Assessment of DNA fibers to track replication dynamics.

DNA replication is a complex and tightly regulated process that must proceed accurately and completely if the cell is to faithfully transmit genetic material to its progeny. Organisms have thus evolved complex mechanisms to deal with the myriad exogenous and endogenous sources of replication impediments to which the cell is subject. These mechanisms are of particular relevance to cancer biology, given that such "replication stress" frequently foreshadows genome instability during cancer pathogenesis, and that many traditional chemotherapies and a number of precision medicines function by interfering with the progress of DNA replication. Visualization of the progress and dynamics of DNA replication in living cells was historically a major challenge, neatly surmounted by the development of DNA fiber assays that utilize the fluorescent detection of halogenated nucleotides to track replication forks at single-molecule resolution. This methodology has been widely applied to study the dynamics of unperturbed DNA replication, as well as the cellular responses to various replication stress scenarios. In recent years, subtle modifications to DNA fiber assays have facilitated assessment of the stability of nascent DNA at stalled replication forks, as well as the detection of single-stranded DNA gaps and their subsequent filling by error-prone polymerases. Here, we present and discuss several iterations of the fiber assay and suggest methodologies for the analysis of the data obtained.

MRNIP is a replication fork protection factor.

The remodeling of stalled replication forks to form four-way DNA junctions is an important component of the replication stress response. Nascent DNA at the regressed arms of these reversed forks is protected by RAD51 and the tumor suppressors BRCA1/2, and when this function is compromised, stalled forks undergo pathological MRE11-dependent degradation, leading to chromosomal instability. However, the mechanisms regulating MRE11 functions at reversed forks are currently unclear. Here, we identify the MRE11-binding protein MRNIP as a novel fork protection factor that directly binds to MRE11 and specifically represses its exonuclease activity. The loss of MRNIP results in impaired replication fork progression, MRE11 exonuclease-dependent degradation of reversed forks, persistence of underreplicated genomic regions, chemosensitivity, and chromosome instability. Our findings identify MRNIP as a novel regulator of MRE11 at reversed forks and provide evidence that regulation of specific MRE11 nuclease activities ensures protection of nascent DNA and thereby genome integrity.

Aircrew radiation dose estimates during recent solar particle events and the effect of particle anisotropy.

A model was developed using a Monte-Carlo radiation transport code, MCNPX, to estimate the additional radiation exposure to aircrew members during solar particle events. The model transports an extrapolated particle spectrum based on satellite measurements through the atmosphere to aircraft altitudes. This code produces the estimated flux at a specific altitude where radiation dose conversion coefficients are applied to convert the particle flux into effective and ambient dose-equivalent rates. A cut-off rigidity model accounts for the shielding effects of the Earth's magnetic field. Comparisons were made between the model predictions and actual flight measurements taken with various types of instruments used to measure the mixed radiation field during ground level enhancements (GLEs) 60 and 65. An anisotropy analysis that uses neutron monitor responses and the pitch angle distribution of energetic solar particles was used to identify particle anisotropy for a solar event in December 2006. In anticipation of future commercial use, a computer code has been developed to implement the radiation dose assessment model for routine analysis.

Use of a dual-labelled oligonucleotide as a DNA dosemeter for radiological exposure detection.

A reporter molecule consisting of a synthetic oligonucleotide is being characterised for a novel damage detection scenario for its potential use as a field-deployable, personal deoxyribonucleic acid (DNA) dosemeter for radiation detection. This dosemeter is devoid of any biological properties other than being naked DNA and therefore has no DNA repair capabilities. It supports biodosimetry techniques, which require lengthy analysis of cells from irradiated individuals, and improves upon inorganic dosimetry, thereby providing for a more relevant means of measuring the accumulated dose from a potentially mixed-radiation field. Radiation-induced single strand breaks (SSBs) within the DNA result in a quantifiable fluorescent signal. Proof of concept has been achieved over 250 mGy-10 Gy dose range in radiation fields from 6°Co, with similar results seen using a linear accelerator X-ray source. Further refinements to both the molecule and the exposure/detection platform are expected to lead to enhanced levels of detection for mixed-field radiological events.

Modelling of radiation exposure at high altitudes during solar storms.

A transport code analysis using Monte Carlo N-Particle eXtended code, MCNPX, has been used to propagate an extrapolated particle spectrum based on satellite measurements through the atmosphere to estimate radiation exposure during solar storms at high altitudes. Neutron monitor count rate data from stations around the world were used to benchmark the model calculations during a ground-level event (GLE). A comparison was made between the model predictions and actual flight measurements taken with various types of instruments used to measure the mixed radiation field during GLE 60. A computer code has been developed to implement the model for routine analysis.

