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The covalent addition of nitric oxide (NO���) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins. The dysregulation of normal S-nitrosylation contributes to degenerative conditions and to cancer. To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the ���biotin switch��� assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of cesium-137 ��-rays. The dose of radiation was delivered to the whole body in the presence or absence of iopamidol, an iodinated contrast agent used during radiological examinations. To investigate whether similar or distinct nitrosylation patterns are induced following high-dose irradiation, mice were exposed in parallel to acute 4 Gy of cesium-137 �� rays. Analysis of modulated S-nitrosothiols (SNO-proteins) in freshly-harvested organs of animals sacrificed 13 days after irradiation revealed radiation dose- and contrast agent-dependent changes. The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose ��-irradiation.

International audience; The induction of non-targeted stressful effects in cell populations exposed to low fluences of high charge (Z) and high energy (E) particles is relevant to estimates of the outcome of hadron therapy and the long term health risks of space radiation. To gain greater knowledge of HZE-particle-induced non-targeted effects, we investigated the regulation of stress markers in confluent normal human fibroblast cultures exposed to 1000 MeV/u iron ions (linear energy transfer (LET) ~151 keV/μm) or 600 MeV/u silicon ions (LET ~50 keV/μm) at mean absorbed doses as low as 0.2 cGy, wherein 1-3 % of the cells were targeted through the nucleus by a primary particle. Within 24 h post-irradiation, significant increases in the levels of phospho-TP53 (S15), p21Waf1, HDM2, phospho-ERK1/2, protein carbonylation and lipid peroxidation were detected, which suggested participation in the stress response of cells not targeted by primary particles. This was supported by in situ studies that indicated greater increases in 53BP1 foci formation, a marker of DNA damage, than expected from the number of primary particle traversals. The effect was expressed as early as 15 min after exposure, peaked at 1 h, and decreased by 24 h. A similar tendency occurred after exposure of the cell cultures to 0.2 cGy of 3.7 MeV α particles (LET ~109 keV/μm) that targets ~1.6 % of nuclei, but not after 0.2 cGy from 290 MeV/u carbon ions (LET ~13 keV/μm) by which, on average, ~13 % of the nuclei were hit, which highlights the importance of radiation quality in the induced effect. Together, the above data suggested participation of bystander cells in the observed responses. To investigate whether secondary particles are a factor in expression of the apparent HZE-particle-induced bystander effects, the tracks of the secondaries were simulated with the multi-particle transport code FLUKA and absorbed doses received by the monolayer of cells adjacent to the targeted cells were assessed. The simulations revealed that fragmentation products, other than electrons, in cell cultures exposed to HZE particles comprise

International audience; We and others have previously shown, in confluent cell cultures exposed to low fluences of a particles, that the proportion of cells that upregulate stress-inducible proteins is much higher that the number of cells irradiated. This phenomenon, called bystander effect, is now well accepted and is thought to impact the health risks of exposure to ionizing radiation. Here, we investigate the kinetics of propagation of signaling events that lead to induction of DNA damage in bystander cells in confluent normal human AG1522 fibroblasts exposed to a mean dose of 0.2 cGy from 3.2 MeV a particles (LET ; 124 keV/lm) or 1 GeV/n iron ions (LET ; 151 keV/lm). We evaluated the formation of 53BP1 foci (p53 binding protein 1), which localizes at sites of DNA double strand breaks, as a function of time after irradiation. The fraction of cells whose nuclei were traversed by an irradiating particle was derived from Poisson statistics and estimates of cell geometry, particle fluence and energy loss. At a mean dose of 0.2 cGy, only 1.4% and 1.2% of the cells are traversed through the nucleus by a particle or iron ion tracks, respectively. The number of 53BP1 foci in control cells was ; 0.61 foci per cell. In a particle-irradiated cell cultures, the mean number of foci per cell was 0.73 (p,0.001) at 15 min; it reached 0.90 by 3h (p,0.001) after irradiation, following which a decrease was observed. The same trend was detected when the fraction of cells with foci was considered: it reached 61% at 3h which is higher than the expected 47.4% of the cells (46% in control + 1.4% traversed). The increase in foci formation over the expected value was eliminated when the cells were incubated with a specific inhibitor of ATM (Ataxia Telangiectasia Mutated protein). Analyses of bystander effects in iron ion-irradiated cell cultures and the effect of partial oxygen tension on the kinetic of 53BP1 foci formation in low fluences alpha particle-irradiated cell populations are in progress.