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Objectives: This review aimed to present studies that prospectively investigated biological effects in patients following diagnostic dentomaxillofacial radiology (DMFR). Methods: Literature was systematically searched to retrieve all studies assessing radiobiological effects of using X-ray imaging in the dentomaxillofacial area, with reference to radiobiological outcomes for other imaging modalities and fields. Results: There is a lot of variability in the reported radiobiological assessment methods and radiation dose measures, making comparisons of radiobiological studies challenging. Most radiological DMFR studies are focusing on genotoxicity and cytotoxicity, data for 2D dentomaxillofacial radiographs, albeit with some methodological weakness biasing the results. For CBCT, available evidence is limited and few studies include comparative data on both adults and children. Conclusions In the future, one will have to strive towards patient-specific measures by considering age, gender and other individual radiation sensitivity-related factors. Ultimately, future radioprotection strategies should build further on the concept of personalized medicine, with patient-specific optimization of the imaging protocol, based on radiobiological variables.

Background Antarctica is a challenging environment for humans. It serves as a spaceflight ground analog, reflecting some conditions of long-duration exploration class space missions. The French-Italian Concordia station in interior Antarctica is a high-fidelity analog, located 1000 km from the coast, at an altitude of 3232 m. The aim of this field study was to characterize the extent, dynamics, and key mechanisms of the immune adaptation in humans overwintering at Concordia for 1 year. Methods This study assessed immune functions in fourteen crewmembers. Quantitative and phenotypic analyses from human blood were performed using onsite flow cytometry together with specific tests on receptor-dependent and receptor-independent functional innate and adaptive immune responses. Transcriptome analyses and quantitative identification of key response genes were assessed. Results Dynamic immune activation and a two-step escalation/activation pattern were observed. The early phase was characterized by moderately sensitized global immune responses, while after 3-4 months, immune responses were highly upregulated. The cytokine responses to an ex vivo stimulation were markedly raised above baseline levels. These functional observations were reflected at the gene transcriptional level in particular through the modulation of hypoxia-driven pathways. Conclusions This study revealed unique insights into the extent, dynamics, and genetics of immune dysfunctions in humans exposed for 1 year to the Antarctic environment at the Concordia station. The scale of immune function was imbalanced toward a sensitizing of inflammatory pathways.

Although immune dysfunction by space conditions has been reported postflight, as well as during ground-based experiments, the cause(s) and nature of the immunological changes are not completely understood. Microgravity has been suggested as one of the factors responsible for the observed immune dysregulation. The goal of this study was to assess immune changes in simulated microgravity (s-μG) using an in vitro cytokine release assay. The effect of s-μG provided by the desktop random positioning machine on cell-mediated immunity was examined by analyzing interleukin 2 (IL-2), interferon-γ (IFN-γ), tumor necrosis factor alpha (TNF-α), and interleukin 10 (IL-10), in response to immune cell stimulation in whole blood samples (n = 10). Stimuli used were bacterial recall antigens, pokeweed mitogen (PWM), lipopolysaccharide (LPS), or heat-killed Listeria monocytogenes (HKLM). S-μG caused an overall inhibition of the IL-2 and IFN-γ responses to recall antigen and mitogen stimulation. More specifically, s-μG most strongly influenced the levels of all four cytokines elicited by bacterial recall antigen stimulation. In contrast, HKLM-induced TNF-α secretion was elevated. The average concentrations of TNF-α in response to PWM and LPS and IL-10 release stimulated by PWM, LPS, and HKLM were not significantly altered by s-μG. However, a variable response between individual subjects could be observed. In conclusion, our results demonstrate that the in vitro cytokine release assay can detect gravity-related immune alterations. Furthermore, the use of multiple stimuli and the associated changes in cytokine secretion has the potential to reveal information on the underlying mechanisms affected by s-μG.

Unlike individuals on Earth who are protected against most types of ionizing radiation by the shielding characteristics of the atmosphere and the electromagnetic field, astronauts aboard a spaceship or on the surface of the moon or another planet are exposed to much higher doses of ionizing radiation. Besides this, the radiation spectrum present in space is very different from that on Earth (see Chap. 20). Terrestrial radiation mainly consists of low-LET radiation such as X- and gamma rays. Space radiation on the other hand is made up of (1) radiation from the solar wind which is trapped in the Earth’s magnetic field (Van Allen belts), (2) galactic cosmic radiation comprised of high-energy protons and heavy ions, and (3) solar particle events mainly consisting of low-energy protons. Recent estimations of the dose equivalent for a round trip to Mars were in the order of 0.66 Sv, which is well beyond the limitations for nuclear workers. Therefore, for manned spaceflight, the biological response to space radiation is of critical concern for risk assessment for astronauts, especially since space agencies like NASA limit the risk of radiation exposure-induced death due to cancer to only 3% (at 95% confidence level). Based on findings from early NASA astronauts, some of whom traveled beyond Low Earth Orbit, no association was found between radiation dose and mortality from cardiovascular disease or cancer risk using a logistic regression model. Nevertheless, for long-duration interplanetary manned missions the uncertainty about the health risks of radiation exposure remains one of the most important limiting factors. For this reason, the identification of (predictive) biomarkers to determine both the received radiation dose (biodosimetry) as well as the radiosensitivity of individuals may be an important aspect for future crew selection. This chapter reviews some of the most frequently used assays for these purposes.

