It is well established that ionizing radiation induces chromosomal damage, both following direct radiation exposure and via non-targeted (bystander) effects, activating DNA damage repair pathways, of which the proteins are closely linked to telomeric proteins and telomere maintenance. Long-term propagation of this radiation-induced chromosomal damage during cell proliferation results in chromosomal instability. Many studies have shown the link between radiation exposure and radiation-induced changes in oxidative stress and DNA damage repair in both targeted and non-targeted cells. However, the effect of these factors on telomeres, long established as guardians of the genome, still remains to be clarified. In this review, we will focus on what is known about how telomeres are affected by exposure to low- and high-LET ionizing radiation and during proliferation, and will discuss how telomeres may be a key player in the process of radiation-induced carcinogenesis.

It is well known that molecular oxygen is a product of the radiolysis of water with high-linear energy transfer (LET) radiation, which is distinct from low-LET radiation wherein O2 radiolytic yield is negligible. Since O2 is a powerful radiosensitizer, this fact is of practical relevance in cancer therapy with energetic heavy ions, such as carbon ions. It has recently been discovered that large doses of ionizing radiation delivered to tumors at very high dose rates (i.e., in a few milliseconds) have remarkable benefits in sparing healthy tissue while preserving anti-tumor activity compared to radiotherapy delivered at conventional, lower dose rates. This new method is called “FLASH radiotherapy” and has been tested using low-LET radiation (i.e., electrons and photons) in various pre-clinical studies and recently in a human patient. Although the exact mechanism(s) underlying FLASH are still unclear, it has been suggested that radiation delivered at high dose rates spares normal tissue via oxygen depletion. In addition, heavy-ion radiation achieves tumor control with reduced normal tissue toxicity due to its favorable physical depth-dose profile and increased radiobiological effectiveness in the Bragg peak region. To date, however, biological research with energetic heavy ions delivered at ultra-high dose rates has not been performed and it is not known whether heavy ions are suitable for FLASH radiotherapy. Here we present the additive or even synergistic advantages of integrating the FLASH dose rates into carbon-ion therapy. These benefits result from the ability of heavy ions at high LET to generate an oxygenated microenvironment around their track due to the occurrence of multiple (mainly double) ionization of water. This oxygen is abundant immediately in the tumor region where the LET of the carbon ions is very high, near the end of the carbon-ion path (i.e., in the Bragg peak region). In contrast, in the “plateau” region of the depth-dose distribution of ions (i.e., in the normal tissue region), in which the LET is significantly lower, this generation of molecular oxygen is insignificant. Under FLASH irradiation, it is shown that this early generation of O2 extends evenly over the entire irradiated tumor volume, with concentrations estimated to be several orders of magnitude higher than the oxygen levels present in hypoxic tumor cells. Theoretically, these results indicate that FLASH radiotherapy using carbon ions would have a markedly improved therapeutic ratio with greater toxicity in the tumor due to the generation of oxygen at the spread-out Bragg peak.

Significance : During deep space travel, astronauts are often exposed to high atomic number (Z) and high-energy (E) (high charge and high energy [HZE]) particles. On interaction with cells, these particles cause severe oxidative injury and result in unique biological responses. When cell populations are exposed to low fluences of HZE particles, a significant fraction of the cells are not traversed by a primary radiation track, and yet, oxidative stress induced in the targeted cells may spread to nearby bystander cells. The long-term effects are more complex because the oxidative effects persist in progeny of the targeted and affected bystander cells, which promote genomic instability and may increase the risk of age-related cancer and degenerative diseases. Recent Advances: Greater understanding of the spatial and temporal features of reactive oxygen species bursts along the tracks of HZE particles, and the availability of facilities that can simulate exposure to space radiations have supported the characterization of oxidative stress from targeted and nontargeted effects. Critical Issues: The significance of secondary radiations generated from the interaction of the primary HZE particles with biological material and the mitigating effects of antioxidants on various cellular injuries are central to understanding nontargeted effects and alleviating tissue injury. Future Directions: Elucidation of the mechanisms underlying the cellular responses to HZE particles, particularly under reduced gravity and situations of exposure to additional radiations, such as protons, should be useful in reducing the uncertainty associated with current models for predicting long-term health risks of space radiation. These studies are also relevant to hadron therapy of cancer. Antioxid. Redox Signal. 20, 1501–1523.

