Fig 2: CBR2 (CBR2-green) confocal microscopic results, showing images of equine primary skin-derived cultures of keratinocytes (A-H) and fibroblasts (I-P). CBR2 immunoreactivity in keratinocytes (A and E) and fibroblasts (I and M). For keratinocyte and fibroblast immunophenotyping, co-labeling with specific epithelial pan-cytokeratin (CK-red) (B) and fibroblast vimentin (Vim-red) (N) markers was performed (yellow arrows indicate double positive cells). In turn, inverse staining of Vim-red on keratinocytes (F) and CK-red on fibroblast (J) was additionally performed to distinguish both primary cell types (gray arrows indicate single positive cells). The DAPI (blue) was used for cell nucleus counterstaining; images were taken under 40x magnification; scale bars = 20 µm

Fig 3: CBR1 (CBR1-green) confocal microscopic results, showing images of equine primary skin-derived cultures of keratinocytes (A-H) and fibroblasts (I-P). CBR1 immunoreactivity in keratinocytes (A and E) and fibroblasts (I and M). For keratinocyte and fibroblast immunophenotyping, co-labeling of CBR1 with specific epithelial pan-cytokeratin (CK-red) (B) and fibroblast vimentin (Vim-red) (N) markers was performed (yellow arrows indicate double positive cells). Inverse staining of Vim-red on keratinocytes (F) and CK-red on fibroblast (J) with CBR1 was additionally performed to distinguish both primary cell types (gray arrows indicate single positive cells). The DAPI (blue) was used for cell nucleus counterstaining; images were taken under 40x magnification; scale bars = 20 µm

Fig 4: Snail suppresses tumor progression and is associated with tumor immunity. a Western blot of HM-1-control and HM-1-shSnail cells. CDH1: E-Cadherin, VIM vimentin. b Invasion of HM-1-control and HM-1-shSnail cells; n = 5. c Wound healing of HM-1-control and HM-1-shSnail cells. Data represent the percentage of wound width (24 h) relative to that at 0 h; n = 6. d Growth curves of HM-1-control and HM-1-shSnail subcutaneous tumors in immunocompetent mice. P-values represent significance between two groups at day 31; n = 6. e Growth curves of HM-1-control and HM-1-shSnail subcutaneous tumors from immunodeficient nude mice; n = 6; NS no significant difference between two groups at day 23. *P < 0.05, **P < 0.01, and ***P < 0.001 (one-way ANOVA with Tukey’s multiple comparisons test in b–e). Averaged data are presented as the mean ± SEM

Fig 5: Direct targeting of miR-17-5p to the 3'UTR of VIM mRNA.a In LoVo and HT29 cell lines, the interaction between vimentin and miR-17-5p was assessed by RNP IP as described in the “Materials and methods” section. The expression of vimentin in the miR-17-5p IP materials was determined by RT-qPCR. b Schematic representation shows the construction of wild-type or mutant, or perfectly matched vimentin 3'UTR in the vector. c Schematic representation shows the plasmid containing the vimentin 3'UTR in luciferase assay. d Luciferase assays show a decrease in reporter activity after co-transfection with miRNA17 mimics and the wild-type vimentin 3'UTR, and the perfect match in LoVo cells. Mutant of vimentin 3'UTR did not show a decrease. The bars represent mean ± SD. **P < 0.001. All the experiments were repeated three times.