Human mesenchymal stromal cells maintain their stem cell traits after high-LET particle irradiation - Potential implications for particle radiotherapy and manned space missions.

The influence of high-linear energy transfer (LET) particle radiation on the functionalities of mesenchymal stromal cells (MSCs) is largely unknown. Here, we analyzed the effects of proton (1H), helium (4He), carbon (12C) and oxygen (16O) ions on human bone marrow-MSCs. Cell cycle distribution and apoptosis induction were examined by flow cytometry, and DNA damage was quantified using γH2AX immunofluorescence and Western blots. Relative biological effectiveness values of MSCs amounted to 1.0-1.1 for 1H, 1.7-2.3 for 4He, 2.9-3.4 for 12C and 2.6-3.3 for 16O. Particle radiation did not alter the MSCs' characteristic surface marker pattern, and MSCs maintained their multi-lineage differentiation capabilities. Apoptosis rates ranged low for all radiation modalities. At 24 h after irradiation, particle radiation-induced ATM and CHK2 phosphorylation as well as γH2AX foci numbers returned to baseline levels. The resistance of human MSCs to high-LET irradiation suggests that MSCs remain functional after exposure to moderate doses of particle radiation as seen in normal tissues after particle radiotherapy or during manned space flights. In the future, in vivo models focusing on long-term consequences of particle irradiation on the bone marrow niche and MSCs are needed.

Quantitative modeling of carcinogenesis induced by single beams or mixtures of space radiations using targeted and non-targeted effects.

Ionizing radiations encountered by astronauts on deep space missions produce biological damage by two main mechanisms: (1) Targeted effects (TE) due to direct traversals of cells by ionizing tracks. (2) Non-targeted effects (NTE) caused by release of signals from directly hit cells. The combination of these mechanisms generates non-linear dose response shapes, which need to be modeled quantitatively to predict health risks from space exploration. Here we used a TE + NTE model to analyze data on APC(1638N/+) mouse tumorigenesis induced by space-relevant doses of protons, 4He, 12C, 16O, 28Si or 56Fe ions, or γ rays. A customized weighted Negative Binomial distribution was used to describe the radiation type- and dose-dependent data variability. This approach allowed detailed quantification of dose-response shapes, NTE- and TE-related model parameters, and radiation quality metrics (relative biological effectiveness, RBE, and radiation effects ratio, RER, relative to γ rays) for each radiation type. Based on the modeled responses for each radiation type, we predicted the tumor yield for a Mars-mission-relevant mixture of these radiations, using the recently-developed incremental effect additivity (IEA) synergy theory. The proposed modeling approach can enhance current knowledge about quantification of space radiation quality effects, dose response shapes, and ultimately the health risks for astronauts.

Circulating tRNA-Derived Small RNAs as Novel Radiation Biomarkers of Heavy Ion, Proton and X-ray Exposure.

The effective and minimally invasive radiation biomarkers are valuable for exposure scenarios in nuclear accidents or space missions. Recent studies have opened the new sight of circulating small non-coding RNA (sncRNA) as radiation biomarkers. The tRNA-derived small RNA (tsRNA) is a new class of sncRNA. It is more abundant than other kinds of sncRNAs in extracellular vesicles or blood, presenting great potential as promising biomarkers. However, the circulating tsRNAs in response to ionizing radiation have not been reported. In this research, Kunming mice were total-body exposed to 0.05-2 Gy of carbon ions, protons, or X-rays, and the RNA sequencing was performed to profile the expression of sncRNAs in serum. After conditional screening and validation, we firstly identified 5 tsRNAs including 4 tRNA-related fragments (tRFs) and 1 tRNA half (tiRNA) which showed a significant level decrease after exposure to three kinds of radiations. Moreover, the radiation responses of these 5 serum tsRNAs were reproduced in other mouse strains, and the sequences of them could be detected in serum of humans. Furthermore, we developed multi-factor models based on tsRNA biomarkers to indicate the degree of radiation exposure with high sensitivity and specificity. These findings suggest that the circulating tsRNAs can serve as new minimally invasive biomarkers and can make a triage or dose assessment from blood sample collection within 4 h in exposure scenarios.

