1. Volume 132, Issue 4, Pages 1420-1431 (April 2007)
The Forkhead Box M1 Transcription Factor Contributes to the Development and Growth of Mouse Colorectal Cancer 
Yuichi Yoshida, I–Ching Wang, Helena M. Yoder, Nicholas O. Davidson, Robert H. Costa 
Gastroenterology 
Volume 132, Issue 4, Pages (April 2007)
DOI: /j.gastro
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2. Figure 1
Foxm1 is overexpressed in human colon adenocarcinomas and cell lines. (A) Foxm1 mRNA levels are increased in a variety of human colon cancer cell lines as determined by Northern blot analysis. (B) Northern blot analysis of colon tumor RNA (t) from 3 individual patients displayed a significant increased expression of Foxm1 mRNA compared with surrounding normal tissues (N).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2007 AGA Institute Terms and Conditions

3. Figure 2
Colon-specific Foxm1 knockout mice show decreased growth and proliferation of colorectal tumors induced by AOM/DSS protocol. (A) Six- to 8-week-old Foxm1fl/fl (n = 13) and Villin-Cre Foxm1-/- male mice (n = 15) were subjected to a single intraperitoneal injection of AOM (10 mg/kg/body weight) followed by 3 cycles of 1 week of administration of 2.5% DSS in the drinking water, each cycle separated by a 2-week period. Mice were killed at 12 weeks after AOM/DSS exposure. Colons were harvested and examined for tumors using a dissecting microscope, and then fixed and paraffin embedded, or used for extraction of total RNA. (B) Scheme of Villin-Cre–mediated recombination of Foxm1 floxed allele. (C) PCR amplification of genomic DNA. Villin-Cre Foxm1-/- colon shows a 510–base pair band specific to recombination allele. Lu, lung; Br, brain; Ki, kidney; Co, colon. (D) The quantitative real-time RT-PCR analysis of Villin-Cre Foxm1-/- colon tissues shows an approximately 90% decrease of Foxm1 mRNA levels compared with the Foxm1fl/fl colon tissues. (E) Macroscopic finding of Foxm1 fl/fl mouse colons and Villin-Cre Foxm1-/- mouse colons. (F) Villin-Cre Foxm1-/- mouse colons show a significant reduction in the total number of tumors compared with Foxm1fl/fl mouse colons. (G, H) H&E staining of the colon tumors. Villin-Cre Foxm1-/- mouse colons showed a reduced tumor size (bar: 1 mm). (I) Villin-Cre Foxm1-/- mouse colons showed an increased number of tumors in which the diameter was smaller than 3.0 mm. Magnification, 400×. (J, K) BrdU staining of adenocarcinomas in Foxm1 fl/fl mouse colons and Villin-Cre Foxm1-/- mouse colons. We counted the number of BrdU-positive cells in 5 random fields (400×) from different colon tumors from each Foxm1 fl/fl mouse and Villin-Cre Foxm1 fl/fl mouse. Magnification of J, 400×. (L) Villin-Cre Foxm1-/- colon adenocarcinomas display a statistically significant reduction in the proliferation rate compared with Foxm1 fl/fl colon tumors. (M) No statistically significant difference was found in normal mucosa between Foxm1 fl/fl mouse colons and Villin-Cre Foxm1-/- mouse colons. (N) Villin-Cre Foxm1-/- colon adenomas show a significant reduction in the proliferation rate compared with Foxm1 fl/fl colon adenomas. *P < .05, **P ≤ .01, and ***P ≤ .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2007 AGA Institute Terms and Conditions

4. Figure 3
Rosa26-Foxm1b TG mice show accelerated growth and proliferation of carcinogen-induced colorectal cancer. (A) Eight WT and 10 Rosa26-Foxm1b TG male (age, 6–8 wk) mice were subjected to the AOM/DSS protocol to induce colon cancer. After 12 weeks of AOM/DSS exposure, the mice were killed. Colons were examined for counting or measuring tumors using a dissecting microscope, and then fixed and paraffin embedded, or used for preparation of total RNA. (B) The qRT-PCR analysis showed a significantly increased expression of Foxm1 mRNA in Rosa26-Foxm1b TG mice compared with WT mice. (C) Macroscopic view of colon tumors in Rosa26-Foxm1b TG and WT AOM/DSS-treated mice. (D) Rosa26-Foxm1b TG mice display a significantly increased number of colon tumors compared with WT mice after AOM/DSS treatment (E) The diameter of colon tumors in Rosa26-Foxm1b TG mice show a statistically significant increase in size compared with those from WT mice. (F) Rosa26-Foxm1bTG mouse colon adenocarcinomas show a significant increase in the BrdU labeling compared with WT mouse colon tumors. (G, H) BrdU staining of colon tumors in Rosa26-Foxm1b TG and WT mice. We counted the number of BrdU-positive cells in 5 random fields (400×) from different mouse colon tumors in each Rosa26-Foxm1b TG and WT mice. Magnification, 200×. (I) No statistically significant difference in normal mucosa between Rosa26-Foxm1b TG colons and WT colons. (J) Rosa26-Foxm1b TG adenomas show a significant increase in the proliferation rate compared with WT adenomas *P < .05, **P ≤ .01, and ***P ≤ .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2007 AGA Institute Terms and Conditions

5. Figure 4
Foxm1-dependent colorectal carcinogenesis is associated with changes in the expression levels of cyclin A2, cyclin B1, survivin, and TCF-4. Quantification of cyclin A2, cyclin B1, survivin, and TCF-4 mRNA of mouse colorectal tumors using qRT-PCR. (A) Cyclin A2, cyclin B1, survivin, and TCF-4 are down-regulated in Villin-Cre Foxm1-/- colorectal cancer compared with the Foxm1fl/fl colorectal cancer. (B) Rosa26-Foxm1b TG colon cancer showed a significantly increased expression of cyclin A2, cyclin B1, survivin, and TCF-4 compared with WT colon tumors. *P < .05, **P ≤ .01, and ***P ≤ .001.
Gastroenterology  , DOI: ( /j.gastro )
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6. Figure 5
Foxm1 is critical for growth, proliferation, and anchorage-independent growth of human and mouse colon adenocarcinoma cell lines in vitro. (A, B) qRT-PCR analysis confirmed a significant reduction in expression of Foxm1 in siFoxm1-transfected DLD1 and CT26 colon cancer cells. (C, D) Foxm1-depleted DLD1 and CT26 colon cancer cells displayed reduced cell growth in culture. (E, H) Depletion of Foxm1 diminishes DNA synthesis of DLD1 and CT26 colon cancer cells as determined by BrdU labeling. (I, J) Reduced number of anchorage-dependent growth of colonies on soft agar with Foxm1-depleted human DLD1 and mouse CT26 colon adenocarcinoma cell lines. *P < .05, **P ≤ .01, and ***P ≤ .001.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2007 AGA Institute Terms and Conditions