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Recent epidemiologic studies support an association between chronic low-dose radiation exposure and the development of cardiovascular disease (CVD). The molecular mechanisms underlying the adverse effect of chronic low dose exposure are not fully understood. To address this issue, we have investigated changes in the heart proteome of ApoE deficient (ApoE−/−) C57Bl/6 female mice chronically irradiated for 300 days at a very low dose rate (1 mGy/day) or at a low dose rate (20 mGy/day), resulting in cumulative whole-body doses of 0.3 Gy or 6.0 Gy, respectively. The heart proteomes were compared to those of age-matched sham-irradiated ApoE−/− mice using label-free quantitative proteomics. Radiation-induced proteome changes were further validated using immunoblotting, enzyme activity assays, immunohistochemistry or targeted transcriptomics. The analyses showed persistent alterations in the cardiac proteome at both dose rates; however, the effect was more pronounced following higher dose rates. The altered proteins were involved in cardiac energy metabolism, ECM remodelling, oxidative stress, and ageing signalling pathways. The changes in PPARα, SIRT, AMPK, and mTOR signalling pathways were found at both dose rates and in a dose-dependent manner, whereas more changes in glycolysis and ECM remodelling were detected at the lower dose rate. These data provide strong evidence for the possible risk of cardiac injury following chronic low dose irradiation and show that several affected pathways following chronic irradiation overlap with those of ageing-associated heart pathology.

PDF file - 2012K, Cre recombinase mediated deletion of the floxed Rb1 allele (S1); Characterisation of Rb1 expression in the cell lines (S2); Cre transgene expression in bone and explant cultures of primary osteoblasts (S3); Rb1 mRNA levels after transduction with shRB1-expressing lentivirus (S4); In vitro Rb1 excision using lentiviral Cre (S5); Assessment of telomere length in LV-Cre infected cells using genomic PCR (S6); Transduction with the LVRB1 leads to recovery of retinoblastoma protein expression (S7); MDM2 copy number measurement (S8); p53 expression in primary osteoblasts (S9); Formation of acentric fragments (micronuclei) after radiation exposure (S10).

PDF file - 103K, Includes list of primers used in this study. Also includes statistical analysis and gene expression assay.

PDF file - 131K

Reliable data on the effects of chronic prenatal exposure to low dose (LD) of ionizing radiation in humans are missing. There are concerns about adverse long-term effects that may persist throughout postnatal life of the offspring. Due to their slow cell cycle kinetics and life-long residence time in the organism, mesenchymal stem cells (MSCs) are more susceptible to low level genotoxic stress caused by extrinsic multiple LD events. The aim of this study was to investigate the effect of chronic, prenatal LD gamma irradiation to the biology of MSCs later in life. C3H mice were exposed in utero to chronic prenatal irradiation of 10 mGy/day over a period of 3 weeks. Two years later, MSCs were isolated from the bone marrow and analyzed in vitro for their radiosensitivity, for cellular senescence and for DNA double-strand break recognition after a second acute gamma-irradiation. In addition to these cellular assays, changes in protein expression were measured using HPLC–MS/MS and dysregulated molecular signaling pathways identified using bioinformatics. We observed radiation-induced proteomic changes in MSCs from the offspring of in utero irradiated mice (leading to ~ 9.4% of all detected proteins being either up- or downregulated) as compared to non-irradiated controls. The proteomic changes map to regulation pathways involved in the extracellular matrix, the response to oxidative stress, and the Wnt signaling pathway. In addition, chronic prenatal LD irradiation lead to an increased rate of in vitro radiation-induced senescence later in life and to an increased number of residual DNA double-strand breaks after 4 Gy irradiation, indicating a remarkable interaction of in vivo radiation in combination with a second acute dose of in vitro radiation. This study provides the first insight into a molecular mechanism of persistent MSC damage response by ionizing radiation exposure during prenatal time and will help to predict therapeutic safety and efficacy with respect to a clinical application of stem cells. Supplementary Information The online version contains supplementary material available at 10.1007/s00411-021-00925-7.

