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Purpose: MicroRNA miR-21 has emerged as a therapeutic target in the treatment of breast cancer. This study was designed to compare the responses of breast cancer cells and non-transformed breast epithelial cells to a combined regimen of miR-21 inhibition and radiation. Materials and methods: The MDA-MB-361 (breast cancer) and MCF-10A (non-transformed mammary epithelial) cell lines were used for the comparison in this in vitro study. The stable knockdown of miR-21 was performed by using lentiviral approach. The response of the cells was monitored 4, 24 and 48 h after the irradiation with 0.25 and 2.5 Gy, using sham-irradiated cells as controls. The response of the cells was established by performing various functional assays – cell viability and cell attachment, clonogenic survival, cell cycle analysis and 3D microtissue formation. Results: The knockdown of miR-21 induced significant increase in apoptosis and growth delay in MDA-MB-361 cancer cells compared to non-transformed MCF-10A cells. After combined radiation and anti-miR-21 treatment, MDA-MB-361 cells show reduced cell growth and viability what is presented in their inability to form colonies. MCF-10A cells were not as sensitive to the combined treatment and that has also been confirmed with colony forming assay. Conclusions: Cellular response to a combined treatment of anti-miR-21 and radiation is different between cancer and non-cancer cells which highly support the idea of linking miR-21 inhibitor and radiation treatment in the future therapeutic approaches for breast cancer.

Epidemiological data from patients undergoing radiotherapy for thoracic tumours clearly show the damaging effect of ionising radiation on cardiovascular system. The long-term impairment of heart function and structure after local high-dose irradiation is associated with systemic inflammatory response, contraction impairment, microvascular damage and cardiac fibrosis. The goal of the present study was to investigate molecular mechanisms involved in this process. C57BL/6J mice received a single X-ray dose of 16 Gy given locally to the heart at the age of 8 weeks. Radiation-induced changes in the heart transcriptome and proteome were investigated 40 weeks after the exposure. The omics data were analysed by bioinformatics tools and validated by immunoblotting. Integrated network analysis of transcriptomics and proteomics data elucidated the signalling pathways that were similarly affected at gene and protein level. Analysis showed induction of transforming growth factor (TGF) beta signalling but inactivation of peroxisome proliferator-activated receptor (PPAR) alpha signalling in irradiated heart. The putative mediator role of mitogen-activated protein kinase (MAPK) cascade linking PPAR alpha and TGF beta signalling was supported by data from immunoblotting and ELISA. This study indicates that both signalling pathways are involved in radiation-induced heart fibrosis, metabolic disordering and impaired contractility, a pathophysiological condition that is often observed in patients that received high radiation doses in thorax.

// Barbara Tanno 1, * , Gabriele Babini 2, * , Simona Leonardi 1 , Paola Giardullo 3, 4 , Ilaria De Stefano 3 , Emanuela Pasquali 1 , Andrea Ottolenghi 2 , Michael J. Atkinson 5 , Anna Saran 1 , Mariateresa Mancuso 1 1 Laboratory of Biomedical Technologies, Agenzia Nazionale per le Nuove Tecnologie, l'Energia e lo Sviluppo Economico Sostenibile (ENEA), Rome, Italy 2 Department of Physics, University of Pavia, Pavia, Italy 3 Department of Radiation Physics, Guglielmo Marconi University, Rome, Italy 4 Department of Sciences, Roma Tre University, Rome, Italy 5 Helmholtz Zentrum Munchen, German Research Center for Environmental Health, Institute of Radiation Biology, Neuherberg, Germany * These authors contributed equally to this work Correspondence to: Mariateresa Mancuso, email: mariateresa.mancuso@enea.it Anna Saran, email: anna.saran@enea.it Keywords: miRNA, X-rays, medulloblastoma, Shh, GCPs Received: June 08, 2016 Accepted: September 05, 2016 Published: September 10, 2016 ABSTRACT It has historically been accepted that incorrectly repaired DNA double strand breaks (DSBs) are the principal lesions of importance regarding mutagenesis, and long-term biological effects associated with ionizing radiation. However, radiation may also cause dysregulation of epigenetic processes that can lead to altered gene function and malignant transformation, and epigenetic alterations are important causes of miRNAs dysregulation in cancer. Patched1 heterozygous ( Ptch1 +/− ) mice, characterized by aberrant activation of the Sonic hedgehog (Shh) signaling pathway, are a well-known murine model of spontaneous and radiation-induced medulloblastoma (MB), a common pediatric brain tumor originating from neural granule cell progenitors (GCPs). The high sensitivity of neonatal Ptch1 +/− mice to radiogenic MB is dependent on deregulation of the Ptch1 gene function. Ptch1 activates a growth and differentiation programme that is a strong candidate for regulation through the non-coding genome. Therefore we carried out miRNA next generation sequencing in ex vivo irradiated and control GCPs, isolated and purified from cerebella of neonatal WT and Ptch1 +/− mice. We identified a subset of miRNAs, namely let-7 family and miR-17~92 cluster members, whose expression is altered in GCPs by radiation alone, or by synergistic interaction of radiation with Shh-deregulation. The same miRNAs were further validated in spontaneous and radiation-induced MBs from Ptch1 +/− mice, confirming persistent deregulation of these miRNAs in the pathogenesis of MB. Our results support the hypothesis that miRNAs dysregulation is associated with radiosensitivity of GCPs and their neoplastic transformation in vivo .

// Omid Azimzadeh 1 , Tamara Azizova 2 , Juliane Merl-Pham 3 , Vikram Subramanian 1 , Mayur V. Bakshi 1 , Maria Moseeva 2 , Olga Zubkova 2 , Stefanie M. Hauck 3 , Natasa Anastasov 1 , Michael J. Atkinson 1, 4 , Soile Tapio 1 1 Helmholtz Zentrum Munchen-German Research Center for Environmental Health (GmbH), Institute of Radiation Biology, Neuherberg, Germany 2 Southern Urals Biophysics Institute, Russian Federation, Ozyorsk, Russia 3 Helmholtz Zentrum Munchen-German Research Center for Environmental Health (GmbH), Research Unit Protein Science, Munich, Germany 4 Chair of Radiation Biology, Technical University of Munich, Munich, Germany Correspondence to: Soile Tapio, email: soile.tapio@helmholtz-muenchen.de Keywords: ionising radiation, proteomics, heart disease, PPAR alpha, mitochondrial dysfunction Received: April 08, 2016 Accepted: June 17, 2016 Published: July 06, 2016 ABSTRACT Epidemiological studies show a significant increase in ischemic heart disease (IHD) incidence associated with total external gamma-ray dose among Mayak plutonium enrichment plant workers. Our previous studies using mouse models suggest that persistent alteration of heart metabolism due to the inhibition of peroxisome proliferator-activated receptor (PPAR) alpha accompanies cardiac damage after high doses of ionising radiation. The aim of the present study was to elucidate the mechanism of radiation-induced IHD in humans. The cardiac proteome response to irradiation was analysed in Mayak workers who were exposed only to external doses of gamma rays. All participants were diagnosed during their lifetime with IHD that also was the cause of death. Label-free quantitative proteomics analysis was performed on tissue samples from the cardiac left ventricles of individuals stratified into four radiation dose groups (0 Gy, 500 mGy). The groups could be separated using principal component analysis based on all proteomics features. Proteome profiling showed a dose-dependent increase in the number of downregulated mitochondrial and structural proteins. Both proteomics and immunoblotting showed decreased expression of several oxidative stress responsive proteins in the irradiated hearts. The phosphorylation of transcription factor PPAR alpha was increased in a dose-dependent manner, which is indicative of a reduction in transcriptional activity with increased radiation dose. These data suggest that chronic external radiation enhances the risk for IHD by inhibiting PPAR alpha and altering the expression of mitochondrial, structural, and antioxidant components of the heart.

