Implications of an elevated sister-chromatid exchange frequency in rat lymphocytes cultured in the absence of erythrocytes

Abstract

One important variable in complex culture systems such as whole blood is the interaction of the cell types present. To investigate the effects of erythrocytes (RBCs) and monocytes on the sister-chromatid exchange (SCE) frequency, Ficoll-Hypaque-separated Fischer-344 rat leukocytes were added to 1.9 ml of culture medium containing either 4 μg phytohemagglutinin or 4–8 μg concanavalin A/ml. Bromodeoxyuridine (BrdU; 2 μM) was added at 24 h, and the cultures were harvested at 54 or 72 h. SCE frequencies in the mononuclear leukocyte cultures were consistently about 1.5- to 2-fold higher than in the whole-blood cultures. The titration of rat or human RBCs (0.05–2.5 × 10 9) into purified rat leukocyte cultures reduced the SCE frequency to that of whole-blood cultures. Monocyte depletion decreased the elevated SCE frequency by / t 50%. Scintillation counting of [ 14C]BrdU uptake in isolated RBCs revealed that <8% of the total amount of BrdU was sequestered. Also, BrdU induced a concentration-dependent increase in SCE in purified leukocytes, but the absolute increase was no greater than in whole-blood lymphocytes. Thus, BrdU had a minor role in the elevated SCE frequency in purified lymphocytes. Neither anti-oxidant enzymes such as catalase and superoxide dismutase nor the hydroxyl radical scavenger, dimethyl sulfoxide, decreased the SCE frequency. Although purified human lymphocytes had a small, but significant increase in SCE compared to whole blood, the magnitude of the dichotomous response between man and rat may represent a fundamental species difference.