Heart rate variability (HRV), the change in the time intervals between adjacent heartbeats, is an emergent property of interdependent regulatory systems that operates on different time scales to adapt to environmental and psychological challenges. Low age-adjusted HRV has been confirmed as a strong, independent predictor of future health problems in both healthy people and patients with a wide range of diseases that correlate with all-cause mortality. Twenty-four–hour HRV recordings are considered the gold standard and have greater predictive power on health risk than short-term recordings. However, it is not always practical or cost effective to obtain 24-hour HRV recordings, and short-term recordings have been widely used in research and clinical applications for many years. This article will report on the first in a series of research investigations on short-term HRV assessments. The first study examined the correlations between a 10-minute resting state, a 1-minute paced deep breathing protocol, response to handgrip, and 24-hour HRV measures in 28 healthy individuals. Based on the results of the initial study, the primary study examined the correlations between the 1-minute paced deep breathing assessment and 24-hour measures in a general population of 805 individuals. Overall, the findings from the studies suggested that the 1-minute paced deep breathing assessments were highly correlated with 24-hour measures of vagally mediated HRV and very-low-frequency power. The findings from this study suggest that the 1-minute paced deep breathing protocol is an ideal short-term assessment that can be used in a health risk screening context. When low values are observed, it is recommended that a 24-hour assessment be conducted.

As high-throughput approaches in biological and biomedical research are transforming the life sciences into information-driven disciplines, modern analytics platforms for big data have started to address the needs for efficient and systematic data analysis and interpretation. We observe that radiobiology is following this general trend, with -omics information providing unparalleled depth into the biomolecular mechanisms of radiation response - defined as systems radiobiology. We outline the design of computational frameworks and discuss the analysis of big data in low-dose ionizing radiation (LDIR) responses of the mammalian brain. Following successful examples and best practices of approaches for the analysis of big data in life sciences and health care, we present the needs and requirements for radiation research. Our goal is to raise awareness for the radiobiology community about the new technological possibilities that can capture complex information and execute data analytics on a large scale. The production of large data sets from genome-wide experiments (quantity) and the complexity of radiation research with multidimensional experimental designs (quality) will necessitate the adoption of latest information technologies. The main objective was to translate research results into applied clinical and epidemiological practice and understand the responses of biological tissues to LDIR to define new radiation protection policies. We envisage a future where multidisciplinary teams include data scientists, artificial intelligence experts, DevOps engineers, and of course radiation experts to fulfill the augmented needs of the radiobiology community, accelerate research, and devise new strategies.

A suite of software tools has been developed for dose estimation (BAT, WinFRAT) and prediction of acute health effects (WinFRAT, H-Module) using clinical symptoms and/or changes in blood cell counts. We constructed a database of 191 ARS cases using the METREPOL (n = 167) and the SEARCH-database (n = 24). The cases ranged from unexposed (RC0), to mild (RC1), moderate (RC2), severe (RC3), and lethal ARS (RC4). From 2015-2019, radiobiology students and participants of two NATO meetings predicted clinical outcomes (RC, H-ARS, and hospitalization) based on clinical symptoms. We evaluated the prediction outcomes using the same input datasets with a total of 32 teams and 94 participants. We found that: (1) unexposed (RC0) and mildly exposed individuals (RC1) could not be discriminated; (2) the severity of RC2 and RC3 were systematically overestimated, but almost all lethal cases (RC4) were correctly predicted; (3) introducing a prior education component for non-physicians significantly increased the correct predictions of RC, ARS, and hospitalization by around 10% (p

The MSc Radiation Biology course is a highly interdisciplinary degree program placing radiation biology at the interface between biology, medicine, and physics, as well as their associated technologies. The goal was to establish an internationally acknowledged program with diverse and heterogeneous student cohorts, who benefit from each other academically as well as culturally. We have completed a Five-Year evaluation of the program to assess our qualification profile and the further direction we want to take.We evaluated the student cohort's data from the last 5 years regarding gender, age, and nationality as well as the highest degree before applying and career path after graduation.Data shows a great diversity regarding nationalty as well as undergraduate background. Cohort sizes could be increased and future prospects mainly aimed to a PhD. Measures after regular quality meetings and students' feedback led to improving the curriculum and workload, teacher's training, and changes to examination regulations.After 5 years, statistics show that our expectations have been met exceedingly. All graduates had excellent career opportunities reflecting the necessity of this MSc and its topics. We are continuously working on improving the program and adapting the curriculum to the requirements in radiation sciences. The future vision includes an expansion of the program as well as undergraduate education opportunities in this field.