Simple Summary Combined chemotherapy and radiotherapy are an effective treatment for invasive breast cancer. However, some studies suggest that such interventions may increase the risk of metastasis. Cell metastatic behavior is highly dependent on RAS-RAF-MEK pathway and its downstream target activation, including miR-221 overexpression and epithelial-to-mesenchymal transition (EMT). By using MEK1 inhibitor (TAK-733) in combination with radiation therapy for breast cancer cells, significant decrease in migration capacity, including reduction of miR-221 and EMT (ZEB1) marker expression was observed. miR-221 holds great potential as therapeutic biomarker and target for new drug developments, however more insight into efficiency of miR-221 inhibition needs to be followed in the future. Abstract The miR-221 expression is dependent on the oncogenic RAS-RAF-MEK pathway activation and influences epithelial-to-mesenchymal transition (EMT). The Cancer Genome Atlas (TCGA) database analysis showed high gene significance for ZEB1 with EMT module analysis and miR-221 overexpression within the triple-negative breast cancer (TNBC) and HER2+ subgroups when compared to luminal A/B subgroups. EMT marker expression analysis after MEK1 (TAK-733) inhibitor treatment and irradiation was combined with miR-221 and ZEB1 expression analysis. The interaction of miR-221 overexpression with irradiation and its influence on migration, proliferation, colony formation and subsequent EMT target activation were investigated. The results revealed that MEK1 inhibitor treatment combined with irradiation could decrease the migratory potential of breast cancer cells including reduction of miR-221 and corresponding downstream ZEB1 (EMT) marker expression. The clonogenic survival assays revealed that miR-221 overexpressing SKBR3 cells were more radioresistant when compared to the control. Remarkably, the effect of miR-221 overexpression on migration in highly proliferative and highly HER2-positive SKBR3 cells remained constant even upon 8 Gy irradiation. Further, in naturally miR-221-overexpressing MDA-MB-231 cells, the proliferation and migration significantly decrease after miR-221 knockdown. This leads to the assumption that radiation alone is not reducing migration capacity of miR-221-overexpressing cells and that additional factors play an important role in this context. The miR-221/ZEB1 activity is efficiently targeted upon MEK1 inhibitor (TAK-733) treatment and when combined with irradiation treatment, significant reduction in migration of breast cancer cells was shown.

Epidemiological studies on workers employed at the Mayak plutonium enrichment plant have demonstrated an association between external gamma ray exposure and an elevated risk of ischemic heart disease (IHD). In a previous study using fresh-frozen post mortem samples of the cardiac left ventricle of Mayak workers and non-irradiated controls, we observed radiation-induced alterations in the heart proteome, mainly downregulation of mitochondrial and structural proteins. As the control group available at that time was younger than the irradiated group, we could not exclude age as a confounding factor. To address this issue, we have now expanded our study to investigate additional samples using archival formalin-fixed paraffin-embedded (FFPE) tissue. Importantly, the control group studied here is older than the occupationally exposed (&gt, 500 mGy) group. Label-free quantitative proteomics analysis showed that proteins involved in the lipid metabolism, sirtuin signaling, mitochondrial function, cytoskeletal organization, and antioxidant defense were the most affected. A histopathological analysis elucidated large foci of fibrotic tissue, myocardial lipomatosis and lymphocytic infiltrations in the irradiated samples. These data highlight the suitability of FFPE material for proteomics analysis. The study confirms the previous results emphasizing the role of adverse metabolic changes in the radiation-associated IHD. Most importantly, it excludes age at the time of death as a confounding factor.

In children, ketamine sedation is often used during radiological procedures. Combined exposure of ketamine and radiation at doses that alone did not affect learning and memory induced permanent cognitive impairment in mice. The aim of this study was to elucidate the mechanism behind this adverse outcome. Neonatal male NMRI mice were administered ketamine (7.5 mg kg&minus, 1) and irradiated (whole-body, 100 mGy or 200 mGy, 137Cs) one hour after ketamine exposure on postnatal day 10. The control mice were injected with saline and sham-irradiated. The hippocampi were analyzed using label-free proteomics, immunoblotting, and Golgi staining of CA1 neurons six months after treatment. Mice co-exposed to ketamine and low-dose radiation showed alterations in hippocampal proteins related to neuronal shaping and synaptic plasticity. The expression of brain-derived neurotrophic factor, activity-regulated cytoskeleton-associated protein, and postsynaptic density protein 95 were significantly altered only after the combined treatment (100 mGy or 200 mGy combined with ketamine, respectively). Increased numbers of basal dendrites and branching were observed only after the co-exposure, thereby constituting a possible reason for the displayed alterations in behavior. These data suggest that the risk of radiation-induced neurotoxic effects in the pediatric population may be underestimated if based only on the radiation dose.

PurposeGlioblastoma is the most common and lethal adult brain tumor. Despite current therapeutic strategies, including surgery, radiation and chemotherapy, the median survival of glioblastoma patients is 15months. The development of this tumor depends on a sub-population of glioblastoma stem cells governing tumor propagation and therapy resistance. SOX3 plays a role in both normal neural development and carcinogenesis. However, little is known about its role in glioblastoma. Thus, the aim of this work was to elucidate the role of SOX3 in glioblastoma.MethodsSOX3 expression was assessed using real-time quantitative PCR (RT-qPCR), Western blotting and immunohistochemistry. MTT, immunocytochemistry and Transwell assays were used to evaluate the effects of exogenous SOX3 overexpression on the viability, proliferation, migration and invasion of glioblastoma cells, respectively. The expression of Hedgehog signaling pathway components and autophagy markers was assessed using RT-qPCR and Western blot analyses, respectively.ResultsHigher levels of SOX3 expression were detected in a subset of primary glioblastoma samples compared to those in non-tumoral brain tissues. Exogenous overexpression of this gene was found to increase the proliferation, viability, migration and invasion of glioblastoma cells. We also found that SOX3 up-regulation was accompanied by an enhanced activity of the Hedgehog signaling pathway and by suppression of autophagy in glioblastoma cells. Additionally, we found that SOX3 expression was elevated in patient-derived glioblastoma stem cells, as well as in oncospheres derived from glioblastoma cell lines, compared to their differentiated counterparts, implying that SOX3 expression is associated with the undifferentiated state of glioblastoma cells.ConclusionFrom our data we conclude that SOX3 can promote the malignant behavior of glioblastoma cells.

Background Mesenchymal stem cells (MSCs) are attracting increasing interest for cell-based therapies, making use of both their immuno-modulating and regenerative potential. For such therapeutic applications, a massive in vitro expansion of donor cells is usually necessary to furnish sufficient material for transplantation. It is not established to what extent the long-term genomic stability and potency of MSCs can be compromised as a result of this rapid ex vivo expansion. In this study, we investigated the DNA damage response and chromosomal stability (indicated by micronuclei induction) after sub-lethal doses of gamma irradiation in murine MSCs at different stages of their in vitro expansion. Methods Bone-marrow-derived tri-potent MSCs were explanted from 3-month-old female FVB/N mice and expanded in vitro for up to 12 weeks. DNA damage response and repair kinetics after gamma irradiation were quantified by the induction of γH2AX/53BP1 DSB repair foci. Micronuclei were counted in post-mitotic, binucleated cells using an automated image analyzer Metafer4. Involvement of DNA damage response pathways was tested using chemical ATM and DNA-PK inhibitors. Results Murine bone-marrow-derived MSCs in long-term expansion culture gradually lose their ability to recognize endogenous and radiation-induced DNA double-strand breaks. This impaired DNA damage response, indicated by a decrease in the number of γH2AX/53BP1 DSB repair foci, was associated with reduced ATM dependency of foci formation, a slower DNA repair kinetics, and an increased number of residual DNA double-strand breaks 7 h post irradiation. In parallel with this impaired efficiency of DNA break recognition and repair in older MSCs, chromosomal instability after mitosis increased significantly as shown by a higher number of micronuclei, both spontaneously and induced by γ-irradiation. Multifactorial regression analysis demonstrates that in vitro aging reduced DNA damage recognition in MSCs after irradiation by a multiplicative interaction with dose (p

High-dose ionizing radiation is known to induce adverse effects such as inflammation and fibrosis in the heart. Transcriptional regulators PPARα and TGFβ are known to be involved in this radiation response. PPARα, an anti-inflammatory transcription factor controlling cardiac energy metabolism, is inactivated by irradiation. The pro-inflammatory and pro-fibrotic TGFβ is activated by irradiation via SMAD-dependent and SMAD-independent pathways. The goal of this study was to investigate how altering the level of PPARα influences the radiation response of these signaling pathways. For this purpose, we used genetically modified C57Bl/6 mice with wild type (+/+), heterozygous (+/-) or homozygous (-/-) PPARα genotype. Mice were locally irradiated to the heart using doses of 8 or 16 Gy; the controls were sham-irradiated. The heart tissue was investigated using label-free proteomics 20 weeks after the irradiation and the predicted pathways were validated using immunoblotting, ELISA, and immunohistochemistry. The heterozygous PPARα mice showed most radiation-induced changes in the cardiac proteome, whereas the homozygous PPARα mice showed the least changes. Irradiation induced SMAD-dependent TGFβ signaling independently of the PPARα status, but the presence of PPARα was necessary for the activation of the SMAD-independent pathway. These data indicate a central role of PPARα in cardiac response to ionizing radiation.