Development of a predictive code for aircrew radiation exposure.

Using the empirical data measured by the Royal Military College with a tissue equivalent proportional counter, a model was derived to allow for the interpolation of the dose rate for any global position, altitude and date. Through integration of the dose-rate function over a great circle flight path or between various waypoints, a Predictive Code for Aircrew Radiation Exposure (PCAire) was further developed to provide an estimate of the total dose equivalent on any route worldwide at any period in the solar cycle.

Modelling of aircrew radiation exposure from galactic cosmic rays and solar particle events.

Correlations have been developed for implementation into the semi-empirical Predictive Code for Aircrew Radiation Exposure (PCAIRE) to account for effects of extremum conditions of solar modulation and low altitude based on transport code calculations. An improved solar modulation model, as proposed by NASA, has been further adopted to interpolate between the bounding correlations for solar modulation. The conversion ratio of effective dose to ambient dose equivalent, as applied to the PCAIRE calculation (based on measurements) for the legal regulation of aircrew exposure, was re-evaluated in this work to take into consideration new ICRP-92 radiation-weighting factors and different possible irradiation geometries of the source cosmic-radiation field. A computational analysis with Monte Carlo N-Particle eXtended Code was further used to estimate additional aircrew exposure that may result from sporadic solar energetic particle events considering real-time monitoring by the Geosynchronous Operational Environmental Satellite. These predictions were compared with the ambient dose equivalent rates measured on-board an aircraft and to count rate data observed at various ground-level neutron monitors.

Characterisation of bubble detectors for aircrew and space radiation exposure.

The Earth's atmosphere acts as a natural radiation shield which protects terrestrial dwellers from the radiation environment encountered in space. In general, the intensity of this radiation field increases with distance from the ground owing to a decrease in the amount of atmospheric shielding. Neutrons form an important component of the radiation field to which the aircrew and spacecrew are exposed. In light of this, the neutron-sensitive bubble detector may be ideal as a portable personal dosemeter at jet altitudes and in space. This paper describes the ground-based characterisation of the bubble detector and the application of the bubble detector for the measurement of aircrew and spacecrew radiation exposure.

Determination of natural and depleted uranium in urine at the ppt level: an interlaboratory analytical exercise.

An analytical exercise was initiated in order to determine those procedures with the capability to measure total uranium and uranium (238U/235U) isotopic ratios in urine samples containing >0.02 microg U kg-1 urine. A host laboratory prepared six identical sets of twelve synthetic urine samples containing total uranium in the range of 25 to 770 ng U kg-1 urine and with 238U/235U isotopic ratios ranging from 138 (100% NU) to 215 (51% DU). Sets of samples were shipped to five testing laboratories (four based in Canada and one based in Europe). Each laboratory utilized one of the following analytical techniques: sector field inductively coupled plasma mass spectrometry (ICP-SF-MS), quadrupole inductively coupled plasma mass spectrometry (ICP-Q-MS), thermal ionization mass spectrometry (TIMS), and instrumental/delayed neutron activation analysis (I/DNAA), in their analyses.

LET dependence of bubble detector response to heavy ions.

A series of experiments have been recently performed at the Heavy Ion Medical Accelerator in Chiba (HIMAC) laboratory to study the response of bubble detectors to high-mass high-energy (HZE) particles. The motivation for this study was to improve our ability to interpret measurements of neutron energy spectra in space. A recent analysis showed that emulsions of light halocarbons display common properties when they are characterised by a quantity called 'reduced superheat'. This quantity evolved from the examination of neutron and gamma responses of many types of detectors. In this study, we describe direct irradiations with N, Ar and Kr charged particles at HIMAC. It was observed that when the linear energy transfer (LET) corresponding to bubble formation was plotted vs. reduced superheat, different curves were obtained for a particular ion for detectors at different temperatures. Different curves were also obtained when data from different ions were plotted. These results confirm that bubble nucleation is not a simple function of particle LET and that an analysis based on track-structure appears warranted.

Bubble detector characterization for space radiation.

In light of the importance of the neutron contribution to the dose equivalent received by space workers in the near-Earth radiation environment, there is an increasing need for a personal dosimeter that is passive in nature and able to respond to this neutron field in real time. Recent Canadian technology has led to the development of a bubble detector, which is sensitive to neutrons, but insensitive to low linear energy transfer (LET) radiation. By changing the composition of the bubble detector fluid (or "superheat"), the detectors can be fabricated to respond to different types of radiation. This paper describes a preliminary ground-based research effort to better characterize the bubble detectors of different compositions at various charged-particle accelerator facilities, which are capable of simulating the space radiation field.

Estimated neutron dose to embryo and foetus during commercial flight.

