Fig. 8: Collagen XII knockdown reduces metastasis in vivo.

a Schematic representation of orthotopic co-implantation of cancer cells and CAFs (knockdown study: shScr and shColXII; overexpression study eGFP-VPR and ColXII-VPR) injected in a 1:3 ratio respectively. b Bulk modulus as determined by unconfined compression analysis of excised primary tumours generated by co-implantation of cancer cells with control (shScr) and collagen XII knockdown (shColXII#1, shColXII#3) CAFs. Mean ± SD, n = 7 biologically independent samples, *p < 0.05, shScr vs shColXII#1 p = 0.036, shScr vs. shColXII#3 p = 0.045; Kruskal–Wallis test with Dunn’s multiple comparisons test. c Quantitation of the number of mice with or without observed metastases in H&E sections in the knockdown study (n = 7 mice per group). d Quantification of the mean number of metastases normalised to the total lung area in mice with metastases present in the knockdown study (n = 7 shScr, n = 4 shColXII#1 and n = 3 shColXII#3 mice with metastases present). *p = 0.0278, **p = 0.0062 two-sided t test with Welch’s correction. Data are presented as mean ± SD. e Representative images of (left) collagen XII IHC (scale bar = 50 µm) and (right) multi-photon second harmonic generation signal (scale bar = 40 µm) acquired from n = 7 primary tumours in the knockdown study. f Quantification of collagen XII-positive area in primary tumours from the knockdown study (n = 7 mice per group). **p < 0.01 One-way ANOVA with a Dunnett’s multiple comparisons test. Data are presented as mean ± SD. g Representative western blot of collagen XII expression in primary tumours at endpoint in the knockdown study of n = 2 biologically independent experiments. *Denotes non-specific band. Vinculin is a loading control. h Quantification of collagen I fibre bundle width in primary tumours from the knockdown study. Data are presented as mean ± SD, n = 7 biologically independent experiments, ****p < 0.0001, one-way ANOVA with Dunnett’s multiple comparisons test. i Quantification of pMLC2 staining positivity (% of tumour area) in primary tumours. Data are presented as mean ± SD, n = 7 mice per group; 3 fields of view per tumour, *p = 0.018, ***p = 0.0006, two-sided Mann-Whitney test. Source data are provided in the Source data file.