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Cyclophilin C-associated protein (CyCAP) knock-out mice spontaneously develop colonic mucosal hyperplasia and exaggerated tumorigenesis after treatment with carcinogen azoxymethane.

Torlakovic EE, Keeler V, Wang C, Lim HJ, Lining LA, Laferté S - BMC Cancer (2009)

Bottom Line: Cyclophilin C-associated protein (CyCAP) has been described to down-modulate endotoxin signaling in colorectal murine mucosa and is a murine orthologue of the tumor-associated antigen 90 K (TAA90K)/mac-2-binding protein.The length of the crypts (r = 0.723, p = 0.018, Spearman Correlation) and the number of colonocytes per crypt (r = 0.863, p = 0.001, Spearman Correlation) in non-tumorous areas were positively associated with azoxymethane-induced number of tumors.We conclude that CyCAP-deficient mice spontaneously and after challenge with carcinogen develop significantly more colorectal mucosal hyperplasia, an early stage in murine colonic carcinogenesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, Royal University Hospital, College of Medicine, University of Saskatchewan, Saskatoon, Canada. emt323@mail.usask.ca

ABSTRACT

Background: The discovery of a "serrated neoplasia pathway" has highlighted the role of hyperplastic lesions of the colon as the significant precursor of colorectal adenocarcinoma. In mice, hyperplasia of the colonic mucosa is a regular phenomenon after a challenge with colonic carcinogens indicating that mucosal hyperproliferation and thickening, even without cytological dysplasia, represents an early pre-malignant change. Cyclophilin C-associated protein (CyCAP) has been described to down-modulate endotoxin signaling in colorectal murine mucosa and is a murine orthologue of the tumor-associated antigen 90 K (TAA90K)/mac-2-binding protein.

Methods: Female Balb/c wild-type (WT) and CyCAP knock-out (KO) mice (6-8 weeks old) were administered 2 or 6 weekly subcutaneous injections of azoxymethane. The animals were evaluated post-injection at six weeks for aberrant crypt foci (ACF) study and at five months for colon tumor measurement. The thickness of the colon crypts was measured in microns and the number of colonocytes per crypt was also determined in well-oriented crypts. Morphometric analyses of the colon mucosa were also performed in untreated 6-8 weeks old KO and WT animals. Formalin-fixed/paraffin-embedded colon sections were also studied by immunohistochemistry to determine the Ki-67 proliferation fraction of the colon mucosa, beta-catenin cellular localization, cyclin D1, c-myc, and lysozyme in Paneth cells.

Results: Cyclophilin C-associated protein (CyCAP)-/- mice, spontaneously developed colonic mucosal hyperplasia early in life compared to wild-type mice (WT) (p < 0.0001, T-test) and crypts of colonic mucosa of the (CyCAP)-/- mice show higher proliferation rate (p = 0.039, Mann-Whitney Test) and larger number of cyclin D1-positive cells (p < 0.0001, Mann-Whitney Test). Proliferation fraction and cyclin D1 expression showed positive linear association (p = 0.019, Linear-by-Linear Association). The hyperplasia was even more pronounced in CyCAP-/- mice than in WT after challenge with azoxymethane (p = 0.005, T-test). The length of the crypts (r = 0.723, p = 0.018, Spearman Correlation) and the number of colonocytes per crypt (r = 0.863, p = 0.001, Spearman Correlation) in non-tumorous areas were positively associated with azoxymethane-induced number of tumors. CyCAP-/- developed larger numbers of tumors than WT animals (p = 0.003, T-Test) as well as overall larger tumor mass (p = 0.016, T-Test). Membranous beta-catenin was focally overexpressed in KO mice including proliferative zone of the crypts.

Conclusion: CyCAP-/- represent the first described model of spontaneous colonic mucosal hyperplasia. We conclude that CyCAP-deficient mice spontaneously and after challenge with carcinogen develop significantly more colorectal mucosal hyperplasia, an early stage in murine colonic carcinogenesis.

