Fig. 4
CSCC-secreted exosomal miR-221-3p promotes lymphangiogenesis and lymphatic metastasis in vivo. a popliteal lymph node metastasis model was established in nude mice by inoculating the footpad with Siha/anti-221-3p (5 × 106) stably expressing mCherry. When footpad tumor size reached 50 mm3, exosomes (10 µg) secreted by Ect1/miR-NC, Ect1/miR-221-3p, Siha/anti-NC, or Siha/anti-221-3p were then injected into the center of the tumors (n = 3/group, repeated twice) twice a week. After five injections, primary tumors reached a comparable size of ~ 150 mm3, and then footpad tumors and popliteal LNs were collected for study. a Staining of miR-221-3p and LYVE1 in serial sections of mice footpad tumors. Representative micrographs of positive staining are shown (left). The tumor cells are indicated by black arrows. The lymphatic vessels are indicated by red arrows. The correlation between miR-221-3p levels and PLVD was statistically analyzed (right). Scale bar, 20 µm. b Staining of mCherry in popliteal LNs from mice treated with the indicated exosomes. Representative micrographs are shown. Metastasis-positive LNs were identified by staining for cancer cell-expressed mCherry. Scale bar, upper panel, 200 µm; lower panel, 20 µm. c The ratio of metastasis-positive to total dissected popliteal LNs from mice treated with the indicated exosomes. ***, P < 0.001
