Radiation-quality-dependent bystander cellular effects induced by heavy-ion microbeams through different pathways.

We investigated the radiation-quality-dependent bystander cellular effects using heavy-ion microbeams with different ion species. The heavy-ion microbeams were produced in Takasaki Ion Accelerators for Advanced Radiation Application, National Institutes for Quantum Science and Technology. Carbon (12C5+, 220 MeV), neon (20Ne7+, 260 MeV) and argon (40Ar13+, 460 MeV) ions were used as the microbeams, collimating the beam size with a diameter of 20 µm. After 0.5 and 3 h of irradiation, the surviving fractions (SFs) are significantly lower in cells irradiated with carbon ions without a gap-junction inhibitor than those irradiated with the inhibitor. However, the same SFs with no cell killing were found with and without the inhibitor at 24 h. Conversely, no cell-killing effect was observed in argon-ion-irradiated cells at 0.5 and 3 h; however, significantly low SFs were found at 24 h with and without the inhibitor, and the effect was suppressed using vitamin C and not dimethyl sulfoxide. The mutation frequency (MF) in cells irradiated with carbon ions was 8- to 6-fold higher than that in the unirradiated control at 0.5 and 3 h; however, no mutation was observed in cells treated with the gap-junction inhibitor. At 24 h, the MFs induced by each ion source were 3- to 5-fold higher and the same with and without the inhibitor. These findings suggest that the bystander cellular effects depend on the biological endpoints, ion species and time after microbeam irradiations with different pathways.

High-Dose Irradiation Inhibits Motility and Induces Autophagy in Caenorhabditis elegans.

Radiation damages many cellular components and disrupts cellular functions, and was previously reported to impair locomotion in the model organism Caenorhabditis elegans. However, the response to even higher doses is not clear. First, to investigate the effects of high-dose radiation on the locomotion of C. elegans, we investigated the dose range that reduces whole-body locomotion or leads to death. Irradiation was performed in the range of 0-6 kGy. In the crawling analysis, motility decreased after irradiation in a dose-dependent manner. Exposure to 6 kGy of radiation affected crawling on agar immediately and caused the complete loss of motility. Both γ-rays and carbon-ion beams significantly reduced crawling motility at 3 kGy. Next, swimming in buffer was measured as a motility index to assess the response over time after irradiation and motility similarly decreased. However, swimming partially recovered 6 h after irradiation with 3 kGy of γ-rays. To examine the possibility of a recovery mechanism, in situ GFP reporter assay of the autophagy-related gene lgg-1 was performed. The fluorescence intensity was stronger in the anterior half of the body 7 h after irradiation with 3 kGy of γ-rays. GFP::LGG-1 induction was observed in the pharynx, neurons along the body, and the intestine. Furthermore, worms were exposed to region-specific radiation with carbon-ion microbeams and the trajectory of crawling was measured by image processing. Motility was lower after anterior-half body irradiation than after posterior-half body irradiation. This further supported that the anterior half of the body is important in the locomotory response to radiation.

RNA sequencing data for gamma radiation response in the extremotolerant tardigrade Ramazzottius varieornatus.

Tardigrades are microscopic animals of which terrestrial species are capable of tolerating extreme environments by entering a desiccated ametabolic state known as anhydrobiosis. Intriguingly, they survive high dosage gamma rays (>4,000 Gy), possibly through a mechanism known as cross-tolerance. We hypothesized that anhydrobiosis genes are also regulated during cross-tolerance, thus we submitted Ramazzottius varieornatus to 500 Gy 60Co gamma-ray and conducted time-course low-input RNA-Seq. The gene expression was quantified with RSEM and differential expression was determined with DEseq2. Differentially expressed genes were submitted to gene ontology enrichment analysis with GOStat. The transcriptome dynamically shifted nine hours post-exposure.

Development of ultra-thin chips for immobilization of Caenorhabditis elegans in microfluidic channels during irradiation and selection of buffer solution to prevent dehydration.

