Fig. 5. Bovine milk-derived EVs increase metastasis of breast cancer cells.

a Schematic representation of intramammary fat pad injections (IMFP) of 4T1.2 breast cancer cells and treatment timeline with milk-derived EVs. b Primary breast tumor volume in immunocompetent mice treated with milk-derived EVs (low dosage: 25 mg/kg; higher dosage: 50 mg/kg; n = 4). c Harvested primary tumor size from mice treated with PBS and milk-derived EVs is shown. d Quantification of ex vivo imaging of lungs from metastatic breast cancer models treated with PBS and milk-derived EVs. Total lung flux to quantify breast cancer metastasis to lungs is depicted (n = 4). e Relative metastatic tumor burden is depicted (n = 4). f Pathological examination of lung metastases on the surface of the tissue. g Primary breast tumor volume in immunocompetent mice orally administered with PBS, sonicated and intact milk-derived CM EVs (higher dosage: 50 mg/kg; n = 5). h Pathological examination of lung metastases and quantification of visible metastases on the surface of the lungs from mice orally administered with PBS, sonicated and intact milk-derived CM EVs (n = 5). i Primary breast tumor volume in immunocompetent mice orally administered with PBS (n = 8) and CM EVs (5 µg; n = 9). j Relative metastatic tumor burden is depicted (n = 3). Mice exclusion reason: for measuring the relative tumor burden, 3 mice lungs were used randomly; the remaining mice lungs were used to isolate mCherry positive cells for a different experiment. All data are represented as mean ± s.e.m. Statistical significance was determined by unpaired two-tailed t-test.