The use of carbon ion therapy for cancer treatment is becoming more widespread due to the advantages of carbon ions compared with X‑rays. Breast cancer patients may benefit from these advantages, as the surrounding healthy tissues receive a lower dose, and the increased biological effectiveness of carbon ions can better control radioresistant cancer cells. Accumulating evidence indicates that the Hedgehog (Hh) pathway is linked to the development and progression of breast cancer, as well as to resistance to X‑irradiation and the migratory capacity of cancer cells. Hence, there is an increasing interest in targeting the Hh pathway in combination with radiotherapy. Several studies have already investigated this treatment strategy with conventional radiotherapy. However, to the best of our knowledge, the combination of Hh inhibitors with particle therapy has not yet been explored. The aim of the present study was to investigate the potential of the Hh inhibitor GANT61 as an effective modulator of radiosensitivity and migration potential in MCF‑7 breast cancer cells, and compare potential differences between carbon ion irradiation and X‑ray exposure. Although Hh targeting was not able to radiosensitise cells to any radiation type used, the combination of GANT61 with X‑rays or carbon ions (energy: 95 MeV/n; linear energy transfer: 73 keV/µm) was more effective in decreasing MCF‑7 cell migration compared with either radiation type alone. Gene expression of the Hh pathway was affected to different degrees in response to X‑ray and carbon ion irradiation, as well as in response to the combination of GANT61 with irradiation. In conclusion, combining Hh inhibition with radiation (X‑rays or carbon ions) more effectively decreased breast cancer cell migration compared with radiation treatment alone. ispartof: INTERNATIONAL JOURNAL OF ONCOLOGY vol:55 issue:6 pages:1339-1348 ispartof: location:Greece status: published

Objectives: Cone-beam CT (CBCT) is a medical imaging technique used in dental medicine. However, there are no conclusive data available indicating that exposure to X-ray doses used by CBCT are harmless. We aim, for the first time, to characterize the potential age-dependent cellular and subcellular effects related to exposure to CBCT imaging. Current objective is to describe and validate the protocol for characterization of cellular and subcellular changes after diagnostic CBCT. Methods: Development and validation of a dedicated two-part protocol: 1) assessing DNA double strand breaks (DSBs) in buccal mucosal (BM) cells and 2) oxidative stress measurements in saliva samples. BM cells and saliva samples are collected prior to and 0.5 h after CBCT examination. BM cells are also collected 24 h after CBCT examination. DNA DSBs are monitored in BM cells via immunocytochemical staining for γH2AX and 53BP1. 8-oxo-7,8-dihydro-2’-deoxyguanosine (8-oxo-dG) and total antioxidant capacity are measured in saliva to assess oxidative damage. Results: Validation experiments show that sufficient BM cells are collected (97.1 ± 1.4 %) and that γH2AX/53BP1 foci can be detected before and after CBCT examination. Collection and analysis of saliva samples, either sham exposed or exposed to IR, show that changes in 8-oxo-dG and total antioxidant capacity can be detected in saliva samples after CBCT examination. Conclusion: The DIMITRA Research Group presents a two-part protocol to analyze potential age-related biological differences following CBCT examinations. This protocol was validated for collecting BM cells and saliva and for analyzing these samples for DNA DSBs and oxidative stress markers, respectively.

Endothelial cells (ECs) grow as single layers on the bottom surface of cell culture flasks under normal (1g) culture conditions. In numerous experiments using simulated microgravity we noticed that the ECs formed three-dimensional, tube-like cell aggregates resembling the intima of small, rudimentary blood vessels. The SPHEROIDS project has now shown that similar processes occur in space. For the first time, we were able to observe scaffold-free growth of human ECs into multicellular spheroids and tubular structures during an experiment in real microgravity. With further investigation of the space samples we hope to understand endothelial 3D growth and to improve the in vitro engineering of biocompatible vessels which could be used in surgery.

Objectives: Cone-beam CT (CBCT), a radiographic tool for diagnosis, treatment, and follow-up in dental practice, was introduced also in pediatric radiology, especially orthodontics. Such patients subjected to repetitive X-rays examinations may receive substantial levels of radiation doses. Ionizing radiation (IR), a recognized carcinogenic factor causing DNA double-strand breaks (DSBs) could be harmful to undifferentiated cells such as dental pulp stem cells (DPSCs) since inaccurately repaired or unrepaired DSBs may lead to malignant transformation. The H2AX and MRE11 proteins generated following DSBs formation and pro-inflammatory cytokines (CKs) secreted after irradiation are relevant candidates to monitor the cellular responses induced by CBCT. Methods: DPSCs were extracted from human exfoliated deciduous teeth and their phenotype was assessed by immunocytochemistry and flow-cytometry. Cells were exposed to IR doses: 5.4-107.7 mGy, corresponding to 0.5-8 consecutive skull exposures, respectively. H2AX and MRE11 were detected in whole cells, while IL-1 alpha, IL-6, IL-8, TNF alpha in supernatants, using enzyme-linked immunosorbent assay (ELISA) at different time points after exposure. Results: The phosphorylation level of H2AX in DPSCs increased considerably at 0.5 h after exposure (p < 0.001 for 3, 5, 8 skull exposures and p < 0.05 for 1 skull exposure, respectively). MRE11 response could only be detected for the highest IR dose (p < 0.001) in the same interval. CKs secretion increased upon CBCT exposure according to doses and time. Conclusions: The DPSCs exposure to CBCT induces transient DNA damage and persistent inflammatory reaction in DPSCs drawing the attention on the potential risks of IR exposures and on the importance of dose monitoring in pediatric population. The research leading to these results has received funding from the European Atomic Energy Community's Seventh Framework Programme FP7/2007-2011 under grant agreement no 604984 (OPERRA: Open Project for the European Radiation Research Area).

