMCs-DA were treated with DMSO or DAPT for 48 h. The mRNA level of Hey1 and contractile-related genes (a) and secretory-related genes (b) was determined using qRT-PCR. The concentration of TNFα and Cxcl10 in the supernatant was measured using ELISA (c) (n = 5). d vSMCs-DA were treated with DMSO or DAPT for 48 h. The protein level of total and phosphorylated IκBα and nuclear p65 was determined using western blotting (n = 3), with lamin A/C as an internal control. e vSMCs-DA were treated with DMSO or DAPT for 48 h. Cell migration was evaluated by Transwell assay (top) (n = 5). Cell proliferation was determined using EdU incorporation (middle) (n = 3). For adhesion, cells were labeled with Dil and incubated with bEnd.3 cells for 2 h, and evaluated after washing (n = 5). f vSMCs-DA cells were treated with DMSO or DAPT for 48 h, followed by collagen gel contraction assay in the presence of DMSO or DAPT, respectively (n = 5). g vSMCs-DA were treated with DMSO or DAPT for 48 h, followed by culturing with normal medium for 48 h. CM was collected and used to culture LLC or B16-F10 cells in a Transwell system for the invasion assay. The invasion of tumor cells was evaluated after 24 h using crystal violet staining (n = 7 for LLC; n = 10 for B16-F10). h LLC cells were treated with vSMCs-derived CM as in (g). Cell proliferation was evaluated using the EdU incorporation assay (n = 5). Bars = means ± SD. *, P < 0.05; **, P < 0.01; and ***, P < 0.001