BACKGROUND: Targeted microbeam irradiation of Caenorhabditis elegans allows the effective knockdown of specific regions, thus helping to identify their roles in processes such as locomotion. We previously employed on-chip immobilization of individuals without anesthesia; however, this method was limited by the thickness of the chip, which prevented the detection of ions passing through the animal, and by dehydration of the animals after prolonged immobilization. NEW METHOD: We developed ultra-thin, ion-penetrable, polydimethylsiloxane microfluidic chips, referred to as Worm Sheets, with and without wettability (hydrophilicity/hydrophobicity), and identified suitable buffer conditions for maintaining moisture in the microfluidic channels. RESULTS: Using a collimating microbeam system, we demonstrated that carbon ions (with a range of ∼1 mm) could pass through the chip, thus allowing the ions to be detected and the applied radiation dose to therefore by measured accurately. We also examined the locomotion of C. elegans following on-chip immobilization in different buffers. Locomotion was decreased in certain buffers on unwettable chips as a result of dehydration due to evaporation, but not on wettable chips. However, locomotion was unaffected on either chip in the presence of a gelatin-based wash buffer. COMPARISON WITH EXISTING METHOD(S): We developed 300-µm-ultra-thin, wettable, ion-penetrable chips for immobilizing C. elegans and provided initial guidance regarding suitable buffer solutions to maintain moisture in microfluidic channels. CONCLUSIONS: This improved, wettable chip, together with the identification of suitable buffer conditions, will become a powerful tool for prolonged immobilizing C. elegans, and is widely applicable not only to microbeam irradiation but also to neurobiological assays.

Region-specific irradiation system with heavy-ion microbeam for active individuals of Caenorhabditis elegans.

Radiation may affect essential functions and behaviors such as locomotion, feeding, learning and memory. Although whole-body irradiation has been shown to reduce motility in the nematode Caenorhabditis elegans, the detailed mechanism responsible for this effect remains unknown. Targeted irradiation of the nerve ring responsible for sensory integration and information processing would allow us to determine whether the reduction of motility following whole-body irradiation reflects effects on the central nervous system or on the muscle cells themselves. We therefore addressed this issue using a collimating microbeam system. However, radiation targeting requires the animal to be immobilized, and previous studies have anesthetized animals to prevent their movement, thus making it impossible to assess their locomotion immediately after irradiation. We developed a method in which the animal was enclosed in a straight, microfluidic channel in a polydimethylsiloxane chip to inhibit free motion during irradiation, thus allowing locomotion to be observed immediately after irradiation. The head region (including the central nervous system), mid region around the intestine and uterus, and tail region were targeted independently. Each region was irradiated with 12 000 carbon ions (12C; 18.3 MeV/u; linear energy transfer = 106.4 keV/µm), corresponding to 500 Gy at a φ20 µm region. Motility was significantly decreased by whole-body irradiation, but not by irradiation of any of the individual regions, including the central nervous system. This suggests that radiation inhibits locomotion by a whole-body mechanism, potentially involving motoneurons and/or body-wall muscle cells, rather than affecting motor control via the central nervous system and the stimulation response.

Histone Deacetylase Inhibitors Sensitize Murine B16F10 Melanoma Cells to Carbon Ion Irradiation by Inducing G1 Phase Arrest.

Epigenetic processes, in addition to genetic abnormalities, play a critical role in refractory malignant diseases and cause the unresponsiveness to various chemotherapeutic regimens and radiotherapy. Herein we demonstrate that histone deacetylase inhibitors (HDACis) can be used to sensitize malignant melanoma B16F10 cells to carbon ion irradiation. The cells were first treated with HDACis (romidepsin [FK228, depsipeptide], trichostatin A [TSA], valproic acid [VPA], and suberanilohydroxamic acid [SAHA, vorinostat]) and were then exposed to two types of radiation (carbon ions and gamma-rays). We found that HDACis enhanced the radiation-induced apoptosis and suppression of clonogenicity that was induced by irradiation, having a greater effect with carbon ion irradiation than with gamma-rays. Carbon ion irradiation and the HDACi treatment induced G2/M and G0/G1 cell cycle arrest, respectively. Thus, it is considered that HDACi treatment enhanced the killing effects of carbon ion irradiation against melanoma cells by inducing the arrest of G1 phase cells, which are sensitive to radiation due to a lack of DNA homologous recombination repair. Based on these findings, we propose that pretreatment with HDACis as radiosensitizers to induce G1 arrest combined with carbon ion irradiation may have clinical efficacy against refractory cancer.

In vivo 3D analysis of systemic effects after local heavy-ion beam irradiation in an animal model.

