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. 2017 Jun 19;36(42):5885–5896. doi: 10.1038/onc.2017.194

Figure 2.

Figure 2

Nuclear localization of p53 in the invasive intestinal tumor cells. (a) Representative photographs of immunohistochemistry for p53 in small intestinal (SI) polyps of the indicated genotype mice. The insets show enlarged images. The arrowheads indicate p53 positive tumor cells. Bars, 200 μm. (b) Representative photographs of fluorescence immunocytochemistry for p53 (green), nuclear counterstaining with DAPI (blue) and merged images (right) of AP-MM6 cells transfected with the wild-type (wt) p53 expression vector (middle and bottom) and control (top). The arrowheads indicate AP-MM6 cells showing both nuclear and cytoplasmic p53 localization. Bars, 50 μm. (c) The ratio of AP-MM6 cells with cytoplasmic p53 distribution after transfection of 0.01 μg (left) or 0.05 μg (right) of wild-type p53 expression vector relative to the control level (mean±s.d.). Asterisks, P<0.05. (d) Relative staining intensity of nuclear p53 in AP-MM6 cells with or without cytoplasmic distribution of p53 after wt p53 vector transfection (blue and red, respectively). Each dot indicates a single cell. Asterisk, P<0.05. (e) Representative photographs of immunohistochemistry for p53 in the invasive region of ApcΔ716 Trp53+/R270H mouse intestinal tumors with low-power magnification (left top) and an enlarged image (left bottom). The arrows indicate tumor cells with nuclear-accumulated p53. Bars, 200 μm. The relative ratio of tumor cells with nuclear-accumulated p53 in the indicated genotypes is shown (mean±s.d.) (right). Asterisk, P<0.05. (f) An LOH analysis for Apc and Trp53 by LMD-based genomic PCR in the non-invasive (Non-inv) and invasive (Inv) areas of ApcΔ716 Trp53+/R270H intestinal tumors and normal villi as the controls. The mutant Trp53 and wild-type Trp53-specific bands are indicated as ‘mutant’ and ‘wt’, respectively. Genomic DNA of Trp53 wild-type mouse tissue and AP-MM6 were used for positive control of wt and mutant Trp53, respectively. Note that wild-type (wt) Apc was lost in both non-invasive and invasive tumor cells, whereas wild-type Trp53 is remained in these cells.