Reduction of arthritic symptoms by low dose radiation therapy (LD-RT) is associated with an anti-inflammatory phenotype

Abstract

Purpose: The aim of this paper was to provide an integrative mechanistic appraisal to account for consistent observations of protective effects of ionizing radiation on the occurrence/ progression of arthritis in multiple animal models.

Materials and methods: A critical analysis of the biomedical literature was undertaken to assess mechanisms by which low doses of ionizing radiation prevent and/or reduce the occurrence of experimental-induced arthritis in animal models.

Results: Detailed mechanistic-related research indicates that low doses of ionizing radiation induce a highly integrated multiple pathway process that results in the formation of a generalized anti-inflammatory phenotype which can both prevent the occurrence of arthritic changes and/or reverse such effects.

Conclusions: The manifestation of the anti-inflammatory features occurred within the context of highly consistent hormetic (i.e., biphasic dose) responses across studies, biological models and mechanisms. The reduction of multiple bioindicators of experimentally-induced arthritis by exposure to low doses of ionizing radiation was associated with the occurrence of a generalized anti-inflammatory phenotype.     

Oxidative and nitrative stress-related changes in human lens epithelial cells following exposure to X-rays

Purpose: There is limited understanding of the mechanistic effects of ionizing radiation (IR) exposure in cataract formation. In this study, we explored the effects of IR on reactive oxygen/nitrogen species (ROS and RNS) generation in human lens epithelial (HLE) cells as an early key event to long-term damage.

Materials and methods: HLE cell-line was exposed to X-rays at varied doses (0–5?Gy) and dose-rates. Cell lysates and supernatants were collected 20?h post-exposure and analysed for viability, cell cycling and metabolites of ROS (p, m-, o-, tyrosines, 3-chlorotyrosine (cl-tyrosine), 8-hydroxy deoxyguanosine, (8-OH-dG) and RNS (3-nitrotyrosine).

Results and conclusions: HLE cell-line exhibited a bi-phasic response in terms of cell viability, ROS and RNS profiles. At doses <0.5?Gy, ROS and RNS levels were lower than control and at higher doses (>0.5?Gy) a steady increase was observed in each metabolite. This response was observed irrespective of dose-rate. Among the associations tested, cl, p, m-tyrosine and 3-nitrotyrosine revealed changes (p?相似文献   

Fractionated exposure to low doses of ionizing radiation results in accumulation of DNA damage in mouse spleen tissue and activation of apoptosis in a p53/Atm-independent manner

Purpose: While the effects of high doses of ionizing radiation (IR) are relatively well characterized, the molecular mechanisms underlying cellular responses to prolonged exposure to low doses of radiation remain largely under-investigated.

Materials and methods: Here, we addressed the DNA damage and apoptotic response in the spleen tissue of C57BL/6 male mice after fractionated exposure to X-rays within the 0.1–0.5?Gy dose range.

Results: The response to initial exposure to 0.1?Gy of IR was characterized by increased DNA damage and elevated levels of apoptosis. Subsequent exposures (cumulative doses of 0.2 and 0.3?Gy) resulted in adaptive response-like changes, represented as increased proliferation and apoptotic response. Cumulative doses of 0.4 and 0.5?Gy were characterized by accumulation of DNA damage and reactivation of apoptosis and apoptosis-related proteins. Additionally, spleen cells with irreversible damage caused by radiation can undergo apoptosis via activation of p38, which does not necessarily involve the Atm/p53 pathway.

Conclusions: Fractionated exposure to low doses of X-rays resulted in accumulation of DNA damage in the murine spleen and induction of apoptotic response in p53/Atm-independent manner. Further studies are needed to understand the outcomes and molecular mechanisms underlying cellular responses and early induction of p38 in response to prolonged exposure to IR.     