Since early April 2020, there has been intense debate over proposed clinical use of ionizing radiation to treat life-threatening pneumonia in Coronavirus Disease 2019 (COVID-19) patients. At least twelve relevant papers appeared by 20 May 2020. The radiation dose proposed for clinical trials are a single dose (0.1-1 Gy) or two doses (a few mGy followed by 0.1-0.25 Gy involving a putative adaptive response, or 1-1.5 Gy in two fractions 2-3 days apart). The scientific rationale for such proposed so-called low dose radiotherapy (LDRT) is twofold (note that only doses below 0.1 Gy are considered as low doses in the field of radiation protection, but here we follow the term as conventionally used in the field of radiation oncology). Firstly, the potentially positive observations in human case series and biological studies in rodent models on viral or bacterial pneumonia that were conducted in the pre-antibiotic era. Secondly, the potential anti-inflammatory properties of LDRT, which have been seen when LDRT is applied locally to subacute degenerative joint diseases, mainly in Germany. However, the human and animal studies cited as supportive evidence have significant limitations, and whether LDRT produces anti-inflammatory effects in the inflamed lung or exacerbates ongoing COVID-19 damage remains unclear. Therefore, we conclude that the available scientific evidence does not justify clinical trials of LDRT for COVID-19 pneumonia, with unknown benefit and known mortality risks from radiogenic cancer and circulatory disease. Despite the significant uncertainties in these proposals, some clinical trials are ongoing and planned. This paper gives an overview of current situations surrounding LDRT for COVID-19 pneumonia.

This letter addresses our concerns regarding a paper by Salomaa et al. entitled ‘Low dose radiation therapy for COVID-19 pneumonia: is there any supportive evidence?’ published in International Jou...

Purpose: Pulmonary inflammation is an adverse consequence of radiation therapy in breast cancer. The aim of this study was to elucidate biological pathways leading to this pathology. Materials and methods: Lung endothelial cells were isolated 24 h after thorax-irradiation (sham or 10 Gy X-ray) from female C57Bl/6 mice and cultivated for 6 days. Results: Quantitative proteomic analysis of lung endothelial cells was done using data independent acquisition (DIA) mass spectrometry. The data were analyzed using Ingenuity Pathway Analysis and STRINGdb. In total, 4220 proteins were identified using DIA of which 60 were dysregulated in the irradiated samples (fold change >= 2.00 or

Radiation exposure can evoke cellular stress responses. Emerging recognition that long non-coding RNAs (lncRNAs) act as regulators of gene expression has broadened the spectra of molecules controlling the genomic landscape upon alterations in environmental conditions. Knowledge of the mechanisms responding to low dose irradiation (LDR) exposure is very limited yet most likely involve subtle ancillary molecular pathways other than those protecting the cell from direct cellular damage. The discovery that transcription of the lncRNA PARTICLE (promoter of MAT2A- antisense radiation-induced circulating lncRNA; PARTICL) becomes dramatically instigated within a day after LDR exposure introduced a new gene regulator onto the biological landscape. PARTICLE affords an RNA binding platform for genomic silencers such as DNA methyltransferase 1 and histone tri-methyltransferases to reign in the expression of tumor suppressors such as its neighboring MAT2A in cis as well as WWOX in trans. In silico evidence offers scope to speculate that PARTICLE exploits the abundance of Hoogsten bonds that exist throughout mammalian genomes for triplex formation, presumably a vital feature within this RNA silencer. PARTICLE may provide a buffering riboswitch platform for S-adenosylmethionine. The correlation of PARTICLE triplex formation sites within tumor suppressor genes and their abundance throughout the genome at cancer-related hotspots offers an insight into potential avenues worth exploring in future therapeutic endeavors.

