Fig. 2

EV-packaged circTLCD4-RWDD3 promotes lymphangiogenesis and LN metastasis in NSCLC in vitro and in vivo. a Representative images and quantification of the tube formation and Transwell migration of HLECs treated with circTLCD4-RWDD3-downregulating or -upregulating A549 cell-derived EVs. Scale bars, 100 µm. b Schematic diagram of the construction of a nude mice popliteal LN metastasis model. Schematic was created with BioRender (www.biorender.com). c Representative images and quantification of bioluminescence of the popliteal LN metastasis in nude mice (n = 12 per group). d Representative image of the footpad primary tumor and popliteal metastatic LN in a nude mouse. e Representative bioluminescence image of excised popliteal LNs from the nude mice (n = 12 per group). f Representative anti-mCherry IHC images of nude mice popliteal LNs (n = 12 per group). Scale bars, 500 µm (black); scale bars, 100 µm (red). g The percentage of metastatic popliteal LN in the nude mice (n = 12 per group). h, i Representative IHC images and percentages of LYVE-1-indicated lymphatic vessel density in intratumoral (h) and peritumoral region (i) of footpad primary tumor tissues. Scale bars, 50 µm. The statistical difference was assessed with one-way ANOVA followed by Dunnett tests in (a); and unpaired Student’s t-test in (a, c, h, and i); and Chi-square test in (g). Error bars show the SD from three independent experiments. *P < 0.05; **P < 0.01