This long-term study examined relationships between solar and magnetic factors and the time course and lags of autonomic nervous system (ANS) responses to changes in solar and geomagnetic activity. Heart rate variability (HRV) was recorded for 72 consecutive hours each week over a five-month period in 16 participants in order to examine ANS responses during normal background environmental periods. HRV measures were correlated with solar and geomagnetic variables using multivariate linear regression analysis with Bonferroni corrections for multiple comparisons after removing circadian influences from both datasets. Overall, the study confirms that daily ANS activity responds to changes in geomagnetic and solar activity during periods of normal undisturbed activity and it is initiated at different times after the changes in the various environmental factors and persist over varying time periods. Increase in solar wind intensity was correlated with increases in heart rate, which we interpret as a biological stress response. Increase in cosmic rays, solar radio flux, and Schumann resonance power was all associated with increased HRV and parasympathetic activity. The findings support the hypothesis that energetic environmental phenomena affect psychophysical processes that can affect people in different ways depending on their sensitivity, health status and capacity for self-regulation.

Age-related changes in vascular functioning are a harbinger of cardiovascular disease but the biological mechanisms during the progression of endothelial senescence have not been studied. We investigated alterations in the proteome and miRNA profiles in the course of replicative senescence using primary human umbilical vein endothelial cells as an in vitro vascular model. Quantitative proteomic profiling from early growth stage to senescence was performed by isotope-coded protein label coupled to LC-ESI-MS/MS analysis. Some proteins consistently changed their expression during the senescence whereas others appeared as deregulated only during the late senescence. The latter was accompanied by alterations in morphology of senescent endothelial cells. MicroRNA expression profiling revealed transient changes in the level of miR-16-5p, miR-28-3p and miR-886-5p in the early senescence, decrease in the level of miR-106b-3p at the late stage, and continuous changes in the expression of miR-181a-5p and miR-376a-3p during the whole senescence process. Integrating data on proteomic and microRNA changes indicated potential crosstalk between specific proteins and non-coding RNAs in the regulation of metabolism, cell cycle progression and cytoskeletal organization in the endothelial senescence. The knowledge of molecular targets that change during the senescence can ultimately contribute to a better understanding and prevention of age-related vascular diseases.

Reversible lysine acetylation is a highly regulated post-translational protein modification that is known to regulate several signaling pathways. However, little is known about the radiation-induced changes in the acetylome. In this study, we analyzed the acute post-translational acetylation changes in primary human cardiac microvascular endothelial cells 4 h after a gamma radiation dose of 2 Gy. The acetylated peptides were enriched using anti-acetyl conjugated agarose beads. A total of 54 proteins were found to be altered in their acetylation status, 23 of which were deacetylated and 31 acetylated. Pathway analyses showed three protein categories particularly affected by radiation-induced changes in the acetylation status: the proteins involved in the translation process, the proteins of stress response, and mitochondrial proteins. The activation of the canonical and non-canonical Wnt signaling pathways affecting actin cytoskeleton signaling and cell cycle progression was predicted. The protein expression levels of two nicotinamide adenine dinucleotide (NAD+)-dependent deacetylases, sirtuin 1 and sirtuin 3, were significantly but transiently upregulated 4 but not 24 h after irradiation. The status of the p53 protein, a target of sirtuin 1, was found to be rapidly stabilized by acetylation after radiation exposure. These findings indicate that post-translational modification of proteins by acetylation and deacetylation is essentially affecting the radiation response of the endothelium.

The idea that an energy exchange of some type occurs between individuals is a central theme in many healing techniques. This concept has often been disputed by Western science due to the lack of a plausible mechanism to explain the nature of this energy or how it could affect or facilitate the healing process. The fact that the heart generates the strongest electromagnetic field produced by the body, coupled with the recent discovery that this field becomes more coherent as the individual shifts to a sincerely loving or caring state prompted us to investigate the possibility that the field generated by the heart may significantly contribute to this energy exchange. We present a sampling of results which provide intriguing evidence that an exchange of electromagnetic energy produced by the heart occurs when people touch or are in proximity. Signal averaging techniques are used to show that one’s electrocardiogram (ECG) signal is registered in another person’s electroencephalogram (EEG) and elsewhere on the other person’s body. While this signal is strongest when people are in contact, it is still detectable when subjects are in proximity without contact. This study represents one of the first successful attempts to directly measure an energy exchange between people, and provides a solid, testable theory to explain the observed effects of many healing modalities that are based upon the assumption that an energy exchange takes place. Nonlinear stochastic resonance is discussed as a mechanism by which weak, coherent electromagnetic fields, such as those generated by the heart of an individual in a caring state, may be detected and amplified by biological tissue, and potentially produce measurable effects in living systems. One implication is that the effects of therapeutic techniques involving contact or proximity between practitioner and patient could be amplified by practitioners consciously adopting a sincere caring attitude, and thus introducing increased coherence into their cardiac field.

Results of a study on the effect of the Heart Lock-In meditation technique on the synchronization between heart rate variability and local magnetic field activity is presented in this paper. A technique based on the near-optimal chaotic attractor embedding was applied in order to evaluate the geometrical synchronization between analyzed time series. The results demonstrate that Heart Lock-In technique had a strong influence on the relationship between cardiac and geomagnetic activity.

Because of the increasing application of ionizing radiation in medicine, quantitative data on effects of low-dose radiation are needed to optimize radiation protection, particularly with respect to cataract development. Using mice as mammalian animal model, we applied a single dose of 0, 0.063, 0.125 and 0.5 Gy at 10 weeks of age, determined lens opacities for up to 2 years and compared it with overall survival, cytogenetic alterations and cancer development. The highest dose was significantly associated with increased body weight and reduced survival rate. Chromosomal aberrations in bone marrow cells showed a dose-dependent increase 12 months after irradiation. Pathological screening indicated a dose-dependent risk for several types of tumors. Scheimpflug imaging of the lens revealed a significant dose-dependent effect of 1% of lens opacity. Comparison of different biological end points demonstrated long-term effects of low-dose irradiation for several biological end points. Electronic supplementary material The online version of this article (10.1007/s00411-017-0728-z) contains supplementary material, which is available to authorized users.

Epidemiological data from radiotherapy patients show the damaging effect of ionizing radiation on heart and vasculature. The endothelium is the main target of radiation damage and contributes essentially to the development of cardiac injury. However, the molecular mechanisms behind the radiation-induced endothelial dysfunction are not fully understood. In the present study, 10-week-old C57Bl/6 mice received local X-ray heart doses of 8 or 16 Gy and were sacrificed after 16 weeks; the controls were sham-irradiated. The cardiac microvascular endothelial cells were isolated from the heart tissue using streptavidin-CD31-coated microbeads. The cells were lysed and proteins were labeled with duplex isotope-coded protein label methodology for quantification. All samples were analyzed by LC–ESI–MS/MS and Proteome Discoverer software. The proteomics data were further studied by bioinformatics tools and validated by targeted transcriptomics, immunoblotting, immunohistochemistry, and serum profiling. Radiation-induced endothelial dysfunction was characterized by impaired energy metabolism and perturbation of the insulin/IGF-PI3K-Akt signaling pathway. The data also strongly suggested premature endothelial senescence, increased oxidative stress, decreased NO availability, and enhanced inflammation as main causes of radiation-induced long-term vascular dysfunction. Detailed data on molecular mechanisms of radiation-induced vascular injury as compiled here are essential in developing radiotherapy strategies that minimize cardiovascular complications.

