Exogenous carbon monoxide suppresses adaptive response induced in zebrafish embryos in vivo by microbeam protons

Dechorionated embryos of the zebrafish, Danio rerio, irradiated at 5 h post-fertilization (hpf) with 30 protons delivered to 10 separate positions each with an energy of 3.4 MeV from the microbeam irradiation facility (Single-Particle Irradiation System to Cell, acronym as SPICE) at the National Institute of Radiological Sciences (NIRS), developed radioadaptive response (RAR) against a subsequent challenging exposure of 2 Gy of X-ray irradiation at 10 hpf, corroborated by reduced apoptotic signals at 25 hpf revealed through terminal dUTP transferase-mediated nick end-labeling assay. \nThe effects of the CO liberator tricarbonylchloro(glycinato)ruthenium (II) (CORM-3) on the induction of RAR were examined by transferring the irradiated embryos to freshly prepared medium with the chemical at different time points after the application of the priming dose. Our results showed that transfer of irradiated embryos into media with CORM-3 at 0, 1, 2 and 3 h after application of priming exposure significantly suppressed RAR, while transfer at 5 h did not suppress RAR. This was attributed to the protection of bystander cells from the released CO, which caused less de novo synthesis of factors and thus less efficient induction of RAR. Once the factors were synthesized, RAR was induced, which would not be further affected by the application of CORM-3 introduced at 5 h after the application of the priming dose

Studies of non-targeted radiation effects through microbeam irradiated embryos of Danio rerio.

Linear no-threshold (LNT) hypothesis, which assumes that the risk from an ionizing-radiation exposure is linearly proportional to the dose normalized by the radiation weighting factor, is commonly adopted for radiation protection considerations. Data in the low-dose regime are relatively scarce, so the detrimental effect from exposure to low-dose radiation is commonlyextrapolated from data obtained in the high-dose regime by using the LNT model. There is a considerable amount of evidence showing that organisms may exhibit different responses to a low-dose exposure from that to a high-dose exposure. As such, the present project was focusing on the study of non targeted radiation effects, including the radiation induced radioadaptive response (RAR) and hormetic effect through the use of proton microbeam. The dose response at low-dose exposure was also studied. A better understanding of these non-targeted radiation effects can give a better estimation on the radiation risk which forms the main purpose of the research. The microbeam irradiation system (Single-Particle Irradiation System to Cell, acronym as SPICE) at the National Institute of Radiological Sciences(NIRS), Japan, was originally designed for radiobiological studies. In our studies, we made use of the SPICE to irradiate the zebrafish (Danio rerio) embryos with a control of the irradiation spots. Protons with an initial energy of 3.4 MeV would first travel through a Si3N4 exit window with a thickness of 100 nm, and then through a 2.5 μm Mylar film with less than 50 μm air gap between the exit window and Mylar film before the protons finally reached the target [1].第４回共用施設（PASTA&SPICE, NASBEE)共同研究成果報告

Response of 5 hpf zebrafish embryos to low-dose microbeam protons

The microbeam irradiation system (single-particle irradiation system to cell, acronym as SPICE) at the National Institute of Radiological Sciences (NIRS), Japan, was employed to irradiate dechorionated embryos of the zebrafish, Danio rerio, at 5 h post-fertilization (hpf) by protons each having an energy of ∼3.4 MeV. Either 1 or 10 positions on the cells were irradiated with different number of protons. The levels of apoptosis in zebrafish embryos at 25 hpf were quantified through terminal dUTP transferase-mediated nick end-labeling assay. Triphasic dose–esponses were obtained (Fig. 1), including (i) a subhormetic zone with an increase in apoptotic signals for a small number of irradiated protons per position, (ii) a hormetic zone with a reduction in the apoptotic signals below the spontaneous level for a larger number of irradiated protons per position and (iii) a toxic zone with an increase in apoptotic signals again if the number of irradiated protons per position was further increased

Roles of nitric oxide in adaptive response induced in zebrafish embryos in vivo by microbeam protons

Radioadaptive response (RAR) was successfully induced in dechorionated (5 h post-fertilization, hpf) embryos of the zebrafish, Danio rerio, by 3.4 MeV protons from the microbeam irradiation facility (Single-Particle Irradiation System to Cell, acronym as SPICE) [ 1] at the National Institute of Radiological Sciences (NIRS), against a challenging exposure of 2 Gy of X-ray irradiation at 10 hpf. The RAR induction was corroborated by reduced apoptotic signals at 25 hpf revealed through terminal dUTP transferase-mediated nick end-labeling assay. If de novo synthesis of factors was required for RAR induction, these should have already been synthesized at 5 h after the priming dose. \nApplication of a nitric oxide scavenger 2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) to the medium at 0, 1, 2, 3 or 5 h after application of priming exposure significantly suppressed RAR. The suppression of RAR with the application of cPTIO to the medium at 5 h after the priming dose irradiation, where de novo synthesis of factors should have been completed, suggested that NO scavenging impaired the repair machineries in the bystander cells. The suppression of RAR with the application of cPTIO to the medium at earlier than 5 h after the priming dose irradiation could be explained by the scavenging of bystander NO signals in the medium and thus deterring the de novo synthesis of factors