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Colorectal hyperplasia and dysplasia due to human carcinoembryonic antigen (CEA) family member expression in transgenic mice.

Chan CH, Camacho-Leal P, Stanners CP - PLoS ONE (2007)

Bottom Line: The colonocyte surface level of integrin alpha5 and the activation of AKT increased progressively with the expression levels of CEA/CEACAM6.Colonic crypts showed a progressive increase in colonocyte proliferation, an increase in crypt fission, and a strong inhibition of both differentiation and anoikis/apoptosis.All transgenic mice showed massively enlarged colons comprising a continuous mosaic of severe hyperplasia, dysplasia and serrated adenomatous morphology.

View Article: PubMed Central - PubMed

Affiliation: McGill Cancer Centre, Department of Biochemistry, McGill University, Montréal, Québec, Canada. carlos.chan@mcgill.ca

ABSTRACT
CEA and CEACAM6 are immunoglobulin family intercellular adhesion molecules that are up-regulated without structural mutations in approximately 70% of human cancers. Results in in vitro systems showing tumorigenic effects for these molecules suggest that this correlation could indicate an instrumental role in tumorigenesis. To test whether this applies in vivo, transgenic mice harboring 187 kb of the human genome containing four CEA family member genes including the CEA and CEACAM6 genes were created and their copy numbers increased by mating until colonocyte expression levels reached levels seen in human colorectal carcinomas. The colonocyte surface level of integrin alpha5 and the activation of AKT increased progressively with the expression levels of CEA/CEACAM6. Colonic crypts showed a progressive increase in colonocyte proliferation, an increase in crypt fission, and a strong inhibition of both differentiation and anoikis/apoptosis. All transgenic mice showed massively enlarged colons comprising a continuous mosaic of severe hyperplasia, dysplasia and serrated adenomatous morphology. These results suggest that up-regulated non-mutated adhesion molecules could have a significant instrumental role in human cancer.

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Subcellular localization of CEACAM and integrin signaling elements in purified colonocytes.A) Immunoblots of different membrane fractions of purified colonocytes derived from sucrose density gradients show a shift in subcellular localization from high density (H) towards low density (L) membrane complexes for integrin α5, ILK, total AKT and Ser473-phosphorylated AKT (pAKT) in purified colonocytes from CEABAC2 and CEABAC10 mice relative to WT littermate mice. No such shift was observed for integrin α2, integrin β1 and FAK. GM1 is a marker for membrane raft-containing membrane fractions. Note that these immunoblots are representative of three independent experiments using at least 6 mice per mouse group. B) Antibody-mediated cross-linking of CEA and CEACAM6 with J22 mAb in the purified colonocytes accentuates the shift of ILK to the less dense membrane fractions shown in A. C) Densitometric analysis of immunoblots for AKT and pAKT shown in A. The mean densitometric ratios (the intensity of band in L divided by the one in H) from three independent experiments show a statistical significant shift of AKT and pAKT to the low density membrane complexes in the CEABAC colonocytes (P<0.001 for AKT and P<0.005 for pAKT). Error bars  =  SEM (N = 3).
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pone-0001353-g003: Subcellular localization of CEACAM and integrin signaling elements in purified colonocytes.A) Immunoblots of different membrane fractions of purified colonocytes derived from sucrose density gradients show a shift in subcellular localization from high density (H) towards low density (L) membrane complexes for integrin α5, ILK, total AKT and Ser473-phosphorylated AKT (pAKT) in purified colonocytes from CEABAC2 and CEABAC10 mice relative to WT littermate mice. No such shift was observed for integrin α2, integrin β1 and FAK. GM1 is a marker for membrane raft-containing membrane fractions. Note that these immunoblots are representative of three independent experiments using at least 6 mice per mouse group. B) Antibody-mediated cross-linking of CEA and CEACAM6 with J22 mAb in the purified colonocytes accentuates the shift of ILK to the less dense membrane fractions shown in A. C) Densitometric analysis of immunoblots for AKT and pAKT shown in A. The mean densitometric ratios (the intensity of band in L divided by the one in H) from three independent experiments show a statistical significant shift of AKT and pAKT to the low density membrane complexes in the CEABAC colonocytes (P<0.001 for AKT and P<0.005 for pAKT). Error bars  =  SEM (N = 3).

Mentions: In agreement with the latter supposition, a shift in subcellular localization towards less dense membrane complexes typical of membrane rafts for integrin α5, ILK and AKT and an increase in phosphorylation for AKT were observed in purified colonocytes from CEABAC2 and CEABAC10 mice relative to WT littermate mice (Figure 3A & C), as was previously seen for these elements in various in vitro systems [21]. No such changes were observed for the integrin α2 subunit, the integrin β1 subunit or FAK (Figure 3A). However, since the overall detection level of integrin β1 (in contrast to integrin α2 and FAK) was very low and no visible signal was detected in the low-density membrane fractions, the failure to see a shift in the subcellular localization of integrin β1 was questionable. In addition, the β1 subunit is shared by many α subunits and so might not be expected to show large changes. Similarly, PI3K was undetectable in this experiment (data not shown).


