Suppression of Apoptosis, Crypt Hyperplasia, and Altered Differentiation in the Colonic Epithelia of Bak-Null Mice  Carrie A. Duckworth, D. Mark Pritchard  Gastroenterology  Volume 136, Issue 3, Pages 943-952 (March 2009) DOI: 10.1053/j.gastro.2008.11.036 Copyright © 2009 AGA Institute Terms and Conditions

Figure 1 Photomicrographs of bak protein expression in C57BL/6 (A) and bak-null (B) distal colonic crypts. Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 2 H&E-stained photomicrographs of C57BL/6 (A) and bak-null (B) distal colon. Bar charts show cell number per hemi-crypt in proximal, middle, and distal thirds of bak-null colon (white) compared with C57BL/6 (black) (C), and cell number per hemi-crypt in distal half of colon of bax-null (hashed) compared with bax-wt (black) mice (D). Significant differences between bak-null and C57BL/6 by t test (*P < .001). Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 3 Apoptotic bodies (black arrows) in the colonic crypt table of C57BL/6 mice after H&E (A) and active caspase-3 immunostaining (C). Apoptotic events per crypt were assessed from H&E-stained (B) and active caspase-3–stained (D) sections of C57BL/6 (black) and bak-null (white) distal colon. The number of nuclei per unit length of colonic crypt table (indicated by the black bar in E) was quantified. Relative number of nuclei per unit length is shown for C57BL/6 (black) and bak-null (white) distal colon (F). Significant differences by t test (**P < .001, *P < .01). Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 4 Mean mitotic index % (A) in C57BL/6 (black) and bak-null (white) distal colon. Cell positional distribution of mitosis (B) in C57BL/6 (solid line) and bak-null (dotted line) distal colon. BrdU-positive cells per hemi-crypt (C) in C57BL/6 (black) and bak-null (white) distal colon. Cell positional distribution of BrdU-labeled cells (D) in C57BL/6 (solid line) and bak-null (dotted line) distal colon. Significant differences by t test (*P < .05), and significantly different cell positions by modified median test are indicated by bars on the cell positional plots. (E) Representative Western blots for cyclin D1, cdk4, p27, p16, p21, and actin in colonic crypt–enriched isolates from untreated and 4.5 hours after 8-Gy, γ-irradiated C57BL/6 and bak-null mice. Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 5 Apoptosis and mitosis 4.5 hours after 1-Gy γ-irradiation or 8 hours after 10 mg/kg AOM in C57BL/6 (black) and bak-null (white) distal colon. Mean apoptotic index after γ-irradiation or AOM (A). Cell positional distribution of apoptotic cells after γ-irradiation (B) or AOM (C) in C57BL/6 (solid line) and bak-null (dotted line) distal colon. Mean mitotic index after γ-irradiation or AOM (D). Cell positional distribution of mitotic figures after γ-irradiation (E) or AOM (F) in C57BL/6 (solid line) and bak-null (dotted line) distal colon. Significant differences by t test (**P < .001, *P < .05), and significantly different cell positions by modified median test are indicated by bars on the cell positional graphs. Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 6 Examples of methylene blue–stained colonic ACF are indicated by black arrows (B and C). Proximal (white), middle (hashed), and distal (black) thirds of C57BL/6 and bak-null colon were assessed for ACF (A). Significantly different by t test (*P = .01). Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 7 Photomicrographs of alcian blue–stained goblet cells in C57BL/6 (A) and bak-null (B) distal colon. Mean goblet cell index % (C) for C57BL/6 (black) and bak-null (white) distal colon. Cell positional distribution of goblet cells (D) for C57BL/6 (solid line) and bak-null distal colon (dotted line). Significantly different by t test (*P < .001) and by modified median test between cell positions 4 and 44. Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions

Figure 8 Photomicrographs of chromogranin A–immunostained endocrine cells in C57BL/6 (A) and bak-null (B) distal colon. Mean endocrine cell index % (C) for C57BL/6 (black) and bak-null (white) distal colon. Cell positional distribution of endocrine cells (D) for C57BL/6 (solid line) and bak-null distal colon (dotted line). Significantly different by t test (*P < .001) and by modified median test between cell positions 4 and 10. (E) Relative fold-change in gene expression of Atoh1, Ngn3, Hes-1, Klf4, Klf9, Neuro D1, Sox9, Pax6, Elf3, and Gfi1 in bak-null compared with C57BL/6 colonic crypts (n = 6). Significantly different by t test (***P < .001, **P < .01, *P < .05). Gastroenterology 2009 136, 943-952DOI: (10.1053/j.gastro.2008.11.036) Copyright © 2009 AGA Institute Terms and Conditions