Figure 1. Excessive angiogenesis occurred in the peri-necrotic tumor region.

4T1 breast cancer cells were orthotopically inoculated into the fat pad of 4th breast of BALB/c mice. 28 days later, the whole tumor tissue from untreated mouse was extracted for further analysis. (A) Representative image (Immunological Histological Chemistry) for showing CD31⁺ vessels distributed in peri-necrotic region, transitional region and the non-necrotic region. Scale bar: 500 μm. (B) The color-reverse image and (C) regional distribution map of panel (A) for revealing the discrepancy of angiogenic ability of vessels distributed in different tumor regions. In the area adjacent to tumor necrosis, vessels exhibited the abnormally high abilities of sprouting and branching. Red tri-angle indicates vessels with decreased CD31 expression in the edge region of tumor necrosis. Scale bar: 500 μm. The peri-necrotic and non-necrotic regions were surrounded by red and yellow dotted lines, respectively. Quantification of (D) microvessel density (MVD, No. of vessels per mm²) and (E) vascular branch density (No. of vascular branches per vessel) of regions having different distance from tumor necrosis (n = 10). Spearman analysis for evaluation of the linear correlation between percentage of tumor necrotic area and MVD of (F) the whole tumor area, (G) peri-necrotic region (PNR), (H) transitional region (TR) and (I) non-necrotic region (NNR). Quantitative data are indicated as mean ± SEM. *p < 0.05; **p < 0.01; ***p < 0.001; ns indicates no significant (P > 0.05).