Date of Award

8-2009

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Legacy Department

Biosystems Engineering

Advisor

Walker, Terry H

Committee Member

Chen , Chin-Fu

Committee Member

Nghiem , Nhuan

Committee Member

Luo , June

Abstract

Accumulating evidences indicate that dietary intake of long-chain polyunsaturated fatty acids (PUFAs) can affect various cellular processes and improve response of cancer cells to chemotherapy. The mechanisms by which PUFAs affect this response are not well understood. P-glycoprotein (P-gp), encoded by the multidrug resistance gene MDR1, is a drug efflux transporter that plays an important role in the bioavailability of anti-cancer drugs. Effects of long-chain polyunsaturated fatty acids on MDR1 gene expression and functional activity in the human colon cancer cell line Caco-2 were studied in this research. Caco-2 cells were treated with different concentrations of three PUFAs: eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and arachidonic acid (AA). All three PUFAs down-regulated the expression of the MDR1 gene (EPA, 34%, DHA, 32% and AA, 27%). The inhibition of gene expression by these PUFAs was accompanied by reduction in protein levels of P-gp. The calcein-AM efflux assay indicated that EPA, DHA, and AA can increase intracellular accumulation (hence decrease the efflux) of calcein-AM (a P-gp substrate) by 25% to 31%. In addition, incubation of cells with PUFAs greatly enhanced the cytotoxicity of the anti-cancer drug paclitaxel. All three PUFAs also induced apoptosis and enhanced paclitaxel-induced apoptosis in Caco-2 cells. Together, these results suggest that inhibition of the multidrug resistance MDR1/P-gp is one mechanism through which dietary polyunsaturated fatty acids exert a positive effect on the response of tumor cells to anti-cancer drugs. In addition, transcriptional promotion of the nuclear receptors CAR and PXR by PUFAs was also observed in this study.
Moreover, to determine whether the eicosapentaenoic acid affects BRCA1 expression through promoter methylation, BRCA1 promoter methylation patterns and gene expression in U937 cells were examined. The methylation status of the BRCA1 promoter was evaluated by methylation-specific PCR (MSP) of bisulfite conversion products. The results indicate that methylation of BRCA1 promoter DNA is reduced in EPA-treated cells. The reduction of methylation in the BRCA1 promoter was accompanied by an increase in mRNA levels obtained by real-time quantitative PCR (qPCR), suggesting that DNA methylation is a possible mechanism by which the dietary polyunsaturated fatty acids mediate gene expression in human cells.
Because of these characteristics, use of PUFAs as adjuvants presents a promising strategy in cancer prevention and therapeutics.

Share

COinS