Due to the advantages of charged particles compared to conventional radiotherapy, a vast increase is noted in the use of particle therapy in the clinic. These advantages include an improved dose deposition and increased biological effectiveness. Metastasis is still an important cause of mortality in cancer patients and evidence has shown that conventional radiotherapy can increase the formation of metastasizing cells. An important pathway involved in the process of metastasis is the Hedgehog (Hh) signaling pathway. Recent studies have demonstrated that activation of the Hh pathway, in response to X-rays, can lead to radioresistance and increased migratory, and invasive capabilities of cancer cells. Here, we investigated the effect of X-rays, protons, and carbon ions on cell survival, migration, and Hh pathway gene expression in prostate cancer (PC3) and medulloblastoma (DAOY) cell lines. In addition, the potential modulation of cell survival and migration by the Hh pathway inhibitor GANT61 was investigated. We found that in both cell lines, carbon ions were more effective in decreasing cell survival and migration as well as inducing more significant alterations in the Hh pathway genes compared to X-rays or protons. In addition, we show here for the first time that the Hh inhibitor GANT61 is able to sensitize DAOY medulloblastoma cells to particle radiation (proton and carbon ion) but not to conventional X-rays. This important finding demonstrates that the results of combination treatment strategies with X-ray radiotherapy cannot be automatically extrapolated to particle therapy and should be investigated separately. In conclusion, combining GANT61 with particle radiation could offer a benefit for specific cancer types with regard to cancer cell survival. KK is a recipient of a SCK.CEN-KUL PhD grant. KK, NB, MV, and BB received a Horizon2020 travel grant (No. 654002) for experiments at GANIL. KK received an EU CONCERT travel grant and Belgian Hadron therapy Consortium (BHTC) travel grant, whereas MM received an FWO travel grant to perform experiments at iThemba LABS in South-Africa. KH is a clinical research fellow of the Research Foundation Flanders, Belgium.

While many efforts have been made to pave the way toward human space colonization, little consideration has been given to the methods of protecting spacefarers against harsh cosmic and local radioactive environments and the high costs associated with protection from the deleterious physiological effects of exposure to high-Linear energy transfer (high-LET) radiation. Herein, we lay the foundations of a roadmap toward enhancing human radioresistance for the purposes of deep space colonization and exploration. We outline future research directions toward the goal of enhancing human radioresistance, including upregulation of endogenous repair and radioprotective mechanisms, possible leeways into gene therapy in order to enhance radioresistance via the translation of exogenous and engineered DNA repair and radioprotective mechanisms, the substitution of organic molecules with fortified isoforms, and methods of slowing metabolic activity while preserving cognitive function. We conclude by presenting the known associations between radioresistance and longevity, and articulating the position that enhancing human radioresistance is likely to extend the healthspan of human spacefarers as well.

MELODI is the European platform dedicated to low-dose radiation risk research. From 7 October through 10 October 2013 the Fifth MELODI Workshop took place in Brussels, Belgium. The workshop offered the opportunity to 221 unique participants originating from 22 countries worldwide to update their knowledge and discuss radiation research issues through 118 oral and 44 poster presentations. In addition, the MELODI 2013 workshop was reaching out to the broader radiation protection community, rather than only the low-dose community, with contributions from the fields of radioecology, emergency and recovery preparedness, and dosimetry. In this review, we summarise the major scientific conclusions of the workshop, which are important to keep the MELODI strategic research agenda up-to-date and which will serve to establish a joint radiation protection research roadmap for the future.

