Radiation - Radiation - Control of radiation risks: In view of the fact that radiation is now assumed to play a role in mutagenic or carcinogenic activity, any procedure involving radiation exposure is considered to entail some degree of risk. Direct exposure to these photons with doses from 0 to 4 Gy showed a clear dose responsive increase in the number of micronuclei and the number of bridges per 1000 binucleated cells for both cell lines (Figure 4). These results will facilitate refined estimates of the risk-benefit ratio of neutron therapy and may be valuable to those who are concerned about the health effects of exposure of space travel. These numbers clearly indicate that the RBEs for neutrons vary with the biological system, neutron energy and the end-point. All neutron irradiations were carried out using the fast clinical neutron therapy system (CNTS) at the University of Washington (Seattle, WA). We cultured GM15510 and GM15036 cells in ICCM obtained from the corresponding cell line that had been irradiated with cobalt-60 at doses equivalent to 5% of the neutron doses. Here we used the cytokinesis-block micronucleus assay to address the question whether neutrons induce a bystander effect in normal human lymphoblastoid cell lines. Different responses to neutrons were observed in the two cell lines we used. [71] reported RBEs of 2.35 and 2.42 for fast neutrons in immature rat hippocampal cells, as determined by two different cell viability assays. The frequencies of nucleoplasmic bridges (Figure 2b) showed considerable variation and no consistent dose-related response was seen for either cell line. Understanding the biological effects of neutrons may also enable more refined evaluations of the standards for radiation protection and safety. Two important stochastic effects of radiation are cancer, which results from mutations in nongerm cells (termed somatic cells), and heritable changes, which result from mutations in germ cells (eggs and sperm). Performed the experiments: IS JLS RDS RE MCJ. https://doi.org/10.1371/journal.pone.0098947.g001. No, Is the Subject Area "Malignant tumors" applicable to this article? No conclusive cytogenetic evidence exists to support or refute the existence of non-targeted effects in cellular responses to neutrons. Yang et al. https://doi.org/10.1371/journal.pone.0098947.g003. 4 Gy. Here we asked whether neutrons also induce a bystander effect in two normal human lymphoblastoid cell lines. Kinashi et al. While much has been learned about x-ray and gamma-ray effects on cells and whole organisms, less is known about the biological effects of neutrons. No, Is the Subject Area "Gamma radiation" applicable to this article? High levels of neutron irradiation occur in patients receiving neutron therapy, while low levels of neutron exposure occur in patients treated with high energy photons and protons. The principal kinds of ionizing radiation are: Yes Η. Chadwick Discussion 161 Differential effects of electrons and neutrons on cell killing and ability of DNA-membrane complex to synthesize DNA in E_. Neutrons are highly energetic uncharged particles that induce more severe DNA damage than photons and are therefore more effective than photons in controlling radioresistant tumors. The cell cultures were then incubated at 37°C for 28 hours. Two flasks per cell line were then shipped overnight to the University of Washington Medical Center (UWMC), Seattle. When the biological responses for both radiation types are linearly related to dose, as seen here, the RBE is also the ratio of the biological effects at the same dose. The same set of analyses were performed for the neutron bystander data which included replicate experiments as an additional variable. Knowing the RBE is important for radiation oncologists to determine the dose prescription and the most effective radiotherapy treatment plan for cancer patients. The inability of fast neutrons to induce a cytogenetic bystander effect as shown here may be due to different types of damage induced at the molecular level compared to photons. https://doi.org/10.1371/journal.pone.0098947.g002. https://doi.org/10.1371/journal.pone.0098947.t002, https://doi.org/10.1371/journal.pone.0098947.t003. Neutron irradiation to high fluence will lead to changes in the properties of the structural materials. Cells grown in ICCM from irradiated cells showed no significant increase in the frequencies of micronuclei or nucleoplasmic bridges compared to cells grown in ICCM from sham irradiated cells for either cell line. here. Although these cells had NDI values similar to the 3 Gy and 4 Gy photon-irradiated cells, they could not be scored because their morphology was not compatible with accurate damage assessments. Cells that were directly irradiated as well as those treated with conditioned media were centrifuged onto ethanol-cleaned microscope slides for 4 minutes at 93 x g using a cytocentrifuge (Statspin, Westwood, MA). Photons have been known to induce bystander effects in human cells