Radiotherapy is widely used in cancer treatment. In addition to inducing effects in the irradiated area, irradiation may induce effects on tissues close to and distant from the irradiated area. Japanese medaka, Oryzias latipes, is a small teleost fish and a model organism for evaluating the environmental effects of radiation. In this study, we applied low-energy carbon-ion (26.7 MeV/u) irradiation to adult medaka to a depth of approximately 2.2 mm from the body surface using an irradiation system at the National Institutes for Quantum and Radiological Science and Technology. We histologically evaluated the systemic alterations induced by irradiation using serial sections of the whole body, and conducted a heart rate analysis. Tissues from the irradiated side showed signs of serious injury that corresponded with the radiation dose. A 3D reconstruction analysis of the kidney sections showed reductions in the kidney volume and blood cell mass along the irradiated area, reflecting the precise localization of the injuries caused by carbon-beam irradiation. Capillary aneurysms were observed in the gill in both ventrally and dorsally irradiated fish, suggesting systemic irradiation effects. The present study provides an in vivo model for further investigation of the effects of irradiation beyond the locally irradiated area.

Nitric oxide-mediated bystander signal transduction induced by heavy-ion microbeam irradiation.

In general, a radiation-induced bystander response is known to be a cellular response induced in non-irradiated cells after receiving bystander signaling factors released from directly irradiated cells within a cell population. Bystander responses induced by high-linear energy transfer (LET) heavy ions at low fluence are an important health problem for astronauts in space. Bystander responses are mediated via physical cell-cell contact, such as gap-junction intercellular communication (GJIC) and/or diffusive factors released into the medium in cell culture conditions. Nitric oxide (NO) is a well-known major initiator/mediator of intercellular signaling within culture medium during bystander responses. In this study, we investigated the NO-mediated bystander signal transduction induced by high-LET argon (Ar)-ion microbeam irradiation of normal human fibroblasts. Foci formation by DNA double-strand break repair proteins was induced in non-irradiated cells, which were co-cultured with those irradiated by high-LET Ar-ion microbeams in the same culture plate. Foci formation was suppressed significantly by pretreatment with an NO scavenger. Furthermore, NO-mediated reproductive cell death was also induced in bystander cells. Phosphorylation of NF-κB and Akt were induced during NO-mediated bystander signaling in the irradiated and bystander cells. However, the activation of these proteins depended on the incubation time after irradiation. The accumulation of cyclooxygenase-2 (COX-2), a downstream target of NO and NF-κB, was observed in the bystander cells 6 h after irradiation but not in the directly irradiated cells. Our findings suggest that Akt- and NF-κB-dependent signaling pathways involving COX-2 play important roles in NO-mediated high-LET heavy-ion-induced bystander responses. In addition, COX-2 may be used as a molecular marker of high-LET heavy-ion-induced bystander cells to distinguish them from directly irradiated cells, although this may depend on the time after irradiation.

Dependence of the bystander effect for micronucleus formation on dose of heavy-ion radiation in normal human fibroblasts.

Ionising radiation-induced bystander effects are well recognised, but its dependence on dose or linear energy transfer (LET) is still a matter of debate. To test this, 49 sites in confluent cultures of AG01522D normal human fibroblasts were targeted with microbeams of carbon (103 keV µm(-1)), neon (375 keV µm(-1)) and argon ions (1260 keV µm(-1)) and evaluated for the bystander-induced formation of micronucleus that is a kind of a chromosome aberration. Targeted exposure to neon and argon ions significantly increased the micronucleus frequency in bystander cells to the similar extent irrespective of the particle numbers per site of 1-6. In contrast, the bystander micronucleus frequency increased with increasing the number of carbon-ion particles in a range between 1 and 3 particles per site and was similar in a range between 3 and 8 particles per site. These results suggest that the bystander effect of heavy ions for micronucleus formation depends on dose.

Endovascular repair with contralateral external-to-internal iliac artery bypass grafting: a case series.

BACKGROUND: To report a technique of keeping unilateral blood flow in the internal iliac artery in cases of an abdominal aortic aneurysm in achieving successful Endovascular abdominal aortic aneurysm repair using an external-to-internal artery bypass. CASE PRESENTATION: 6 japanese patients with infra-renal abdominal aortic aneurysms were treated using the retroperitoneal approach via a left (right) paramedian incision followed by an external-to-internal artery bypass. Endovascular abdominal aortic aneurysm repair was conducted on mean postoperative day 29 ± 18 and was performed because the contralateral internal iliac artery, which was not involved in the external-to-internal artery bypass, was treated with a coil embolization. No complications developed during the postoperative follow-up period (17 ± 1.5 months). In all 6 patients, patent grafts were evident on computed tomography angiography scans even after 1-3 months. CONCLUSIONS: Endovascular abdominal aortic aneurysm repair with unilateral internal iliac artery embolization and contralateral external-to-internal artery bypass is feasible with a relatively low risk. It is a safe procedure and reduces the incidence of postoperative complications.

