An evaluation of factors influencing sister chromatid exchange frequency in human chromosomes (1980)
AuthorsMorgan, William F.show all
Technical variables in lymphocyte culture, harvest and differential staining were investigated for their effect on baseline sister chromatid exchange (SCE) levels. The culture medium, 5 Bromodeoxyuridine (BrdUrd) concentration and method of differential staining all influenced SCE frequency, whereas the method of lymphocyte separation, the mitogen utilized and the timing of BrdUrd addition did not. Repeat cultures scored for SCE at varying time intervals during the course of this study showed that for most individuals, baseline SCE levels were constant over time. SCE induction was studied using a range of chemical and physical agents differing in their interactions with DNA. The results suggested that a variety of DNA interactions culminated in SCE formation. SCE analysis appeared a useful technique for assessing many, but not all, classes of chromosome damaging agents and therefore is best used in conjunction with other test systems in assessing DNA damaging agents. Different experimental protocols were utilized to investigate SCE induction. Induced exchange levels varied considerably between resting (Go) lymphocytes and replicating lymphocytes, and between replicating lymphocytes that had incorporated BrdUrd prior to chemical or physical challenge and those lymphocytes that had not. It appeared that incorporation of BrdUrd into chromosomal DNA increased lymphocyte sensitivity to damage by most agents. These factors should be considered when assessing induced SCEs and a testing schedule is described which would take these variables into account. The effect of X irradiation on baseline SCE frequency was studied in detail. X irradiation caused a small but significant increase in SCEs. Although incorporation of BrdUrd sensitizes chromosomes to damage by ionizing radiation, increased BrdUrd concentrations in the culture medium did not lead to an increase in X-ray induced SCEs. Results from post irradiation holding experiments suggested that SCEs represent a form of DNA repair. This SCE repair was thought to operate on residual damage not repaired by other DNA repair mechanisms. A comparison was made between the frequency of induced SCEs in human lymphocytes and Chinese hamster ovary (CHO) cells. CHO cells showed significantly higher SCE levels, suggesting that the results of SCE analysis in CHO cells was not a good indicator of potential SCE induction in man.