Genomic Maintenance & Repair

Monday, October 17, 2005 3:00 PM-5:00 PM Exhibit Hall

(PP320) A role for bystander-like factors in maintaining genomic instability.

Laiakis, Evagelia*,1, 2, Morgan, William1, 3, 1 Radiation Oncology Research Laboratory/ Department of Radiation Oncology, Baltimore, Maryland, USA2 Graduate Program in Human Genetics, Baltimore, Maryland, USA3 Greenebaum Cancer Center, Baltimore, Maryland, USA

ABSTRACT- Radiation-induced genomic instability manifests as chromosomal rearrangements, micronuclei formation, gene amplifications, gene mutations, and/or reduced plating efficiency. Possible reasons for this phenomenon have been attributed to bystander effects and/or clastogenic factors that can be transmitted to neighboring cells through cell-to-cell gap junctions or are secreted into the media. Recently, we have developed an assay, in which ionizing radiation can induce or turn off green fluorescence protein (GFP) expression due to recombination or mutation of a GFP based vector. Recombination/mutation results in the presence of colonies with a mixture of GFP+ and GFP- cells. This assay was utilized to examine genetic changes due to bystander effects. In addition, we looked at differences in cell killing, determined as alterations in the plating efficiency of cells. Experiments using co-cultures of GFP+ stable cells with GFP- stable or unstable cell lines were conducted. Media from non-transfected stable or unstable cells was transferred to GFP+ transfected cells and changes of GFP expression were examined using fluorescence microscopy. Results indicate that there is variability in the plating efficiency among cell lines and that it can be further altered by media from certain unstable cell lines. In addition, media from unstable cells was able to induce genomic instability in GFP cells, demonstrated by the presence of mixed GFP+ and GFP- colonies. The factors responsible for this phenomenon have not yet been identified. However, our unstable clones demonstrate persistently increased levels of reactive oxygen species (ROS) compared to the parental stable cell line. The presence of higher ROS levels represents a condition similar to inflammation and implicates cytokines as the mediators for genomic instability. Expression analysis of cytokines between stable and unstable cell lines reveals differences at both the cDNA and protein level. These differences in cytokine levels could reveal a connection between cytokines and ROS and the initiation and/or perpetuation of radiation induced genomic instability.

Key words: Radiation Induced Genomic Instability, Reactive Oxygen Species, Cytokines

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2005 RRS