Changes in geomagnetic conditions have been shown to affect the rhythms produced by the brain and heart and that human autonomic nervous system activity reflected in heart rate variability (HRV) over longer time periods can synchronize to changes in the amplitude of resonant frequencies produced by geomagnetic field-line and Schumann resonances. During a 15-day period, 104 participants located in California, Lithuania, Saudi Arabia, New Zealand, and England underwent continuous ambulatory HRV monitoring. The local time varying magnetic field (LMF) intensity was obtained using a time synchronized and calibrated network of magnetometers located at five monitoring sites in the same geographical locations as the participant groups. This paper focuses on the results of an experiment conducted within the larger study where all of the participants simultaneously did a heart-focused meditation called a Heart Lock-In (HLI) for a 15-min period. The participant’s level of HRV coherence and HRV synchronization to each other before, during and after the HLI and the synchronization between participants’ HRV and local time varying magnetic field power during each 24-h period were computed for each participant and group with near-optimal chaotic attractor embedding techniques. In analysis of the participants HRV coherence before, during and after the HLI, most of the groups showed significantly increased coherence during the HLI period. The pairwise heart rhythm synchronization between participants’ in each group was assessed by determining the Euclidean distance of the optimal time lag vectors of each participant to all other participants in their group. The group member’s heart rhythms were significantly more synchronized with each other during the HLI period in all the groups. The participants’ daily LMF-HRV-synchronization was calculated for each day over an 11-day period, which provided a 5-day period before, the day of and 5-days after the HLI day. The only day where all the groups HRV was positively correlated with the LMF was on the day of the HLI and the synchronization between the HRV and LMF for all the groups was significantly higher than most of the other days.

Multi-station observations of Schumann resonance (SR) intensity document common behavior in the evolution of continental-scale lightning activity in two super El Niño events, occurring in 1997/98 and 2015/16. The vertical electric field component of SR at Nagycenk, Hungary and the two horizontal magnetic field components in Rhode Island, USA in 1997, and in 2014–2015, the two horizontal magnetic field components at Hornsund, Svalbard and Eskdalemuir, United Kingdom as well as in Boulder Creek, California and Alberta, Canada exhibit considerable increases in SR intensity from some tens of percent up to a few hundred percents in the transition months preceding the two super El Niño events. The UT time distribution of anomalies in SR intensity indicates that in 1997 the lightning activity increases mainly in Southeast Asia, the Maritime Continent and India, i.e. the Asian chimney region. On the other hand, a global response in lightning is indicated by the anomalies in SR intensity in 2014 and 2015. SR-based results are strengthened by comparison to independent lightning observations from the Optical Transient Detector and the World Wide Lightning Location Network, which also exhibit increased lightning activity in the transition months. The increased lightning is attributable to increased instability due to thermodynamic disequilibrium between the surface and the mid-troposphere during the transition. The main conclusion is that variations in SR intensity may act as a precursor for the occurrence and magnitude of these extreme climate events, and in keeping with earlier findings, as a precursor to maxima in global surface air temperature.

Extracellular vesicles (EVs) are freely circulating nano/micrometer-sized membrane-bound vesicles released by various cell types. Their cargo consists of proteins, lipids, metabolites, and different types of RNA molecules reflecting the origin of the secreting cell type or tissue. Since the EV cargo is constantly changing in response to pathological status or different environmental stressors, it has been extensively studied in the quest for biomarkers, especially in the cancer research. Mass spectrometry (MS)-based proteome analysis is a powerful tool to elucidate the protein cargo in EVs. This chapter describes and characterizes three MS-compatible lysis methods, namely by using urea, guanidium hydrochloride, and radioimmunoprecipitation buffer for isolating proteins from EVs.

Clinical tissue archives represent an invaluable source of biological information. Formalin-fixed, paraffin-embedded (FFPE) tissue can be used for retrospective investigation of biomarkers of diseases and prognosis.Recently, the number of studies using proteome profiling of samples from clinical archives has markedly increased. However, the application of conventional quantitative proteomics technologies remains a challenge mainly due to the harsh fixation process resulting in protein cross-linking and protein degradation. In the present chapter, we demonstrate a protocol for label-free proteomic analysis of FFPE tissue prepared from human cardiac autopsies. The data presented here highlight the applicability and suitability of FFPE heart tissue for understanding the molecular mechanism of cardiac injury using a proteomics approach.

Extracellular vesicles (EVs) are freely circulating nano/micrometer-sized membrane-bound vesicles released by various cell types. Their cargo consists of proteins, lipids, metabolites, and different types of RNA molecules reflecting the origin of the secreting cell type or tissue. Since the EV cargo is constantly changing in response to pathological status or different environmental stressors, it has been extensively studied in the quest for biomarkers, especially in the cancer research. Mass spectrometry (MS)-based proteome analysis is a powerful tool to elucidate the protein cargo in EVs. This chapter describes and characterizes three MS-compatible lysis methods, namely by using urea, guanidium hydrochloride, and radioimmunoprecipitation buffer for isolating proteins from EVs.

