This work investigated the role of the transcription factor NF-B in the radiation response of directly irradiated cells and of cells treated with conditioned medium of radiation exposed cells. Genetically impeding NF-B activation by removal of NEMO enables a mechanistic analysis of the NF-B signaling pathway (SCHMIDT-SUPPRIAN et al., 2000). For that reason a knock-out cell culture model was chosen. MEF NEMO ko cells are NF-B (-) due to the removal of NEMO, one of the main components of the NF-B signaling cascade (ADHIKARI et al., 2007; COURTOIS, FAUVARQUE, 2018; MAUBACH et al., 2017; MCCOOL, MIYAMOTO, 2012; WANG et al., 2017b). Studies of molecular mechanisms using pharmacological inhibition suffer a series of limitations: degree of inhibition efficiency, specificity to the target protein, potential interactions with secondary targets, and toxicity of the compound (KNIGHT, SHOKAT, 2007). Several inhibitors of NF-B have been tested for efficiency and cytotoxicity before (HELLWEG et al., 2009). The most promising of those substances was MG-132, a proteasome inhibitor that enables NF-B cytoplasmic retention through IB. The target specificity on the other hand made MG-132 unsuitable for investigation of NF- B-dependent mechanisms, as the proteasome plays a role in many cellular processes. Another substance, BMS-345541, inhibits IKK and thereby blocks NF-B activity similar to the NEMO ko model (BURKE et al., 2003; WU et al., 2011; YANG et al., 2006). Preliminary tests showed effective inhibition of NF-B at 10 µM BMS-345541, but long incubation times, necessary for the study of the radiation-induced bystander effect, increased the cytotoxicity of the compound to levels that rendered the use of BMS-345541 in future experiments inapplicable (data not shown).
The radiationinduced incorporation of CO at ele vated pressure (up to 15 bar) in pure as well as aqueous m ethanol was studied in view of a future utilization of CO. The yields of the final products: formaldehyde, glycolic aldehyde, formic and oxalic acids were measured. In aqueous methanol solutions, in addition to the CO -concentration the pH is a deter mining factor of the process.
Abstract: Irreparable double-strand breaks (DSBs) in response to ionizing radiation (IR) trigger prolonged DNA damage response (DDR) and induce premature senescence. Profound chromatin reorganization with formation of senescence-associated heterochromatin foci (SAHF) is an essential epigenetic mechanism for controlling the senescence-associated secretory phenotype (SASP). To decipher molecular mechanisms provoking continuous DDR leading to premature senescence, radiation-induced DSBs (53BP1-foci) and dynamics of histone variant H2A.J incorporation were analyzed together with chromatin re-modeling in human fibroblasts after IR exposure. High-resolution imaging by transmission electron microscopy revealed that persisting 53BP1-foci developed into DNA segments with chromatin alterations reinforcing senescence (DNA-SCARS), consistently located at the periphery of SAHFs. Quantitative immunogold-analysis by electron microscopy revealed that H2A.J, steadily co-localizing with 53BP1, is increasingly incorporated into DNA-SCARS during senescence progression. Strikingly, shRNA-mediated H2A.J depletion in fibroblasts modified senescence-associated chromatin re-structuring and abolished SASP, thereby shutting down the production of inflammatory mediators. These findings provide mechanistic insights into biological phenomena of SASP and suggest that H2A.J inhibition could ablate SASP, without affecting the senescence-associated growth arrest.