Optimizing proton minibeam radiotherapy by interlacing and heterogeneous tumor dose on the basis of calculated clonogenic cell survival.

Proton minibeam radiotherapy (pMBRT) is a spatial fractionation method using sub-millimeter beams at center-to-center (ctc) distances of a few millimeters to widen the therapeutic index by reduction of side effects in normal tissues. Interlaced minibeams from two opposing or four orthogonal directions are calculated to minimize side effects. In particular, heterogeneous dose distributions applied to the tumor are investigated to evaluate optimized sparing capabilities of normal tissues at the close tumor surrounding. A 5 cm thick tumor is considered at 10 cm depth within a 25 cm thick water phantom. Pencil and planar minibeams are interlaced from two (opposing) directions as well as planar beams from four directions. An initial beam size of σ0 = 0.2 mm (standard deviation) is assumed in all cases. Tissue sparing potential is evaluated by calculating mean clonogenic cell survival using a linear-quadratic model on the calculated dose distributions. Interlacing proton minibeams for homogeneous irradiation of the tumor has only minor benefits for the mean clonogenic cell survival compared to unidirectional minibeam irradiation modes. Enhanced mean cell survival, however, is obtained when a heterogeneous dose distribution within the tumor is permitted. The benefits hold true even for an elevated mean tumor dose, which is necessary to avoid cold spots within the tumor in concerns of a prescribed dose. The heterogeneous irradiation of the tumor allows for larger ctc distances. Thus, a high mean cell survival of up to 47% is maintained even close to the tumor edges for single fraction doses in the tumor of at least 10 Gy. Similar benefits would result for heavy ion minibeams with the advantage of smaller minibeams in deep tissue potentially offering even increased tissue sparing. The enhanced mean clonogenic cell survival through large ctc distances for interlaced pMBRT with heterogeneous tumor dose distribution results in optimum tissue sparing potential. The calculations show the largest enhancement of the mean cell survival in normal tissue for high-dose fractions. Thus, hypo-fractionation or even single dose fractions become possible for tumor irradiation. A widened therapeutic index at big cost reductions is offered by interlaced proton or heavy ion minibeam therapy.

Targeting the DNA replication stress phenotype of KRAS mutant cancer cells.

Mutant KRAS is a common tumor driver and frequently confers resistance to anti-cancer treatments such as radiation. DNA replication stress in these tumors may constitute a therapeutic liability but is poorly understood. Here, using single-molecule DNA fiber analysis, we first characterized baseline replication stress in a panel of unperturbed isogenic and non-isogenic cancer cell lines. Correlating with the observed enhanced replication stress we found increased levels of cytosolic double-stranded DNA in KRAS mutant compared to wild-type cells. Yet, despite this phenotype replication stress-inducing agents failed to selectively impact KRAS mutant cells, which were protected by CHK1. Similarly, most exogenous stressors studied did not differentially augment cytosolic DNA accumulation in KRAS mutant compared to wild-type cells. However, we found that proton radiation was able to slow fork progression and preferentially induce fork stalling in KRAS mutant cells. Proton treatment also partly reversed the radioresistance associated with mutant KRAS. The cellular effects of protons in the presence of KRAS mutation clearly contrasted that of other drugs affecting replication, highlighting the unique nature of the underlying DNA damage caused by protons. Taken together, our findings provide insight into the replication stress response associated with mutated KRAS, which may ultimately yield novel therapeutic opportunities.

Photon versus proton neurotoxicity: Impact on mitochondrial function and 8-OHdG base-excision repair mechanism in human astrocytes.

