EcN facilitated anti-invasion and anti-metastasis of TGF-β blockade in tumor-bearing mice and cancer cell xenografted zebrafish. (A) Representative images of the lungs from mice bearing subcutaneous tumor (n = 4-6). The white arrows pointed the metastatic nodules. (B) Histological analysis of the lung sections by H&E staining at 100× (n = 3). (C) Expression of CD31, an angiogenesis marker, in representative tumor sections by immunostaining at 200× (n = 3). (D) Wet weight of the lungs from mice bearing subcutaneous tumor (n = 4-6). (E) Numbers of superficial metastatic nodules in the lung tissues from mice bearing subcutaneous tumor (n = 4-6). (F) Analysis of CD31⁺ cells in tumor tissues from different treated tumor-bearing mice. (G) Quantification of the percentage of xenografts with micrometastasis. (H) Fluorescence images (100×) of zebrafish tail at 4 dpi. (I) Schematic representation of xenotransplantation in zebrafish, 4T1 tumor cell were labeled with Dio and injected into the perivitelline space of zebrafish embryos. All data are shown as mean ± SD. (* P < 0.05, ** P < 0.01)