Clinical tissue archives represent an invaluable source of biological information. Formalin-fixed, paraffin-embedded (FFPE) tissue can be used for retrospective investigation of biomarkers of diseases and prognosis.Recently, the number of studies using proteome profiling of samples from clinical archives has markedly increased. However, the application of conventional quantitative proteomics technologies remains a challenge mainly due to the harsh fixation process resulting in protein cross-linking and protein degradation. In the present chapter, we demonstrate a protocol for label-free proteomic analysis of FFPE tissue prepared from human cardiac autopsies. The data presented here highlight the applicability and suitability of FFPE heart tissue for understanding the molecular mechanism of cardiac injury using a proteomics approach.

Radiation-induced inflammation leading to the permeability of the endothelial barrier may increase the risk of cardiovascular disease. The aim of this study was to investigate potential mechanisms in vitro at the level of the proteome in human coronary artery endothelial cells (HCECest2) that were exposed to radiation doses of 0, 0.25, 0.5, 2.0 and 10 Gy (60Co-&gamma, ). Proteomics analysis was performed using mass spectrometry in a label-free data-independent acquisition mode. The data were validated using bioinformatics and immunoblotting. The low- and moderate-dose-irradiated samples (0.25 Gy, 0.5 Gy) showed only scarce proteome changes. In contrast, an activation of DNA-damage repair, inflammation, and oxidative stress pathways was seen after the high-dose treatments (2 and 10 Gy). The level of the DNA damage response protein DDB2 was enhanced early at the 10 Gy dose. The expression of proteins belonging to the inflammatory response or cGAS-STING pathway (STING, STAT1, ICAM1, ISG15) increased in a dose-dependent manner, showing the strongest effects at 10 Gy after one week. This study suggests a connection between the radiation-induced DNA damage and the induction of inflammation which supports the inhibition of the cGAS-STING pathway in the prevention of radiation-induced cardiovascular disease.

In the event of a mass casualty radiological or nuclear scenario, it is important to distinguish between the unexposed (worried well), low-dose exposed individuals and those developing the hematological acute radiation syndrome (HARS) within the first three days postirradiation. In previous baboon studies, we identified altered gene expression changes after irradiation, which were predictive for the later developing HARS severity. Similar changes in the expression of four of these genes were observed using an in vitro human whole blood model. However, these studies have provided only limited information on the time frame of the changes after exposure in relationship to the development of HARS. In this study we analyzed the time-dependent changes in mRNA expression after in vitro irradiation of whole blood. Changes in the expression of informative mRNAs (FDXR, DBB2, POU2AF1 and WNT3) were determined in the blood of eight healthy donors (6 males, 2 females) after irradiation at 0 (control), 0.5, 2 and 4 Gy using qRT-PCR. FDXR expression was significantly upregulated (P < 0.001) 4 h after ≥0.5 Gy irradiation, with an 18-40-fold peak attained 4-12 h postirradiation which remained elevated (4-9-fold) at 72 h. DDB2 expression was upregulated after 4 h (fold change, 5-8, P < 0.001 at ≥ 0.5 Gy) and remained upregulated (3-4-fold) until 72 h (P < 0.001). The earliest time points showing a significant downregulation of POU2AF1 and WNT3 were 4 h (fold change = 0.4, P = 0.001, at 4 Gy) and 8 h (fold change = 0.3-0.5, P < 0.001, 2-4 Gy), respectively. These results indicate that the diagnostic window for detecting HARS-predictive changes in gene expression may be opened as early as 2 h for most (75%) and at 4 h postirradiation for all individuals examined. Depending on the RNA species studied this may continue for at least three days postirradiation.

The triple-negative breast tumor boundary is made of aligned, linear collagen. The pro-oncogenic impact of linear collagen is well established; however, its mechanism of formation is unknown. An in vitro analogue of the tumor border is created by a co-culture of MDA-MB-231 cells, adipose derived stem cells, and dermal fibroblasts. Decellularization of this co-culture after seven days reveals an extracellular matrix that is linear in fashion, high in pro-oncogenic collagen type VI, and able to promote invasion of reseeded cells. Further investigation revealed linear collagen VI is produced by fibroblasts in response to a paracrine co-culture of adipose derived stem cells and MDA-MB-231, which together secrete high levels of the chemokine CCL5. The addition of monoclonal antibody against CCL5 to the co-culture results in an unorganized matrix with dramatically decreased collagen VI. Importantly, reseeded cells do not exhibit pro-oncogenic behavior. These data illustrate a cellular mechanism, which creates linear extracellular matrix (ECM) in vitro, and highlight a potential role of CCL5 for building striated tumor collagen in vivo.