This study assesses and compares the neurotoxic effects of proton and photon radiation on mitochondrial function and DNA repair capabilities of human astrocytes. Human astrocytes received either proton (0.5 Gy and 3 Gy), photon (0.5 Gy and 3 Gy), or sham-radiation treatment. The mRNA expression level of the DNA repair protein OGG1 was determined via RT-qPCR. The levels of 8-OHdG in the cell media were measured via ELISA. Real-time kinetic analysis of extracellular oxygen consumption rates was performed to assess mitochondrial function. Radiation-induced changes in mitochondrial mass and oxidative activity were assessed using fluorescent imaging with MitoTracker™ Green FM and MitoTracker™ Orange CM-H2TMRos dyes respectively. PCR was used to quantify the alteration in the mitochondrial DNA content, measured as the mitochondrial to nuclear DNA ratio. A significant increase in mitochondrial mass and levels of reactive oxygen species was observed after radiation treatment. Additionally, real-time PCR analysis indicated a significant depletion of mitochondrial DNA content in the irradiated cells when compared to the control. This was accompanied by a decreased gene expression of the DNA base-excision repair protein OGG1 and reduced clearance of 8-OHdG adducts from the genome. Photon radiation treatment was associated with a more detrimental cellular impact when compared to the same dose of proton radiation. These results are indicative of a radiation-induced dose-dependent decrease in mitochondrial function, an increase in senescence and astrogliosis, and impairment of the DNA repair capabilities in healthy glial cells. Photon irradiation was associated with a more significant disruption in mitochondrial function and base-excision repair mechanisms in vitro in comparison to proton treatment.

Simulation of Proton-Induced DNA Damage Patterns Using an Improved Clustering Algorithm.

Simulations of deoxyribonucleic acid (DNA) molecular damage use the traversal algorithm that has the disadvantages of being time-consuming, slowly converging, and requiring high-performance computer clusters. This work presents an improved version of the algorithm, "density-based spatial clustering of applications with noise" (DBSCAN), using a KD-tree approach to find neighbors of each point for calculating clustered DNA damage. The resulting algorithm considers the spatial distributions for sites of energy deposition and hydroxyl radical attack, yielding the statistical probability of (single and double) DNA strand breaks. This work achieves high accuracy and high speed at calculating clustered DNA damage that has been induced by proton treatment at the molecular level while running on an i7 quad-core CPU. The simulations focus on the indirect effect generated by hydroxyl radical attack on DNA. The obtained results are consistent with those of other published experiments and simulations. Due to the array of chemical processes triggered by proton treatment, it is possible to predict the effects that different track structures of various energy protons produce on eliciting direct and indirect damage of DNA.

Protons and High-Linear Energy Transfer Radiation Induce Genetically Similar Lymphomas With High Penetrance in a Mouse Model of the Aging Human Hematopoietic System.

PURPOSE: Humans are exposed to charged particles in different scenarios. The use of protons and high-linear energy transfer (LET) in cancer treatment is steadily growing. In outer space, astronauts will be exposed to a mixed radiation field composed of both protons and heavy ions, in particularly the long-term space missions outside of earth's magnetosphere. Thus, understanding the radiobiology and transforming potential of these types of ionizing radiation are of paramount importance. METHODS AND MATERIALS: We examined the effect of 10 or 100 cGy of whole-body doses of protons or 28Si ions on the hematopoietic system of a genetic model of aging based on recent studies that showed selective loss of the MLH1 protein in human hematopoietic stems with age. RESULTS: We found that Mlh1 deficient animals are highly prone to develop lymphomas when exposed to either low doses of protons or 28Si ions. The lymphomas that develop are genetically indistinguishable, in spite of different types of damage elicited by low- and high-LET radiation. RNA sequencing analyses reveal similar gene expression patterns, similar numbers of altered genes, similar numbers of single nucleotide variants and insertions and deletions, and similar activation of known leukemogenic loci. CONCLUSIONS: Although the incidence of malignancy is related to radiation quality, and increased due to loss of Mlh1, the phenotype of the tumors is independent of LET.

Effects of single-dose protons or oxygen ions on function and structure of the cardiovascular system in male Long Evans rats.