The increased use of radiation-based medical imaging methods such as computer tomography is a matter of concern due to potential radiation-induced adverse effects. Efficient protection against such detrimental effects has not been possible due to inadequate understanding of radiation-induced alterations in signaling pathways. The aim of this study was to elucidate the molecular mechanisms behind learning and memory deficits after acute low and moderate doses of ionizing radiation. Female C57BL/6J mice were irradiated on postnatal day 10 (PND10) with gamma doses of 0.1 or 0.5 Gy. This was followed by evaluation of the cellular proteome, pathway-focused transcriptome, and neurological development/disease-focused miRNAome of hippocampus and cortex 24 h postirradiation. Our analysis showed that signaling pathways related to mitochondrial and synaptic functions were changed by acute irradiation. This may lead to reduced mitochondrial function paralleled by enhanced number of dendritic spines and neurite outgrowth due to elevated long-term potentiation, triggered by increased phosphorylated CREB. This was predominately observed in the cortex at 0.1 and 0.5 Gy and in the hippocampus only at 0.5 Gy. Moreover, a radiation-induced increase in the expression of several neural miRNAs associated with synaptic plasticity was found. The early changes in signaling pathways related to memory formation may be associated with the acute neurocognitive side effects in patients after brain radiotherapy but might also contribute to late radiation-induced cognitive injury.

Summary We did a lifetime study in irradiated and control mice with a follow up time until 24 months post irradiation with repeated in vivo analysis of the lens density and the retina. Mice of different genetic constitution (wild-type, Ercc2+/-) were acutely whole body irradiated with low doses of ionising radiation (0, 63, 125 and 500 mGy) and a low dose rate (63 mGy/min). Lens opacification was analysed monthly by Scheimpflug imaging and the retinal fundus and thickness was analysed by OCT. At different time points (4 and 24 hours, 12, 18 and 24 months post irradiation) mice were sacrificed and eyes were analysed by histology and immunohistochemistry. Mice of all groups, even the DNA repair helicase deficient mice (Ercc2+/−), did not develop clinically relevant radiation-dependent lens opacities. Only an age-dependent increase of lens density was identified. Consequently, no major morphological changes were visible in the eye histology of all mice. Immunohistochemical staining against DNaseIIb revealed no differences between the groups as well as 8-OHG staining resulted in similar ROS levels in all irradiation groups. However, non-irradiated Ercc2+/- mice showed an increased level of oxidative stress compared to wild-type mice.

Radiation is the most common treatment of cancer. Minimizing the normal tissue injury, especially the damage to vascular endothelium, remains a challenge. This study aimed to analyze direct and indirect radiation effects on the endothelium by investigating mechanisms of signal transfer from irradiated to nonirradiated endothelial cells by means of secreted proteins. Human coronary artery endothelial cells (HCECest2) undergo radiation-induced senescence in vitro 14 days after exposure to 10 Gy X-rays. Proteomics analysis was performed on HCECest2 14 days after irradiation with X-ray doses of 0 Gy (control) or 10 Gy using label-free technology. Additionally, the proteomes of control and radiation-induced secretomes, and those of nonirradiated HCECest2 exposed for 24 h to secreted proteins of either condition were measured. Key changes identified by proteomics and bioinformatics were validated by immunoblotting, ELISA, bead-based multiplex assays, and targeted transcriptomics. The irradiated cells, their secretome, and the nonirradiated recipient cells showed similar inflammatory response, characterized by induction of interferon type I-related proteins and activation of the STAT3 pathway. These data indicate that irradiated endothelial cells may adversely affect nonirradiated surrounding cells via senescence-associated secretory phenotype. This study adds to our knowledge of the pathological background of radiation-induced cardiovascular disease.

Epidemiological studies indicate that radiation doses as low as 0.5 Gy increase the risk of cardiovascular disease decades after the exposure. The aim of the present study was to investigate whether this radiation dose causes late molecular alterations in endothelial cells that could support the population-based data.Human coronary artery endothelial cells were irradiated at 0.5 Gy (X-ray) and radiation-induced changes in the proteome were investigated after different time intervals (1, 7 and 14 d) using ICPL technology. Key changes identified by proteomics and bioinformatics were validated by immunoblotting and ELISA.The radiation-induced alteration of the endothelial proteome was characterized by sustained perturbation of Rho GDP-dissociation inhibitor (RhoGDI) and nitric oxide (NO) signalling pathways. At later time-points, this was accompanied by reduced proteasome activity, enhanced protein carbonylation indicating augmented oxidative stress, and senescence.These molecular changes are indicative of long-term premature endothelial dysfunction and provide a mechanistic framework to the epidemiological data showing increased risk of cardiovascular disease at 0.5 Gy.

Radiation is a highly efficient therapy in squamous head and neck carcinoma (HNSCC) treatment. However, local recurrence and metastasis are common complications. Recent evidence shows that cancer-cell-derived exosomes modify tumour cell movement and metastasis. In this study, we link radiation-induced changes of exosomes to their ability to promote migration of recipient HNSCC cells. We demonstrate that exosomes isolated from irradiated donor cells boost the motility of the HNSCC cells BHY and FaDu. Molecular data identified enhanced AKT-signalling, manifested through increased phospho-mTOR, phospho-rpS6 and MMP2/9 protease activity, as underlying mechanism. AKT-inhibition blocked the pro-migratory action, suggesting AKT-signalling as key player in exosome-mediated migration. Proteomic analysis of exosomes isolated from irradiated and non-irradiated BHY donor cells identified 39 up- and 36 downregulated proteins. In line with the observed pro-migratory effect of exosomes isolated from irradiated cells protein function analysis assigned the deregulated exosomal proteins to cell motility and AKT-signalling. Together, our findings demonstrate that exosomes derived from irradiated HNSCC cells confer a migratory phenotype to recipient cancer cells. This is possibly due to radiation-regulated exosomal proteins that increase AKT-signalling. We conclude that exosomes may act as driver of HNSCC progression during radiotherapy and are therefore attractive targets to improve radiation therapy strategies.

PARTICLE (Gene PARTICL- ‘Promoter of MAT2A-Antisense RadiaTion Induced Circulating LncRNA) expression is transiently elevated following low dose irradiation typically encountered in the workplace and from natural sources. This long non-coding RNA recruits epigenetic silencers for cis-acting repression of its neighbouring Methionine adenosyltransferase 2A gene. It now emerges that PARTICLE operates as a trans-acting mediator of DNA and histone lysine methylation. Chromatin immunoprecipitation sequencing (ChIP-seq) and immunological evidence established elevated PARTICLE expression linked to increased histone 3 lysine 27 trimethylation. Live-imaging of dbroccoli-PARTICLE revealing its dynamic association with DNA methyltransferase 1 was confirmed by flow cytometry, immunoprecipitation and direct competitive binding interaction through electrophoretic mobility shift assay. Acting as a regulatory docking platform, the long non-coding RNA PARTICLE serves to interlink epigenetic modification machineries and represents a compelling innovative component necessary for gene silencing on a global scale.

