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Open Access Methodology article

A methodology for distinguishing divergent cell fates within a common progenitor population: adenoma- and neuroendocrine-like cells are confounders of rat ileal epithelial cell (IEC-18) culture

Phillip V Gordon1*, Jessica B Paxton1 and Nena S Fox2

Author affiliations

1 Department of Pediatrics, University of Virginia, PO Box 800386, Charlottesville, VA 22908, USA

2 Department of Microbiology, University of Virginia, PO Box 800734, Charlottesville, VA 22908, USA

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Citation and License

BMC Cell Biology 2005, 6:2  doi:10.1186/1471-2121-6-2

Published: 18 January 2005

Abstract

Background

IEC-18 cells are a non-transformed, immortal cell line derived from juvenile rat ileal crypt cells. They may have experimental advantages over tumor-derived gastrointestinal lineages, including preservation of phenotype, normal endocrine responses and retention of differentiation potential. However, their proclivity for spontaneous differentiation / transformation may be stereotypical and could represent a more profound experimental confounder than previously realized. We hypothesized that IEC-18 cells spontaneously diverge towards a uniform mixture of epigenetic fates, with corresponding phenotypes, rather than persist as a single progenitor lineage.

Results

IEC-18 cells were cultured for 72 hours in serum free media (SFM), with and without various insulin-like growth factor agonists to differentially boost the basal rate of proliferation. A strategy was employed to identify constitutive genes as markers of divergent fates through gene array analysis by cross-referencing fold-change trends for individual genes against crypt cell abundance in each treatment. We then confirmed the cell-specific phenotype by immunolocalization of proteins corresponding to those genes. The majority of IEC-18 cells in SFM alone had a loss in expression of the adenomatous polyposis coli (APC) gene at the mRNA and protein levels, consistent with adenoma-like transformation. In addition, a small subset of cells expressed the serotonin receptor 2A gene and had neuroendocrine-like morphology.

Conclusions

IEC-18 cells commonly undergo a change in cell fate prior to reaching confluence. The most common fate switch that we were able to detect correlates with a down regulation of the APC gene and transformation into an adenoma-like phenotype.