Fig. 4

ZEB2 induces a transition towards a quiescent/slow cycling and mesechymal-like state in CRC xenografts. a Left: Xenograft volume of SW480 cells transduced with pLenti-GFP (Vector, black line/triangles) or with pLenti-GFP-ZEB2 (ZEB2, red line/squares). Mean ± SEM, 6 tumors/group. **P < 0.01 (two-tailed t test). Middle: representative confocal images of Vector- and GFP-ZEB2-transduced SW480 xenografts stained with anti-ZEB2 (red) and anti-Ki67 (white) antibodies. Scale bar 60 μm. Right: quantification of Ki67-, ZEB2- and GFP-positive cells performed on 3 sets composed of 5 fields/group. *P < 0.05 and **P < 0.01. Mean ± SD (two-tailed t test, n.s. = not significant). AU, arbitrary units. b Cell cycle analysis of GFP⁺ cells from Vector- and ZEB2-transduced tumors. c qRT-PCR analysis of GFP⁺ cells from Vector- and ZEB2-transduced tumors, n = 3 pools of 2 tumor each. *P < 0.05 and **P < 0.01 (two-tailed t test). Mean ± SD. d mRNA levels of cell cycle genes in GFP⁺ cells from Vector- and GFP-ZEB2-transduced tumors. Mean ± SD, n = 3. *P < 0.05 (two-tailed t test). e Volume of xenografts expressing pLenti-GFP (Vector, black line) or GFP-ZEB2 (ZEB2, red line), untreated or treated (Vector, gray line/triangles and ZEB2 yellow line/diamonds) with 5-fluorouracil plus oxaliplatin (5FU + OXA). Mean ± SEM, 6 tumors per group. *P < 0.05 and **P < 0.01 from one-way ANOVA and Bonferroni post-tests. f Upper panels: immunoblot analysis of ZEB2, CRAF pS338 and ASK1 pS83 on whole tumor lysates derived from SW480 xenografts. Lower panels: densitometry analysis of western blots, n = 3, *P < 0.05, **P < 0.01 and ***P < 0.001 (two-tailed t test). g Left: immunoblot analysis of EMT-related proteins on whole xenograft lysates. Every sample is a pool of 2 tumors. Right: densitometry analysis, n = 3, *P < 0.05, **P < 0.01 and ***P < 0.001 (two-tailed t test)