Normal tissue toxicity is a dose-limiting factor in radiation therapy. Therefore, a detailed understanding of the normal tissue response to radiation is necessary to predict the risk of normal tissue toxicity and to development strategies for tissue protection. One component of normal tissue that is continuously exposed during therapeutic irradiation is the circulating population of peripheral blood mononuclear cells (PBMC). PBMCs are highly sensitive to ionizing radiation (IR), however, little is known about how IR affects the PBMC response on a systemic level. It was the aim of this study to investigate whether IR was capable to induce changes in the composition and function of extracellular vesicles (EVs) secreted from PBMCs after radiation exposure to different doses. Therefore, whole blood samples from healthy donors were exposed to X-ray radiation in the clinically relevant doses of 0, 0.1, 2 or 6 Gy and PBMC-secreted EVs were isolated 72 h later. Proteome and miRNome analysis of EVs as well as functional studies were performed. Secreted EVs showed a dose-dependent increase in the number of significantly deregulated proteins and microRNAs. For both, proteome and microRNA data, principal component analysis showed a dose-dependent separation of control and exposed groups. Integrated pathway analysis of the radiation-regulated EV proteins and microRNAs consistently predicted an association of deregulated molecules with apoptosis, cell death and survival. Functional studies identified endothelial cells as an efficient EV recipient system, in which irradiation of recipient cells further increased the uptake. Furthermore an apoptosis suppressive effect of EVs from irradiated PBMCs in endothelial recipient cells was detected. In summary, this study demonstrates that IR modifies the communication between PBMCs and endothelial cells. EVs from irradiated PBMC donors were identified as transmitters of protective signals to irradiated endothelial cells. Thus, these data may lead to the discovery of biomarker candidates for radiation dosimetry and even more importantly, they suggest EVs as a novel systemic communication pathway between irradiated normal, non-cancer tissues.

Schumann resonances (SR) are the global electromagnetic resonances of the Earth-ionosphere cavity and constitute the extremely low frequency (< 100 Hz) radiation of the worldwide lightning activity (Schumann, 1952). The recording of SR intensity at a few distant SR stations is an efficient tool to monitor the global lightning. We present the variations of SR intensity in the transition months preceding the warm ENSO episodes for the two super El Niño events in 1997/98 and 2015/16 as well as for the two medium size El Niño periods in 2001/02 and 2008/09 based on SR observations at multiple locations.: Nagycenk, Hungary (47.6N, 16.7E); Hornsund, Svalbard (77.0N, 15.6E); Eskdalemuir, UK (55.3N, 3.2W); Alberta, Canada (51.9N, 111.5W); Boulder Creek, USA (37.2N, 122.1W).A remarkable increase in SR intensity is documented two-three months before or just at the beginning of El Niño episodes as compared with the SR intensity in the same months of the preceding La Niña (or non-ENSO) phase for all cases studied here. The percentage increase in SR intensity depends on the amplitude of the warm ENSO period, and is consistently higher for the two super El Niño events. The enhanced SR intensity indicates a worldwide response of global lightning activity. Increased atmospheric instability due to the land-ocean thermal interaction during the transition interval could be responsible for the intensification of lightning activity. This systematic behavior may have been overlooked in earlier studies that compared lightning activity in the integrated ‘cold’ and the ‘warm’ phases, but without exploring the transitional variation. Our results suggest that the SR intensity variation on the interannual time scale acts a precursor for the occurrence of warm ENSO episode.

We assessed students' expectations to a full two-year Master of Science course regarding workload, extracurricular activities, learning methods, and career plans.A questionnaire was handed out to all students in the MSc radiation biology course. Questions evaluated the time for study and lectures expected a desire for specific teaching and testing formats, expectations from extracurricular activities as well as the motivation to study the subject and the future career plans. All students (100%) enrolled in the first semester were handed out and completed the questionnaire.Most students had learned about the course from the internet (68.75%) or received information from teachers or professors (25%). Two students stated that all disciplines were equally relevant (25%). Others students made clear preferences: fourteen voted molecular biology (87.5%) as relevant, radiation protection in 93.75%, 81.25% consider physics the most important topic, followed by immunology (62.5%). Tutorials and lectures were preferred teaching formats. Generally, a workload of 20 hours per week is preferred.An ongoing feedback loop is important in designing a modern Master of Science course in the context of the Bologna process. Valuable information is given by students and should be integrated continuously in the design and continuation process.