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In many early adenomas after azoxymethane treatment, hyperplastic-like changes were also present. Adenomatous changes without (A) and with (B) evidence of cytologic dysplasia and cystically dilated crypts and hyperplastic changes in close proximity to muscularis mucosae were found in tumors after azoxymethane treatment (20 × magnification, H&E).
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Figure 5: In many early adenomas after azoxymethane treatment, hyperplastic-like changes were also present. Adenomatous changes without (A) and with (B) evidence of cytologic dysplasia and cystically dilated crypts and hyperplastic changes in close proximity to muscularis mucosae were found in tumors after azoxymethane treatment (20 × magnification, H&E).

Mentions: All available well-oriented crypts were evaluated for the number of cells per crypt as well as for the absolute length of the crypt. The number of observations (N) per animal varied from 60 to 246 per animal depending on the number of available well-oriented crypts. The results are summarized in Tables 1 and 2 and Figure 1, 2, 3, 4, 5, 6, and 7. The length of the crypts in CyCAP mice and WT mice (r = 0.723, p = 0.018, Spearman Correlation) as well as the number of colonocytes per crypt (r = 0.863, p = 0.001, Spearman Correlation) in non-tumorous areas of the colon were positively associated with total number of tumors induced by azoxymethane (Figure 1 and 2). Also, the length of the crypts (r = 0.782, p = 0.008, Spearman Correlation) and the number of colonocytes per crypt (r = 0.927, p < 0.0001, Spearman Correlation) in non-tumorous areas of the colon of CyCAP mice and WT mice correlated with the total size of the tumors. In addition, untreated 8-week old CyCAP mice had significantly thicker mucosa and significantly larger numbers of colonocytes per crypt (hyperplasia) (p < 0.0001, T-test), as well as higher proliferation fraction as measured by Ki-67 immunohistochemical test (p = 0.032, Mann-Whitney Test) even before azoxymethane treatment was initiated (Figure 1, 2A, and 3) indicating spontaneous development of mucosal hyperplasia. The difference in diffuse mucosal thickness was even more pronounced after the treatment with carcinogen (p < 0.0001, T-test, Figure 2A, 3, 4, and 5). Similarly, the mice had larger numbers of cells per crypt both before (p = 0.032, Linear Regression; plots not shown) and after treatment with carcinogen (p = 0.012, Linear Regression; plots not shown) compared with WT animals.


Cyclophilin C-associated protein (CyCAP) knock-out mice spontaneously develop colonic mucosal hyperplasia and exaggerated tumorigenesis after treatment with carcinogen azoxymethane.

Torlakovic EE, Keeler V, Wang C, Lim HJ, Lining LA, Laferté S - BMC Cancer (2009)

In many early adenomas after azoxymethane treatment, hyperplastic-like changes were also present. Adenomatous changes without (A) and with (B) evidence of cytologic dysplasia and cystically dilated crypts and hyperplastic changes in close proximity to muscularis mucosae were found in tumors after azoxymethane treatment (20 × magnification, H&E).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2724547&req=5

Figure 5: In many early adenomas after azoxymethane treatment, hyperplastic-like changes were also present. Adenomatous changes without (A) and with (B) evidence of cytologic dysplasia and cystically dilated crypts and hyperplastic changes in close proximity to muscularis mucosae were found in tumors after azoxymethane treatment (20 × magnification, H&E).
Mentions: All available well-oriented crypts were evaluated for the number of cells per crypt as well as for the absolute length of the crypt. The number of observations (N) per animal varied from 60 to 246 per animal depending on the number of available well-oriented crypts. The results are summarized in Tables 1 and 2 and Figure 1, 2, 3, 4, 5, 6, and 7. The length of the crypts in CyCAP mice and WT mice (r = 0.723, p = 0.018, Spearman Correlation) as well as the number of colonocytes per crypt (r = 0.863, p = 0.001, Spearman Correlation) in non-tumorous areas of the colon were positively associated with total number of tumors induced by azoxymethane (Figure 1 and 2). Also, the length of the crypts (r = 0.782, p = 0.008, Spearman Correlation) and the number of colonocytes per crypt (r = 0.927, p < 0.0001, Spearman Correlation) in non-tumorous areas of the colon of CyCAP mice and WT mice correlated with the total size of the tumors. In addition, untreated 8-week old CyCAP mice had significantly thicker mucosa and significantly larger numbers of colonocytes per crypt (hyperplasia) (p < 0.0001, T-test), as well as higher proliferation fraction as measured by Ki-67 immunohistochemical test (p = 0.032, Mann-Whitney Test) even before azoxymethane treatment was initiated (Figure 1, 2A, and 3) indicating spontaneous development of mucosal hyperplasia. The difference in diffuse mucosal thickness was even more pronounced after the treatment with carcinogen (p < 0.0001, T-test, Figure 2A, 3, 4, and 5). Similarly, the mice had larger numbers of cells per crypt both before (p = 0.032, Linear Regression; plots not shown) and after treatment with carcinogen (p = 0.012, Linear Regression; plots not shown) compared with WT animals.