for many years. For the gamma radiation experiments, flasks were transported to and from the laboratory and the radiation center, a 5-minute car ride, in an insulated container as described above. Bystander effects have been observed repeatedly in mammalian cells following photon and alpha particle irradiation. RBE values are known to depend on factors such as linear energy transfer, tissue type, the extent of biological damage, and dose [60]. This observation is in agreement with other reports [55], [68], [69]. As expected, GM15510 and GM15036 cells cultured in ICCM obtained from cells exposed to high doses of cobalt-60 gamma rays showed a 2 to 3 fold increase in micronuclei frequencies compared to sham treated controls, clearly indicating induction of a bystander effect (Figure 5a). Citation: Seth I, Schwartz JL, Stewart RD, Emery R, Joiner MC, Tucker JD (2014) Neutron Exposures in Human Cells: Bystander Effect and Relative Biological Effectiveness. [56] suggested that the gamma ray-induced bystander effect might have been suppressed by the neutron exposures. Since neutrons are scattered and absorbed by air, neutron radiation effects drop off rapidly with distance in air. These analyses were performed using JMP software version 6.0, SAS Institute Inc. Chi squared analyses were used to evaluate changes in the frequencies of micronuclei and bridges in the irradiated cells as a pooled group compared to the unirradiated (0-dose control) cells for the high dose photon experiments, and for the low dose photon contamination experiments. Exposure to very high levels of radiation, such as being close to an atomic blast, … Another possible explanation for the lack of a bystander effect with neutrons observed in this study may be the presence of dimethyl sulfoxide (DMSO), a scavenger of reactive oxygen species [64], [65], which was used to dissolve cytochalasin B that is required for the cytokinesis-block micronucleus assay. These cells were exposed to fast neutrons produced by targeting a near-monoenergetic 50.5 MeV proton beam at a Be target (17 MeV average neutron energy), and irradiated-cell conditioned media (ICCM) was transferred to unirradiated cells. Here, radiation-induced cell cycle delays precluded obtaining sufficient numbers of scoreable binucleated cells (Table 1). The average energy and neutron mean free path varies with field size and water-equivalent depth (nominal depth of maximum dose is 1.5 cm). However, since the neutron exposures and the low-dose photon exposures did not induce a bystander effect, it was important to verify that the cells and the experimental conditions in these experiments, including the serum used in the culture media, are capable of demonstrating a bystander effect if one exists. For cells irradiated with high doses of photons, a considerable amount of damage was attributed to the bystander component. [57] studied a neutron-induced bystander effect in boron neutron capture therapy with a cell survival assay as well as cloning and sequencing methods. Neutrons, depending on their energy, might be more effective in controlling certain tumor types where conventional photon therapy is ineffective [63] because the oxygen enhancement ratio, i.e. However, if the damage is not repaired correctly, a cell may die or eventually become cancerous. Nuclear radiation can ionise chemicals within a body, which changes the way the cells behave. There is also the possibility that a neutron-induced bystander effect, if any, might depend on cell type, the endpoint being evaluated [61], [62], and the energy of the neutrons. The Nuclear Division Indices (NDI) for all experimental conditions were high enough to enumerate micronuclei and bridges, with the exception of cells irradiated with the highest two neutron doses (3 Gy and 4 Gy). Ionizing radiation which can damage living tissue is emitted as the unstable atoms (radionuclides) change ('decay') spontaneously to become different kinds of atoms. Bridges were required to span the entire distance between the two nuclei. Copyright: © 2014 Seth et al. Some of the gamma radiation signs can be nausea, vomiting, bleeding, mouth sores, nausea, diarrhea and … Ionizing radiation is a form of energy that acts by removing electrons from atoms and molecules of materials that include air, water, and living tissue. Observation of an absence of a bystander effect in the present study may help explain the sensitivity of radioresistant tumor cells to neutrons, because there is a possibility that the protection otherwise provided by the bystander effect on the tumor in response to neutrons is absent or not strong enough in magnitude, thereby causing tumor cells to be killed. Cellular recognition of DNA damage and the subsequent repair processes may differ between neutrons and photons. [56] have suggested that neutrons might suppress gamma ray-induced bystander signaling. Biomedical/nuclear engineer ely5: While gamma or Xray radiation is just photons that have to rely on funny electric and quantum