Reliable data on the effects of chronic prenatal exposure to low dose (LD) of ionizing radiation in humans are missing. There are concerns about adverse long-term effects that may persist throughout postnatal life of the offspring. Due to their slow cell cycle kinetics and life-long residence time in the organism, mesenchymal stem cells (MSCs) are more susceptible to low level genotoxic stress caused by extrinsic multiple LD events. The aim of this study was to investigate the effect of chronic, prenatal LD gamma irradiation to the biology of MSCs later in life. C3H mice were exposed in utero to chronic prenatal irradiation of 10 mGy/day over a period of 3 weeks. Two years later, MSCs were isolated from the bone marrow and analyzed in vitro for their radiosensitivity, for cellular senescence and for DNA double-strand break recognition after a second acute gamma-irradiation. In addition to these cellular assays, changes in protein expression were measured using HPLC–MS/MS and dysregulated molecular signaling pathways identified using bioinformatics. We observed radiation-induced proteomic changes in MSCs from the offspring of in utero irradiated mice (leading to ~ 9.4% of all detected proteins being either up- or downregulated) as compared to non-irradiated controls. The proteomic changes map to regulation pathways involved in the extracellular matrix, the response to oxidative stress, and the Wnt signaling pathway. In addition, chronic prenatal LD irradiation lead to an increased rate of in vitro radiation-induced senescence later in life and to an increased number of residual DNA double-strand breaks after 4 Gy irradiation, indicating a remarkable interaction of in vivo radiation in combination with a second acute dose of in vitro radiation. This study provides the first insight into a molecular mechanism of persistent MSC damage response by ionizing radiation exposure during prenatal time and will help to predict therapeutic safety and efficacy with respect to a clinical application of stem cells.

The long non-coding RNA PARTICLE (Gene PARTICL- ‘Promoter of MAT2A-Antisense RadiaTion Induced Circulating LncRNA) partakes in triple helix (triplex) formation, is transiently elevated following low dose irradiation and regulates transcription of its neighbouring gene - Methionine adenosyltransferase 2A. It now emerges that PARTICLE triplex sites are predicted in many different genes across all human chromosomes. In silico analysis identified additional regions for PARTICLE triplexes at >1600 genomic locations. Multiple PARTICLE triplexes are clustered predominantly within the human and mouse tumor suppressor WW Domain Containing Oxidoreductase (WWOX) gene. Surface plasmon resonance diffraction and electrophoretic mobility shift assays were consistent with PARTICLE triplex formation within human WWOX with high resolution imaging demonstrating its enrichment at this locus on chromosome 16. PARTICLE knockdown and over-expression resulted in inverse changes in WWOX transcripts levels with siRNA interference eliminating PARTICLEs elevated transcription to irradiation. The evidence for a second functional site of PARTICLE triplex formation at WWOX suggests that PARTICLE may form triplex-mediated interactions at multiple positions in the human genome including remote loci. These findings provide a mechanistic explanation for the ability of lncRNAs to regulate the expression of numerous genes distributed across the genome.

Mesenchymal stem cells (MSCs) are multilineage adult stem cells with considerable potential for cell���based regenerative therapies. In vitro expansion changes their epigenetic and cellular properties, with a poorly understood impact on DNA damage response (DDR) and genome stability. We report here results of a transcriptome���based pathway analysis of in vitro���expanded human bone marrow���derived mesenchymal stem cell (hBM���MSCs), supplemented with cellular assays focusing on DNA double���strand break (DSB) repair. Gene pathways affected by in vitro aging were mapped using gene ontology, KEGG, and GSEA, and were found to involve DNA repair, homologous recombination (HR), cell cycle control, and chromosomal replication. Assays for the recognition (�����H2AX + 53BP1 foci) and repair (pBRCA1 + �����H2AX foci) of X���ray���induced DNA DSBs in hBM���MSCs show that over a period of 8 weeks of in vitro aging (i.e., about 10 doubling times), cells exhibit a reduced DDR and a higher fraction of residual DNA damage. Furthermore, a distinct subpopulation of cells with impaired DNA DSB recognition was observed. Several genes that participate in DNA repair by HR (e.g., Rad51, Rad54, BRCA1) show a 2.3��� to fourfold reduction of their mRNA expression by qRT���PCR. We conclude that the in vitro expansion of hMSCs can lead to aging���related impairment of the recognition and repair of DNA breaks., publishedVersion