// Valerie Brid O'Leary 1 , Jan Smida 1 , Martina Matjanovski 1 , Corinna Brockhaus 1 , Klaudia Winkler 1 , Simone Moertl 1 , Saak Victor Ovsepian 2, 3 and Michael John Atkinson 1, 4 1 Institute of Radiation Biology, Helmholtz Zentrum Munich - German Research Center for Environmental Health, Neuherberg, Germany 2 Institute of Biological and Medical Imaging, Helmholtz Zentrum Munich - German Research Center for Environmental Health, Neuherberg, Germany 3 Faculty for Electrical Engineering and Information Technology, Technical University Munich, Munich, Germany 4 Chair of Radiation Biology, Technical University Munich, Munich, Germany Correspondence to: Valerie Brid O'Leary, email: olearyv@yahoo.co.uk Keywords: circRNA, WWOX, QUAKING, KIRKOS, exosomes Received: May 07, 2017 Accepted: June 10, 2017 Published: July 13, 2017 ABSTRACT Generated by Quaking (QKI), circular RNAs (circRNAs) are newly recognised non-coding RNA (ncRNA) members characterised by tissue specificity, increased stability and enrichment within exosomes. Studies have shown that ionizing radiation (IR) can influence ncRNA transcription. However, it is unknown whether circRNAs or indeed QKI are regulated by IR. Microarray circRNA profiling and next generation sequencing revealed that circRNA expression was altered by low and medium dose exposure sourced predominantly from genes influencing the p53 pathway. CircRNAs KIRKOS-71 and KIRKOS-73 transcribed from the WWOX ( WW Domain Containing Oxidoreductase ) tumor suppressor (a p53 regulator) responded within hours to IR. KIRKOS-71 and KIRKOS-73 were present in exosomes yet exhibited differential transcript clearance between irradiated cell lines. Dual-quasar labelled probes and in-situ hybridization demonstrated the intercellular distribution of KIRKOS-71 and KIRKOS-73 predominantly within the perinucleus. QKI knockdown removed nuclear expression of these circRNAs with no significant effect on cytosolic KIRKOS-71 and KIRKOS-73. Distinct QKI transcription between cell lines and its augmented interaction with KIRKOS-71 and KIRKOS-73 was noted post IR. This foremost study provides evidence that QKI and circRNAs partake in the cellular irradiation response. KIRKOS-71 and KIRKOS-73 as stable secreted circRNAs may afford vital characteristics worth syphoning as promising diagnostic radiotherapy biomarkers.

The non-coding transcriptome, in particular microRNAs (miRNA), influences cellular survival after irradiation. However, the underlying mechanisms of radiation-induced miRNA expression changes and consequently target expression changes are poorly understood. In this study we show that a single dose of 5Gy ɣ-radiation decreases expression of the miR-23a~27a~24-2 cluster in the human endothelial cell-line EA.hy926 and the mammary epithelial cell-line MCF10A. In the endothelial cells this was facilitated through transcriptional regulation by promoter methylation and also at the post-transcriptional level by reduced miRNA processing through phosphorylation of Argonaute (AGO). Furthermore, we demonstrate that all three mature cluster miRNAs reduce apoptosis by increasing expression of the common target protein XIAP. These findings link a temporal succession of transcriptional and post-transcriptional regulatory mechanisms of the miR~23a~24-2~27a cluster, enabling a dynamic stress response and assuring cellular survival after radiation exposure.

The tumour suppressor gene (Rb1) is necessary for the maintenance of telomere integrity in osteoblastic cells. We now show that the compaction of telomeric chromatin and the appropriate histone modifications of telomeric DNA are both dependent upon Rb1-mediated transcription of the telomere-derived long non-coding RNA TERRA. Expression of TERRA was reduced in Rb1 haploinsufficient cells, and further decreased by shRNA-mediated reduction of residual Rb1 expression. Restoration of Rb1 levels through lentiviral transduction was sufficient to reestablish both transcription of TERRA and condensation of telomeric chromatin. The human chromosome 15q TERRA promoter contains predicted retinoblastoma control elements, and was able to confer Rb1-dependent transcription upon a promoterless reporter gene. Chromatin immunoprecipitation revealed preferential binding of phosphorylated over non-phosphorylated Rb1 at the TERRA promoter. As Rb1-deficient cells show increased genomic instability we suggest that this novel non-canonical action of Rb1 may contribute to the tumour suppressive actions of Rb1.

International audience; In 2009, the European High Level and Expert Group identified key policy and scientific questions to be addressed through a strategic research agenda for low-dose radiation risk. This initiated the establishment of a European Research Platform, called MELODI (Multidisciplinary European Low Dose Research Initiative). In 2010, the DoReMi Network of Excellence was launched in the Euratom 7th Framework Programme. DoReMi has acted as an operational tool for the sustained development of the MELODI platform during its early years. A long-term Strategic Research Agenda for European low-dose radiation risk research has been developed by MELODI. Strategic planning of DoReMi research activities is carried out in close collaboration with MELODI. The research priorities for DoReMi are designed to focus on objectives that are achievable within the 6-y lifetime of the project and that are in areas where stimulus and support can help progress towards the longer-term strategic objectives. © The Author 2014.

Lentiviral vectors (LV) are widely used to successfully transduce cells for research and clinical applications. Lentiviral vectors pseudotyped with the vesicular stomatitis virus glycoprotein (VSV-G) can be produced to high titers and mediate high transduction efficiencies in vitro. For clinical applications the need for optimized transduction protocols and the limited activity of retronectin as LV enhancer, results in the application of a high multiplicity of infection (MOI) to achieve effective transduction efficiencies for a number of therapeutically relevant cells, e.g. CD34(+) hematopoietic stem cells, T- and B-cells. Our study describes an optimized LV infection protocol including a non-toxic poloxamer-based adjuvant combined with antibody-retargeted lentiviral particles, improving transduction efficiency at low MOI. Cell specificity of lentiviral vectors was increased by displaying different ratios of scFv-fused VSV-G glycoproteins on the viral envelope. The system was validated with difficult to transduce human CD30(+) lymphoma cells, and EGFR(+) tumor cells. Highly efficient transduction of lymphoma cells was achieved, >50% of cells were transduced when MOI 1 was used. The scFv displaying lentiviral particles gained relative specificity for transduction of target cells. Preferential gene delivery to CD30(+) or EGFR(+) cells was increased 4-fold in mixed cell cultures by presenting scFv antibody fragments binding to respective surface markers. A combination of spinoculation, poloxamer-based chemical adjuvant, and LV displaying scFv fragments increases transduction efficiencies of hard-to-transduce suspension lymphoma cells, and promises new chances for the future development of improved clinical protocols.

Chronic exposure to low-dose ionizing radiation is associated with an increased risk of cardiovascular disease. Alteration in energy metabolism has been suggested to contribute to radiation-induced heart pathology, mitochondrial dysfunction being a hallmark of this disease. The goal of this study was to investigate the regulatory role of acetylation in heart mitochondria in the long-term response to chronic radiation. ApoE-deficient C57Bl/6J mice were exposed to low-dose-rate (20 mGy/day) gamma radiation for 300 days, resulting in a cumulative total body dose of 6.0 Gy. Heart mitochondria were isolated and analyzed using quantitative proteomics. Radiation-induced proteome and acetylome alterations were further validated using immunoblotting, enzyme activity assays, and ELISA. In total, 71 proteins showed peptides with a changed acetylation status following irradiation. The great majority (94%) of the hyperacetylated proteins were involved in the TCA cycle, fatty acid oxidation, oxidative stress response and sirtuin pathway. The elevated acetylation patterns coincided with reduced activity of mitochondrial sirtuins, increased the level of Acetyl-CoA, and were accompanied by inactivation of major cardiac metabolic regulators PGC-1 alpha and PPAR alpha. These observations suggest that the changes in mitochondrial acetylation after irradiation is associated with impairment of heart metabolism. We propose a novel mechanism involved in the development of late cardiac damage following chronic irradiation.

Background: MiR-221/-222 are frequently overexpressed in breast cancer and are associated with increased malignancy. The specific modification of microRNAs (miRNAs) expression could be a promising strategy in breast cancer therapy, leading to the suppression of tumourigenic processes in tumour cells. Methods: MiR-221/-222 expressions were analysed in 86 breast cancer tissues by quantitative RT–PCR and tested for correlation with immunohistochemistry data and clinical follow-up. In vitro assays were conducted using human breast cancer cell lines with lentiviral overexpression of miR-221/-222. Results: In tumour tissues, miR-221/-222 were associated with the occurrence of distant metastases. In particular, high levels of miR-221 were revealed to have a high prognostic impact for the identification of significantly different groups with advanced tumours. MiR-221/-222 overexpression strongly increased cell proliferation and invasion in vitro. Following miR-221/-222 overexpression an increased uPAR expression and cell invasion were observed. Conclusion: This study demonstrates a significant role for highly expressed miR-221/-222 in advanced breast cancers allowing for the identification of significantly different prognostic groups, particularly for HER2-positive and lymph-node-positive breast cancers. Considering that miR-221/-222 are strongly involved in cell invasion, these miRNAs may be promising markers for breast cancer prognosis and therapy.