There is one particular idiosyncrasy of ionizing radiation (IR) inherent within radiation itself which makes its subsistence as a carcinogen outstanding, and shows how it greatly differs in its physical make-up when compared with other exogenous agents such as chemical toxicants, or other physical carcinogens, which are, on the whole, tissue specific in their action (Hall 2000; Schecker & Reynolds 1995). Not being hampered by the usual cellular walls that chemical agents produce, radiation has the power to deposit energy randomly within the biological material it so aptly penetrates (Nias 1998). Even though all the cells of the human body are, in such an event, likely to suffer damage through IR, the damage consistency is relative to the physical parameters that determine the dose of radiation which those particular cells, or tissue, actually receive. In recent years, cellular systems have been developed in order to be able to study better the malignant transformation of an individual cell in vitro (Liber et al. 1989). Conclusions resulting from these studies have confirmed several of the corroborative factors derived from the in vivo experiments (Pfeiffer et al. 2004). Despite the fact that the overall characteristics of radiation-induced cancer in vivo and the basic aspects of cellular transformation in vitro had already been quite clearly identified by the 1980s, only a limited quota of research has been undertaken in this specific field since then. In fact, IR has become a widely used tool for studying cellular and molecular responses to DNA damage, as these serve to enhance better understanding of the mutations and carcinogenic transformation of the cells. IR includes, sparsely ionizing events resulting from X-rays and γ-rays (electromagnetic) and densely ionizing events induced by α-particles, neutrons and atomic nuclei such as helium, carbon or argon (particulate), which can be accelerated in nuclear physics machines (Hall 2000).
Introduction: Early integration of science and ex- ploration concerns into the design of the Deep Space Gateway (DSG) is essential to maximizing its science and exploration potential. The proposed concept, char- acterization of outer space radiationinduced changes in microbial extremophiles, requires the DSG as infra- structure supplying power, communications, etc. to otherwise autonomous systems. Survival and prolifera- tion of life beyond low earth orbit (LBLEO) can be accomplished by exposing extremophilic microorgan- isms in outer space radiation (OSR) conditions using DSG system. Extremophilic microbial survival, adap- tation, biological functions, and molecular mechanisms associated with outer space radiation can be tested by exposing them onto DSG hardware (inside/outside) utilizing the traditional microbiology methods and state-of-the-art molecular biology techniques.
Oral mucositis is the most frequently occurring early side effect of radiotherapy in patients treated for head-and-neck cancers. The majority of patients undergoing curative treat- ment develop a severe reaction, manifesting as painful con- fluent lesions and/or ulcerations of the epithelial membranes lining the oral cavity [ 1 ]. Oral mucositis significantly re- duces the patient’s quality of life due to mucositis-associ- ated pain and speaking and swallowing difficulties, which often necessitate parenteral nutrition. Additionally, radia- tion-induced oral mucositis often leads to local and sys- temic infections because of disturbed mucosal barrier func- tion. Besides the socio-economic factor due to mucositis-as- sociated hospitalizations [ 2 ], these radiation-induced com- plications might lead to treatment interruptions thus signif- icantly lowering the tumor control probability.
Pentoxifylline (PTX), a nonspecific phosphodiesterase inhibitor, may improve tumor oxygenation due to rheo- logic effects, but may also ameliorate treatment-associated normal tissue morbidity through modulation of inflamma- tory changes [ 9 ]. When tested in a preclinical model of radiation-induced early oral mucositis, PTX was found to significantly reduce the incidence of mucosal ulceration in daily fractionation studies in the established mouse tongue model. PTX treatment yielded the most pronounced ra- dioprotective effect, when the administration interval in- cluded the second treatment week of fractionation. This is the time when repopulation, the adaptive epithelial ra- diation response [ 10 ], is already fully active. Time course parameters of oral mucositis, i. e., latent time to the onset of mucosal ulcerations and their respective duration, however, were found virtually unchanged. [ 11 ].