Feuerecker, Matthias, Brian Crucian, Alex P. Salam, Ales Rybka, Ines Kaufmann, Marjan Moreels, Roel Quintens, Gustav Schelling, Manfred Thiel, Sarah Baatout, Clarence Sams, and Alexander Choukèr. Early adaption in the Antarctic environment at Dome C: Consequences on stress-sensitive innate immune functions. High Alt Med Biol 15:341-348, 2014.-Purpose/Aims: Medical reports of Antarctic expeditions indicate that health is affected under these extreme conditions. The present study at CONCORDIA-Station (Dome C, 3233 m) seeks to investigate the early consequences of confinement and hypobaric hypoxia on the human organism.Nine healthy male participants were included in this study. Data collection occurred before traveling to Antarctica (baseline), and at 1 week and 1 month upon arrival. Investigated parameters included basic physiological variables, psychological stress tests, cell blood count, stress hormones, and markers of innate immune functions in resting and stimulated immune cells. By testing for the hydrogen peroxide (H2O2) production of stimulated polymorphonuclear leukocytes (PMNs), the effects of the hypoxia-adenosine-sensitive immune modulatory pathways were examined.As compared to baseline data, reduced oxygen saturation, hemoconcentration, and an increase of secreted catecholamines was observed, whereas no psychological stress was seen. Upon stimulation, the activity of PMNs and L-selectin shedding was mitigated after 1 week. Endogenous adenosine concentration was elevated during the early phase. In summary, living conditions at high altitude influence the innate immune system's response. After 1 month, some of the early effects on the human organism were restored.As this early adaptation is not related to psychological stress, the changes observed are likely to be induced by environmental stressors, especially hypoxia. As hypoxia is triggering ATP-catabolism, leading to elevated endogenous adenosine concentrations, this and the increased catecholamine concentration might contribute to the early, but reversible downregulation of innate immune functions. This indicates the slope of innate immune adaptation to hypoxia.

Despite the generalized use of photon-based radiation (i.e., gamma rays and X-rays) to treat different cancer types, particle radiotherapy (i.e., protons and carbon ions) is becoming a popular, and more effective tool to treat specific tumors due to the improved physical properties and biological effectiveness. Current scientific evidence indicates that conventional radiation therapy affects the tumor immunological profile in a particular manner, which in turn, might induce beneficial effects both at local and systemic (i.e., abscopal effects) levels. The interaction between radiotherapy and the immune system is being explored to combine immune and radiation (including particles) treatments, which in many cases have a greater clinical effect than any of the therapies alone. Contrary to localized, clinical irradiation, astronauts are exposed to whole body, chronic cosmic radiation, where protons and heavy ions are an important component. The effects of this extreme environment during long periods of time, e.g., a potential mission to Mars, will have an impact on the immune system that could jeopardize the health of the astronauts, hence the success of the mission. To this background, the purpose of this mini review is to briefly present the current knowledge in local and systemic immune alterations triggered by particle irradiation and to propose new lines of future research. Immune effects induced by particle radiation relevant to clinical applications will be covered, together with examples of combined radiotherapy and immunotherapy. Then, the focus will move to outer space, where the immune system alterations induced by cosmic radiation during spaceflight will be discussed.

Space travel is a major challenge for human beings. Especially, the mechanisms through which space conditions might alter animal development have been questioned for a long time. The two major physical stress factors that are of relevance in this context are space radiation and weightlessness. While it has been extensively shown that high doses of ionizing radiation induce deleterious effects on embryonic development, so far, little is known about the potential harmful effects of radiation in combination with microgravity on the developing organism. In the present study, we investigated the effects of simulated microgravity on irradiated STO mouse fetal fibroblast cells using a random positioning machine (RPM). Radiation-induced cell cycle changes were not affected when cells were subjected to simulated microgravity for 24 h. Moreover, no morphological differences were observed in irradiated samples exposed to simulated microgravity compared to cells that were exclusively irradiated. However, microgravity simulation significantly decreased the level of apoptosis at all doses as measured by caspase-3 activity and it prevented cells from undergoing radiation-induced size increase up to 1 Gy.

A simple method is described to establish primary cultures of kidney proximal tubule cells (PTC) on membranes. The permeable membranes represent a unique culture surface, allowing a high degree of differentiation since both apical and basolateral membranes are accessible for medium. Proximal tubule (PT) segments from collagenase-digested mouse renal cortices were grown for 7 days, by which time cells were organized as a confluent monolayer. Electron microscopic evaluation revealed structurally polarized epithelial cells with numerous microvilli, basolateral invaginations, and apical tight junctions. Immunoblotting for markers of distinct parts of the nephron demonstrated that these primary cultures only expressed PT-specific proteins. Moreover immunodetection of distinct components of the receptor-mediated endocytic pathway and uptake of FITC-albumin indicated that these cells expressed a functional endocytotic apparatus. In addition, primary cultures possessed the PT brush-border enzymes, alkaline phosphatase, and γ-glutamyl-transferase, and a phloridzin-sensitive sodium-dependent glucose transport at their apical side. Electrophysiological measurements show that the primary cultured cells have a low transepithelial resistance and high short-circuit current that was completely carried by Na+ similar to a leaky epithelium like proximal tubule cells. This novel method established well-differentiated PTC cultures.

Oligodendrocytes are glial cells responsible for the synthesis and maintenance of myelin in the central nervous system (CNS). Oligodendrocytes are vulnerable to damage occurring in a variety of neurological diseases. Understanding oligodendrocyte biology is crucial for the dissemination of de- and remyelination mechanisms. The goal of the present study is the construction of a protein database of mature rat oligodendrocytes. Post-mitotic oligodendrocytes were isolated from mature Wistar rats and subjected to immunocytochemistry. Proteins were extracted and analyzed by means of two-dimensional gel electrophoresis and two-dimensional liquid chromatography, both coupled to mass spectrometry. The combination of the gel-based and gel-free approach resulted in confident identification of a total of 200 proteins. A minority of proteins were identified in both proteomic strategies. The identified proteins represent a variety of functional groups, including novel oligodendrocyte proteins. The results of this study emphasize the power of the applied proteomic strategy to study known or to reveal new proteins and to investigate their regulation in oligodendrocytes in different disease models.