Rapidly developing postgenome research has made proteins an attractive target for biological analysis. The well-established term of proteome is defined as the complete set of proteins expressed in a given cell, tissue or organism. Unlike the genome, a proteome is rapidly changing as it tends to adapt to microenvironmental signals. The systematic analysis of the proteome at a given time and state is referred to as proteomics. This technique provides information on the molecular and cellular mechanisms that regulate physiology and pathophysiology of the cell. Applications of proteome profiling in radiation research are increasing. However, the large-scale proteomics data sets generated need to be integrated into other fields of radiation biology to facilitate the interpretation of radiation-induced cellular and tissue effects. The aim of this review is to introduce the most recent developments in the field of radiation proteomics.

Germline mutations of the retinoblastoma gene (RB1) predispose to both sporadic and radiation-induced osteosarcoma, tumors characterized by high levels of genomic instability, and activation of alternative lengthening of telomeres. Mice with haploinsufficiency of the Rb1 gene in the osteoblastic lineage reiterate the radiation susceptibility to osteosarcoma seen in patients with germline RB1 mutations. We show that the susceptibility is accompanied by an increase in genomic instability, resulting from Rb1-dependent telomere erosion. Radiation exposure did not accelerate the rate of telomere loss but amplified the genomic instability resulting from the dysfunctional telomeres. These findings suggest that telomere maintenance is a noncanonical caretaker function of the retinoblastoma protein, such that its deficiency in cancer may potentiate DNA damage-induced carcinogenesis by promoting formation of chromosomal aberrations, rather than simply by affecting cell-cycle control. Cancer Res; 73(14); 4247–55. ©2013 AACR.

The fourth workshop of the Multidisciplinary European Low Dose Initiative (MELODI) was organised by STUK—Radiation and Nuclear Safety Authority of Finland. It took place from 12 to 14 September 2012 in Helsinki, Finland. The meeting was attended by 179 scientists and professionals engaged in radiation research and radiation protection. We summarise the major scientific findings of the workshop and the recommendations for updating the MELODI Strategic Research Agenda and Road Map for future low dose research activities.

PURPOSE: Ionizing radiation induces cardiovascular disease, the endothelium being the main target. The exact mechanism of the damage is unclear but the involvement of multiple signaling pathways is probable. Reversible lysine acetylation is a posttranslational protein modification that regulates activity across a broad range of signaling pathways. The goal of this study was to determine if a low radiation dose results in acetylome alteration in endothelial cells. MATERIALS AND METHODS: Human coronary artery endothelial cell line was irradiated with Cs-137 gamma-rays (0.5 Gy) and proteomics analysis was performed using enriched acetylated peptides and all peptides. Data were validated using immunoblotting, deacetylase activity assay, and RhoA activity assay. RESULTS: Nearly a hundred proteins were found to have an altered acetylation status 24 h after irradiation, primarily due to an overall decrease in acetylation. The expression of specific deacetylases was significantly increased, coinciding with an enhancement in global deacetylase activity. Proteins changed in their acetylation status belonged to several pathways including protein synthesis, cytoskeleton-related processes, protein folding and calcium signaling. The predicted changes in the RhoA/actin cytoskeleton pathway were validated by immunoassay. CONCLUSIONS: This study shows that protein acetylation is an important mediator of radiation responses in human cardiac coronary endothelial cells. Increased knowledge of the endothelial response to radiation is crucial for the development of normal tissue sparing modalities during radiation therapy.

PURPOSE: Multiple cell types secrete exosome-like extracellular vesicles (ELVs) to the extracellular environment. Pathological conditions can produce characteristic changes to the vesicle cargo. We investigated if ionizing radiation is capable of inducing changes in the protein and microRNA (miRNA) cargo of ELVs. MATERIALS AND METHODS: Whole blood samples from healthy donors were irradiated with 2 Gy gamma rays and then peripheral blood mononuclear cells and plasma were separated from residual blood and co-cultivated for 24 h. The released ELVs were collected by differential ultracentrifugation from irradiated and non-irradiated samples. microRNAs and proteins were quantified by qPCR and label-free proteomics. RESULTS: Here we report a first characterization of radiation-induced changes in the protein and miRNA cargo of ELVs isolated from plasma. Proteome analysis of ELVs identified 214 proteins, of which nine significantly changed their abundance after irradiation. The radiation-induced down-regulation of afamin and serpine peptidase F1 was confirmed by immunoblotting. miRNA expression profiling identified 58 different exosomal miRNAs, the expression of miR-204-5p, miR-92a-3p and miR-31-5p was significantly increased in ELVs from irradiated samples. CONCLUSIONS: This study provides evidence that radiation-induced changes occur in the protein and miRNA cargo of plasma ELVs. These data imply a novel systemic communication pathway between irradiated and non-irradiated cells and tissues.

Asthma is a disease affecting millions of people with high economic burden, because of many variations in clinical presentation and response to treatment. MiRNAs modulate gene expression in response to various stimuli and their role in all aspects of asthma remains to be fully elucidated. We investigated the changes in miRNA expression in human bronchial epithelial cell line (BEAS-2B) in response to montelukast (MNT), widely used as asthma therapy. As bronchial epithelial cells contribute to innate immunity, we have also chosen to analyze changes in miRNA expression in response to lipopolysaccharide (LPS) stimulation with or without pre-treatment with MNT. Total RNA was extracted and analyzed by Taqman Low Density miRNA array. The obtained results were normalized to RNU44 and statistically evaluated. We present the results on expression of mir-183 cluster (miR-183, -96 and -182). The members of this cluster have been implicated in processes important for lung function, such as response to high CO 2 concentrations and development of lung cancer. The expression of miR-183 was significantly elevated after treatment of BEAS-2B cells with LPS (2 -ddCt =5.8), but its expression decreased when the cells were pre-treated with MNT (2 -ddCt =3.1). When the cells were treated with MNT alone there was a significant down-regulation (2 -ddCt = 0.64) in miR-183 expression compared to control. MiR-182 expression was increased in comparison to control in MNT treated (2 -ddCt =3.9) and pre-treated (2 -ddCt =1.7) cells, but was down-regulated after stimulation with LPS (2 -ddCt =0.65). MiR-96 was not detected. The role of miRNAs needs further investigation in both asthma pathology and response to therapy and miR-183 cluster could have some clinical significance.

Prenatal exposure to stress such as increased level of reactive oxygen species or antiviral therapy are known factors leading to adult heart defects. The risks following a radiation exposure during fetal period are unknown, as are the mechanisms of any potential cardiac damage. The aim of this study was to gather evidence for possible damage by investigating long-term changes in the mouse heart proteome after prenatal exposure to low and moderate radiation doses. Pregnant C57Bl/6J mice received on embryonic day 11 (E11) a single total body dose of ionizing radiation that ranged from 0.02 Gy to 1.0 Gy. The offspring were sacrificed at the age of 6 months or 2 years. Quantitative proteomic analysis of heart tissue was performed using Isotope Coded Protein Label technology and tandem mass spectrometry. The proteomics data were analyzed by bioinformatics and key changes were validated by immunoblotting. Persistent changes were observed in the expression of proteins representing mitochondrial respiratory complexes, redox and heat shock response, and the cytoskeleton, even at the low dose of 0.1 Gy. The level of total and active form of the kinase MAP4K4 that is essential for the embryonic development of mouse heart was persistently decreased at the radiation dose of 1.0 Gy. This study provides the first insight into the molecular mechanisms of cardiac impairment induced by ionizing radiation exposure during the prenatal period.

