Effect of Antioxidants on X-ray–induced γ-H2AX Foci in Human Blood Lymphocytes: Preliminary Observations

Abstract

To investigate the effect of a radioprotective oral agent containing a formulation of antioxidants and glutathione-elevating compounds on the extent of x-ray-induced γ-H2AX foci formation. The study was approved by local ethics committee and informed consent was obtained from each subject. In vitro experiments with blood lymphocytes of 25 healthy volunteers were performed without antioxidants and with antioxidants added either before or immediately after irradiation (10 mGy). For in vivo/in vitro tests, blood samples were obtained before, 15, 30, and 60 minutes (n=17) after, and 2, 3, and 5 hours (n=11) after oral ingestion of antioxidant pills and were irradiated (10 mGy). DNA double-strand breaks (DSBs) were quantified in isolated lymphocytes 5 minutes (in vitro and in vivo/in vitro) and 15 minutes (in vitro) after irradiation by enumerating γ-H2AX foci. To validate the data, additional in vitro experiments with use of 53BP1 as another independent marker for DSBs were performed. Nonirradiated samples served as controls. Statistical analyses were performed by using Wilcoxon rank-sum tests (in vitro), repeated-measures test, and Dunnett test (in vivo/in vitro). In the in vitro experiments, 15-minute preincubation with antioxidants significantly reduced mean γ-H2AX foci levels by 23% (P<.0001), whereas addition of antioxidants immediately after irradiation did not lead to a reduction of x-ray-induced foci (P=.6905). Mean 53BP1 foci were also reduced by preincubation with the radioprotectant. In the in vivo/in vitro tests, oral pretreatment with antioxidants also led to a significant reduction of γ-H2AX foci formation; administration 60 minutes before irradiation resulted in a mean foci reduction of 58% (P<.0001). The tested formulation of antioxidants significantly reduced formation of γ-H2AX and 53BP1 foci after irradiation at a radiologic radiation dose typical for computed tomographic imaging; administration 60 minutes prior to irradiation seems to be appropriate and leads to a significant reduction in foci.