The aim is to establish dose response relations for radiation mutation, including chromosome breakage, deletions and loss of an entire chromosome in cultured mammalian cells at low doses (10 rad or less). We have developed and will employ two methodologies which are 10-100 times more sensitive for detention of radiation damage than other culture methods. The first system measures mutation in a human-hamster cell line (AL) which stably retains human chromosome no. 11 as its sole human chromosome. Cell surface antigens result from a set of at least three genes located on opposite arms of chromosome 11. Exposure of AL cells to mutagens increase mutation in specific regions to chr. 11. Mutants can form colonies in the presence of complement and specific antisera that kills the non-mutant cells. The frequency and pattern of marker loss quantitates gene mutation, deletions and loss of the entire chr. 11. These large genic damages are implicated in genetic disease, teratogenesis and cancer but go undetected in other systems where they are lethal. Such aberrations are measurable in the AL method because no known function for survival of the hybrid is supplied by chr. 11. Thus this entire chromosome can be a target and the mutation rate is at least 100 times greater than in other cell systems. The other technique, premature chromosome condensation (PCC) allows assessment of chromosome breakage in interphase human cultured cell lines or in human bone marrow cells immediately after irradiation. PCC is obtained by fusing irradiated interphase cells with mitotic cells which causes the chromosomes of the former to condense so that breaks may be scored microscopically. Measurements in both systems will be made at high and low dose rates, at different stages of the cell cycle, and in the presence of agents that inhibit genome repair. Kinetics of chromosome break rejoining will be studied. Correlations will be sought between damage as measured by cytogenetics on the one hand and by mutation on the other. Attempts will be made to construct new hybrids with improved properties for mutation analysis and also to devise ways of quantitating mutation by flow cytometry. Even in their present state these two methodologies promise unprecedented sensitivity in assessing the spectrum of x-ray genetic damage and should provide new insight into how mutation, repair and chromosome breakage and rejoining are related.
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