Access Type

Open Access Thesis

Date of Award

1-1-2010

Degree Type

Thesis

Degree Name

M.S.

Department

Biological Sciences

First Advisor

James D. Tucker

Abstract

This work evaluated the radiation-induced bystander effect in mitochondrial mutant cells and in normal cells treated with mitochondrial inhibitors. Although much research has been performed on the bystander effect, the underlying molecular mechanisms remain largely unknown. Cells without intact mitochondrial DNA have been shown to lack the bystander effect, which is an energy-dependent process. Based on these findings, cells harboring mutations in the mitochondrial genes responsible for ATP synthesis, and normal cells treated with mitochondrial inhibitors, were hypothesized to show a decreased bystander effect when compared to normal cells that were not treated with the mitochondrial inhibitors.

Radiation-induced bystander effects were analyzed in normal and mitochondrial mutant human lymphoblastoid cells using the cytokinesis block micronucleus assay. The results indicate that the mitochondrial mutant cells do not exhibit the bystander effect while the normal cells do. Both the normal and mitochondrial mutant cells were found to show a dose-responsive increase in DNA damage upon direct exposure to radiation.

The generation of and response to bystander signals in the normal and mitochondrial mutant cells were then analyzed by evaluating the radiation-induced bystander effects between each possible pair-wise combination of normal and mutant cell lines. Normal cells were found to generate and to respond to bystander signals, while the mitochondrial mutant cells neither generated nor responded to bystander signals.

To determine whether normal cells behave similar to the mutant cells when their mitochondrial ATP synthesis is inhibited, the radiation-induced bystander effect in normal cells treated with mitochondrial inhibitors was evaluated. The results indicate that the normal cells treated with rotenone or oligomycin do not exhibit the bystander effect. To determine the time period during which the inhibition was effective, normal cells were treated with the inhibitors at different times before, during and after radiation. The results indicate that normal cells do not exhibit the bystander effect when the inhibitors were added 24 hours before radiation and were removed immediately after radiation. Similarly, normal cells treated with the inhibitors after radiation did not exhibit the bystander effect. However, when the inhibitors were added just prior to radiation and were removed immediately after radiation, normal cells did not show an inhibition of bystander effect, indicating that this short period of treatment might not be sufficient to cause the inhibition.

These results indicate that mitochondrial ATP synthesis in human lymphoblastoid cells plays an important role in the generation and response to bystander signals and thus, the bystander effect.

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