Fig. 3.

LncRNA VIM-AS1 competes with ZEB1 for miR-655 binding to modulate bladder cancer metastasis. a, b T24 and RT24 cells were stimulated with 0, 2, 5 and 10 ng/ml TGFβ1 and examined for the protein (upper) and mRNA (under) expression levels of ZEB1. c T24 cells were transfected with si-VIM-AS1 and RT24 cells were transfected with VIM-AS1 vector and examined for the protein (upper) and mRNA (under) levels of ZEB1. d The expression of miR-655 was determined in 35 paired bladder cancer and noncancerous tissue samples. e The expression of miR-655 was analyzed in 15 nonmetastatic and 20 metastatic bladder cancer tissues. f miR-655 was overexpressed or inhibited in T24 and RT24 cells by transfection of miR-655 mimics or miR-655 inhibitor, as confirmed by real-time PCR. g Wild-type and mutant ZEB1 3′UTR and VIM-AS1 luciferase reporter vectors were constructed as described. h The wt-VIM-AS1 or mut-VIM-AS1 vector were co-transfected into 293T cells with miR-655 mimics or miR-655 inhibitor and the luciferase activity was determined. i The wt-ZEB1 3′UTR or mut-ZEB1 3′UTR vector was co-transfected into 293T cells with si-VIM-AS1, and the luciferase activity was determined. j–l T24 cells were co-transfected with miR-655 inhibitor and si-VIM-AS1 and examined for the level of VIM-AS1 (j), miR-655 (k) and expression of ZEB1 protein and mRNA (l). m–o T24 cells were co-transfected with miR-655 mimics and VIM-AS1 vector and examined for the level of VIM-AS1 (m), miR-655 (n) and expression of ZEB1 protein and mRNA (O). *P < 0.05, **P < 0.01 compared to control, si-NC, NC, NC mimics, NC inhibitor, inhibitor NC + si-NC or mimics NC + vector group. ^##P < 0.01 compared to miR-655 inhibitor + si-NC or miR-655 mimics + vector group