Reactive oxygen species mediates 50-Hz magnetic field-induced EGF receptor clustering via acid sphingomyelinase activation

Purpose: Exposure to extremely low frequency electromagnetic fields (ELF-EMFs) could elicit biological effects including carcinogenesis. However, the detailed mechanisms by which these ELF-EMFs interact with biological system are currently unclear. Previously, we found that a 50-Hz magnetic field (MF) exposure could induce epidermal growth factor receptor (EGFR) clustering and phosphorylation on cell membranes. In the present experiment, the possible roles of reactive oxygen species (ROS) in MF-induced EGFR clustering were investigated.

Materials and methods: Human amnion epithelial (FL) cells were exposed to a 50-Hz MF with or without N-acetyl-l-cysteine (NAC) or pyrrolidine dithiocarbamate (PDTC). EGFR clustering on cellular membrane surface was analyzed using confocal microscopy after indirect immunofluorescence staining. The intracellular ROS level and acid sphingomyelinase (ASMase) activity were detected using an ROS assay kit and an Amplex^® Red Sphingomyelinase Assay Kit, respectively.

Results: Results showed that exposure of FL cells to a 50-Hz MF at 0.4?mT for 15?min significantly enhanced the ROS level, induced EGFR clustering and increased ASMase activity. However, pretreatment with NAC or PDTC, the scavenger of ROS, not only counteracted the effects of a 50-Hz MF on ROS level and AMS activity, but also inhibited the EGFR clustering induced by MF exposure.

Conclusions: The present and previous data suggest that ROS mediates the MF-induced EGFR clustering via ASMase activation.     

A small fish model for quantitative analysis of radiation effects using visualized thymus responses in GFP transgenic medaka

Abstract

Purpose: The aim of this study was to establish a new method of real-time, in vivo detection of radiation damage and recovery.

Methods: The thymus was observed under fluorescent light in a green fluorescent protein transgenic medaka. After irradiation, medaka thymus images were analyzed to quantify the effects of radiation by measuring changes in thymus size. A single acute irradiation of X-rays (0–30?Gy) or heavy Fe ions (0–10?Gy) was delivered to the medaka. Images were captured 0, 1, 2, 3, 5, 7, 11, and 21 d after irradiation. Dose-response assessment was conducted to provide a direct measurement of the effects of the radiation.

Conclusion: A biomonitoring system to detect the effects of radiation in real time was established. Using this system, the threshold doses for the induction of thymic atrophy by acute X-rays and Fe ions were 2–5?Gy and 0.5–1?Gy, respectively. The Relative Biological Effectiveness (RBE) of Fe-ion to X-rays was estimated to be around 3. This system may be used to evaluate the risk from concurrent exposure to hazards, such as chemicals and radiation, and for aging research.     

Ferulic acid (FA) abrogates ionizing radiation-induced oxidative damage in murine spleen

Purpose: The spleen is a crucial organ manifesting immune functions. Thus, radiation-induced oxidative challenge is vulnerable for the spleen. Our major objective was to protect the spleen from radiation-induced anomalous situations and to identify the signaling pathways involved.

Materials and methods: Swiss albino mice were treated with ferulic acid (FA) once in a day at a dose of 50?mg/kg body weight for 5 consecutive days before exposing them to single dose of 10?Gy irradiation. The ROS generation and MMP change were determined by flow cytometry. The expression of different signaling proteins was investigated by immunoblotting and immunocytochemistry.

Results: FA pretreatment significantly prevented radiation-induced oxidative stress by downregulating TBARS formation and by upregulating SOD and catalase activity. FA scavenged ROS, prevented the alteration of MMP and downregulated the expression of stress marker Cdc42 and apoptotic markers p⁵³, p²¹, Bax and PTEN. Cell cycle analysis showed DNA damage induced arrest of cells at subG0/G1 phase. Moreover, pretreatment with FA augmented Bcl2 expression and also increased the level of p-PI3K.

Conclusion: FA prevented the activation of apoptotic signaling events in the spleen by interfering with the free radical chain reaction and by scavenging superfluous ROS. This is perhaps the first comprehensive study with a mechanistic viewpoint that FA can protect the spleen from ionizing radiation.     