Multiple sclerosis is a chronic inflammatory disease of the central nervous system. Myelin and oligodendrocytes are considered the major targets of injury caused by a cell-mediated immune response. There is circumstantial evidence that proinflammatory cytokines like tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) could have disease-promoting roles in multiple sclerosis (MS). In the present study, the cytotoxic effects of IFN-gamma and TNF-alpha on the human oligodendroglial cell lines human oligodendroglioma (HOG) and MO3.13 were analyzed. When the oligodendroglial cell lines were cultured in the presence of IFN-gamma or TNF-alpha, apoptotic cell death was observed in both cell lines after >24 hr incubation. Apoptosis was evidenced by a decrease in cell viability, apoptotic changes in cell and nucleus morphology, and disruption of the membrane asymmetry. Our data show that TNF-alpha and IFN-gamma induce apoptosis in a dose-dependent fashion in both oligodendroglial cell lines and that their synergistic effect results in enhanced cell death. Understanding the regulation of cell death pathways in oligodendrocytes is critical for protecting myelin-producing cells and their associated axons during injury in patients with MS.

The synovial membrane (SM) is a source of multipotent mesenchymal stem cells (MSCs), which appeared microscopically to be a relatively homogeneous population of fibroblast-like cells (FCs) in culture (De Bari et al., 2001). The aim of this study was to investigate phenotypic characteristics of the SM-derived FCs (SD-FCs) that could elucidate their origin inside the synovial tissue. Morphological characterization of SD-FCs was assessed by electron microscopy and by expression of surfactant protein A (SP-A). This study, yielded substantial evidence that SD-FCs show ultrastructural and immunocytochemical features of type B synoviocytes; they contained characteristic lamellar bodies (LBs) that are secreted by exocytosis. LB secretion ability was maintained upon passaging (P3-P10). Immunocytochemistry showed that SD-FCs express surfactant protein A (SP-A). Taken together, these results indicate that multipotent SD-MSCs may originate from the synovial lining, having a phenotype highly similar to that of type B synoviocytes. We believe our data highlight the potent ability of type B synoviocytes to have a multilineage differentiation potential.

Hadrontherapy is an advanced form of radiotherapy that uses beams of charged particles (such as protons and carbon ions). Compared with conventional radiotherapy, the main advantages of carbon ion therapy are the precise absorbed dose localization, along with an increased relative biological effectiveness (RBE). This high ballistic accuracy of particle beams deposits the maximal dose to the tumor, while damage to the surrounding healthy tissue is limited. Currently, hadrontherapy is being used for the treatment of specific types of cancer. Previous in vitro studies have shown that, under certain circumstances, exposure to charged particles may inhibit cell motility and migration. In the present study, we investigated the expression of four motility-related genes in prostate (PC3) and colon (Caco-2) cancer cell lines after exposure to different radiation types. Cells were irradiated with various absorbed doses (0, 0.5 and 2 Gy) of accelerated (13)C-ions at the GANIL facility (Caen, France) or with X-rays. Clonogenic assays were performed to determine the RBE. RT-qPCR analysis showed dose- and time-dependent changes in the expression of CCDC88A, FN1, MYH9 and ROCK1 in both cell lines. However, whereas in PC3 cells the response to carbon ion irradiation was enhanced compared with X-irradiation, the effect was the opposite in Caco-2 cells, indicating cell-type-specific responses to the different radiation types.

Hadrontherapy is a form of external radiation therapy, which uses beams of charged particles such as carbon ions. Compared to conventional radiotherapy with photons, the main advantage of carbon ion therapy is the precise dose localization along with an increased biological effectiveness. The first results obtained from prostate cancer patients treated with carbon ion therapy showed good local tumor control and survival rates. In view of this advanced treatment modality we investigated the effects of irradiation with different beam qualities on gene expression changes in the PC3 prostate adenocarcinoma cell line. For this purpose, PC3 cells were irradiated with various doses (0.0, 0.5 and 2.0 Gy) of carbon ions (LET=33.7 keV/μm) at the beam of the Grand Accélérateur National d'Ions Lourds (Caen, France). Comparative experiments with X-rays were performed at the Belgian Nuclear Research Centre. Genome-wide gene expression was analyzed using microarrays. Our results show a downregulation in many genes involved in cell cycle and cell organization processes after 2.0 Gy irradiation. This effect was more pronounced after carbon ion irradiation compared with X-rays. Furthermore, we found a significant downregulation of many genes related to cell motility. Several of these changes were confirmed using qPCR. In addition, recurrence-free survival analysis of prostate cancer patients based on one of these motility genes (FN1) revealed that patients with low expression levels had a prolonged recurrence-free survival time, indicating that this gene may be a potential prognostic biomarker for prostate cancer. Understanding how different radiation qualities affect the cellular behavior of prostate cancer cells is important to improve the clinical outcome of cancer radiation therapy.