PURPOSE: Studies are required to determine whether exposures to radiation encountered during manned missions in deep space may have adverse effects on the cardiovascular system. Most of the prior studies on effects of simulated space radiation on the heart and vasculature have been performed in mouse models. To provide data from a second animal species, two studies were performed to assess effects of high-energy charged particle radiation on the heart and abdominal aorta in a rat model. MATERIALS AND METHODS: In study A, male Long Evans rats were exposed to whole-body protons (250 MeV, 0.5 Gy) or oxygen ions (16O, 600 MeV/n, 0.5 Gy), and ultrasonography was used to measure in vivo cardiac function and blood flow parameters at 3, 5, 9 and 12 months after radiation, followed by tissue collection at 12 months. In study B, male Long Evans rats were exposed to 16O (1 GeV/n, 0.01-0.25 Gy), and hearts collected at 6 to 7 and 12 months for histology and western-blots. RESULTS: Both protons (250 MeV) and 16O (600 MeV/n) caused a decrease in left ventricular posterior wall thickness at 3-5 months, but did not change echocardiographic measures of cardiac function. In Pulsed-wave Doppler assessment of the abdominal aorta, an increase was seen in mean velocity, peak velocity, and velocity time integral at 12 months after 16O (600 MeV/n), suggesting a change in vascular function. There were no significant changes in histopathology or histological quantification of total collagens in heart or aorta. On the other hand, an increase was seen in a 75 kDa peptide of collagen type III in the left ventricle of rats exposed to protons (250 MeV) and 16O (600 MeV/n and 1 GeV/n), suggesting that radiation caused remodeling of existing collagens in the heart. 16O (600 MeV/n and 1 GeV/n) caused increases in left ventricular protein levels of immune cell markers CD2, CD4, CD8, and CD68. CONCLUSION: A single low dose of whole body protons or 16O in male Long Evans rats did not change cardiac function or induce gross pathological changes in the heart or aorta, but induced mild changes in vascular function and remodeling of existing collagens in the heart. Altogether, studies in prior mouse models and the current work in rats indicate minor changes in cardiac function and structure after a low dose of single-ion radiation.

Increase of mtDNA number and its mutant copies in rat brain after exposure to 150 MeV protons.

Proton beam therapy is widely used for treating brain tumor. Despite the efficacy of treatment, the use of this therapy has met some limitations associated with possible damage to normal brain tissues located beyond the tumor site. In this context, the exploration of the harmful effects of protons on the normal brain tissues is of particular interest. We have investigated changes in the total mitochondrial DNA (mtDNA) copy number and identified mtDNA mutant copies in three brain regions (the hippocampus, cortex and cerebellum) of rats after irradiation their whole-head with 150 MeV protons at doses of 3 and 5 Gy. The study was performed in 2-months old male Spraque Dawley rats (n = 5 each group). The mtDNA copy numbers were determined by real-time PCR. The level of mtDNA heteroplasmy was estimated using Surveyor nuclease technology. Our results show that after head exposure to protons, levels of mtDNA copy number in three rat brain regions increase significantly as the levels of mtDNA mutant copies increase. The most significant elevation is observed in the hippocampus. In conclusion, an increase in mtDNA mutant copies may contribute to mitochondrial dysfunction accompanied by increased oxidative stress in different brain regions and promote the development of neurodegenerative diseases and the induction of carcinogenesis.

Particle radiation-induced dysregulation of protein homeostasis in primary human and mouse neuronal cells.

Space particle radiations may cause significant damage to proteins and oxidative stress in the cells within the central nervous system and pose a potential health hazard to humans in long-term manned space explorations. Dysregulation of the ubiquitin-proteasome system as evidenced by abnormal accumulation of polyubiquitin (pUb) chain linkages has been implicated in several age-related neurodegenerative disorders by mechanisms that may involve the inter-neuronal spread of toxic misfolded proteins, the induction of chronic neuroinflammation, or the inappropriate inhibition or activation of key enzymes, which could lead to dysfunction in, for example, proteolysis, or the accumulation of post-translationally-modified substrates.In this study, we employed a quantitative proteomics method to evaluate the impact of particle-radiation induced alterations in three major pUb-linked chains at lysine residues Lys-48 (K-48), Lys-63 (K-63), and Lys-11 (K-11), and probed for global proteomic changes in mouse and human neural cells that were irradiated with low doses of 250 MeV proton, 260 MeV/u silicon or 1 GeV/u iron ions. We found significant accumulation in K-48 linkage after 1 Gy protons and K-63 linkage after 0.5 Gy iron ions in human neural cells. Cells derived from different regions of the mouse brain (cortex, striatum and mesencephalon) showed differential sensitivity to particle radiation exposure. Although none of the linkages were altered after proton exposure, both K-48 and K-63 linkages in mouse striatal neuronal cells were elevated after 0.5 Gy of silicon or iron ions. Changes were also seen in proteins commonly used as markers of neural progenitor and stem cells, in DNA binding/damage repair and cellular redox pathways. In contrast, no significant changes were observed at the same time point after proton irradiation. These results suggest that the quality of the particle radiation plays a key role in the level, linkage and cell type specificity of protein homeostasis in key populations of neuronal cells.