Effects of ionizing radiation on DNA methylation: from experimental biology to clinical applications

Purpose: Ionizing radiation (IR) is a ubiquitous environmental stressor with genotoxic and epigenotoxic capabilities. Terrestrial IR, predominantly a low-linear energy transfer (LET) radiation, is being widely utilized in medicine, as well as in multiple industrial applications. Additionally, an interest in understanding the effects of high-LET irradiation is emerging due to the potential of exposure during space missions and the growing utilization of high-LET radiation in medicine.

Conclusions: In this review, we summarize the current knowledge of the effects of IR on DNA methylation, a key epigenetic mechanism regulating the expression of genetic information. We discuss global, repetitive elements and gene-specific DNA methylation in light of exposure to high and low doses of high- or low-LET IR, fractionated IR exposure, and bystander effects. Finally, we describe the mechanisms of IR-induced alterations to DNA methylation and discuss ways in which that understanding can be applied clinically, including utilization of DNA methylation as a predictor of response to radiotherapy and in the manipulation of DNA methylation patterns for tumor radiosensitization.     

Temporal variability of the quality of Taraxacum officinale seed progeny from the East-Ural radioactive trace: is there an interaction between low level radiation and weather conditions?

Hypothesis: The multiple stressors, in different combinations, may impact differently upon seed quality, and low-level doses of radiation may enhance synergistic or antagonistic effects.

Results: During 1991–2014 we investigated the quality of the dandelion (Taraxacum officinale s.l.) seed progeny growing under low-level radiation exposure at the East-Ural Radioactive Trace (EURT) area (result of the Kyshtym accident, Russia), and in plants from areas exposed to background radiation. The viability of the dandelion seed progeny was assessed according to chronic radiation exposure, accounting for the variability of weather conditions among years. Environmental factors (temperature, precipitation, and their ratio in different months) can modify the radiobiological effects. We found a wide range of possible responses to multiple stressors: inhibition, stimulation, and indifferent effects in different seasons.

Conclusion: The intraspecific variability of the quality of dandelion seed progeny was greatly increased under conditions of low doses of chronic irradiation. Temperature was the most significant factor for seed progeny formation in the EURT zone, whereas the sums of precipitation and ratios of precipitation to temperature dominantly affected organisms from the background population.     

Radiation hormesis: Autophagy and other cellular mechanisms

Abstract

Purpose: To review the cellular mechanisms of hormetic effects induced by low dose and low dose rate ionising radiation in model systems, and to call attention to the possible role of autophagy in some hormetic effects.

Results and conclusions: Very low radiation doses stimulate cell proliferation by changing the equilibrium between the phosphorylated and dephosphorylated forms of growth factor receptors. Radioadaptation is induced by various weak stress stimuli and depends on signalling events that ultimately decrease the molecular damage expression at the cellular level upon subsequent exposure to a moderate radiation dose. Ageing and cancer result from oxidative damage under oxidative stress conditions; nevertheless, ROS are also prominent inducers of autophagy, a cellular process that has been shown to be related both to ageing retardation and cancer prevention. A balance between the signalling functions and damaging effects of ROS seems to be the most important factor that decides the fate of the mammalian cell when under oxidative stress conditions, after exposure to ionising radiation. Not enough is yet known on the pre-requirements for maintaining such a balance. Given the present stage of investigation into radiation hormesis, the application of the conclusions from experiments on model systems to the radiation protection regulations would not be justified.     

The soluble receptor ST2 is positively associated with occupational exposure to radiation

Purpose Radiation exposure, besides the risk of cancer, may also increase the risk of non-cancer diseases, including cardiovascular disease (CVD). This study investigates whether the soluble form of the ST2 receptor (sST2), an emerging prognostic marker in patients with CVD, can be used to monitor the CVD risk in individuals occupationally exposed to radiation.

Materials and methods sST2 in blood plasma from 69 individuals, 45 workers from the nuclear industry and 24 controls, was analyzed using enzyme-linked assay (ELISA). Total antioxidant status (TAS) of blood plasma and levels of reactive oxygen species (ROS) in lymphocytes were determined by colorimetric and fluorescence assays.

