Date of Award

1-2009

Document Type

Thesis

Degree Name

Master of Science (MS)

College/School

College of Science and Mathematics

Department/Program

Biology

Thesis Sponsor/Dissertation Chair/Project Chair

Reginald Halaby

Committee Member

John J. Gaynor

Committee Member

Quinn C. Vega

Abstract

Ceramide is a second messenger involved in apoptosis, cell differentiation, and growth arrest. We hypothesize that in tumor cells, insufficient sphingomyelin (SM) limits the amount of ceramide which can be generated for the probagation of the apoptotic signal resulting from treatment with chemotherapy. As a result, we postulated that exogenous SM will facilitate apoptosis and synergize with chemotherapeutic agents.

The current studies examined the response of the Panel human pancreatic cancer cell line to gemcitabine, a nucleoside analog, in the presence and absence of sub-toxic doses of SM. Cytotoxic dose response relationships demonstrated an increased chemosensitivity to gemcitabine with SM inclusion which was synergistic. Treatment with gemcitabine and/or SM led to an increased percentage of apoptotic cells with combination treatment as compared to the single agents. Gemcitabine treatment caused a decrease in the expression of anti-apoptotic bcl-2 family proteins in Panel cells, irrespective of the inclusion of SM. However, treatment of Panel cells with a combination of gemcitabine and exogenous SM resulted in a greater percentage of cells accumulating in the S phase when compared to single agent treatments. Changes in morphology and /3-galactosidase activity in cells treated with gemcitabine were consistent with senescence and gemcitabine-induced senescence was abrogated by concomitant SM treatment. Moreover, exposure of Panel cells to Cg-ceramide demonstrated ceramide induces both apoptosis and senescence in a concentration dependent manner with senescence occurring at lower concentrations and apoptosis at high concentrations.

These findings support our proposed model which suggests a cell’s ability to progress through the cell cycle, undergo apoptosis, or enter into senescence is associated with intracellular ceramide levels. By supporting ceramide generation via exogenous SM addition, it is possible to increase apoptosis signaling in tumor cells and increase gemcitabine cytotoxicity. However, failure to generate sufficient signaling ceramide will redirect cells to senescence.

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