Our long-term objective is to identify and characterize populations of intestinal stem cells (SC). Developing the ability to identity SC in colonic crypts will make possible investigation into their putative role in the origin of colon cancer. Our immediate objective is to develop markers for colonic stem cell enriched (SCE) preparations. Array-based gene expression profiling will be used to characterize colonic SCE preparations made based on anatomical and functional properties of SC (location at the bottom of the crypt & clonogenicity). We will investigate SCE preparations from purified normal crypts (Aim 1) & from crypts that contain mutant APC (from familial adenomatous polyposis [FAP] patients;
Aim 2) to compare their gene expression profiles. We will test two hypothesis: H1: Genes selectively expressed in crypt base cells (compared to the whole crypt) are also selectively expressed in clonogenic cells (compared to the whole crypt). H2: Genes selectively expressed in crypt base cells and/or clonogenic cells have increased expression in FAP crypts compared to normal crypts. Tissue samples from surgical colectomy specimens will be collected, colonic crypts purified, and microarray analysis done to characterize SCE preparations.
AIM 1 : To determine gene expression profiles for SCE preparations from normal human crypts. Task 1.a: evaluate microarray profiles for SCE preparations from the bottom of purified crypts. Task 1.b evaluate microarray profiles for SCE preparations from crypt clonogenic cells from colonies in soft agarose. Task 1.c determine which genes are common to profiles in both I.a and 1.b and build a targeted array.
AIM 2 : To determine gene expression patterns and levels for SCE marker genes in abnormal crypts, (FAP crypts). Task 2: compare gene expression levels and patterns in purified whole crypts from FAP vs. normal individuals using the targeted array (1 .c). We predict: i) there is a unique gene expression pattern for each SCE preparation (base & clonogenic cells), and ii) these patterns are similar. We also predict that genetic markers for SCE preparations have increased levels of expression in FAP crypts compared to normal crypts. Our results will provide gene expression profiles that might serve as specific markers for colonic SC populations or at least for SCE preparations. With markers for SCE preparations, future research could lead, through various experimental approaches, to specific markers for colonic SC. With specific markers for SC, we could directly test the idea that colon cancer initiation occurs because APC mutation leads to SC overproduction. ? ?
| Bhatlekar, Seema; Addya, Sankar; Salunek, Moreh et al. (2014) Identification of a developmental gene expression signature, including HOX genes, for the normal human colonic crypt stem cell niche: overexpression of the signature parallels stem cell overpopulation during colon tumorigenesis. Stem Cells Dev 23:167-79 |
| Zhang, Tao; Fields, Jeremy Z; Opdenaker, Lynn et al. (2010) Survivin-induced Aurora-B kinase activation: A mechanism by which APC mutations contribute to increased mitoses during colon cancer development. Am J Pathol 177:2816-26 |
| Huang, Emina H; Hynes, Mark J; Zhang, Tao et al. (2009) Aldehyde dehydrogenase 1 is a marker for normal and malignant human colonic stem cells (SC) and tracks SC overpopulation during colon tumorigenesis. Cancer Res 69:3382-9 |
