Fig 1: CXCL5-activated neutrophils induce lymphangiogenesis.A Chemotaxis of neutrophils induced by conditioned medium from BCa cells. B VEGF-C expression in neutrophils induced by conditioned medium was detected by qRT-PCR. C Neutrophils in footpad tumors in the indicated mouse group were evaluated by HE and IHC staining. Black arrows indicate the infiltrated neutrophils in footpad tumors. Scale bars, 50 µm. D Tube formation and Transwell migration of HLEC cultured with conditioned medium from neutrophils induced by indicated BCa cells. Scale bars, 100 µm. Error bars indicate standard deviations of three independent experiments. *p < 0.05 and **p < 0.01.
Fig 2: circDHTKD1 facilitates LN metastasis by targeting CXCL5 and sponges miR-149-5p in BCa.A A heatmap showing mRNA levels of 30 genes co-expressed with circDHTKD1 based on sequencing analysis. B and C The mRNA and protein expression of CXCL5 in BCa cells with overexpression or knockdown of circDHTKD1. D and E ELISA of CXCL5 in BCa cells with overexpression or knockdown of circDHTKD1. F Images of HE and IHC staining showing LN metastasis and lymphatic vessel quantities in footpad tumors in the indicated mouse group. Scale bars: red, 200 µm; black, 50 µm. G Schematic diagram showing overlapping of binding miRNAs of circDHTKD1 and CXCL5 predicted by miRanda, TargetScan, and starBase. H and I The enrichment of circDHTKD1 in BCa cell lysates with circDHTKD1 probe was detected by qRT-PCR. Relative level of circDHTKD1 was normalized to input. J and K Relative levels of 8 miRNA candidates in circDHTKD1 pull-down complexes were detected by qRT-PCR. L qRT-PCR analysis of circDHTKD1 captured by biotinylated miR-149-5p. M and N FISH showing the location of circDHTKD1 and miR-149-5p in BCa cells. Scale bars, 20 µm. Error bars indicate standard deviations of three independent experiments. *p < 0.05 and **p < 0.01.
Fig 3: CXCL5 reverses the inhibitory effects of miR-432-5p on colorectal cancer cell invasion and migration of SW480 and HT-29 cells. (A) CXCL5 protein expression was detected in SW480 and HT-29 cells after co-transfection of miR-432-5p mimics and pcDNA3.1-CXCL5 (pc-CXCL5) plasmids. (B and C) The effect of CXCL5 on SW480 and HT-29 cell migration as determined by the wound healing assay. (D-F) The effect of CXCL5 on SW480 and HT-29 cell migration and invasion determined by transwell assay. Bar, 200 µm. *P<0.05; **P<0.01. CXCL, C-X-C motif chemokine ligand 5; miR, microRNA; NC, negative control.
Fig 4: circDHTKD1 antagonizes the repression of miR-149-5p on CXCL5.A and B Tube formation and Transwell migration of HLEC cultured with conditioned medium from miR-149-5p-upregulated or control BCa cells. C and D Tube formation and Transwell migration of HLEC cultured with conditioned medium from miR-149-5p-downregulated or control BCa cells. Scale bars, 100 µm. E Predictive binding sequence of miR-149-5p in the 3’UTR of CXCL5. F Luciferase activities of CXCL5-3’UTR co-transfected with miR-149-5p or control mimics. G and H Western blot showing the CXCL5 levels after miR-149-5p overexpression and knockdown in BCa cells. I Western blot showing the effect of miR-149-5p upregulation on circDHTKD1-induced CXCL5 expression in BCa cells. Error bars indicate standard deviations of three independent experiments. *p < 0.05 and **p < 0.01.
Fig 5: miR-145-3p downregulates CXCL5 expression. (a) The diagram of binding sequences of CXCL5-3'UTR-WT/MUT and miR-145-3p. (b) The luciferase activity of EC9706 cells in different treatment groups. (c, d) CXCL5 mRNA and protein expression in varying treatment groups. The difference comparisons are compared with the miR-NC group. **p < 0.01 and ***p < 0.001.
Supplier Page from Abcam for Anti-CXCL5 antibody [EPR4450(2)]