Free radical-initiated and gap junction-mediated bystander effect due to nonuniform distribution of incorporated radioactivity in a three-dimensionaltissue culture model

To investigate the biological effects of nonuniform distribution of radioac tivity in mammalian cells, we have developed a novel three-dimensional tiss ue culture model. Chinese hamster V79 cells were labeled with tritiated thy midine and mixed with unlabeled cells, and multicellular clusters (similar to1.6 mm in diameter) were formed by gentle centrifugation. The short-range beta particles emitted by H-3 impart only self-irradiation of labeled cell s without significant cross-irradiation of unlabeled bystander cells. The c lusters were assembled in the absence or presence of 10% dimethyl sulfoxide (DMSO) and/or 100 muM lindane. DMSO is a hydroxyl radical scavenger, where as lindane is an inhibitor of gap junctional intercellular communication. T he clusters were maintained at 10.5 degreesC for 72 h to allow 3H decays to accumulate and then dismantled, and the cells were plated for colony forma tion When 100% of the cells were labeled, the surviving fraction was expone ntially dependent on the mean level of radioactivity per labeled cell. A tw o-component exponential response was observed when either 50 or 10% of the cells were labeled. Though both DMSO and lindane significantly protected th e unlabeled or bystander cells when 50 or 10% of the cells were labeled, th e effect of lindane was greater than that of DMSO. In both cases, the combi ned treatment (DMSO + lindane) elicited maximum protection of the bystander cells. These results suggest that the bystander effects caused by nonunifo rm distributions of radioactivity are affected by the fraction of cells tha t are labeled. Furthermore, at least a part of these bystander effects are initiated by free radicals and are likely to be mediated by gap junctional intercellular communication. (C) 2001 by Radiation Research Society.