Results The data suggest a 5-fold increase in the number of subjects with sST2 levels above the clinical threshold and a 10-fold increase in the number of subjects with TAS levels outside the reference range in the exposed group when compared to the group of non-exposed individuals. The strongest up-regulation of TAS was measured in the group of younger workers with cumulative doses not exceeding 50 mSv.

Conclusion The present study may represent an initial step towards the establishment of sST2 as a biomarker for CVD risk estimation in the context of radiation exposure.     

Mapping the research trends on the biological effects of radiation less than 100 mSv: a bibliometric analysis for 30 years publication

Abstract

Purpose: The purpose of this study is to evaluate the global scientific publication of biological research of low dose radiation for the past 30 years and provide the insights into the characteristics of research activities and major topics regarding biological effects of low dose radiation exposure.

Materials and methods: We bibliometrically investigated the biological research publication of radiation exposure less than 100 mSv. References published from 1987 to 2016 were achieved from the Scopus database and filtered by several criteria such as publication types, research fields, and radiation dose range.

Results: Total 753 references were assembled for the bibliometric analysis on the biological studies of radiation effect less than 100 mSv. It provided fundamental knowledge of research, including production tendency, contribution, impact journals, and major research themes. Based on the keyword analysis, we found that specific topics on the biological response to radiation exposure have been changed from the examination of low dose radiation-induced phenomena to the investigation of how to induce a physiological response. In addition, featured articles showed the various views on the biological effects of radiation less than 100 mSv in 30 years publication, depending on radiation doses and types.

Conclusions: Continuous studies in large programs of low dose radiation led to the increment of research achievements in accordance with societal needs in radiation safety regulation for health protection. Our findings can surely help radiation researchers to gain insights and penetration in low dose risk research for radiation protection, and establish a further research direction.     

Mechanisms of inflammatory responses to radiation and normal tissues toxicity: clinical implications

Purpose: Cancer treatment is one of the most challenging diseases in the present era. Among a few modalities for cancer therapy, radiotherapy plays a pivotal role in more than half of all treatments alone or combined with other cancer treatment modalities. Management of normal tissue toxicity induced by radiation is one of the most important limiting factors for an appropriate radiation treatment course. The evaluation of mechanisms of normal tissue toxicity has shown that immune responses especially inflammatory responses play a key role in both early and late side effects of exposure to ionizing radiation (IR). DNA damage and cell death, as well as damage to some organelles such as mitochondria initiate several signaling pathways that result in the response of immune cells. Massive cell damage which is a common phenomenon following exposure to a high dose of IR cause secretion of a lot of inflammatory mediators including cytokines and chemokines. These mediators initiate different changes in normal tissues that may continue for a long time after irradiation. In this study, we reviewed the mechanisms of inflammatory responses to IR that are involved in normal tissue toxicity and considered as the most important limiting factors in radiotherapy. Also, we introduced some agents that have been proposed for management of these responses.

Conclusions: The early inflammation during the radiation treatment is often a limiting factor in radiotherapy. In addition to the limiting factors, chronic inflammatory responses may increase the risk of second primary cancers through continuous free radical production, attenuation of tumor suppressor genes, and activation of oncogenes. Moreover, these effects may influence non-irradiated tissues through a mechanism named bystander effect.     

Autophagy suppresses radiation damage by activating PARP-1 and attenuating reactive oxygen species in hepatoma cells

Abstract

Purpose: To investigate the relationship between autophagy and radiation damage of human hepatoma cells and to explore the role of reactive oxygen species (ROS).

Materials and methods: HepG2 cells were exposed to X-rays, then the protein expressions of microtubule-associated protein 1 light chain 3 (LC3) and poly ADP-ribose polymerase-1 (PARP-1) were measured by Western blot assay, the formation of autophagosomes was detected by an autophagy detection kit, the intracellular ROS level was measured by flow cytometer, and DNA damage was evaluated by the incidence of micronuclei (MN). A CCK-8 kit was used to measure the proliferation ability of irradiated cells with or without N-acetyl-l-cysteine (NAC) treatment. In some experiments, the hepatoma cells were transferred with LC3 siRNA or PARP-1 siRNA before irradiation.