Proton and carbon ion radiation therapy for locally advanced pancreatic cancer: A phase I dose escalation study.

OBJECTIVE: To determine the maximum tolerated dose (MTD) of proton and carbon ion radiation therapy (PCRT) for locally advanced pancreatic cancer (LAPC). METHODS: A single-institution, phase I dose escalation study was performed. The proton dose of 50.4 GyE in 28 fractions was delivered to clinical target volume, and carbon ion as a boost dose to gross tumor volume escalated from 12 GyE to 18 GyE with 3 GyE per fraction in 3 dose levels. The dose limiting toxicity (DLT) was defined as any treatment-related grade (G)3 or higher of non-hematological toxicity. The MTD was exceeded if ≥2 patients in a dose level developed DLT. RESULTS: From May 2015 to July 2016, ten patients were enrolled, 3 in dose level 1, 4 in dose level 2, and 3 in dose level 3. With a median follow-up of 17.4 months, no patient developed a DLT, and the acute G1-2 of gastrointestinal (GI) and hepatic toxicity occurred in 40% of patients, and G1 of GI late toxicity, in 30%. The median overall survival was 17.3 months. CONCLUSION: Higher than 50.4 GyE could be given by PCRT with slight toxicity and good tolerance for LAPC, and the tumor control and survival had been improved, but not significantly. Better outcome may be achieved using carbon ion radiation therapy with higher biological equivalent dose.

Computational feasibility of simulating changes in blood flow through whole-organ vascular networks from radiation injury.

Vasculature is necessary to the healthy function of most tissues. In radiation therapy, injury of the vasculature can have both beneficial and detrimental effects, such as tumor starvation, cardiac fibrosis, and white-matter necrosis. These effects are caused by changes in blood flow due to the vascular injury. Previously, research has focused on simulating the radiation injury of vasculature in small volumes of tissue, ignoring the systemic effects of local damage on blood flow. Little is known about the computational feasibility of simulating the radiation injury to whole-organ vascular networks. The goal of this study was to test the computational feasibility of simulating the dose deposition to a whole-organ vascular network and the resulting change in blood flow. To do this, we developed an amorphous track-structure model to transport radiation and combined this with existing methods to model the vasculature and blood flow rates. We assessed the algorithm's computational scalability, execution time, and memory usage. The data demonstrated it is computationally feasible to calculate the radiation dose and resulting changes in blood flow from 2 million protons to a network comprising 8.5 billion blood vessels (approximately the number in the human brain) in 87 hours using a 128-node cluster. Furthermore, the algorithm demonstrated both strong and weak scalability, meaning that additional computational resources can reduce the execution time further. These results demonstrate, for the first time, that it is computationally feasible to calculate radiation dose deposition in whole-organ vascular networks. These findings provide key insights into the computational aspects of modeling whole-organ radiation damage. Modeling the effects radiation has on vasculature could prove useful in the study of radiation effects on tissues, organs, and organisms.

Role of Endoplasmic Reticulum and Mitochondrion in Proton Microbeam Radiation-Induced Bystander Effect.