Bottom Line: Cyclophilin C-associated protein (CyCAP) has been described to down-modulate endotoxin signaling in colorectal murine mucosa and is a murine orthologue of the tumor-associated antigen 90 K (TAA90K)/mac-2-binding protein.The length of the crypts (r = 0.723, p = 0.018, Spearman Correlation) and the number of colonocytes per crypt (r = 0.863, p = 0.001, Spearman Correlation) in non-tumorous areas were positively associated with azoxymethane-induced number of tumors.We conclude that CyCAP-deficient mice spontaneously and after challenge with carcinogen develop significantly more colorectal mucosal hyperplasia, an early stage in murine colonic carcinogenesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, Royal University Hospital, College of Medicine, University of Saskatchewan, Saskatoon, Canada. emt323@mail.usask.ca

ABSTRACT

Background: The discovery of a "serrated neoplasia pathway" has highlighted the role of hyperplastic lesions of the colon as the significant precursor of colorectal adenocarcinoma. In mice, hyperplasia of the colonic mucosa is a regular phenomenon after a challenge with colonic carcinogens indicating that mucosal hyperproliferation and thickening, even without cytological dysplasia, represents an early pre-malignant change. Cyclophilin C-associated protein (CyCAP) has been described to down-modulate endotoxin signaling in colorectal murine mucosa and is a murine orthologue of the tumor-associated antigen 90 K (TAA90K)/mac-2-binding protein.

Methods: Female Balb/c wild-type (WT) and CyCAP knock-out (KO) mice (6-8 weeks old) were administered 2 or 6 weekly subcutaneous injections of azoxymethane. The animals were evaluated post-injection at six weeks for aberrant crypt foci (ACF) study and at five months for colon tumor measurement. The thickness of the colon crypts was measured in microns and the number of colonocytes per crypt was also determined in well-oriented crypts. Morphometric analyses of the colon mucosa were also performed in untreated 6-8 weeks old KO and WT animals. Formalin-fixed/paraffin-embedded colon sections were also studied by immunohistochemistry to determine the Ki-67 proliferation fraction of the colon mucosa, beta-catenin cellular localization, cyclin D1, c-myc, and lysozyme in Paneth cells.

Results: Cyclophilin C-associated protein (CyCAP)-/- mice, spontaneously developed colonic mucosal hyperplasia early in life compared to wild-type mice (WT) (p < 0.0001, T-test) and crypts of colonic mucosa of the (CyCAP)-/- mice show higher proliferation rate (p = 0.039, Mann-Whitney Test) and larger number of cyclin D1-positive cells (p < 0.0001, Mann-Whitney Test). Proliferation fraction and cyclin D1 expression showed positive linear association (p = 0.019, Linear-by-Linear Association). The hyperplasia was even more pronounced in CyCAP-/- mice than in WT after challenge with azoxymethane (p = 0.005, T-test). The length of the crypts (r = 0.723, p = 0.018, Spearman Correlation) and the number of colonocytes per crypt (r = 0.863, p = 0.001, Spearman Correlation) in non-tumorous areas were positively associated with azoxymethane-induced number of tumors. CyCAP-/- developed larger numbers of tumors than WT animals (p = 0.003, T-Test) as well as overall larger tumor mass (p = 0.016, T-Test). Membranous beta-catenin was focally overexpressed in KO mice including proliferative zone of the crypts.

Conclusion: CyCAP-/- represent the first described model of spontaneous colonic mucosal hyperplasia. We conclude that CyCAP-deficient mice spontaneously and after challenge with carcinogen develop significantly more colorectal mucosal hyperplasia, an early stage in murine colonic carcinogenesis.

Show MeSH
Related in: MedlinePlus