Fig. 3

Comparison of the phenotype of colorectal cancer (CRC) cells at depth 4 and depth 5. A, B Representative topoisomerase 1 (TOPO1)-immunohistochemistry (IHC) in CRC at depth 4 (A) and depth 5 (B). 90.6% TOPO1-positive (TOPO1⁺) CRC cells proliferated at a depth of 4 (A, 164 TOPO1⁺ cells/181 CRC cells). Tumor buds were negative for TOPO1 at depth 5 (B, yellow arrows). C Boxplot of TOPO1 positivity in CRC cells at depth 4 and depth 5 (n = 20). In TOPO1-IHC, two areas (depth 4 and depth 5) were selected from each of the twenty cases and counted. The TOPO1 positivity rate in CRC cells at depth 5 was lower than that at depth 4 (depth 4, 27.5 ± 32.6%; depth 5, 5.7 ± 6.7%; *, P < 0.05). D, E Representative CD205-IHC in CRC at depth 4 (D) and depth 5 (E). 95.9% CD205-positive CRC cells proliferated at a depth of 4 (D, 186 CD205⁺ cells/194 CRC cells). Tumor buds were negative for CD205 at depth 5 (E, yellow arrows). F, G Representative double immunostaining for AE1/AE3 and Ki-67 in CRC at depth 4 (F) and depth 5 (G). 43.8% Ki-67-positive CRC cells proliferated at a depth of 4 (F, 92 Ki-67⁺ cells/210 AE1/AE3⁺ cells). Tumor buds were negative for Ki-67 at depth 5 (G, yellow arrows). H Boxplot of Ki-67 positivity in CRC cells at depth 4 and depth 5 (n = 5). In double immunostaining for AE1/AE3 and Ki-67, three areas were selected from each of the five cases and counted. The Ki-67 positivity rate in CRC cells at depth 5 was lower than that at depth 4 (depth 4, 24.2 ± 12.1%; depth 5, 5.3 ± 5.3%; **P < 0.01). Bars, 50 μm