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. 2022 Mar 11;11(9):1925–1936. doi: 10.1002/cam4.4533

FIGURE 2.

FIGURE 2

Cooperation of Smad4 and p53 in regulating normal intestinal mucosal growth. VSP, Villin‐Cre; Smad4F / F ; Trp53F / F . VS, Villin‐Cre; Smad4F / F . VP, Villin‐Cre; Trp53F / F . (A) Villous length of normal intestinal mucosa from each genotype mice (n = 5) at 20 weeks of age based on H&E staining slides. (B) BrdU staining results in BrdU‐labeled cryptic cells of intestinal mucosa from each genotype (n = 5). Mice were intraperitoneally injected with BrdU at 20 mg/kg body weight. Two hours later, the intestinal mucosa was collected. (C) The percentage of PCNA‐positive cells in cryptic cells of normal intestinal mucosa from each genotype mice (n = 5) at 20 weeks of age. (D) p21 mRNA expression in mouse normal intestinal epithelium from each genotype mice (n = 5) at 20 weeks of age based on quantitative real‐time PCR analysis. (E) mRNA expression of p21 in human colorectal cancer TCGA dataset, according to Smad4 and p53 status. (F) The percentages of p21‐, p27‐, and p15‐positive cells in cryptic cells from 10‐week‐old Villin‐Cre; Smad4F / F ; Trp53F / F (n = 5) and Villin‐Cre‐negative control mice (n = 5) after 24 h of 10 Gy of IR treatment based on IHC staining results. (G) Representative IHC images for p21 expression in IR‐treated intestinal mucosa from Villin‐Cre; Smad4F / F ; Trp53F / F and Villin‐Cre‐negative control mice. Red dotted lines indicate crypt cells. (H) Representative images for TUNEL assay in IR‐treated intestinal mucosa from of Villin‐Cre; Smad4F / F ; Trp53F / F and Villin‐Cre‐negative control mice. Nucleus, blue (DAPI); Apoptotic cells, green. (I) Quantitative real‐time PCR analysis for the measurement of etoposide‐induced p21 mRNA expression in primarily cultured Villin‐Cre; Smad4F / F ; Trp53F / F intestinal cancer cells (primary #1) after the restoration of Smad4 and/or p53 using lentiviral system. (Right) Western blotting analysis. BrdU, bromodeoxyuridine; H&E, hematoxylin and eosin; IHC, immunohistochemistry; IR, ionizing radiation; NS, No significance; PCNA, proliferating cell nuclear antigen. *p < 0.05