Results: The protein expressions of LC3 and PARP-1 and the inductions of autophagosomes and intracellular ROS were increased in the irradiated HepG2 cells. Pretreatment of cells with NAC relieved the irradiation-induced inhibition of cell proliferation. When HepG2 cells were transfected with the LC3 siRNA, the over-expression of PARP-1 was diminished in the irradiated cells. Compared with the control group, the inhibitions of LC3 and PARP-1 increased ROS level in the irradiated HepG2 cells and hence sensitized radiation responses of both proliferation inhibition and MN induction.

Conclusion: Autophagy upregulates the expression of PARP-1 and relieves radiation damage by reducing the generation of ROS.     

Gossypetin,a naturally occurring hexahydroxy flavone,ameliorates gamma radiation-mediated DNA damage

Abstract

Purpose: To evaluate the protective effect of gossypetin (GTIN) against gamma (γ)-radiation-mediated DNA damage.

Materials and methods: Increasing concentrations (10–150 μM) of GTIN were incubated with supercoiled DNA 1 h prior exposure to γ-radiation in the range of 5-Gy absorbed dose from Co⁶⁰ γ source. To establish the effective protective concentration of GTIN, supercoiled DNA was pre-incubated with 50 μM of GTIN for 1 h followed by exposure of 5, 10 and 20 Gy doses of γ-radiation. Moreover, 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical, hydroxyl radical, nitric oxide (NO) scavenging, metal chelating activity and ferric reducing antioxidant power (FRAP) of GTIN were measured and compared with standards. The flowcytometric analysis and radiation-induced genomic DNA damage by comet assay were employed to estimate the level of intracellular reactive oxygen species (ROS) using isolated murine hepatocytes.

Results: GTIN was able to effectively scavenge different free radicals in in vitro situations. It could significantly prevent radiation induced supercoiled and genomic DNA damage with reduced comet parameters. It also acted as a potent scavenger of the radiation induced ROS.

Conclusions: GTIN ameliorated radiation-induced oxidative stress and DNA damage by its free-radical scavenging activity.     

Late health effects of radiation exposure: New statistical,epidemiological, and biological approaches*

Abstract

Purpose: The 2012 Conference on Radiation and Health in Kennebunkport, Maine, USA, brought together epidemiologists, statisticians, basic scientists, and clinical scientists interested in the health effects of radiation exposure due to medical, diagnostic, occupational, and non-medical sources, to review the current status of epidemiologic and clinical research on radiation exposure in relation to risk of breast, thyroid cancer, and leukemia, cardiopulmonary events, and other late effects. Topics discussed included synergy between radiation exposure and genetic background; late effects of radiation therapy in childhood cancer survivors and several other medically exposed cohorts; leukemia risk seen in Russian and Chernobyl studies, and leukemia risk from computed tomography scans in childhood.

Results and conclusions: This report summarizes the presentations at the meeting and discusses their significance in light of earlier studies and of other ongoing research.     

The effects of chronic gamma irradiation on oxidative stress response and the expression of anthocyanin biosynthesis-related genes in wheat (Triticum aestivum)

Abstract

Purpose: To investigate the mechanisms of adaptation and tolerance to ionizing radiation using chronic radiation in wheat.

Materials and methods: We exposed wheat plants to chronic gamma irradiation (50 Gy) for 2, 4, and 6 weeks and measured various biological parameters.

Results: Plant height was reduced by exposure to gamma irradiation; this effect increased with increasing exposure time. Photosynthetic pigment levels decreased with increasing exposure time, while anthocyanin levels significantly increased after exposure to gamma rays. The activities of antioxidant enzymes (superoxide dismutase [SOD], ascorbate peroxidase [APX], catalase [CAT], and peroxidase [POD]) and malondialdehyde (MDA) levels increased with increasing duration of exposure to gamma irradiation. Electron spin resonance (ESR) signals were strongly detected in wheat that was gamma-irradiated for two weeks and then gradually decreased with increasing exposure time. The expression of anthocyanin biosynthesis genes (flavanone 3-hydroxylase [F3H], dihydroflavonol reductase [DFR], anthocyanin reductase [ANS], and UDPG-flavonoid glucosyl transferase [UFGT]) and sugar contents increased after exposure to gamma rays.