Monoclonal antibodies inhibit the EGFR by blocking the extracellular ligand binding domain of the EGFR. When the monoclonal antibodies bind to the EGFR, the receptor is blocked from binding extracellular ligands and the ligand dependent activation of EGFR is disrupted. Additionally, due to the molecular size and properties of the antibodies, the dimerization of the EGFR, which is required for activation of the intracellular tyrosine kinase domain, is inhibited. As a consequence, radiation-induced EGFR activation is impaired as well. In addition to the direct inhibition of the EGFR, monoclonal antibodies are thought to induce downregulation of the inhibited EGFR, leading to reduced EGFR expression (Jutten et al., 2009). It is also possible that the binding of monoclonal antibodies to the EGFR causes an immunomodulatory effect by homing immune cells and their cytotoxic immunologic reactions onto the antibody marked cancer cells. A drawback of EGFR inhibition via monoclonal antibodies is the required parenteral application, while due to their long half-life, the antibodies can be administered on a weekly basis during treatment (Guarino et al., 2009). A commonly used monoclonal antibody for EGFR inhibition is cetuximab. Various studies were performed to asses the effectiveness of cetuximab as an EGFR inhibitor. Radiosensitization was achieved in vitro as well as in xenografts using cetuximab with cancer cells overexpressing the EGFR, including lung cancer cell lines (Raben et al., 2005; Krause et al., 2005).
2 Nanophotonic applications of fs-laser radiationinduced nanostructures and their theoretical description
Abstract: We present the analysis of the optical properties of two different fs-laser- induced nanostructures which possess great potential for near-field applications due to their sub-100 nm features. The first structures are standing gold nanojets generated by irradiation of thin gold films with single pulses of fs-laser radiation. Due to their upright standing orientation, the characteristic extinction resonances of these nanojets are studied by grazing incidence infrared spectroscopy with p- polarized radiation. The resonance wavelength strongly depends on the lateral extent of the nanojet and can be tailored by adjusting the focusing numerical aperture and the applied pulse energy. The observed correlation between the lateral extent of the nanojet and the resonance wavelength is described by the monopole model of a linear antenna. The second type of structures under investigation are fs-laser- induced sub-wavelength ripples on silicon. Ripple structures of different geometrical shape are generated and coated with a thin gold film. Here, the metallic cover layer allows for the excitation of surface plasmons. Based on the generated surface pro- files, the corresponding plasmonic resonances are simulated. Angle-resolved UV-VIS spectroscopy measurements are performed and the resulting plasmonic dispersion relation of these structures is compared to the calculated resonance conditions. The plasmonic resonances and additional experiments of the surface structures reveal insights into the localized or propagation nature of the plasmonic modes excited on generated surface profiles.
Radiation-induced bystander effects (RIBE) are an acknowledged issue of radiation therapy. Radiation of tumor tissue has been shown to affect non-irradiated neighboring cells in a paracrine and endocrine manner. Transduction of bystander signaling though remains to be investigated in detail. A part of the transduction is the receptor-initiated activation of signaling pathways by secreted factors of the irradiated cell during irradiation damage response. This work focusses on the activation of the transcription factor Nuclear Factor κB (NF-κB) in bystander cells after irradiation. NF-κB is a well- known contributor to inflammatory processes like cyto- / chemokine production as well as to stress reactions such as the DNA damage response and cell cycle regulation. Using a mouse embryonic fibroblasts (MEF) in vitro model with a genetic knock-out of an NF-κB regulator (NEMO, NF-κB essential modulator), clonogenic survival and cell cycle distribution was determined in directly irradiated cells and in cells incubated with conditioned medium from X-irradiated cells (bystander treatment). Directly irradiated NEMO ko cells, plated for clonogenic survival immediately after X- irradiation, display the same dose-effect curve as the wildtype (wt) (a/bNEMO ko = 13.92 ± 2.4 vs. a/bwt = 12.37 ± 2.6). But when allowed to recover for 24 h, the wt cells show a broader shoulder in the curve (a/b =3.5 ± 2.9), indicating a role of NF-kB in the repair of radiationinduced DNA damages. Looking into the survival of bystander cells, the survival curves show a statistically different slope, with NEMO ko cells surviving better than wt cells (S16 Gy: NEMO ko = 1.66 vs wt = 0.83). The different behavior may correlate with NF-kB dependent DNA repair in bystander cells for NEMO ko and wt cells. Cell cycle analysis revealed a 6 hour delayed arrest in G2/M phase in directly irradiated NEMO ko cells compared to wt cells. This indicates that NF-kB regulated DNA repair pathways are important for recovery of radiationinduced damages. Bystander NEMO ko show an even further delayed arrest at 48 h, while wt bystander cells show no G2/M arrest at all. This supports the assumption that damages have to overcome a certain threshold to be recognized as repair-worthy. As NFkB has been reported to be involved in homologous recombination; cells with impairment in NFkB pathways, such as NEMO ko, register damages caused by bystander treatment differently from wt cells. This leads to G2/M arrest extending time for repair in NEMO ko bystander cells.