Background: Cancer-associated fibroblasts (CAFs) are a major component of the cancer stroma, and their response to therapeutic treatments likely impacts the outcome. We tested the hypothesis that CAFs develop unique characteristics that enhance their resistance to ionizing radiation.Methods: CAFs were generated through intimate coculture of normal human fibroblasts of skin or lung origin with various human cancer cell types using permeable microporous membrane inserts. Fibroblasts and cancer cells are grown intimately, yet separately, on either side of the insert’s membrane for extended times to generate activated fibroblast populations highly enriched in CAFs.Results: The generated CAFs exhibited a decrease in Caveolin-1 protein expression levels, a CAF biomarker, which was further enhanced when the coculture was maintained under in-vivo-like oxygen tension conditions. The level of p21^Waf1was also attenuated, a characteristic also associated with accelerated tumor growth. Furthermore, the generated CAFs experienced perturbations in their redox environment as demonstrated by increases in protein carbonylation, mitochondrial superoxide anion levels, and modulation of the activity of the antioxidants, manganese superoxide dismutase and catalase. Propagation of the isolated CAFs for 25 population doublings was associated with enhanced genomic instability and a decrease in expression of the senescence markers β-galactosidase and p16^INK4a. With relevance to radiotherapeutic treatments, CAFs in coculture with cancer cells of diverse origins (breast, brain, lung, and prostate) were resistant to the clastogenic effects of¹³⁷Cs γ rays compared to naïve fibroblasts. Addition of repair inhibitors of single- or double-stranded DNA breaks attenuated the resistance of CAFs to the clastogenic effects of γ rays, supporting a role for increased ability to repair DNA damage in CAF radioresistance.Conclusions: This study reveals that CAFs are radioresistant and experience significant changes in indices of oxidative metabolism. The CAFs that survive radiation treatment likely modulate the fate of the associated cancer cells. Identifying them together with their mode of communication with cancer cells, and eradicating them, particularly when they may exist at the margin of the radiotherapy planning target volume, may improve the efficacy of cancer treatments.

Background: Glioblastoma multiforme is an aggressive primary brain tumor that is characterized by local invasive growth and resistance to therapy. The role of the microenvironment in glioblastoma invasiveness remains unclear. While carcinomas release CD147, a protein that signals for increased matrix metalloproteinase (MMP) release by fibroblasts, glioblastoma does not have a significant fibroblast component. We hypothesized that astrocytes release MMPs in response to CD147 contained in glioblastoma-derived extracellular vesicles (EVs) and that ionizing radiation, part of the standard treatment for glioblastoma, enhances this release. Methods: Astrocytes were incubated with EVs released by irradiated or non-irradiated human glioblastoma cells wild-type, knockdown, or knockout for CD147. Levels of CD147 in glioblastoma EVs and MMPs secreted by astrocytes were quantified. Levels of proteins in the mitogen activated protein kinase (MAPK) pathway, which can be regulated by CD147, were measured in astrocytes incubated with EVs from glioblastoma cells wild-type or knockdown for CD147. Immunofluorescence was performed on the glioblastoma cells to identify changes in CD147 localization in response to irradiation, and to confirm uptake of the EVs by astrocytes. Results: Immunoblotting and mass spectrometry analyses showed that CD147 levels in EVs were transiently increased when the EVs were from glioblastoma cells that were irradiated with γ rays. Specifically, the highly-glycosylated 45 kDa form of CD147 was preferentially present in the EVs relative to the cells themselves. Immunofluorescence demonstrated that astrocytes incorporate glioblastoma EVs and subsequently increase their secretion of active MMP9. The increase was greater if the EVs were from irradiated glioblastoma cells. Testing MAPK pathway activation, which also regulates MMP expression, showed that JNK signaling, but not ERK1/2 or p38, was increased in astrocytes incubated with EVs from irradiated compared to non-irradiated glioblastoma cells. Knockout of CD147 in glioblastoma cells blocked the increased JNK signaling and the rise in secreted active MMP9 levels. Conclusions: The results support a tumor microenvironment-mediated role of CD147 in glioblastoma invasiveness, and reveal a prominent role for ionizing radiation in enhancing the effect. They provide an improved understanding of glioblastoma intercellular signaling in the context of radiotherapy, and identify pathways that can be targeted to reduce tumor invasiveness. Graphical

During interplanetary missions, astronauts are exposed to mixed types of ionizing radiation. The low ‘flux’ of the high atomic number and high energy (HZE) radiations relative to the higher ‘flux’ of low linear energy transfer (LET) protons makes it highly probable that for any given cell in the body, proton events will precede any HZE event. Whereas progress has been made in our understanding of the biological effects of low-LET protons and high-LET HZE particles, the interplay between the biochemical processes modulated by these radiations is unclear. Here we show that exposure of normal human fibroblasts to a low mean absorbed dose of 20 cGy of 0.05 or 1-GeV protons (LET ∼ 1.25 or 0.2 keV/μm, respectively) protects the irradiated cells (P < 0.0001) against chromosomal damage induced by a subsequent exposure to a mean absorbed dose of 50 cGy from 1 GeV/u iron ions (LET ∼ 151 keV/μm). Surprisingly, unirradiated (i.e. bystander) cells with which the proton-irradiated cells were co-cultured were also significantly protected from the DNA-damaging effects of the challenge dose. The mitigating effect persisted for at least 24 h. These results highlight the interactions of biological effects due to direct cellular traversal by radiation with those due to bystander effects in cell populations exposed to mixed radiation fields. They show that protective adaptive responses can spread from cells targeted by low-LET space radiation to bystander cells in their vicinity. The findings are relevant to understanding the health hazards of space travel.