Microgravity and cosmic rays as found in space are difficult to recreate on earth. However, ground-based models exist to simulate space flight experiments. In the present study, an experimental model was utilized to monitor gene expression changes in fetal skin fibroblasts of murine origin. Cells were continuously subjected for 65 h to a low dose (55 mSv) of ionizing radiation (IR), comprising a mixture of high‑linear energy transfer (LET) neutrons and low-LET gamma-rays, and/or simulated microgravity using the random positioning machine (RPM), after which microarrays were performed. The data were analyzed both by gene set enrichment analysis (GSEA) and single gene analysis (SGA). Simulated microgravity affected fetal murine fibroblasts by inducing oxidative stress responsive genes. Three of these genes are targets of the nuclear factor‑erythroid 2 p45-related factor 2 (Nrf2), which may play a role in the cell response to simulated microgravity. In addition, simulated gravity decreased the expression of genes involved in cytoskeleton remodeling, which may have been caused by the downregulation of the serum response factor (SRF), possibly through the Rho signaling pathway. Similarly, chronic exposure to low-dose IR caused the downregulation of genes involved in cytoskeleton remodeling, as well as in cell cycle regulation and DNA damage response pathways. Many of the genes or gene sets that were altered in the individual treatments (RPM or IR) were not altered in the combined treatment (RPM and IR), indicating a complex interaction between RPM and IR.

Ionizing radiation can elicit harmful effects on the cardiovascular system at high doses. Endothelial cells are critical targets in radiation-induced cardiovascular damage. Astronauts performing a long-term deep space mission are exposed to consistently higher fluences of ionizing radiation that may accumulate to reach high effective doses. In addition, cosmic radiation contains high linear energy transfer (LET) radiation that is known to produce high values of relative biological effectiveness (RBE). The aim of this study was to broaden the understanding of the molecular response to high LET radiation by investigating the changes in gene expression in endothelial cells. For this purpose, a human endothelial cell line (EA.hy926) was irradiated with accelerated nickel ions (Ni) (LET, 183 keV/mu m) at doses of 0.5, 2 and 5 Gy. DNA damage was measured 2 and 24 h following irradiation by gamma-H2AX foci detection by fluorescence microscopy and gene expression changes were measured by microarrays at 8 and 24 h following irradiation. We found that exposure to accelerated nickel particles induced a persistent DNA damage response up to 24 h after treatment. This was accompanied by a downregulation in the expression of a multitude of genes involved in the regulation of the cell cycle and an upregulation in the expression of genes involved in cell cycle checkpoints. In addition, genes involved in DNA damage response, oxidative stress, apoptosis and cell-cell signaling (cytokines) were found to be upregulated. An in silico analysis of the involved genes suggested that the transcription factors, E2F and nuclear factor (NF)-kappa B, may be involved in these cellular responses.

In case of accidental radiation exposure or a nuclear incident, physical dosimetry is not always complete. Therefore, it is important to develop tools that allow dose estimates and determination that are based on biological markers of radiation exposure. Exposure to ionizing radiation triggers a large-scale activation of specific DNA signaling and repair mechanisms. This includes the phosphorylation of γH2AX in the vicinity of a double-strand break (DSB). A DNA DSB is a cytotoxic form of DNA damage, and if not correctly repaired can initiate genomic instability, chromosome aberrations, mutations or apoptosis. Measurements of DNA DSBs and their subsequent repair after in vitro irradiation has been suggested to be of potential use to monitor cellular responses. The bone marrow and the blood are known to be the most radiosensitive tissues of the human body and can therefore be of particular importance to find radiation-induced biological markers. In the present study, changes in H2AX phosphorylation and apoptosis of irradiated human peripheral blood mononuclear cells (PBMCs) were analyzed. Freshly isolated PBMCs from healthy donors were irradiated with X-rays (0.1, 0.25, 0.5, 1, 2 and 4 Gy). The phosphorylation of γH2AX was measured at different time points (0, 0.25, 1, 2, 4, 6 and 24 h) after irradiation. We detected a linear dose-dependency of γH2AX phosphorylation measured by γH2AX foci scoring using immunofluorescence microscopy as well as by γH2AX fluorescence detection using flow cytometry. Apoptosis was detected by measuring DNA fragmentation at different time points (0, 24, 48, 72, 96 h) after X-irradiation using DNA ladder gel electrophoresis. The apoptotic DNA fragmentation increased in a dose-dependent manner. In conclusion, DNA DSBs and subsequent apoptotic DNA fragmentation monitoring have potential as biomarkers for assessing human exposure in radiation biodosimetry.