The radiationinduced formation of E t3SiBr and H B r from E t3SiH using various concentrations of E tB r in absence of air was investigated as a func tion of radiation dose. The highest product yields, G j(E t 3 SiBr) = 140 and G,(HBr) = 44, were achieved by using 1.30 mol/dm 3 EtB r. Taking into considera
DS exists either as soluble free glycosaminoglycan or constitutes an abundantly expressed proteoglycan, when covalently linked to a protein core. Sulfated glycans are key players in molecular and cellular events of inflamma- tion, chemokine regulation and leukocyte guidance [ 18 , 21 ], which could influence the inflammatory response in irradi- ated tissues [ 16 ]. Additionally, DS has a strong anticoagu- latory activity [ 20 ], which may target the early onset of hy- poxic conditions, which have been demonstrated in irradi- ated oral mucosa [ 17 ]. Furthermore, DS exerts functions in extracellular matrix assembly, fibroblast activity and wound repair processes [ 19 , 29 – 31 ]. Therefore, DS treatment could effectively modify various radiation-induced or -altered cel- lular or cell-matrix interactions [ 32 – 34 ]. To date, clinical applications of DS are scarce, although several preclinical and clinical studies revealed an array of potential indica- tions. Specifically, the antithrombotic properties of DS have been shown to be of clinical relevance. The studies revealed a predictable concentration-dependent dose response and overall clinical safety. To date, no DS therapy-associated bleeding complications have been reported, which makes DS a promising anticoagulation candidate for patients with bleeding disorders [ 35 , 36 ].
The bulk o f the radiation-induced degradation products o f 2'-deoxyguanosine in oxygen-free aqueous solution have been separated by high perform ance liquid chrom atography and charac terized by various spectroscopic techniques including fast atom bom bardm ent m ass spectrom etry, ‘H N M R and circular dichroism. The two main decom position products result from the opening o f the im idazole ring and further rearrangement o f the sugar m oiety. In addition, the form ation o f six other nucleosides was shown to involve sugar radicals with subsequent epim erization, re arrangement or cyclization reactions.
received N ovem ber 2. 1995/February 14, 1996 A poptosis. pH. Thymocytes, Irradiation
Thymocytes were used as a m odel system to study the effect of microenvironmental pH changes on the radia- tion-induced apoptosis. We found that the sensitivity of thym ocytes toward radiationinduced apoptosis is increased by increasing the pH of the incubation m e dium. The major sensitivity change occurs between pH 7 and 8.