A mathematical model of radiation-induced responses in a cellular population including cell-to-cell communications.

Cell-to-cell communication is an important factor for understanding the mechanisms of radiation-induced responses such as bystander effects. In this study, a new mathematical model of intercellular signalling between individual cells in a cellular population is proposed. The authors considered two types of transmission of signals: via culture medium and via gap junction. They focus on the effects that radiation and intercellular signalling have on cell-cycle modification. The cell cycle is represented as a virtual clock that includes several checkpoint pathways within a cyclic process. They also develop a grid model and set up diffusion equations to model the propagation of signals to and from spatially located cells. The authors have also considered the role that DNA damage plays in the cycle of cells which can progress through the cell cycle or stop at the G1, S, G2 or M-phase checkpoints. Results of testing show that the proposed model can simulate intercellular signalling and cell-cycle progression in individual cells during and after irradiation.

The bystander cell-killing effect mediated by nitric oxide in normal human fibroblasts varies with irradiation dose but not with radiation quality.

PURPOSE: To investigate the dependence of the bystander cell-killing effect on radiation dose and quality, and to elucidate related molecular mechanisms. MATERIALS AND METHODS: Normal human fibroblast WI-38 cells were irradiated with 0.125 - 2 Gy of γ-rays or carbon ions and were co-cultured with non-irradiated cells. Survival rates of bystander cells were investigated using the colony formation assays, and nitrite concentrations in the medium were measured using the modified Saltzman method. RESULTS: Survival rates of bystander cells decreased with doses of γ-rays and carbon ions of ≤ 0.5 Gy. Treatment of the specific nitric oxide (NO) radical scavenger prevented reductions in survival rates of bystander cells. Moreover, nitrite concentrations increased with doses of less than 0.25 Gy (γ-rays) and 1 Gy (carbon ions). The dose responses of increased nitrite concentrations as well as survival reduction were similar between γ-rays and carbon ions. In addition, negative relationships were observed between survival rates and nitrite concentrations. CONCLUSION: The bystander cell-killing effect mediated by NO radicals in normal human fibroblasts depends on irradiation doses of up to 0.5 Gy, but not on radiation quality. NO radical production appears to be an important determinant of γ-ray- and carbon-ion-induced bystander effects.

Early detection of vulnerable atherosclerotic plaque for risk reduction of acute aortic rupture and thromboemboli and atheroemboli using non-obstructive angioscopy.

The mortality rate due to rupture of aortic dissection and aortic aneurysm is approximately 90%. Acute aortic rupture can be fatal prior to hospitalization and has proven difficult to diagnose correctly or predict. The in-hospital mortality rate of ruptured aortic aneurysm ranges from 53 to 66%. Emergency surgical and endovascular treatments are the only options for ruptured aortic dissection and aortic aneurysm. No method of systematic early detection or inspection of vessel injury is available at the prevention stage. Regardless of the improvement in many imaging modalities, aortic diameter has remained a major criterion for recommending surgery in diagnosed patients. Previous reports have suggested a relationship between vulnerable plaque and atherosclerotic aortic aneurysm. Non-obstructive angioscopy is a new method for evaluating intimal injury over the whole aorta. It has been used to identify many advanced atherosclerotic plaques that were missed on traditional imaging modalities before aneurysm formation. Non-obstructive angioscopy has shown that atherosclerosis of the aorta begins before that of the coronary artery, which had been noted on autopsy "in vivo". Strong or repetitive aortic injuries might cause sudden aortic disruption. Aortic atheroma is also a risk factor of stroke and perivascular embolism. Detecting aortic vulnerable atherosclerotic plaque on non-obstructive angioscopy may not only clarify the pathogenesis of acute aortic rupture and "aortogenic" thromboemboli and atheroemboli but also play a role in the pre-emptive medicine.

Nonhomologous end-joining repair plays a more important role than homologous recombination repair in defining radiosensitivity after exposure to high-LET radiation.