The impact of ionizing radiation on developing organisms has been widely studied for risk assessment purposes. Even though efforts have been made to decrease received doses to as low as reasonably achievable, the possibility of accidental exposure has to be considered as well. Mammalian gestation is usually divided into three periods. Radiation exposure during the 'pre-implantation period' may essentially result in embryonic mortality while exposure during the 'organogenesis period' may characteristically lead to malformations. In humans, the 'fetal period' is one of particular sensitivity to radiation induction of mental retardation, especially if the exposure occurs between weeks 8-15 of gestation. It is also admitted that prenatal irradiation may increase the risk of leukemia and childhood cancer, with an equal risk over the whole pregnancy. The aim of this study was to investigate the effects of moderate to high doses of X-irradiation on mouse skin fetal fibroblasts, one of the cell types subjected to the highest dose of radiation. Exposure of the cells to X-rays led to a rapid and significant increase in γ-H2AX foci, indicative of high levels of DNA double strand breaks. High doses (2 Gy) also led to a pronounced G2-arrest and a decrease in the number of cells in the S phase, which was followed after 24 h by a decrease in cell survival and an increase in the level of apoptosis and necrosis. This study shows that mouse fetal skin fibroblasts are sensitive to high doses of X-irradiation. Furthermore, we report a better repair for higher doses than lower, which seems to indicate that little DNA damage is not necessarily repaired immediately. However, more sensitive approaches are necessary to identify the risk associated with low doses of radiation.

Unlike individuals on Earth who are protected against most types of radiation by the shielding characteristics of the atmosphere and the electromagnetic field, astronauts aboard a spaceship or on the surface of the moon or another planet will receive much higher doses of ionizing radiation. Besides this, the radiation quality present in space is very different from the radiation on Earth. Whereas typical terrestrial radiation mainly consists of low-LET radiation such as X and gamma rays, space radiation is comprised of high-energy protons and high-charge (Z) and energy (E) nuclei. Therefore, for manned spaceflight, the biological response after cosmic ray exposure is of critical concern for risk assessment for astronauts (Legner 2004). Ongoing research focuses on the identification of (predictive) biomarkers to determine the radiosensitivity of an individual. This chapter reviews some of the most frequently used cytogenetic assays. In addition, emerging technologies for high-throughput screening of interindividual differences in radiosensitivity are discussed.

In contrast to individuals on Earth, astronauts receive much higher doses of ionizing radiation during spaceflight. Besides this, the type of radiation in space is quite different from terrestrial radiation, and consists mainly of high-energy protons and highly charged and energetic particles. It is well known that all types of ionizing radiation induce a large spectrum of DNA lesions and the global response of a cell to DNA damage triggers multiple pathways involved in sensing DNA damage, activating cell cycle checkpoints and inducing DNA repair. However, when damage is severe, apoptosis, also known as “programmed cell death”, can be induced. Ionizing radiation exposure induces biological effects, depending on the type of radiation, dose, and exposure time. In this context, radiation dosimetry on board spacecraft can be useful to estimate the cumulative equivalent doses to which astronauts are exposed. So far, the precise impact of radiation on the deterioration of the immune system in astronauts is still under investigation. Several ground-based experiments on various radiation facilities have been performed and have contributed to our general knowledge of radiation-induced immune changes. Crucially, radiation interacts with other spaceflight stressors, such as microgravity, thereby resulting in a joint effect on the immune system.

Space travel exposes astronauts to a plethora of potentially detrimental conditions, such as cosmic radiation and microgravity. As both factors are hard to simulate on Earth, present knowledge remains limited. However, this knowledge is of vital importance, making space flight experiments a necessity for determining the biological effects and the underlying biochemical processes, especially when keeping future long-term interplanetary missions in mind. Instead of estimating the long-term effects, which usually implicate severe endpoints (e.g., cancer) and which are often difficult to attribute, research has shifted to finding representative biomarkers for rapid and sensitive detection of individual radiosensitivity. In this context, an appealing set of candidate markers is the group of secreted proteins, as they exert an intercellular signaling function and are easy to assess. We screened a subset of secreted proteins in cells exposed to space travel by means of multiplex bead array analysis. To determine the cell-specific signatures of the secreted molecules, we compared the conditioned medium of normal fibroblast cells to fibroblasts isolated from a patient with Hutchinson-Gilford Progeria syndrome, which are known to have a perturbed nuclear architecture and DNA damage response. Out of the 88 molecules screened, 20 showed a significant level increase or decrease, with a differential response to space conditions between the two cell types. Among the molecules that were retained, which may prove to be valuable biomarkers, are apolipoprotein C-III, plasminogen activator inhibitor type 1, beta-2-microglobulin, ferritin, MMP-3, TIMP-1 and VEGF.

This study investigated whether neonatal rat cardiomyocytes (NRCM), when co-cultured, can induce transdifferentiation of either human mesenchymal stromal cells (MSC) or hematopoietic stem cells (HSC) into cardiomyocytes. Stem cells were obtained from patients with ischemic heart disease.Ex vivo-expanded MSC or freshly isolated HSC were used to set-up a co-culture system between NRCM and MSC or HSC. 5-azacytidin (5-aza) or dimethylsulfoxide (DMSO) was used as differentiation-inducing factor. Co-cultured stem cells were separated from NRCM by flow sorting, and cardiac gene expression was analyzed by reverse transcriptase-polymerase chain reaction. Cellular morphology was analyzed by immunofluorescence and transmission electron microscopy (TEM).Co-culturing MSC induced expression of troponin T and GATA-4. However, no expression of alpha-actinin, myosin heavy chain or troponin I was detected. In the case of HSC, only expression of troponin T could be induced. Immunofluorescence and TEM confirmed the absence of sarcomeric organization in co-cultured MSC and HSC. Adding 5-aza or DMSO to the co-cultures did not influence differentiation.This in vitro co-culture study obtained no convincing evidence of transdifferentiation of either MSC or HSC into functional cardiomyocytes. Nevertheless, induction of troponin T was observed in MSC and HSC, and GATA-4 in MSC. However, no morphologic changes could be detected by immunofluorescence or by TEM. These data could explain why only limited functional improvement was reported in clinical stem cell trials.

