Transcriptional responses to ionising radiation for biological dosimetry purposes

This thesis was submitted for the degree of Doctor of Philosophy and awarded by Brunel University LondonExposures to ionising radiation resulting from natural sources or medical diagnostics are generally very low. In contrast, exposures to therapeutic radiation, although, they are often partial exposures, represent much higher doses. Similar levels of exposure may also occur as a consequence of a radiological accident, where it would be necessary to quickly separate individuals requiring urgent medical attention from the “worried-well”. The well-established biodosimetry techniques based on cytogenetics, particularly scoring dicentric chromosomes, are accurate and sensitive, yet, they are unsuitable for mass screening due to limited capacity and the time required for providing dose estimates. Measuring gene expression changes following radiation exposure was suggested over a decade ago to be an alternative method for dose estimation, as it is a quick, sensitive and suitable technique for high throughput application. The qPCR protocol was extensively optimised for increased reproducibility and sensitivity in order to be suitable for biodosimetry purposes. Radiation-responsive transcripts were identified and characterised in terms of temporal- and dose-responses. Finally, candidate transcripts were validated in human blood irradiated ex vivo and in vivo in blood samples obtained from cancer patients undergoing radiotherapy treatment. The data generated here serve as a proof of principle that qPCR-based gene expression assays can be used for radiation biodosimetry purposes to aid classical cytogenetics tools in an event of mass causality.Public Health Englan

A lightweight planar antenna element with optimized feed for use onboard spacecraft, Journal of Telecommunications and Information Technology, 2005, nr 2

This paper is a report on low gain antennas (LGAs) manufactured with bonded lightweight materials. These antennas can sustain large temperature variations and are capable of functioning in modern miniature spacecraft, mainly in the telemetry, command and ranging (TC&R) links. When made of cheaper materials, the proposed circularly polarized antenna can be widely used in the base stations of short-range wireless systems. The recommended operating frequencies are between 1 and 8 GHz. One major technical consideration is obtaining the required high quality of circular polarization with as low space demands as possible. A lightweight 90◦ polarizer, printed on a dielectric membrane and operating over a broad bandwidth, is proposed for the antenna feed. Owing to the bandwidth advantage of the polarizer and the use of carefully designed aperture coupled feed, the electrical characteristics maintain good properties over a wide frequency range (15%)

Optimizing circular polarization within a beam of patch antenna elements, Journal of Telecommunications and Information Technology, 2007, nr 1

The paper presents the results of an investigation into patch antenna elements that would be capable of pro- viding good circular polarization not only in the broadside direction, but also over a wide range of elevation angles

A broadband uniplanar quasi-Yagi antenna – parameter study in application to a spatial power combiner, Journal of Telecommunications and Information Technology, 2001, nr 4

A parameter study is performed of a broadband uniplanar quasi-Yagi antenna with regard to its design and use in a spatial power combiner. A 3D full-wave electromagnetic field analysis is applied to identify parameters, which mostly affect the design frequency and operational bandwidth of this antenna. Optimal design conditions are determined. Using these design criteria a passive spatial power combiner employing trays of back-to-back connected quasi-Yagi antennas is developed. This combiner is investigated in terms of insertion losses and field uniformity, which are key factors in obtaining high power combining efficiency

Comparison of established and emerging biodosimetry assays

Rapid biodosimetry tools are required to assist with triage in the case of a large-scale radiation incident. Here, we aimed to determine the dose-assessment accuracy of the well-established dicentric chromosome assay (DCA) and cytokinesis-block micronucleus assay (CBMN) in comparison to the emerging γ-H2AX foci and gene expression assays for triage mode biodosimetry and radiation injury assessment. Coded blood samples exposed to 10 X-ray doses (240 kVp, 1 Gy/min) of up to 6.4 Gy were sent to participants for dose estimation. Report times were documented for each laboratory and assay. The mean absolute difference (MAD) of estimated doses relative to the true doses was calculated. We also merged doses into binary dose categories of clinical relevance and examined accuracy, sensitivity and specificity of the assays. Dose estimates were reported by the first laboratories within 0.3-0.4 days of receipt of samples for the γ-H2AX and gene expression assays compared to 2.4 and 4 days for the DCA and CBMN assays, respectively. Irrespective of the assay we found a 2.5-4-fold variation of interlaboratory accuracy per assay and lowest MAD values for the DCA assay (0.16 Gy) followed by CBMN (0.34 Gy), gene expression (0.34 Gy) and γ-H2AX (0.45 Gy) foci assay. Binary categories of dose estimates could be discriminated with equal efficiency for all assays, but at doses ≥1.5 Gy a 10% decrease in efficiency was observed for the foci assay, which was still comparable to the CBMN assay. In conclusion, the DCA has been confirmed as the gold standard biodosimetry method, but in situations where speed and throughput are more important than ultimate accuracy, the emerging rapid molecular assays have the potential to become useful triage tools

