Fig. 4

Immunofluorescence analysis of early-stage Caco-2 cell tumors. A-E Immunofluorescence analysis of various types of Caco-2 cell tumors one month after transplantation. A Tumor foci were predominantly located within the region corresponding to the original epithelial layer, with their basal side supported by a collagen layer. However, small invasive foci were occasionally observed in the submucosa beyond the collagen layer, as indicated by the white arrowhead. B Distribution of vimentin (VIM)-positive mesenchymal cells surrounding Caco-2 cell tumors. C Cell proliferation analysis in different types of Caco-2 cell tumors by EdU staining. Various types of mScarlet-positive Caco-2 cell tumors, such as flat monolayers (middle, FM), ACF-like (top, ACF), protruded (top, PR) and invasive tumors (top, IV), exhibited a significant presence of EdU-positive tumor cells. By contrast, normal epithelial cells adjacent to tumors were deficient in cell proliferation (top and middle, white arrowheads). (bottom) Control mouse cecum with many proliferating normal epithelial cells. D Positivity of Caco-2 tumors for the cell proliferation marker, Ki-67. Ki-67-positive cells were present abundantly in tumor regions, whereas adjacent normal epithelial cells were mostly quiescent. E, F Staining of differentiated goblet cell granules with rhodamine-UEA-I. Tumors were deficient in UEA-I-positive cells, while they were abundant in the adjacent normal epithelium. F Percentage of UEA-I-positive goblet cells in the normal mucosa and tumor-adjacent mucosa (mean ± SEM). *, p < 0.05 (Student’s t test)