Radiotherapy (RT) alone or in combination with surgery and chemotherapy is a central component of curative or palliative treatment for many cancer patients. For example, patients suffering from advanced non–small cell lung cancer (NSCLC) receive standard treatment with fractionated RT to the thoracic region or concurrent platinum-based radiochemotherapy (RCT), yielding local control rates of 40–66% ( 1 – 3 ). Yet, intratumoral heterogeneity and high intrinsic or acquired radioresistance can lead to relapse, whereas a pronounced radiosensitivity of coirradiated normal lung tissue causes adverse effects in sensitive patients, thereby limiting the application of curative RT doses and therapy intensification efforts of RT or RCT ( 3 , 4 ). Instead tolerable radiation doses are often linked to suboptimal tumor control, despite accepting side effects that decrease quality of life ( 2 , 3 , 5 ). Current efforts to improve RT outcome therefore aim at combining highly conformal RT with molecularly tailored treatments to increase efficacy of tumor cell killing or reduce adverse effects to normal tissues, respectively. Based on exciting findings about RT-induced support of local and systemic antitumor immunity particularly in combination with immunotherapy ( 6 – 11 ), further clinical trials evaluate the use of combining RT or RCT with immune checkpoint inhibitors (ICIs); yet despite encouraging results and durable responses obtained, for example, in NSCLC patients, only a fraction of patients respond to multimodal therapies with RCT and inhibitors of the PD-1/PD-L1 immune checkpoint, or patients develop resistance ( 12 , 13 ). Because RT-induced immune effects can also contribute to radiation-induced adverse effects ( 14 –
The enzyme activity and the electrophoretic characteristics of lactic dehydrogenase (LDH) in the plasma and different tissues from the mice with radiation-induced lymphoid leukaemia were examined and compared with those obtained from the normal mice. The LDH activity was ab normally increased in the leukaemic plasma. A considerable elevation of the enzyme level was also found both in the leukaemic spleen and in the lymphocytes isolated from it, but not in the liver, kidney, heart and muscle. LDH isozyme patterns were obtained by agar-gel electrophoresis. Each tissue exhibited a characteristic distribution of LDH activity among the separated isozymes. Leukaemic plasma contained much more LDH-5 than normal plasma, showing the pat tern similar to that obtained from leukaemic spleen. Kidney and heart from leukaemic mice indi cated no difference in the LDH-patterns as compared with those from normal mice.
In this study efforts were made to analyze global 5mC as well as 5hmC changes after irradiation over time using colorimetric ELISA-based techniques. While reasonable results were obtained for 5mC abundance after irradiation with X-rays (Figure 5-1 A), measurements of 5hmC abundance did not show reliable results and varied considerably (Figure 5-1 B). On the one hand, this may be caused by low 5hmC abundance in the cell lines investigated. As already mentioned, the total amount of 5hmC can be very limited (between 0.009 and 0.7 % of all bases [Globisch et al., 2010, Ito et al., 2011]) and therefore analysis may be difficult. On the other hand, in the experiments conducted in this work, ELISA-based colorimetric analysis might not provide sufficient sensitivity to detect small 5hmC amounts and only minor differences in 5hmC abundance induced by IR. Therefore those analyses can only give an estimate of global DNA hydroxymethylation levels. However this might not be generally valid since the colorimetric assay was successfully employed to assess global 5hmC changes in cells overexpressing TET2 catalytic domain, where a pronounced change of 5hmC abundance was expected. For the analysis of endogenous 5hmC levels, that is, in absence of TET2CD-GFP, immunofluorescence was preferred.
Ionising radiation was applied for therapeutic purposes soon after the discovery of x-rays by William Roentgen in 1895 (Durovic and Spasic-Jokic, 2008). In the early 1900’s the use widened with applications in clinical diagnostics, and treatment of diseases, use of radiation therapy to cure cancer in the 1920’s was to some extent successful (O'Farrell, 1975, Perluigi et al., 2009, Preston L.Dale 1994). Apart from therapeutic applications ionising radiation has been used for occupational purposes in the form of radium painting of luminous watch dials, nuclear panels, aircrafts etc (Herrera et al., 2010). One other non-therapeutic breakthrough in the field of ionising radiation was the development of nuclear power generation plants. The first reported adverse effects of radiation were noticed to be skin lesions after exposure (Gilchrist, 1897). The adverse consequences of all of these applications have come to the forefront, as in the case of over- exposure during radiation therapy, atomic bomb explosions or in the case of nuclear accidents (Chernobyl). The harmful effects of exposure to ionising radiation are dependent on the quality and dose of ionising radiation received (Beebe, 1982).