A study has been carried out to assess the radiation exposure from cosmic-ray neutrons to the embryo and foetus of pregnant aircrew and air travellers in consideration of the radiation exposure from cosmic-ray neutrons to the embryo and foetus. A Monte Carlo analysis was performed to determine the equivalent dose from neutrons to the brain and body of an embryo at 8 weeks and to the foetus at the 3, 6 and 9 month periods. Neutron fluence-to-absorbed dose conversion coefficients for the foetal brain and for the entire foetal body (isotropic irradiation geometry) have been determined at the four developmental stages. The equivalent dose rate to the foetus during commercial flights has been further evaluated considering the fluence-to-absorbed dose conversion coefficients, a neutron spectrum measured at an altitude of 11.3 km and an ICRP-92 radiation-weighting factor for neutrons. This study indicates that the foetus can exceed the annual dose limit of 1 mSv for the general public after, for example, 15 round trips on commercial trans-Atlantic flights.

Aircrew dosimetry using the Predictive Code for Aircrew Radiation Exposure (PCAIRE).

During 2003, a portable instrument suite was used to conduct cosmic radiation measurements on 49 jet-altitude flights, which brings the total number of in-flight measurements by this research group to over 160 flights since 1999. From previous measurements, correlations have been developed to allow for the interpolation of the dose-equivalent rate for any global position, altitude and date. The result was a Predictive Code for Aircrew Radiation Exposure (PCAIRE), which has since been improved. This version of the PCAIRE has been validated against the integral route dose measurements made at commercial aircraft altitudes during the 49 flights. On most flights, the code gave predictions that agreed to the measured data (within +/- 25%), providing confidence in the use of PCAIRE to predict aircrew exposure to galactic cosmic radiation. An empirical correlation, based on ground-level neutron monitoring data, has also been developed for the estimation of aircrew exposure from solar energetic particle (SEP) events. This model has been used to determine the significance of SEP exposure on a theoretical jet altitude flight during GLE 42.

Assessment of aircrew radiation exposure by further measurements and model development.

A methodology is presented for collecting and analysing exposure measurements from galactic cosmic radiation using a portable equipment suite and encapsulating these data into a semi-empirical model/Predictive Code for Aircrew Radiation Exposure (PCAIRE) for the assessment of aircrew radiation exposure on any flight over the solar cycle. The PCAIRE code has been validated against integral route dose measurements at commercial aircraft altitudes during experimental flights made by various research groups over the past 5 y with code predictions typically within +/-20% of the measured data. An empirical correlation, based on ground-level neutron monitoring data, is detailed further for estimation of aircrew exposure from solar particle events. The semi-empirical models have been applied to predict the annual and career exposure of a flight crew member using actual flight roster data, accounting for contributions from galactic radiation and several solar energetic-particle events over the period 1973-2002.

Development of neutron radioscopy for the inspection of CF188 flight control surfaces.

Neutron radioscopy, using a cooled charged coupled device (CCD) camera and a neutron-sensitive scintillation screen, was developed at the SLOWPOKE-2 Facility at the Royal Military College (RMC) to detect water ingress into the composite layers and the aluminium honeycomb core in flight control surfaces on the CF188 Hornet aircraft. The response of the CCD camera system was tested at different neutron fluxes utilising the SLOWPOKE-2 at RMC and the Breazeale Nuclear Reactor at Pennsylvania State University.

Galactic and solar radiation exposure to aircrew during a solar cycle.

An on-going investigation using a tissue-equivalent proportional counter (TEPC) has been carried out to measure the ambient dose equivalent rate of the cosmic radiation exposure of aircrew during a solar cycle. A semi-empirical model has been derived from these data to allow for the interpolation of the dose rate for any global position. The model has been extended to an altitude of up to 32 km with further measurements made on board aircraft and several balloon flights. The effects of changing solar modulation during the solar cycle are characterised by correlating the dose rate data to different solar potential models. Through integration of the dose-rate function over a great circle flight path or between given waypoints, a Predictive Code for Aircrew Radiation Exposure (PCAIRE) has been further developed for estimation of the route dose from galactic cosmic radiation exposure. This estimate is provided in units of ambient dose equivalent as well as effective dose, based on E/H x (10) scaling functions as determined from transport code calculations with LUIN and FLUKA. This experimentally based treatment has also been compared with the CARI-6 and EPCARD codes that are derived solely from theoretical transport calculations. Using TEPC measurements taken aboard the International Space Station, ground based neutron monitoring, GOES satellite data and transport code analysis, an empirical model has been further proposed for estimation of aircrew exposure during solar particle events. This model has been compared to results obtained during recent solar flare events.

An examination of uranium levels in Canadian forces personnel who served in the Gulf War and Kosovo.