Colorectal hyperplasia and dysplasia due to human carcinoembryonic antigen (CEA) family member expression in transgenic mice.

Chan CH, Camacho-Leal P, Stanners CP - PLoS ONE (2007)

Subcellular localization of CEACAM and integrin signaling elements in purified colonocytes.A) Immunoblots of different membrane fractions of purified colonocytes derived from sucrose density gradients show a shift in subcellular localization from high density (H) towards low density (L) membrane complexes for integrin α5, ILK, total AKT and Ser473-phosphorylated AKT (pAKT) in purified colonocytes from CEABAC2 and CEABAC10 mice relative to WT littermate mice. No such shift was observed for integrin α2, integrin β1 and FAK. GM1 is a marker for membrane raft-containing membrane fractions. Note that these immunoblots are representative of three independent experiments using at least 6 mice per mouse group. B) Antibody-mediated cross-linking of CEA and CEACAM6 with J22 mAb in the purified colonocytes accentuates the shift of ILK to the less dense membrane fractions shown in A. C) Densitometric analysis of immunoblots for AKT and pAKT shown in A. The mean densitometric ratios (the intensity of band in L divided by the one in H) from three independent experiments show a statistical significant shift of AKT and pAKT to the low density membrane complexes in the CEABAC colonocytes (P<0.001 for AKT and P<0.005 for pAKT). Error bars  =  SEM (N = 3).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2131779&req=5

pone-0001353-g003: Subcellular localization of CEACAM and integrin signaling elements in purified colonocytes.A) Immunoblots of different membrane fractions of purified colonocytes derived from sucrose density gradients show a shift in subcellular localization from high density (H) towards low density (L) membrane complexes for integrin α5, ILK, total AKT and Ser473-phosphorylated AKT (pAKT) in purified colonocytes from CEABAC2 and CEABAC10 mice relative to WT littermate mice. No such shift was observed for integrin α2, integrin β1 and FAK. GM1 is a marker for membrane raft-containing membrane fractions. Note that these immunoblots are representative of three independent experiments using at least 6 mice per mouse group. B) Antibody-mediated cross-linking of CEA and CEACAM6 with J22 mAb in the purified colonocytes accentuates the shift of ILK to the less dense membrane fractions shown in A. C) Densitometric analysis of immunoblots for AKT and pAKT shown in A. The mean densitometric ratios (the intensity of band in L divided by the one in H) from three independent experiments show a statistical significant shift of AKT and pAKT to the low density membrane complexes in the CEABAC colonocytes (P<0.001 for AKT and P<0.005 for pAKT). Error bars  =  SEM (N = 3).
Mentions: In agreement with the latter supposition, a shift in subcellular localization towards less dense membrane complexes typical of membrane rafts for integrin α5, ILK and AKT and an increase in phosphorylation for AKT were observed in purified colonocytes from CEABAC2 and CEABAC10 mice relative to WT littermate mice (Figure 3A & C), as was previously seen for these elements in various in vitro systems [21]. No such changes were observed for the integrin α2 subunit, the integrin β1 subunit or FAK (Figure 3A). However, since the overall detection level of integrin β1 (in contrast to integrin α2 and FAK) was very low and no visible signal was detected in the low-density membrane fractions, the failure to see a shift in the subcellular localization of integrin β1 was questionable. In addition, the β1 subunit is shared by many α subunits and so might not be expected to show large changes. Similarly, PI3K was undetectable in this experiment (data not shown).

Bottom Line: The colonocyte surface level of integrin alpha5 and the activation of AKT increased progressively with the expression levels of CEA/CEACAM6.Colonic crypts showed a progressive increase in colonocyte proliferation, an increase in crypt fission, and a strong inhibition of both differentiation and anoikis/apoptosis.All transgenic mice showed massively enlarged colons comprising a continuous mosaic of severe hyperplasia, dysplasia and serrated adenomatous morphology.

View Article: PubMed Central - PubMed

Affiliation: McGill Cancer Centre, Department of Biochemistry, McGill University, Montréal, Québec, Canada. carlos.chan@mcgill.ca

ABSTRACT
CEA and CEACAM6 are immunoglobulin family intercellular adhesion molecules that are up-regulated without structural mutations in approximately 70% of human cancers. Results in in vitro systems showing tumorigenic effects for these molecules suggest that this correlation could indicate an instrumental role in tumorigenesis. To test whether this applies in vivo, transgenic mice harboring 187 kb of the human genome containing four CEA family member genes including the CEA and CEACAM6 genes were created and their copy numbers increased by mating until colonocyte expression levels reached levels seen in human colorectal carcinomas. The colonocyte surface level of integrin alpha5 and the activation of AKT increased progressively with the expression levels of CEA/CEACAM6. Colonic crypts showed a progressive increase in colonocyte proliferation, an increase in crypt fission, and a strong inhibition of both differentiation and anoikis/apoptosis. All transgenic mice showed massively enlarged colons comprising a continuous mosaic of severe hyperplasia, dysplasia and serrated adenomatous morphology. These results suggest that up-regulated non-mutated adhesion molecules could have a significant instrumental role in human cancer.

Show MeSH
Related in: MedlinePlus