Aims Rapid and extensive activation of astrocytes occurs subsequent to many forms of central nervous system (CNS) injury. Recent studies have revealed that the expression profile of reactive astrocytes comprises antigens present during astrocyte development. Elevated levels of the injury-related cytokine transforming growth factor-beta 1 (TGF-beta1) secreted by microglial cells and invading macrophages have been correlated with the reactive astrocyte phenotype and glial scar formation. Methods In the present study, the expression profile of alpha-smooth muscle actin (alpha-SMA) and nestin, two cytoskeletal proteins expressed during astrocyte development, was studied in multiple sclerosis (MS) lesions. In addition, alpha-SMA and nestin organization and expression were analysed in rat primary astrocyte cultures in response to TGF-beta1. Results In active lesions and in the hypercellular margin of chronic active MS lesions, immunostaining for alpha-SMA revealed a subpopulation of reactive astrocytes, whereas the majority of reactive astrocytes expressed nestin. alpha-SMA and nestin expressing reactive astrocytes were in close relationship with TGF-beta1 expressing macrophages or microglia. In addition, TGF-beta1 expression within alpha-SMA or nestin expressing astrocytes was also detected. Our in vitro experiments showed that TGF-beta1 regulated the organization and expression of alpha-SMA and nestin in astrocytes. Conclusions Reactive astrocytes in active MS lesions re-express alpha-SMA and nestin. We suggest that the in vivo re-expression might be under regulation of TGF-beta1. These results further clarify the regulation of astrocyte activity after CNS injury, which is important for the astroglial adaptation to pathological situations.

Oligodendrocytes are glial cells responsible for the synthesis and maintenance of myelin in the central nervous system (CNS). Oligodendrocytes are vulnerable to damage occurring in a variety of neurological diseases. Understanding oligodendrocyte biology is crucial for the dissemination of de- and remyelination mechanisms. The goal of the present study is the construction of a protein database of mature rat oligodendrocytes. Post-mitotic oligodendrocytes were isolated from mature Wistar rats and subjected to immunocytochemistry. Proteins were extracted and analyzed by means of two-dimensional gel electrophoresis and two-dimensional liquid chromatography, both coupled to mass spectrometry. The combination of the gel-based and gel-free approach resulted in confident identification of a total of 200 proteins. A minority of proteins were identified in both proteomic strategies. The identified proteins represent a variety of functional groups, including novel oligodendrocyte proteins. The results of this study emphasize the power of the applied proteomic strategy to study known or to reveal new proteins and to investigate their regulation in oligodendrocytes in different disease models.

Radial glial cells are transiently bipolar cells in the developing central nervous system, best known for their role in guiding migrating neurons. The aim of the present study was to investigate phenotypic characteristics of these bipolar precursor cells in a mixed glial cell culture system derived from the rat neonatal spinal cord. Morphological characterization was assessed by cell-specific immunocytochemical markers (nestin, vimentin, 3CB2) and transmission electron microscopy. Our study yielded substantial evidence showing that the bipolar cells exhibit immunocytochemical and ultrastructural features of radial glial cells. Immunohistochemistry of the neonatal rat spinal cord using the same cell-specific markers suggested these cells are likely derived from the subependymal zone, ventral commissure, and dorsomedial septum. We believe our data recommend this mixed glial culture system to be a valuable tool in studying radial glial cells in vitro.

In multiple sclerosis (MS), damage to oligodendrocytes is believed to be caused by an aberrant immune response initiated by autoreactive T cells. Increasing evidence indicates that these T cells are not exclusively detrimental but might also exert protective effects. We report for the first time that myelin-reactive T-cell clones from eight MS patients (6/19) and five healthy controls (4/11) produce leukemia inhibitory factor (LIF), a member of the neuropoietic family of neurotrophins. In addition, T-cell clones specific for tetanus toxoid, CD4(+) and CD8(+) T cells, and monocytes, but not B cells, secreted LIF LIF-producing T lymphocytes and macrophages were also identified immunohistochemically in both active and chronic-active MS lesions. We further demonstrated dose-dependent protective effects of LIF on tumor necrosis factor-alpha-induced apoptosis of oligodendrocytes. In conclusion, our data demonstrate that peripheral and CNS-infiltrating T cells from MS patients produce LIF, a protective factor for oligodendrocytes. This study emphasizes that secretion of LIF may contribute to the neuroprotective effects of auto reactive T cells. (C) 2006 Wiley- Liss, Inc.