γ-H2AX/53BP1/pKAP-1 foci and their linear tracks induced by in vitro exposure to radon and its progeny in human peripheral blood lymphocytes

Abstract

The biodosimetric information is critical for evaluating the human health hazards caused by radon and its progeny. Here, we demonstrated that the formation of phosphorylated histone variant H2AX (γ-H2AX), p53-binding protein 1 (53BP1) and phosphorylated KRAB-associated protein 1 (pKAP-1) foci and their linear tracks in human peripheral blood lymphocytes (HPBLs) in vitro exposed to radon and its progeny were dependent on the cumulative absorbed dose of radon exposure but was unrelated to the concentration of radon. Among them, γ-H2AX foci and its linear tracks were the most sensitive indicators with the lowest estimable cumulative absorbed dose of 1.74 mGy from their linear dose-response curves and sustained for 12 h after termination of radon exposure. In addition, three types of foci showed an overdispersed non-Poisson distribution in HPBLs. The ratios of pKAP-1/γ-H2AX foci co-localization, 53BP1/γ-H2AX foci co-localization and 53BP1/pKAP-1 foci co-localization were significantly increased in HPBLs exposed to radon while they were unrelated to the cumulative dose of radon exposure, suggesting that γ-H2AX, pKAP-1 and 53BP1 play an important role in the repair of heterochromatic double-strand breaks. Altogether, our findings provide an experimental basis for estimating the biological dose of internal α-particle irradiation from radon and its progeny exposure in humans.

Nanostructure of Clustered DNA Damage in Leukocytes after In-Solution Irradiation with the Alpha Emitter Ra-223

“…To reveal the DNA damage-and repair-associated protein distribution within the damage-loaded chromatin tracks, we applied SMLM ( Figure 1B,C) and quantitative analyses of chromatin conformations including statistical physics, graph theory and cluster algorithms to study repair protein arrangements. As noted by wide-field microscopy ( Figure 1A), alpha-induced DNA damage tracks in leukocyte nuclei showed several, more-or-less linearly aligned super-foci or continuous tracks that consisted of highly concentrated γ-H2AX fluorescent tags that represented the most abundant damage marker ( Figures 1B and 2A), as expected from earlier analysis [8,35,46,51,52]. At an even higher resolution, the super-foci could be subdivided into further subunits with characteristic signal accumulations (Figures 1 and 2).…”

“…Active, monomeric ATM phosphorylates, among other downstream targets, the histone 2 variant H2AX at its Serine 139 moiety, forming γ-H2AX [21]. The latter is in various cell types the most prominent and persisting chromatin mark along alpha particle and accelerated ion tracks [31,51,53,64,65].…”

DNA damage in leukocytes after internal ex-vivo irradiation of blood with the α-emitter Ra-223

“…DNA damage induced by α-particles, on the other hand, involves the generation of closely stacking DSBs and DNA modifications along the ionisation tracks that can be visualised in exposed nuclei as long tracks of γ-H2AX-positive chromatin and repair associated proteins when the ionisation tracks in the nuclei lie parallel to the focal plane of the observer 30 – 32 . It has been shown that the quantification of these tracks can be informative for biological dose estimation 24 , 33 , 34 . So far, this method has been used to quantify DNA damage to cultured human blood cells after external α-irradiation 24 and after radon gas exposure 33 , but not after internal irradiation with the clinically used α-emitter Ra-223.…”

“…It has been shown that the quantification of these tracks can be informative for biological dose estimation 24 , 33 , 34 . So far, this method has been used to quantify DNA damage to cultured human blood cells after external α-irradiation 24 and after radon gas exposure 33 , but not after internal irradiation with the clinically used α-emitter Ra-223.…”

MODELLING Γ-H2ax FOCI INDUCTION TO MIMIC LIMITATIONS IN THE SCORING TECHNIQUE

“…has been adopted to better reproduce the saturation, with b and c free parameters of the fit. Linear fits have been applied in many studies to X-rayinduced foci yields at low doses (9) . The saturation following photon irradiation was instead assessed in the same dose range of this study when ICC is performed for the scoring (2) , while higher saturation thresholds were observed when the overall fluorescence intensity from the whole nucleus is recorded by means of flow cytometry (3) .…”