Conclusions: This suggests that exposure to ionizing radiation according to increase of exposure time has led to efficient induction of anthocyanin and antioxidant enzyme activities. This study indicates that reactive oxygen species (ROS) is eliminated by biosynthesis of anthocyanin and antioxidant enzymes. This study helps elucidate the biological effects of various durations of low-dose exposure to chronic gamma radiation in wheat plants.     

Exposure to electromagnetic field attenuates oxygen-glucose deprivation-induced microglial cell death by reducing intracellular Ca2+ and ROS

Purpose The aim of this research was to demonstrate the protective effects of electromagnetic field (EMF) exposure on the human microglial cell line, HMO6, against ischemic cell death induced by in vitro oxygen-glucose deprivation (OGD).

Materials and methods HMO6 cells were cultured for 4?h under OGD with or without exposure to EMF with different combinations of frequencies and intensities (10, 50, or 100?Hz/1?mT and 50?Hz/0.01, 0.1, or 1?mT). Cell survival, intracellular calcium and reactive oxygen species (ROS) levels were measured.

Results OGD caused significant HMO6 cell death as well as elevation of intracellular Ca²⁺ and ROS levels. Among different combinations of EMF frequencies and intensities, 50?Hz/1?mT EMF was the most potent to attenuate OGD-induced cell death and intracellular Ca²⁺ and ROS levels. A significant but less potent protective effect was also found at 10?Hz/1?mT, whereas no protective effect was found at other combinations of EMF. A xanthine oxidase inhibitor reversed OGD-induced ROS production and cell death, while NADPH oxidase and mitochondrial respiration chain complex II inhibitors did not affect cell death.

Conclusions 50?Hz/1?mT EMF protects human microglial cells from OGD-induced cell death by interfering with OGD-induced elevation of intracellular Ca²⁺ and ROS levels, and xanthine oxidase is one of the main mediators involved in OGD-induced HMO6 cell death. Non-invasive treatment of EMF radiation may be clinically useful to attenuate hypoxic-ischemic brain injury.     

低温等离子体联合辐射对三种癌细胞系放射增敏效应的研究

目的 探讨低温等离子体对人肝癌细胞系HepG2、非小细胞肺癌细胞系A549及人宫颈癌细胞系HeLa的放射增敏作用及其机制。方法 应用克隆形成实验观察低温等离子体对3种细胞的放射增敏作用;流式细胞仪分析3种细胞周期分布、凋亡率及活性氧含量;Western blot法检测单纯照射(R组)、单纯等离子体作用(P组)及其联合辐射(P+R组)对3种细胞中Bcl-2、Caspase-3蛋白表达的影响。结果 低温等离子体对HepG2、A549及HeLa细胞均有放射增敏作用,放射增敏比(SER_D0)分别为1.28、1.32、1.29。HepG2、A549及HeLa细胞P+R组G₂/M期比例、凋亡率及活性氧含量与R组比较均明显增高(t_G2/M期=9.52、8.24、9.53,P<0.05;t_凋亡率=10.67、38.56、6.74,P<0.05;t_{活性氧含量}=9.41、15.42、13.53,P<0.05)。HepG2和A549细胞P+R组G₂/M期比例、凋亡率及活性氧含量与P组比较均明显升高(t_G2/M期=8.75、20.37,P<0.05;t_凋亡率=8.43、9.99,P<0.05;t_{活性氧含量}=4.82、5.27,P<0.05)。3种癌细胞中P+R组Bcl-2蛋白表达较R组降低,而Caspase-3蛋白表达升高。结论 低温等离子体可提高HepG2、A549及HeLa细胞系的放射敏感性,其对HepG2及A549细胞系的放射增敏机制可能与抑制亚致死损伤修复,使细胞周期阻滞在G₂/M期,以及提高细胞内ROS水平诱导细胞凋亡有关。     

Role of Nrf2-antioxidant in radioprotection by root extract of Inula racemosa

Abstract

Purpose: Inula racemosa, a Trans-Himalayan plant is an important medicinal herb. In this study, the radio-modulatory efficacy of aqueous root extract of I. racemosa was investigated.