effects to manipulate the "macro" word, neutrons are large enough to act like actual projectiles to molecules of smaller atoms (e.g. This is also known as radiation poisoning. The cytokinesis-block micronucleus assay was used to quantify genetic damage in radiation-naïve cells exposed to ICCM from cultures that received 0 (control), 0.5, 1, 1.5, 2, 3 or 4 Gy neutrons. Radiation exists all around us, from both natural and manmade sources, and is in two forms: ionizing and non-ionizing radiation. These results may be relevant to radiation therapy with fast neutrons and for regulatory agencies setting standards for neutron radiation protection and safety. Since the doses of photons that contaminated these neutron exposures exceeded the minimum threshold for inducing a bystander effect [29], [58], Wang et al. Committee I of the International Commission on Radiological Protection has recommended that the Quality Factor for neutrons should be changed from 10 to 20. Department of Biological Sciences, Wayne State University, Detroit, Michigan, United States of America, Affiliation The percent contribution by the bystander exposure to the direct exposure was highest at the lowest dose delivered (0.5 Gy) and then it appears to saturate as dose increases, perhaps because there is saturation either of the bystander signals or the cellular responses to those signals [67]. To ensure that no cells were transferred along with the media, the supernatant was then passed through 0.22 µm polyethylsulfonate syringe filters (Nalgene). Ionizing radiation is also an effective method for treating tumors because it can be localized to the tumor and is a potent inducer of DNA double-strand breaks, a highly toxic form of DNA damage. For each treatment condition, at least 1000 binucleated cells were scored by trained observers. To ensure that the bystander effect observed was actually due to signals produced by the irradiated cells rather than an effect of exposure of the culture medium or an artifact of the media transfer process, a media-only control was included in which fresh media without cells was irradiated at the highest dose (4 Gy) prior to being transferred to non-irradiated cells. There are no available cytogenetic data concerning the bystander effect in human cells in response to neutrons. [72] used the cytokinesis-block micronucleus assay and a cell survival assay in a human colon carcinoma cell line and observed an RBE of 4.4 for neutrons in boron neutron capture therapy. How can ionizing radiation cause cancer? All slides were coded prior to scoring to prevent observer bias. However, the neutron beam has a photon dose-contamination of 5%, which may modulate a neutron-induced bystander effect. Contributed reagents/materials/analysis tools: IS JLS RDS RE MCJ JDT. This study used cells that lack gap junctions. Yes Our findings are in agreement with previous studies that have reported the lack of a bystander effect on neutron exposure using clonogenic cell survival assay in a human skin cell line [29] and zebrafish [56]. Neutron effects in humans: protection considerations. RBEs of 2.0±0.13 for micronuclei and 5.8±2.9 for nucleoplasmic bridges relative to cobalt-60 suggest that different kinds of genetic damage may be associated with different RBEs. Neutrons were listed as a carcinogen for the first time in the Eleventh Report on Carcinogens [2]. Yes If no bystander effect is induced in these cell lines, then it may be likely that neutrons do not have any ability to induce a bystander effect. Following exposure the flasks were returned immediately to the laboratory and incubated for 28 hours. Analyzed the data: IS JDT. We also included a media-only control containing fresh media without cells, which was irradiated at the highest dose, i.e. 0 (sham), 0.025, 0.05, 0.075, 0.10, 0.15 and 0.20 Gy. The authors thank Zainab Hammoud for assisting with the microscopy and Geoffrey Linn for his technical support. Conceived and designed the experiments: IS JDT RDS MCJ. Since serotonin has been reported to play a role in the bystander effect [49] and is light sensitive, the bottles containing the culture media were wrapped in aluminum foil and stored in the dark. Click through the PLOS taxonomy to find articles in your field. This change in quality of a mixed-spectrum field is significant since the biological damages produced by high-LET and low-LET radiations are not equivalent. Nuclear radiation arises from hundreds of different kinds of unstable atoms. This box was the same as that used to carry the flasks to and from the laboratory and the radiation center (a 2 minute walk). There is clear evidence of a bystander effect in response to high doses of photons when we used the same serum and cell lines as for the neutron experiments. For both cell lines and for both end-points, i.e. Wrote the paper: IS JLS RDS RE MCJ JDT. As expected, the media-only control and the 0 Gy control were not statistically different (p>0.05). The involvement of intracellular signaling molecules