It is well known that mitochondria and the endoplasmic reticulum (ER) play important roles in radiation response, but their functions in radiation-induced bystander effect (RIBE) are largely unclear. In this study, we found that when a small portion of cells in a population of human lung fibroblast MRC-5 cells were precisely irradiated through either the nuclei or cytoplasm with counted microbeam protons, the yield of micronuclei (MN) and the levels of intracellular reactive oxygen species (ROS) in nonirradiated cells neighboring irradiated cells were significantly increased. Mito/ER-tracker staining demonstrated that the mitochondria were clearly activated after nuclear irradiation and ER mass approached a higher level after cytoplasmic irradiation. Moreover, the radiation-induced ROS was diminished by rotenone, an inhibitor of mitochondria activation, but it was not influenced by siRNA interference of BiP, an ER regulation protein. While for nuclear irradiation, rotenone-enhanced radiation-induced ER expression, and BiP siRNA eliminated radiation-induced activation of mitochondria, these phenomena were not observed for cytoplasmic irradiation. Bystander MN was reduced by rotenone but enhanced by BiP siRNA. When the cells were treated with both rotenone and BiP siRNA, the MN yield was reduced for nuclear irradiation but was enhanced for cytoplasmic irradiation. Our results suggest that the organelles of mitochondria and ER have different roles in RIBE with respect to nuclear and cytoplasmic irradiation, and the function of ER is a prerequisite for mitochondrial activation.

Influence of Cranial Irradiation with High-Energy Protons on the Visuomotor Behavior in Monkeys.

The visually driven instrumental conditioning of a single monkey (Macaca mulatta) was conducted after single-dose cranial irradiation with high-energy protons. The monkey executed saccades toward the visual stimuli and then responded by manually pressing right or left lever for stimuli in right or left half-field, respectively. The percentage of correct responses with dominant right hand exceeded the percentage of such responses with left hand and temporarily decreased two months after irradiation. A month later, the percentage of correct right-hand responses returned to the level before irradiation. No significant dynamic was found for changes in percentage of correct left-hand responses. The proton irradiation effect on right-hand responses suggests possible short-term disturbances in the eye-hand coordination for right handedness while the visual perception remains unaffected.

Fractionated and Acute Proton Radiation Show Differential Intestinal Tumorigenesis and DNA Damage and Repair Pathway Response in ApcMin/+ Mice.

PURPOSE: Proton radiation is a major component of the radiation field in outer space and is used clinically in radiation therapy of resistant cancers. Although epidemiologic studies in atom bomb survivors and radiologic workers have established radiation as a risk factor for colorectal cancer (CRC), we have yet to determine the risk of CRC posed by proton radiation owing to a lack of sufficient human or animal data. The purpose of the current study was to quantitatively and qualitatively characterize differential effects of acute and fractionated high-energy protons on colorectal carcinogenesis. METHODS AND MATERIALS: We used ApcMin/+ mice, a well-studied CRC model, to examine acute versus fractionated proton radiation-induced differences in intestinal tumorigenesis and associated signaling pathways. Mice were exposed to 1.88 Gy of proton radiation delivered in a single fraction or in 4 equal daily fractions (0.47 Gy × 4). Intestinal tumor number and grade were scored 100 to 110 days after irradiation, and tumor and tumor-adjacent normal tissues were harvested to assess proliferative ß-catenin/Akt pathways and DNA damage response and repair pathways relevant to colorectal carcinogenesis. RESULTS: Significantly higher intestinal tumor number and grade, along with decreased differentiation, were observed after acute radiation relative to fractionated radiation. Acute protons induced upregulation of ß-catenin and Akt pathways with increased proliferative marker phospho-histone H3. Increased DNA damage along with decreased DNA repair factors involved in mismatch repair and nonhomologous end joining were also observed after exposure to acute protons. CONCLUSIONS: We show increased γH2AX, 53BP1, and 8-oxo-dG, suggesting that increased ongoing DNA damage along with decreased DNA repair factors and increased proliferative responses could be triggering a higher number of intestinal tumors after acute relative to fractionated proton exposures in ApcMin/+ mice. Taken together, our data suggest greater carcinogenic potential of acute relative to fractionated proton radiation.

Sensitive to Proton Rhizotoxicity1 Regulates Salt and Drought Tolerance of Arabidopsis thaliana through Transcriptional Regulation of CIPK23.

