Ionizing radiation (IR) induces a variety of DNA lesions among which DNA double-strand breaks (DSBs) are the biologically most significant. It is currently unclear if DSB repair is equally efficient after low and high doses. Here, we use γ-H2AX, phospho-ATM (pATM), and 53BP1 foci analysis to monitor DSB repair. We show, consistent with a previous study, that the kinetics of γ-H2AX and pATM foci loss in confluent primary human fibroblasts are substantially compromised after doses of 10 mGy and lower. Following 2.5 mGy, cells fail to show any foci loss. Strikingly, cells pretreated with 10 μM H 2 O 2 efficiently remove all γ-H2AX foci induced by 10 mGy. At the concentration used, H 2 O 2 produces single-strand breaks and base damages via the generation of oxygen radicals but no DSBs. Moreover, 10 μM H 2 O 2 up-regulates a set of genes that is also up-regulated after high (200 mGy) but not after low (10 mGy) radiation doses. This suggests that low radical levels induce a response that is required for the repair of radiation-induced DSBs when the radiation damage is too low to cause the induction itself. To address the in vivo significance of this finding, we established γ-H2AX and 53BP1 foci analysis in various mouse tissues. Although mice irradiated with 100 mGy or 1 Gy show efficient γ-H2AX and 53BP1 foci removal during 24 h post-IR, barely any foci loss was observed after 10 mGy. Our data suggest that the cellular response to DSBs is substantially different for low vs. high radiation doses.

The highly sensitive gammaH2AX foci assay shows that CM-enhanced radiation damage incurred in peripheral lymphocytes during CT. However, it is unknown whether long-term bioeffects of low-dose ionizing radiation from CT examinations, such as cancer, are increased by using CM.

Purpose: Radiotherapy is an effective cancer treatment, but a few patients suffer severe radiation toxicities in neighboring normal tissues.There is increasing evidence that the variable susceptibility to radiation toxicities is caused by the individual genetic predisposition, by subtle mutations, or polymorphisms in genes involved in cellular responses to ionizing radiation. Doublestrand breaks (DSB) are the most deleterious form of radiation-induced DNA damage, and DSB repair deficiencies lead to pronounced radiosensitivity. Using a preclinical mouse model, the highly sensitive gH2AX-foci approach was tested to verify even subtle, genetically determined DSB repair deficiencies known to be associated with increased normal tissue radiosensitivity. Experimental Design: By enumerating gH2AX-foci in blood lymphocytes and normal tissues (brain, lung, heart, and intestine), the induction and repair of DSBs after irradiation with therapeutic doses (0.1-2 Gy) was investigated in repair-proficient and repair-deficient mouse strains in vivo and blood samples irradiated ex vivo. Results: gH2AX-foci analysis allowed to verify the different DSB repair deficiencies; even slight impairments caused by single polymorphisms were detected similarly in both blood lymphocytes and solid tissues, indicating that DSB repair measured in lymphocytes is valid for different and complex organs. Moreover, gH2AX-foci analysis of blood samples irradiated ex vivo was found to reflect repair kinetics measured in vivo and, thus, give reliable information about the individual DSB repair capacity. Conclusions: gH2AX analysis of blood and tissue samples allows to detect even minor genetically defined DSB repair deficiencies, affecting normal tissue radiosensitivity. Future studies will have to evaluate the clinical potential to identify patients more susceptible to radiation toxicities before radiotherapy.

γ-H2AX immunofluorescence microscopy allows one to determine dose-related effects on x-ray-induced DSB levels and to consider individual factors which cannot be monitored by physical dose measurements.

gamma-H2AX immunofluorescence microscopy is a reliable and sensitive method for measuring the induction and repair of DNA damage caused by ionizing radiation during angiography. To estimate radiation doses delivered during procedures and to consider patients individual repair capacity, postangiography DSB-yields should be compared with DSB-yields after fractioned in vitro irradiation imitating examination conditions.