High doses of ionising radiation significantly increase the risk of cardiovascular disease (CVD), the vascular endothelium representing one of the main targets. Whether radiation doses lower than 500 mGy induce cardiovascular damage is controversial. The aim of this study was to investigate radiation-induced expression changes on protein and microRNA (miRNA) level in primary human coronary artery endothelial cells after a single 200 mGy radiation dose (Co-60). Using a multiplex gel-based proteomics technology (2D-DIGE), we identified 28 deregulated proteins showing more than ±1.5-fold expression change in comparison with non-exposed cells. A great majority of the proteins showed up-regulation. Bioinformatics analysis indicated "cellular assembly and organisation, cellular function and maintenance and molecular transport" as the most significant radiation-responsive network. Caspase-3, a central regulator of this network, was confirmed to be up-regulated using immunoblotting. We also analysed radiation-induced alterations in the level of six miRNAs known to play a role either in CVD or in radiation response. The expression of miR-21 and miR-146b showed significant radiation-induced deregulation. Using miRNA target prediction, three proteins found differentially expressed in this study were identified as putative candidates for miR-21 regulation. A negative correlation was observed between miR-21 levels and the predicted target proteins, desmoglein 1, phosphoglucomutase and target of Myb protein. This study shows for the first time that a low-dose exposure has a significant impact on miRNA expression that is directly related to protein expression alterations. The data presented here may facilitate the discovery of low-dose biomarkers of radiation-induced cardiovascular damage.

Research suggests that police work is among the most stressful occupations in the world and officers typically suffer a variety of physiological, psychological, and behavioral effects and symptoms. Officers operating under severe or chronic stress are likely to be at greater risk of error, accidents, and overreactions that can compromise their performance, jeopardize public safety, and pose significant liability costs to the organization. Therefore, this study explored the nature and degree of physiological activation typically experienced of officers on the job and the impact of the Coherence Advantage resilience and performance enhancement training on a group of police officers from Santa Clara County, California. Areas assessed included vitality, emotional well-being, stress coping and interpersonal skills, work performance, workplace effectiveness and climate, family relationships, and physiological recalibration following acute stressors. Physiological measurements were obtained to determine the real-time cardiovascular impact of acutely stressful situations encountered in highly realistic simulated police calls used in police training and to identify officers at increased risk of future health challenges. The resilience-building training improved officers' capacity to recognize and self-regulate their responses to stressors in both work and personal contexts. Officers experienced reductions in stress, negative emotions, depression, and increased peacefulness and vitality as compared to a control group. Improvements in family relationships, more effective communication and cooperation within work teams, and enhanced work performance also were noted. Heart rate and blood pressure measurements taken during simulated police call scenarios showed that acutely stressful circumstances typically encountered on the job result in a tremendous degree of physiological activation, from which it takes a considerable amount of time to recover. Autonomic nervous system assessment based on heart rate variability (HRV) analysis of 24-hour electrocardiogram (ECG) recordings revealed that 11% of the officers were at higher risk for sudden cardiac death and other serious health challenges. This is more than twice the percentage typically found in the general population and is consistent with epidemiological data indicating that police officers have more than twice the average incidence of cardiovascular-related disease. The data suggest that training in resilience building and self-regulation skills could significantly benefit police organizations by improving judgment and decision making and decreasing the frequency of onthe-job driving accidents and the use of excessive force in high-stress situations. Potential outcomes include fewer citizens' complaints, fewer lawsuits, decreased organizational liabilities, and increased community safety. Finally, this study highlights the value of 24-hour HRV analysis as a useful screening tool to identify officers who are at increased risk, so that efforts can be made to reverse or prevent the onset of disease in these individuals.研究显示，警察是全球压力最大 的职业之一，警员通常要承受各 种生理、心理和行为影响，并出 现相关症状。如警员在严重或慢 性压力下工作，则其犯错、出事 故和反应过度的风险可能升高， 这会影响他们的绩效表现、危害 公众安全并给组织带来显著的责 任成本。因此，本研究探究警员 在工作中通常会体验到的生理活 性的性质和程度，以及协调优势 复原和绩效增强培训对加利福尼 亚州圣克拉拉郡一组警员的影响。评估范围包括活力、情感健康、 压力应对和人际关系技巧、工作 表现、工作场所效益和氛围、家 庭关系和严重压力因素后的生理 调整等各方面。获取生理测定， 以确定在警察训练采用的高仿真 模拟警讯中，遭遇严重压力情况 对心血管的实时影响，及确定在 未来会有更高健康风险的警员。构建复原能力的训练可提高警员 的认识和自我调整能力，对来自 工作和个人环境中的压力因素做 出反应。相比对照组，警员的压 力、负面情绪、抑郁有所减少， 而平静和活力则有所增加。另外 还发现家庭关系改善、工作团队 内部沟通和协作的效率提高，以 及工作绩效改善。模拟警讯期间获得的心率和血压 测量结果显示，通常工作中遇到 的严重压力环境会导致很大程度 的生理活性，这需要很长时间才 能恢复。基于 24 小时心电图 (ECG) 记录 心率变异性分析的自主神经系统 评估显示，11% 的警员出现心脏 性猝死和其他严重健康问题的风 险较高。这是普通人群中典型百 分比的两倍多，与显示警员心血 管相关疾病发病率超过平均水平 两倍的流行病学数据相符。数据显示，构建复原能力和自我 调整技能的训练非常有助于改善 判断和决策、降低执勤所致事故 的频率及减少在高压力状况下使 用过多人力，从而给警察组织带 来好处。潜在结果包括市民投诉 减少、诉讼减少、组织责任减少 及社会安全提高。最后，本研究 突出 24 小时心率变异性 (HRV) 分析的价值，其作为实用的筛选 工具，确定出哪些警员面临较高 风险，从而可据此采取措施逆转 或预防这些人士的疾病发作。.La investigación sugiere que el trabajo de policía se cuenta entre las ocupaciones más estresantes del mundo y que los oficiales de policía suelen sufrir diversos efectos y síntomas fisiológicos, psicológicos y conductuales. Los oficiales de policía que trabajan en condiciones de estrés grave o crónico tienen más probabilidades de cometer errores, sufrir accidentes y sobrerreaccionar, lo que puede poner en peligro su rendimiento y la seguridad pública, además de suponer costes por responsabilidad civil importantes para la organización. Por consiguiente, este estudio exploró la naturaleza y el grado de la activación fisiológica que suelen experimentar los oficiales de policía en el trabajo y el impacto de la formación Coherence Advantage para la mejora del rendimiento y de la resistencia sobre un grupo de oficiales de policía del condado de Santa Clara (California, Estados Unidos).Las áreas evaluadas incluyeron la vitalidad, el bienestar emocional, la capacidad de resistir el estrés y las destrezas interpersonales, el rendimiento laboral, la efectividad y el clima en el lugar de trabajo, las relaciones familiares y la recalibración fisiológica después de las situaciones de estrés agudo. Se obtuvieron medidas fisiológicas para determinar el impacto cardiovascular en tiempo real de las situaciones causantes de estrés agudo en casos de llamadas simuladas de gran realismo a la policía que se utilizan en la formación policial y para identificar a los oficiales de policía que presentaban un aumento del riesgo de futuros retos de salud.La formación en el aumento de la resistencia mejoró la capacidad de los oficiales de policía para reconocer y autorregular sus respuestas a las situaciones de estrés, tanto en el contexto laboral como personal. En comparación con el grupo control, los oficiales de policía obtuvieron reducciones del estrés, de las emociones negativas y de la depresión, además de un aumento de la tranquilidad y la vitalidad. También se observaron mejoras en las relaciones familiares, más efectividad en la comunicación y cooper-ación en los equipos de trabajo y mejora del rendimiento laboral.Las medidas de la frecuencia cardíaca y la presión arterial tomadas durante las simulaciones de las llamadas a la policía mostraron que las circunstancias de estrés agudo que suelen encontrarse en el trabajo resultan en un grado tremendo de activación fisiológica, cuya recuperación tarda un tiempo considerable.La evaluación del sistema nervioso autónomo basada en el análisis de la variabilidad de la frecuencia cardíaca en los registros de electro-cardiogramas (ECG) durante 24 horas reveló que el 11 % de los oficiales de policía presentaba una elevación del riesgo de muerte cardíaca súbita y otros retos graves para la salud. Esto es más del doble del porcentaje que suele hallarse en la población en general y coincide con los datos epidemiológicos que indican que los oficiales de policía tienen más del doble de la incidencia media de la enfermedad relacionada con causas cardiovasculares.Los datos indican que la formación en el aumento de la resistencia y en las destrezas de autorregulación podría beneficiar de forma significativa a las organizaciones policiales mediante la mejora del criterio y de la toma de decisiones, la reducción de la frecuencia de los accidentes de tráfico en el trabajo y del uso excesivo de la fuerza en situaciones de gran estrés. Los posibles resultados incluyen la reducción de las quejas de los ciudadanos, la reducción de los pleitos, la reducción de la responsabilidad civil de la organización y el aumento de la seguridad de la comunidad. Por último, este estudio resalta el valor del análisis de la variabilidad de la frecuencia cardíaca en 24 horas como una herramienta útil de cribado para identificar a los oficiales de policía que presentan un aumento del riesgo, de modo que puedan realizarse esfuerzos para invertir o prevenir el inicio de la enfermedad en estas personas.