The rate of X-ray-induced DNA double-strand break repair in the embryonic mouse brain is unaff ected by exposure to 50 Hz magnetic fi elds

Following in utero exposure to low dose radiation (10 – 200 mGy), we recently observed a linear induction of DNA double-strand breaks (DSB) and activation of apoptosis in the embryonic neuronal stem/progenitor cell compartment. No signifi cant induction of DSB or apoptosis was observed following exposure to magnetic fi elds (MF). In the present study, we exploited this in vivo system to examine whether exposure to MF before and after exposure to 100 mGy X-rays impacts upon DSB repair rates. Materials and methods : 53BP1 foci were quantifi ed following combined exposure to radiation and MF in the embryonic neuronal stem/progenitor cell compartment. Embryos were exposed in utero to 50 Hz MF at 300 m T for 3 h before and up to 9 h after exposure to 100 mGy X-rays. Controls included embryos exposed to MF or X-rays alone plus sham exposures. Results : Exposure to MF before and after 100 mGy X-rays did not impact upon the rate of DSB repair in the embryonic neuronal stem cell compartment compared to repair rates following radiation exposure alone. Conclusions : We conclude that in this sensitive system MF do not exert any signifi cant level of DNA damage and do not impede the repair of X-ray induced damage

Combining CDKN1A gene expression and genome-wide SNPs in a twin cohort to gain insight into the heritability of individual radiosensitivity

Individual variability in response to radiation exposure is recognised and has often been reported as important in treatment planning. Despite many efforts to identify biomarkers allowing the identification of radiation sensitive patients, it is not yet possible to distinguish them with certainty before the beginning of the radiotherapy treatment. A comprehensive analysis of genome-wide single-nucleotide polymorphisms (SNPs) and a transcriptional response to ionising radiation exposure in twins have the potential to identify such an individual. In the present work, we investigated SNP profile and CDKN1A gene expression in blood T lymphocytes from 130 healthy Caucasians with a complex level of individual kinship (unrelated, mono- or dizygotic twins). It was found that genetic variation accounts for 66% (95% CI 37-82%) of CDKN1A transcriptional response to radiation exposure. We developed a novel integrative multi-kinship strategy allowing investigating the role of genome-wide polymorphisms in transcriptomic radiation response, and it revealed that rs205543 (ETV6 gene), rs2287505 and rs1263612 (KLF7 gene) are significantly associated with CDKN1A expression level. The functional analysis revealed that rs6974232 (RPA3 gene), involved in mismatch repair (p value = 9.68e-04) as well as in RNA repair (p value = 1.4e-03) might have an important role in that process. Two missense polymorphisms with possible deleterious effect in humans were identified: rs1133833 (AKIP1 gene) and rs17362588 (CCDC141 gene). In summary, the data presented here support the validity of this novel integrative data analysis strategy to provide insights into the identification of SNPs potentially influencing radiation sensitivity. Further investigations in radiation response research at the genomic level should be therefore continued to confirm these findings.Peer reviewe

Epigenetic clock for skin and blood cells applied to Hutchinson Gilford Progeria Syndrome and ex vivo studies

DNA methylation (DNAm)-based biomarkers of aging have been developed for many tissues and organs. However, these biomarkers have sub-optimal accuracy in fibroblasts and other cell types used in ex vivo studies. To address this challenge, we developed a novel and highly robust DNAm age estimator (based on 391 CpGs) for human fibroblasts, keratinocytes, buccal cells, endothelial cells, lymphoblastoid cells, skin, blood, and saliva samples. High age correlations can also be observed in sorted neurons, glia, brain, liver, and even bone samples. Gestational age correlates with DNAm age in cord blood. When used on fibroblasts from Hutchinson Gilford Progeria Syndrome patients, this age estimator (referred to as the skin & blood clock) uncovered an epigenetic age acceleration with a magnitude that is below the sensitivity levels of other DNAm-based biomarkers. Furthermore, this highly sensitive age estimator accurately tracked the dynamic aging of cells cultured ex vivo and revealed that their proliferation is accompanied by a steady increase in epigenetic age. The skin & blood clock predicts lifespan and it relates to many age-related conditions. Overall, this biomarker is expected to become useful for forensic applications (e.g. blood or buccal swabs) and for a quantitative ex vivo human cell aging assay