The impact of low-dose ionizing radiation (IR) on the human brain has recently attracted attention due to the increased use of IR for diagnostic purposes. The aim of this study was to investigate low-dose radiation response in the hippocampus. Female B6C3F1 mice were exposed to total body irradiation with 0 (control), 0.063, 0.125, or 0.5 Gy. Quantitative label-free proteomic analysis of the hippocampus was performed after 24 months. CREB signaling and CREB-associated pathways were affected at all doses. The lower doses (0.063 and 0.125 Gy) induced the CREB pathway, whereas the exposure to 0.5 Gy deactivated CREB. Similarly, the lowest dose (0.063 Gy) was anti-inflammatory, reducing the number of activated microglia. In contrast, induction of activated microglia and reactive astroglia was found at 0.5 Gy, suggesting increased inflammation and astrogliosis, respectively. The apoptotic markers BAX and cleaved CASP-3 and oxidative stress markers were increased only at the highest dose. Since the activated CREB pathway plays a central role in learning and memory, these data suggest neuroprotection at the lowest dose (0.063 Gy) but neurodegeneration at 0.5 Gy. The response to 0.5 Gy resembles alterations found in healthy aging and thus may represent radiation-induced accelerated aging of the brain.

Development of new technologies has resulted in the significant expansion of biological research, among which studies in the area of genomics, transcriptomics, proteomics, and metabolomics are the leading ones. In the majority of omics studies, the goal is to identify reliable molecular biomarkers and pathways associated with the examined process. In almost all cases, a list of differentially expressed genes or proteins is constructed, which is not easy to obtain for some experimental designs. In our work, we mainly focus on the experiments with small sample size. The goal was to determine the robust proteomic signature of radiation exposure in the mouse model. Our selection algorithm combines mathematical modelling of signal and its fold change distributions with the comprehensive effect size analysis. Thanks to the data-driven automated thresholding of the protein absolute or relative (fold change) expressions, and Cohens effect size based filters, the obtained proteomic signature demonstrated a higher level of consistency and functional coherency. The additional, intuitively expected, signalling pathways were identified when compared to the standard statistical approach.

Pancreatic cancer has a poor prognosis. New treatment options are urgently required to improve patient outcomes. One promising new class of anticancer drugs are synthetic histone deacetylase inhibitors (HDACi) which modulate chromatin structure and gene expression by blocking histone deacetylation. In this study, we aimed at comparing the in vitro capacities of the HDACi SAHA and CUDC-101 to increase radiosensitivity of human pancreatic tumor cell lines. Therefore, three pancreatic cancer cell lines (Su.86.86, MIA Paca-2, T3M-4) were treated with SAHA (1.5&ndash, 5 &micro, M) or CUDC-101 (0.25&ndash, 3 &micro, M) and after 24 h irradiated. Cell proliferation, clonogenic survival and apoptosis was determined. Additionally, cell lysates were investigated for the expression of apoptosis-related proteins. CUDC-101 and SAHA increased the radiation sensitivity of pancreatic tumor cell lines in a dose-dependent manner. This was evidenced by cell proliferation and clonogenic survival. Furthermore, enhanced radiation sensitivity after CUDC-101 or SAHA treatment was confirmed for Su.86.86 and T3M-4 cells in a 3-D microtissue approach. Increased amounts of subG1 cells and diminished full length PARP-1 suggest increased radiation-induced apoptosis after SAHA or CUDC-101 treatment. The comparison of both inhibitors in these assays manifested CUDC-101 as more potent radiosensitizer than SAHA. In line, western blot quantification of the apoptosis-inhibitory proteins XIAP and survivin showed a stronger down-regulation in response to CUDC-101 treatment than after SAHA application. These proteins may contribute to the synergy between HDAC inhibition and radiation response. In conclusion, these preclinical results suggest that treatment with the HDAC inhibitors CUDC-101 or SAHA can enhance radiation-induced cytotoxicity in human pancreatic cells. However, comparison of both inhibitors identified the multi target inhibitor CUDC-101 as more potent radiosensitizer than the HDAC inhibitor SAHA.

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.