Materials and methods: Normal Kidney Epithelium cells were treated with extract (50–200?μg/ml) and exposed to 3?Gy of γ-radiation, while C57BL/6 mice were administered with extract (300–600?mg/kg BW) intraperitoneally prior to exposure to 7.5?Gy of γ-radiation to assess radiation modulatory efficacy.

Results: The administration of extract (30?min and 1?h) prior to radiation exposure improved the survival of NKE cells (as measured by proliferation), restored MMP and ROS levels as compared to radiation-exposed alone cells. These cells showed up-regulated Nrf2 protein levels at 7?h and increased expression of HO-1 and NOQ1 protein at 24?h In mice, the 30 days whole body survival study demonstrated that extract pre-treatment increases survival or delays the onset of radiation-induced mortality as against 70% mortality of 7.5?Gy of γ-radiation.

Conclusions: The aqueous extract of roots of I. racemosa enhanced the survival of irradiated NKE cells and rescued C57BL/6 mice against WBI-induced mortality. The radiation modulation efficacy was mediated through cumulative activation of HO-1 and NQO1 downstream of Nrf2 translocation in NKE cells.

Abbreviations:

ARE: Antioxidant Response Element; FITC: Fluorescein isothiocyanate; GCS: Glutamylcysteine synthase; HO-1: Heme oxygenase-1; LPS: Lipopolysacharide; MRP: Multidrug Resistance-Associated Proteins; NQO1: NAD(P)H Quinone Dehydrogenase 1; NRH: Quinone Oxidoreductase 2 (NQO2); PBS: Phosphate Buffer Saline; PKA: Protein Kinase A; PKC: Protein Kinase C; PI3-kinase: Phosphatidylinositol 3-Kinase; SRB: Sulforhodamine B; UV: Ultra-Violet radiation     

Evaluation of the premature chromosome condensation scoring protocol after proton and X-ray irradiation of human peripheral blood lymphocytes at high doses range

Purpose of the study: One of the main difficulties in radiation dose assessment is cells inability to reach mitosis after exposure to acute radiation. Premature chromosome condensation (PCC) has become an important method used in biological dosimetry in case of exposure to high doses. Various ways to induce PCC including mitotic cells fusion, chemical stimulation with calyculin A or okadaic acid give wide spectrum of application. The main goal of this study was to evaluate the utility of drug-induced PCC scoring procedure by testing 2 experimental modes where 150 and 75 G2/M-PCC phase cells were analyzed after exposure to high dose proton and X-ray radiation. Another aim is to determine the differences in cellular response induced by proton and photon radiation using a HPBL in vitro model as a further extension of our previous studies involving doses up to 4.0?Gy.

Materials and methods: Total body exposure was simulated by irradiating whole blood collected from a healthy donor. Whole blood samples were exposed to two radiation types: 60?MeV protons and 250 kVp X-rays in the dose range of 5.0–20.0?Gy, the dose rate for protons was 0.075 and 0.15?Gy/s for X-rays. Post 48?h of human peripheral blood lymphocytes (HPBL) culture, calyculin A was added. After Giemsa staining, chromosome spreads were photographed and manually analyzed by scorers in the G2/M-PCC phase. In order to check the consistency of obtained results all scorers followed identical scoring criteria. Additionally, PCC index kinetics was evaluated for first 500 cells scored.

Conclusions: Here we provide a different method of results analysis. Presented dose-response curves were obtained by calculating the value of counted excess chromosome fragments. The results indicated that obtained dose estimates as adequate in the high dose range till 18.0?Gy for both studied radiation types, giving an opportunity to further improve PCC assay procedure and shorten the analysis time i.e. in case of partial-body exposure. Moreover, the study presents preliminary results of HPBL cellular response after proton irradiation at high doses range showing differences of PCC index kinetics for different cell classes and cell distribution.