including mitogen-activated protein kinases (MAPK) and their downstream proteins [42], [43], protein kinase C (PKC) isoforms [44], tumor protein 53 (p53) [45], [46], cyclin-dependent kinase inhibitor 1A (CDKN1A, p21) [47], ataxia telangiectasia mutated protein (ATM) [44], and ataxia telangiectasia and Rad3 related (ATR) DNA-dependent protein kinase (DNA-PK) [44], [48] have also been implicated. The dependent variables were micronuclei and bridges. A bystander effect following neutron exposure has been observed in Chinese hamster ovary cells [57], but no effect was seen in zebrafish irradiated in vivo [56]. In health physics, neutron radiation is a type of radiation hazard. Bryant Our observation of an absence of a bystander effect following doses of photons that are associated with exposure to neutrons confirmed that there is a lack of a bystander effect in response to neutrons, regardless of the presence of photons. The 1 Gy data are missing for GM15510 cells replicate 2 because the sample was lost. For more information about PLOS Subject Areas, click For each cell line, the neutron bystander experiment was performed twice, once each on different days. The slides were then mounted with 1X PBS and 25×25 mm2 glass coverslips. However, this possibility seems unlikely in the work described here because we observed a bystander effect with an identical procedure involving DMSO when the same cell lines were exposed to photons. È­ÛMž#: ÑòKúXÜÏáõ‡Wp© zx—t©6×‡ÏŠ(Όp¼À8›®©ÄÉUhh@p]´Bñ³©Ëɘ–.é؍N­û#>®¢ð®"a-¹qyußׂqßZ¹•’w{¤×7dôSú0Þw~y .eŸù4¡ŒQÏlD¡â1XVMÏp?N«Ý‹˜Od &¸V~”ÕnD^æqÓWl ± ÏôÀZ¼.¸. Some of the reactions occur quickly, while others occur slowly. Micronuclei were required to be no more than one-third the size of the nuclei, and to be round or oval with smooth edges and stained the same color as the nuclei. There is a possibility that a neutron-induced bystander effect requires physical contact between cells, which could be tested by performing experiments using cell lines such as fibroblasts and keratinocytes that have gap junctions. [59] found that transplantation of a mixture of neutron irradiated and unirradiated bone marrow cells into mice induced instability in the descendants of unirradiated cells as confirmed by measuring chromosomal aberrations, indicating that neutrons induce a bystander effect. The serum used in this study was prescreened for its ability to support a bystander effect with the cytokinesis-block micronucleus assay in these same cell lines. The effects of radiation on the human body Radioactive materials are hazardous. https://doi.org/10.1371/journal.pone.0098947.t004. Nucleoplasmic bridges (Figure 5b) showed a weak bystander effect for GM15510 (p = 0.052), and for GM15036 the effect was highly significant (p<0.0001). The Tukey HSD test was used for post-hoc evaluations. The cell cultures were then incubated at 37°C for 28 hours. C. Medvedovsky and B. V. Worgul (1991) Neutron Effects on the Lens.Radiation Research: October 1991, Vol. Although we did not measure apoptosis or necrosis, these outcomes may influence the responses seen in these cell lines. There is a large difference in the magnitude of the biological effects of nonionizing radiation (for example, light and microwaves) and ionizing radiation, emissions energetic enough to knock electrons out of molecules (for example, α and β particles, γ rays, X-rays, and high-energy ultraviolet radiation) (Figure \(\PageIndex{2}\)). 47–51 In these five reports, most patients were treated for severe laryngeal necrosis (Chandler grade 3 or 4). However, if a very small bystander effect was in fact induced by neutrons, then it may have been obscured by the DMSO, whereas the bystander effect induced by high levels of photons was too large to be masked by DMSO. Ionizing radiationhas so much energy it can literally knock the electrons out of any atom it … Even though a bystander effect was not observed with the neutron exposures, we sought to determine whether photons might cause a bystander effect at the doses employed in these experiments. To determine whether these low doses of contaminating photons can induce a bystander effect, cells were irradiated with cobalt-60 at doses equivalent to the percent contamination for each neutron dose. Recently, some laboratories have suggested that the presence of serotonin in the serum is one of the key factors involved in the bystander effect [49]–[51], however this finding has been disputed [52]. But the neutronic flash does not last. The neutron beam is shaped by a primary collimator composed of iron and a secondary collimator made up of individually movable leaves composed of iron with cylindrical polyethylene inserts. Space radiation also has very different effects on human DNA, cells and tissues. For both cell lines, direct irradiation with neutrons resulted in clear dose-responsive increases in the number of micronuclei (Figure 1a) and nucleoplasmic bridges (Figure 1b) per 1000 binucleated cells. 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