DNA double-strand breaks (DSBs) induced by ionizing radiation pose a major threat to cell survival. The cell can respond to the presence of DSBs through two major repair pathways: homologous recombination (HR) and nonhomologous end joining (NHEJ). Higher levels of cell death are induced by high-linear energy transfer (LET) radiation when compared to low-LET radiation, even at the same physical doses, due to less effective and efficient DNA repair. To clarify whether high-LET radiation inhibits all repair pathways or specifically one repair pathway, studies were designed to examine the effects of radiation with different LET values on DNA DSB repair and radiosensitivity. Embryonic fibroblasts bearing repair gene (NHEJ-related Lig4 and/or HR-related Rad54) knockouts (KO) were used and their responses were compared to wild-type cells. The cells were exposed to X rays, spread-out Bragg peak (SOBP) carbon ion beams as well as with carbon, iron, neon and argon ions. Cell survival was measured with colony-forming assays. The sensitization enhancement ratio (SER) values were calculated using the 10% survival dose of wild-type cells and repair-deficient cells. Cellular radiosensitivity was listed in descending order: double-KO cells > Lig4-KO cells > Rad54-KO cells > wild-type cells. Although Rad54-KO cells had an almost constant SER value, Lig4-KO cells showed a high-SER value when compared to Rad54-KO cells, even with increasing LET values. These results suggest that with carbon-ion therapy, targeting NHEJ repair yields higher radiosensitivity than targeting homologous recombination repair.

Increase in cell motility by carbon ion irradiation via the Rho signaling pathway and its inhibition by the ROCK inhibitor Y-27632 in lung adenocarcinoma A549 cells.

This study aimed to investigate the effect of carbon ion (C-ion) irradiation on cell motility through the ras homolog gene family member (Rho) signaling pathway in the human lung adenocarcinoma cell line A549. Cell motility was assessed by a wound-healing assay, and the formation of cell protrusions was evaluated by F-actin staining. Cell viability was examined by the WST-1 assay. The expression of myosin light chain 2 (MLC2) and the phosphorylation of MLC2 at Ser19 (P-MLC2-S19) were analyzed by Western blot. At 48 h after irradiation, the wound-healing assay demonstrated that migration was significantly greater in cells irradiated with C-ion (2 or 8 Gy) than in unirradiated cells. Similarly, F-actin staining showed that the formation of protrusions was significantly increased in cells irradiated with C-ion (2 or 8 Gy) compared with unirradiated cells. The observed increase in cell motility due to C-ion irradiation was similar to that observed due to X-ray irradiation. Western-blot analysis showed that C-ion irradiation (8 Gy) increased P-MLC2-S19 expression compared with in unirradiated controls, while total MLC2 expression was unchanged. Exposure to a non-toxic concentration of Y-27632, a specific inhibitor of Rho-associated coiled-coil-forming protein kinase (ROCK), reduced the expression of P-MLC2-S19 after C-ion irradiation (8 Gy), resulting in a significant reduction in migration. These data suggest that C-ion irradiation increases cell motility in A549 cells via the Rho signaling pathway and that ROCK inhibition reduces that effect.

Quadricuspid aortic valve complicated with infective endocarditis: report of a case.

Congenital quadricuspid aortic valve is a rare cardiac malformation with an unknown risk of infective endocarditis. We report a case of quadricuspid aortic valve complicated with infective endocarditis. A 53-year-old Japanese woman was hospitalized with leg edema and a fever of unknown origin. Corynebacterium striatum was detected in the blood culture. Echocardiography demonstrated a quadricuspid aortic valve with vegetation and severe functional regurgitation. The condition was diagnosed as a quadricuspid aortic valve with infective endocarditis, for which surgery was performed. The quadricuspid aortic valve had three equal-sized cusps and one smaller cusp (type B according to Hurwitz classification). We dissected the vegetation and infectious focus and implanted a mechanical valve. Following the case report, we review the literature.

A branching process model for the analysis of abortive colony size distributions in carbon ion-irradiated normal human fibroblasts.

A single cell can form a colony, and ionizing irradiation has long been known to reduce such a cellular clonogenic potential. Analysis of abortive colonies unable to continue to grow should provide important information on the reproductive cell death (RCD) following irradiation. Our previous analysis with a branching process model showed that the RCD in normal human fibroblasts can persist over 16 generations following irradiation with low linear energy transfer (LET) γ-rays. Here we further set out to evaluate the RCD persistency in abortive colonies arising from normal human fibroblasts exposed to high-LET carbon ions (18.3 MeV/u, 108 keV/µm). We found that the abortive colony size distribution determined by biological experiments follows a linear relationship on the log-log plot, and that the Monte Carlo simulation using the RCD probability estimated from such a linear relationship well simulates the experimentally determined surviving fraction and the relative biological effectiveness (RBE). We identified the short-term phase and long-term phase for the persistent RCD following carbon-ion irradiation, which were similar to those previously identified following γ-irradiation. Taken together, our results suggest that subsequent secondary or tertiary colony formation would be invaluable for understanding the long-lasting RCD. All together, our framework for analysis with a branching process model and a colony formation assay is applicable to determination of cellular responses to low- and high-LET radiation, and suggests that the long-lasting RCD is a pivotal determinant of the surviving fraction and the RBE.

