Gastroenterology

Gastroenterology

Volume 141, Issue 3, September 2011, Pages 1003-1013.e10
Gastroenterology

Original Research
Basic and Translational—Alimentary Tract
Mutant Kras Promotes Hyperplasia and Alters Differentiation in the Colon Epithelium but Does Not Expand the Presumptive Stem Cell Pool

https://doi.org/10.1053/j.gastro.2011.05.007Get rights and content

Background & Aims

Adenomatous polyps are precursors to colorectal cancer (CRC), whereas hyperplastic polyps (HPPs) have low risk of progression to CRC. Mutations in KRAS are found in ∼40% of CRCs and large adenomas and a subset of HPPs. We investigated the reasons why HPPs with KRAS mutations lack malignant potential and compared the effects of Kras/KRAS activation with those of Apc/APC inactivation, which promotes adenoma formation.

Methods

We activated a KrasG12D mutant allele or inactivated Apc alleles in mouse colon epithelium and analyzed phenotypes and expression of selected genes and proteins. The mouse data were validated using samples of human HPPs and adenomas. Signaling pathways and factors contributing to Kras/KRAS-induced phenotypes were studied in intestinal epithelial cells.

Results

Activation of Kras led to hyperplasia and serrated crypt architecture akin to that observed in human HPPs. We also observed loss of Paneth cells and increases in goblet cell numbers. Abnormalities in Kras-mediated differentiation and proliferation required mitogen-activated protein kinase signaling and were linked to activation of the Hes1 transcription factor. Human HPPs also had activation of HES1. In contrast to Apc/APC inactivation, Kras/KRAS activation did not increase expression of crypt stem cell markers in colon epithelium or colony formation in vitro. Kras/KRAS activation was not associated with substantial induction of p16INK4a protein expression in mouse colon epithelium or human HPPs.

Conclusions

Although Kras/KRAS mutation promotes serrated and hyperplastic morphologic features in colon epithelium, it is not able to initiate adenoma development, perhaps in part because activated Kras/KRAS signaling does not increase the number of presumptive stem cells in affected crypts.

Section snippets

Mice

KrasLSL-G12D/+ mice have been described previously.15 We crossed KrasLSL-G12D/+ mice to CDX2P9.5-G22Cre (G22Cre) transgenic mice to activate the mutant Kras allele in the intestinal tract.14 To monitor Cre expression, G22Cre mice or G22Cre KrasLSL-G12D/+ mice were intercrossed with mice carrying the Gt(ROSA)26Sortm1(EYFP)Cos/J reporter allele (Jackson Laboratory, Bar Harbor, ME). To target Apc alleles, G22Cre mice or CDX2P 9.5-NLS Cre mice were intercrossed with mice homozygous for Apc targeted

Activation of KrasG12D Mutant Allele in Mouse Colon Epithelium Promotes HPP-like Features

We used the CDX2P-G22Cre transgenic mouse line (referred to as G22Cre),14 which expresses Cre in epithelial cells in terminal ileum, cecum, colon, and rectum, to activate a latent oncogenic KrasLSL-G12D allele.15 Cre-mediated recombination removes transcriptional stop elements, resulting in expression of the mutant KrasLSL-G12D allele under control of its endogenous regulatory elements. Using mice carrying a gene for enhanced yellow fluorescence protein (EYFP) at the Gt(ROSA)26Sor locus19 to

Discussion

Roughly 40% of human CRCs and larger adenomas carry KRAS-activating mutations.5, 35 Mutant KRAS alleles are functionally significant in advanced CRC cells.8 In light of these data on KRAS mutations in CRC pathogenesis, it has not previously been clear why human HPPs with somatic KRAS (or BRAF) mutations have nominal risk of progressing to CRC.2 One well-documented difference between colon adenomas, which are precursor lesions for CRC, and HPPs is that most adenomas have APC inactivation and

Acknowledgments

G.T.B.'s current affiliation is: De Duve Institute, BCHM/GRM, Universite Catholique de Louvain, Belgium.

The authors thank Dr Hideyuki Okano for generously providing the Msi1 antibody and Dr Joel Greenson for providing human colon tissue, Dr Shuling Fan for help with immunofluorescence microscopy, Cameron Bothner for graphics assistance, and Brandon Fishman for comments on the paper.

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    Conflicts of interest The authors disclose no conflicts.

    Funding Supported by National Institutes of Health grant CA082223 and Department of Defense award W81XWH-09-2-0014.

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