Date of Award

January 2018

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Sciences

First Advisor

Othman Ghribi

Abstract

Colorectal Cancer (CRC) is a major health concern with more than 600,000 deaths worldwide each year. Several risk factors for CRC exist including a diet rich in fat and cholesterol. However, the link between cholesterol and CRC is controversial. It is possible that oxysterols, which are active cholesterol metabolites, may correlate better with CRC than cholesterol itself. The most abundant oxysterol in the plasma is 27-hydroxycholesterol (27-OHC), which is a ligand for the nuclear receptors estrogen receptor (ER) and liver-x-receptor (LXR). 27-OHC has been shown to be involved in breast and prostate cancers, but its role in colon cancer remains unexplored. Therefore, we sought to determine the role of 27-OHC in colon cancer cell lines. We also determined the impact of 27-OHC on cellular differentiation through measurement of the protein SLFN12. Using two colon cancer cell lines (Caco2 and SW620), an MTT assay and CyQUANT® proliferation assays, an LDH cytotoxicity assay, a TUNEL assay, western blotting, immunofluorescence, and a scratch assay, we determined 27-OHC’s effects on cellular proliferation, migration, cytotoxicity, and apoptosis. We also determined the role of ER and LXR in cellular proliferation. We found that treatment of Caco2 and SW620 cells with 27-OHC induced a decrease in cell proliferation without a detectable amount of cellular cytotoxicity or apoptosis. Additionally, we found that the observed decrease in cell proliferation is independent of ER and LXR, as 27-OHC did not activate the LXR target genes ABCG1 and ABCA1 nor lead to the nuclear localization of ER. Additionally, we found that 27-OHC leads to a decrease in the phosphorylated and activated form of AKT, a molecule involved in cell cycle progression, protein synthesis, and cellular survival. We also found that 27-OHC increased cellular migration through a scratch assay. 27-OHC decreased the epithelial marker E-cadherin without changes to the mesenchymal markers N-cadherin. In order for cells to migrate, they need to stop proliferating. In conclusion, our data shows that 27-OHC treatment leads to an increase in cellular migration through a loss of proliferation.

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