The transcription factor sensitive to proton rhizotoxicity 1 (STOP1) regulates multiple stress tolerances. In this study, we confirmed its involvement in NaCl and drought tolerance. The root growth of the T-DNA insertion mutant of STOP1 (stop1) was sensitive to NaCl-containing solidified MS media. Transcriptome analysis of stop1 under NaCl stress revealed that STOP1 regulates several genes related to salt tolerance, including CIPK23. Among all available homozygous T-DNA insertion mutants of the genes suppressed in stop1, only cipk23 showed a NaCl-sensitive root growth phenotype comparable to stop1. The CIPK23 promoter had a functional STOP1-binding site, suggesting a strong CIPK23 suppression led to NaCl sensitivity of stop1. This possibility was supported by in planta complementation of CIPK23 in the stop1 background, which rescued the short root phenotype under NaCl. Both stop1 and cipk23 exhibited a drought tolerant phenotype and increased abscisic acid-regulated stomatal closure, while the complementation of CIPK23 in stop1 reversed these traits. Our findings uncover additional pleiotropic roles of STOP1 mediated by CIPK23, which regulates various ion transporters including those regulating K+-homeostasis, which may induce a trade-off between drought tolerance and other traits.

Risks of cognitive detriments after low dose heavy ion and proton exposures.

Purpose: Heavy ion and proton brain irradiations occur during space travel and in Hadron therapy for cancer. Heavy ions produce distinct patterns of energy deposition in neuron cells and brain tissues compared to X-rays leading to large uncertainties in risk estimates. We make a critical review of findings from research studies over the last 25 years for understanding risks at low dose. Conclusions: A large number of mouse and rat cognitive testing measures have been reported for a variety of particle species and energies for acute doses. However, tissue reactions occur above dose thresholds and very few studies were performed at the heavy ion doses to be encountered on space missions (<0.04 Gy/y) or considered dose-rate effects, such that threshold doses are not known in rodent models. Investigations of possible mechanisms for cognitive changes have been limited by experimental design with largely group specific and not subject specific findings reported. Persistent oxidative stress and activated microglia cells are common mechanisms studied, while impairment of neurogenesis, detriments in neuron morphology, and changes to gene and protein expression were each found to be important in specific studies. Future research should focus on estimating threshold doses carried out with experimental designs aimed at understating causative mechanisms, which will be essential for extrapolating rodent findings to humans and chronic radiation scenarios, while establishing if mitigation are needed.

Stress responses in Crassostrea gasar exposed to combined effects of acute pH changes and phenanthrene.

Ocean acidification is a result of the decrease in the pH of marine water, caused mainly by the increase in CO2 released in the atmosphere and its consequent dissolution in seawater. These changes can be dramatic for marine organisms especially for oysters Crassostrea gasar if other stressors such as xenobiotics are present. The effect of pH changes (6.5, 7.0 and 8.2) was assessed on the transcript levels of biotransformation [cytochromes P450 (CYP2AU1, CYP2-like2) and glutathione S-transferase (GSTΩ-like)] and antioxidant [superoxide dismutase (SOD-like), catalase (CAT-like) and glutathione peroxidase (GPx-like)] genes, as well as enzyme activities [superoxide dismutase, (SOD), catalase (CAT), glutathione reductase (GR), glutathione-S-transferases transferase (GST) and glucose-6-phosphate dehydrogenase (G6PDH)] and lipid peroxidation (MDA) in the gills of Crassostrea gasar exposed to 100 µg·L-1 of phenanthrene (PHE) for 24 and 96 h. Likewise, the PHE burdens was evaluated in whole soft tissues of exposed oysters. The accumulation of PHE in oysters was independent of pH. However, acidification promoted a significant decrease in the transcript levels of some protective genes (24 h exposure: CYP2AU1 and GSTΩ-like; 96 h exposure: CAT-like and GPx-like), which was not observed in the presence of PHE. Activities of GST, CAT and SOD enzymes increased in the oysters exposed to PHE at the control pH (8.2), but at a lower pH values, this activation was suppressed, and no changes were observed in the G6PDH activity and MDA levels. Biotransformation genes showed better responses after 24 h, and antioxidant-coding genes after 96 h, along with the activities of antioxidant enzymes (SOD, CAT), probably because biotransformation of PHE increases the generation of reactive oxygen species. The lack of change in MDA levels suggests that antioxidant modulation efficiently prevented oxidative stress. The effect of pH on the responses to PHE exposure should be taken into account before using these and any other genes as potential molecular biomarkers for PHE exposure.