There is no clear evidence proving or disproving that ionising radiation is causally linked with neurodegenerative diseases such as Parkinson’s and Alzheimer’s. However, it is known that high doses of ionising radiation to the head (20–50 Gy) lead to severe learning and memory impairment which is characteristical for Alzheimer’s. The cumulative doses of ionising radiation to the Western population are accruing, mostly due to the explosive growth of medical imaging procedures. Children are in particular prone to ionising radiation as the molecular processes within the brain are not completely finished. Furthermore, they have a long lifespan under risk. We wish to open a debate if such low doses of radiation exposure may lead to delayed long-term cognitive and other defects, albeit at a lower frequency than those observed during application of high doses. Further, we want to sensitise the society towards the risks of ionising radiation. To achieve these aims, we will recapitulate the known symptoms of Parkinson’s and Alzheimer’s on the molecular level and incorporate data of mainly low- and moderate-ionising radiation (

Secondary bone tumours arising in the field of a preceding radiotherapy are a serious late effect, in particular considering the increasing survival times in patients treated for paediatric malignancies. In general, therapy associated tumours are known to show a more aggressive behaviour and a limited response to chemotherapy compared with their primary counterparts. It is not clear however whether this less favourable outcome is caused by inherent genetic factors of the tumour cells or by a general systemic condition of the patient. To elucidate this we analysed a series of bone sarcomas with a history of prior irradiation for the presence of genomic alterations and compared them with the alterations identified earlier in primary osteosarcomas. We analysed seven radiation induced bone sarcomas for genome-wide losses of heterozygosity (LOH) using Affymetrix 10K2 high-density single nucleotide polymorphism (SNP) arrays. Additionally, copy number changes were analysed at two distinct loci on 10q that were recently found to be of major prognostic significance in primary osteosarcomas. All the investigated tumours showed a LOH at 10q21.1 with 86% of cases (6/7) revealing a total genome-wide LOH score above 2400 and more than 24% of the genome being affected. Our results indicate similar genetic alterations in radiation induced sarcomas of bone and primary osteosarcomas with a poor prognosis. We speculate that the high degree of genomic instability found in these tumours causes the poor prognosis irrespective of the initiating event.

Background Although lentiviral transduction methods are widely used, their broader application is dependent upon the optimization of lentiviral transduction efficiency for a broad range of cell types. In the present study, we focus on the evaluation of two chemical classes with respect to their ability to increase lentiviral transduction without cytotoxicity. Methods We compared the activity of adjuvants that are already used for lentivirus delivery with that of novel adjuvants selected on the basis of their chemical and physical characteristics. Results The novel poloxamer synperonic F108 demonstrated superior characteristics for enhancing lentiviral transduction over the best-in-class polybreneassisted transduction. The results revealed that poloxamer synperonic F108 exhibited the dual benefits of low toxicity and a high efficiency of lentiviral gene delivery into a range of different primary cell cultures. In the presence of poloxamer synperonic F108, cells showed an increased propidium dye influx indicating a re-organization of membrane microstructures accompanying lentivirus uptake. The administration of a mixture of poloxamer synperonic F108 with polybrene further enhanced lentiviral transduction rates. Conclusions The results obtained in the present study indicate that a contribution to efficiency is made by each adjuvant, with polybrene acting as a charge protector and poloxamer synperonic F108 as a membrane modulator. Therefore, poloxamer synperonic F108, either alone or in combination, can lead to the optimization of large-scale lentiviral transduction approaches. Copyright © 2012 John Wiley & Sons, Ltd.

Osteosarcomas are genetically complex tumors with abundant structural and numerical alterations. The molecular pathogenesis of the disease is, however, still poorly understood. Aside from various oncogenes and tumor suppressor genes, deregulated microRNAs (miRNAs) are known to influence tumor development and biology. We therefore investigated six well-established osteosarcoma cell lines (HOS58, U2-OS, Saos-2, MNNG/HOS, SJSA-1, and MG-63) for genome-wide miRNA expression (miRBase Version 15.0, http://www.mirbase.org/) and correlated our findings with gene expression. Cultured osteoblasts (hFOB 1.19) and mesenchymal stem cells (L87/4) were used as normal references. Focusing only on miRNAs that were deregulated in the majority of osteosarcoma cell lines, we identified several miRNAs with oncogenic and tumor suppressor properties, including various members of the oncogenic miR-17-92 cluster. In addition, several genes involved in differentiation (RGMB, LRRC17), cell cycle control (CCNE1), and apoptosis (LIMA1, CAMK2N1) were found to be deregulated in osteosarcoma cell lines, most likely due to altered miRNA expression patterns. Our findings indicate a crucial impact of deregulated miRNAs with consecutive changes in gene expression in osteosarcomas, which strongly suggests pathogenetic and potentially therapeutic implications.

Genetic factors can modify susceptibility to the carcinogenic effect of ionising radiation. To establish if radioiodine-induced thyroid cancer is similarly genetically influenced, we studied F1 hybrid crosses between inbred mouse strains. Mice were perinatally exposed to iodine-131 and thyroid tissues examined after 18 months. Differences in the incidence and distribution of histological subtypes were quantified in relation to genetic background. As expected, the occurrence of thyroid lesions was significantly higher in irradiated mouse hybrids than in unirradiated controls. The most frequent alterations were the simple and the complex hyperplasias, followed by follicular adenoma and, less frequently, follicular carcinoma. Both the incidence and distribution of the histiotype were different between the hybrid mouse crosses. Crosses using JF1 mice (M. m. molossinus) produced F1 offspring that were more resistant to radiation-induced thyroid lesions. Sequence analysis of Braf, Ret, Hras, Kras, Kit and Trp53, all genes that are commonly mutated in human thyroid cancers, did not show any evidence of mutation in the tumours. However, microsatellite analysis of genomic DNA revealed frequent allelic imbalances in complex hyperplasia and follicular adenoma. We conclude that genetic background, in particular the JF1 genotype, confer differences in susceptibility to the carcinogenic effects of radioiodine on the thyroid.

High doses of ionising radiation damage the heart by an as yet unknown mechanism. A concern for radiological protection is the recent epidemiological data indicating that doses as low as 100–500 mGy may induce cardiac damage. The aim of this study was to identify potential molecular targets and/or mechanisms involved in the pathogenesis of low-dose radiation-induced cardiovascular disease. The vascular endothelium plays a pivotal role in the regulation of cardiac function and is therefore a potential target tissue. We report here that low-dose radiation induced rapid and time-dependent changes in the cytoplasmic proteome of the human endothelial cell line EA.hy926. The proteomes were investigated at 4 and 24 h after irradiation at two different dose rates (Co-60 gamma ray total dose 200 mGy; 20 mGy/min and 190 mGy/min) using 2D-DIGE technology. Differentially expressed proteins were identified, after in-gel trypsin digestion, by MALDI-TOF/TOF tandem mass spectrometry, and peptide mass fingerprint analyses. We identified 15 significantly differentially expressed proteins, of which 10 were up-regulated and 5 down-regulated, with more than ± 1.5-fold difference compared with unexposed cells. Pathways influenced by the low-dose exposures included the Ran and RhoA pathways, fatty acid metabolism and stress response.