Quantitative analysis of coronary vessels with optimized intracoronary CT number.

BACKGROUND: Variability in intracoronary computed tomography (CT) number may influence vessel quantification. We confirmed the feasibility of a novel method for measuring vessel diameter and area using coronary CT angiography (CCTA) with an optimized intracoronary CT number, 350 HU. METHODS: We performed intravascular ultrasound (IVUS) imaging in 52 patients with significant stenosis detected by coronary CT angiography targeting 350 HU using a CT number-controlling system. We measured 0-to-0 HU distances in the cross-sectional coronary images of 32 patients. We analyzed the ratio of 0-to-0 HU distances in CT images to media-to-media distances in IVUS images (C:I ratio). The area of ≥0 HU for 103 representative points in the remaining 20 patients was compared to the area of the traced external elastic membrane (EEM) in IVUS images. RESULTS: There was a strong correlation between 0-to-0 HU distance in CT images and media-to-media diameter in IVUS images (r = 0.97, p<0.001). The C:I ratio was 1.1. EEM area was estimated by dividing the area of ≥0 HU by the square of C:I. There was also a strong correlation between the estimated EEM area and the EEM area in IVUS images (r = 0.95, p<0.001). CONCLUSIONS: Media-to-media diameter and EEM area can be estimated by CCTA targeting the optimized intracoronary CT number when blood vessel borders are defined at 0 HU.

Nonobstructive angioscopy in patient with atherosclerotic renal artery stenosis.

Few applications of angioscopy for evaluating atherosclerosis of the abdominal aorta have been described. We report the demonstration of atherosclerotic yellow plaque by nonobstructive angioscopy in a patient with left renal artery stenosis. Computed tomography angiography showed stenosis in one of the left renal arteries in a 65-year-old man who presented with renal impairment and hypertension. Invasive selective renal angiography indicated severe stenosis in the proximal portion of the inferior left renal arteries. Intravascular ultrasound demonstrated eccentric plaque with predominant low-density plaque with calcification as the culprit. Percutaneous transluminal renal angioplasty with stent implantation of the left renal artery was performed. Nonobstructive angioscopy demonstrated a grade 3 yellow culprit plaque at the proximal end of the stent, and grade 2 and grade 1 yellow plaques as the culprit plaques at the middle and distal portions of the artery, respectively. .

Gap junction communication and the propagation of bystander effects induced by microbeam irradiation in human fibroblast cultures: the impact of radiation quality.

Understanding the mechanisms underlying the bystander effects of low doses/low fluences of low- or high-linear energy transfer (LET) radiation is relevant to radiotherapy and radiation protection. Here, we investigated the role of gap-junction intercellular communication (GJIC) in the propagation of stressful effects in confluent normal human fibroblast cultures wherein only 0.036-0.144% of cells in the population were traversed by primary radiation tracks. Confluent cells were exposed to graded doses from monochromatic 5.35 keV X ray (LET ~6 keV/µm), 18.3 MeV/u carbon ion (LET ~103 keV/µm), 13 MeV/u neon ion (LET ~380 keV/µm) or 11.5 MeV/u argon ion (LET ~1,260 keV/µm) microbeams in the presence or absence of 18-α-glycyrrhetinic acid (AGA), an inhibitor of GJIC. After 4 h incubation at 37°C, the cells were subcultured and assayed for micronucleus (MN) formation. Micronuclei were induced in a greater fraction of cells than expected based on the fraction of cells targeted by primary radiation, and the effect occurred in a dose-dependent manner with any of the radiation sources. Interestingly, MN formation for the heavy-ion microbeam irradiation in the absence of AGA was higher than in its presence at high mean absorbed doses. In contrast, there were no significant differences in cell cultures exposed to X-ray microbeam irradiation in presence or absence of AGA. This showed that the inhibition of GJIC depressed the enhancement of MN formation in bystander cells from cultures exposed to high-LET radiation but not low-LET radiation. Bystander cells recipient of growth medium harvested from 5.35 keV X-irradiated cultures experienced stress manifested in the form of excess micronucleus formation. Together, the results support the involvement of both junctional communication and secreted factor(s) in the propagation of radiation-induced stress to bystander cells. They highlight the important role of radiation quality and dose in the observed effects.