Abstract

As part of our programme for developing predictive tests for normal tissue response to radiotherapy, we have investigated the efficacy of the cytokinesis-block micronucleus (MN) assay as a means of detecting interindividual differences in cellular radiosensitivity. A study was made of nine fibroblast strains established from vaginal biopsies of pretreatment cervical cancer patients and an ataxia telangiectasia (A-T) cell strain. Cells were irradiated in plateau phase, replated and treated with cytochalasin B 24 h later. MN formation was examined 72 h after irradiation as the number of MN in 100 binucleate cells. The method yielded low spontaneous MN yields (<7 per 100 cells), and mean induced MN frequencies after 3.5 Gy varied between cell strains from 18 to 144 per 100 cells. However, in repeat experiments, considerable intrastrain variability was observed (CV = 32%), with up to twofold differences in MN yields, although this was less than interstrain variability (CV = 62%). An analysis was made of the relationship between MN results and previously obtained clonogenic survival data. There was a significant correlation between MN yields and clonogenic survival. However, when the A-T strain was excluded from the analysis, the correlation lost significance, mainly because of one slow-growing strain which was the most sensitive to cell killing but had almost the lowest MN frequency. With current methodology, the MN assay on human fibroblasts does not appear to have a role in predictive testing of normal tissue radiosensitivity.

Highlights

  • Nine diploid fibroblast cell strains were denrved from vaginal biopsies of pretreatment cervical cancer patients with full informed consent (Kiltie et al, 1997)

  • The results of this study have shown that there is a relationship between MN and clonogenic measurements of radiosensitivity in untransformed fibroblasts. but only when an ataxia telangiectasia (A-T) strain is included in the analyses

  • The MN assay on human fibroblasts does not appear to have a role in predictive testing of normal tissue radiosensitivity

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Summary

Methods

Nine diploid fibroblast cell strains were denrved from vaginal biopsies of pretreatment cervical cancer patients with full informed consent (Kiltie et al, 1997). Included was an ataxia telangiectasia (A-T) cell strain (ATI). All strains (passage 7-22) were maintained as monolayer cultures in minimum essential medium Medium was supplemented with 100 IU ml-' penicillin. 0.1 mg ml-' streptomycin and 1% glutamine (all from Gibco). 3x105 cells were seeded onto 25-cm flasks (Falcon) and incubated at 37°C in a humidified 5% carbon dioxide atmosphere. They were harvested in plateau phase by addition of 0.02% EDTA UK) followed by 10 min incubation at 37°C in 0.0 1% trypsin

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