Mutation induction after low-dose carbon-ion beam irradiation of frozen human cultured cells

Abstract

To study the genetic influence of low-dose ionizing radiation at the chromosomal level, frozen human lymphoblastoid TK6 cells were exposed to a 10 cGy dose delivered by a carbon-ion (24.5 ± 2.0 keV/μm) beam. Mutation assays were performed within a few days or after about one month of preservation at -80°C following irradiation. The results showed an increase in mutation frequencies at the thymidine kinase (TK) gene locus of 1.6-fold (2.5 × 10-6 to 3.9 × 10-6) and 2.1-fold (2.5 × 10-6 to 5.3 × 10-6), respectively. The relative distributions of the observed mutation classes were not changed by the radiation exposure in either assay. Multilocus analysis using two TK locus markers and eleven microsatellite loci spanning chromosome 17 were used for the analysis of TK mutants exhibiting a loss of heterozygosity (LOH). An interesting characteristic was detected using this system; interstitial deletion patterns were observed in hemizygous LOH mutants, which were specific for radiation exposure, and considered to be the result of end-joining repair of carbon-ion-induced DNA double-strand breaks. The relative increase in TK mutation frequency of the irradiated cells after the longer preservation at -80°C is probably due to the lower cell-viability compared to the unirradiated level. These results clearly demonstrate that this type of analysis can be used for the detection of low-dose ionizing radiation effects in frozen cells.