A uranium bioassay program was conducted involving 103 active and retired Canadian Forces personnel. The total uranium concentrations in each of two 24-h urine collections were analyzed separately at independent commercial laboratories by inductively coupled plasma mass spectrometry (ICP-MS) and by instrumental neutron activation analysis (INAA). The mean and median concentrations were determined to be 4.5 ng L(-1) and 2.8 ng L(-1), respectively, from ICP-MS and 17 ng L(-1) and 15 ng L(-1), respectively, from INAA. The total uranium concentrations were sufficiently low so that isotopic (238U:235U ratio) assays could not be performed directly from urine samples. Isotopic assays were performed on hair samples from 19 of the veterans participating in the testing. The isotopic hair assays were scattered around the natural 238U:235U ratio of 137.8, ranging from 122 +/- 21 to 145 +/- 16 (1sigma). Due to concern expressed in the media over possible depleted uranium exposure and long-term retention in bone, a single bone sample (vertebrate bone marrow) from a deceased member of the Canadian Forces was also analyzed for total uranium content and isotopic ratio by ICP-MS. The sample was shown to have 16.0 +/- 0.3 microg kg(-1) uranium by dry weight and a 238U:238U isotopic ratio of 138 +/- 4, consistent with natural uranium.

Aircrew exposure from cosmic radiation on commercial airline routes.

As a result of the recent recommendations of the ICRP 60, and in anticipation of possible regulation on occupational exposure of Canadian-based aircrew, an extensive study was carried out by the Royal Military College of Canada over a one-year period to measure the cosmic radiation at commercial jet altitudes. A tissue-equivalent proportional counter was used to measure the ambient total dose equivalent rate on 62 flight routes, resulting in over 20,000 data points at one-minute intervals at various altitudes and geomagnetic latitudes (i.e. which span the full cut-off rigidity of the Earth's magnetic field). These data were then compared to similar experimental work at the Physikalisch Technische Bundesanstalt, using a different suite of equipment, to measure separately the low and high linear energy transfer components of the mixed radiation field, and to predictions with the LUIN transport code. All experimental and theoretical results were in excellent agreement. From these data, a semiempirical model was developed to allow for the interpolation of the dose rate for any global position, altitude and date (i.e. heliocentric potential). Through integration of the dose rate function over a great circle flight path, a computer code was developed to provide an estimate of the total dose equivalent on any route worldwide at any period in the solar cycle.

Assessment of the cosmic radiation exposure on Canadian-based routes.

As a result of the recent recommendations of the ICRP-60 and in anticipation of possible regulation on occupational exposure of commercial aircrew, a two-phase investigation was carried out over a 1-y period to determine the total dose equivalent on representative Canadian-based flight routes. In the first phase of the study, dedicated scientific flights on a Northern round-trip route between Ottawa and Resolute Bay provided the opportunity to characterize the complex mixed-radiation field and to intercompare various instrumentation using both a conventional suite of powered detectors and passive dosimetry. In the second phase, volunteer aircrew carried (passive) neutron bubble detectors during their routine flight duties. From these measurements, the total dose equivalent was derived for a given route with a knowledge of the neutron fraction as determined from the scientific flights and computer code (CARI-3C) calculations. This study has yielded an extensive database of over 3,100 measurements providing the total dose equivalent for 385 different routes. By folding in flight frequency information and the accumulated flight hours, the annual occupational exposures of 20 flight crew have been determined. This study has indicated that most Canadian-based domestic and international aircrew will exceed the proposed annual ICRP-60 public limit of 1 mSv y(-1) but will be well below the occupational limit of 20 mSv y(-1).

Cosmic radiation exposure on Canadian-based commercial airline routes.

As a result of the recent recommendations of ICRP 60 and in anticipation of possible regulation on occupational exposure of commercial aircrew, a two-part investigation was carried out over a one-year period to determine the total dose equivalent on representative Canadian-based flight routes. As part of the study, a dedicated scientific measurement flight (using both a conventional suite of powered detectors and passive dosimetry) was used to characterise the complex mixed radiation field and to intercompare the various instrumentation. In the other part of the study, volunteer aircrew carried (passive) neutron bubble detectors during their routine flight duties. From these measurements, the total dose equivalent was derived for a given route with a knowledge of the neutron fraction as determined from the scientific flight and computer code (CARI-LF) calculations. This investigation has yielded an extensive database of over 3100 measurements providing the total dose equivalent for 385 different routes. By folding in flight frequency information and the accumulated flight hours, the annual occupational exposures of 26 flight crew have also been determined. This study has indicated that most Canadian-based domestic and international aircrew will exceed the proposed annual ICRP 60 public limit of 1 mSv.y-1, but will he well below the occupational limit of 20 mSv.y-1.