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Biomedical subjects

M Folkard

Publications and source records attributed to M Folkard.

At least 37 records · Page 2Linked to original sources
PubMed2000

Critical energies for SSB and DSB induction in plasmid DNA by low-energy photons: action spectra for strand-break induction in plasmid DNA irradiated in vacuum.

PURPOSE: To measure action spectra for the induction of single-strand breaks (SSB) and double-strand breaks (DSB) in plasmid DNA by low-energy photons and provide estimates for the energy dependence of strand-break formation important for track-structure simulations of DNA damage. MATERIALS AND METHODS: Plasmid pMSG-CAT was irradiated as a monolayer, under vacuum, with 7 150eV photons produced by a synchrotron source. Yields of SSB and DSB were determined by the separation of the three plasmid forms by gel electrophoresis. RESULTS: The yields of SSB per incident photon increased from 1.4x 10(-15) SSB per plasmid per photon/cm2 at 7eV to 7.5 x 10(-14) SSB per plasmid per photon/cm2 at 150 eV. Direct induction of DSB was also detected increasing from 3.4 x 10(-17) DSB per plasmid per photon/cm2 at 7eV to 4.1 x 10(-15) DSB per plasmid per photon/cm2 at 150eV. When the absorption cross-section of the DNA was considered, the quantum efficiency for break formation increased over the energy range studied. Over the entire energy range, the ratio of SSB to DSB remained constant. CONCLUSIONS: These studies provide evidence for the ability of photons as low as 7 eV to induce both SSB and DSB. The common action spectrum for both lesions suggests that they derive from the same initial photoproducts under conditions where the DNA is irradiated in vacuum and a predominantly direct effect is being observed. The spectral and dose-effect behaviour indicates that DSB are induced predominantly by single-event processes in the energy range covered.

DNA↗
PubMed1998

Studies of bystander effects in human fibroblasts using a charged particle microbeam.

PURPOSE: To determine the role of cell-to-cell mediated effects in the response of cells to different radiations by using a charged particle microbeam capable of targeting individual cells with counted particles. MATERIALS AND METHODS: Primary human fibroblasts were irradiated conventionally with X-rays or alpha-particles and scored for the induction of micronucleated or apoptotic cell formation at various times after irradiation. Cells were also individually irradiated with helium-3 particles from a microbeam and the distribution of damaged cells between hit and non-hit cells scored. RESULTS: Conventionally exposed fibroblasts showed a dose-dependent production of micronucleated and apoptotic cells 3 days after irradiation for both X-rays and alpha-particles, with a high RBE value for alpha-particles at low doses. Targeting individual alpha-particles, using a microbeam, to four cells within a population produced more micronucleated and apoptotic cells than expected on the basis of a direct effect only. CONCLUSIONS: This study provides direct evidence for the production of transmissible, cell-to-cell effects between hit and non-hit cells individually exposed to charged particles.

Alpha Particles↗
PubMed1998

A study of DNA fragmentation patterns in cells irradiated with charged particles: evidence for non-random distributions.

Many studies have shown that the effectiveness of radiations of varying LET is similar when yields of dsb have been measured, despite large differences in biological response. Recent evidence has suggested however, that current techniques underestimate the yields of dsb. By monitoring the fragmentation of DNA over a wide range of fragment sizes (< 10 kbp > 6 Mbp) by pulsed field electrophoresis, RBE values greater than 1.0 for radiations of around 100 keV/mm have been determined. The data provide evidence for the production of correlated breaks produced within cells as particle tracks traverse the nucleus. The highly ordered structure of DNA within mammalian cells may lead to clustering of breaks over distances related to the repeating unit structures of the chromatin. As well as these regionally damaged sites, a major contributor to radiation effectiveness will be the localised clustering of damage in the 1-20 bp region. A major effort is required to elucidate the relative importance of these levels of clustering and their importance in biological response.

Alpha Particles↗
PubMed1997

A charged-particle microbeam: I. Development of an experimental system for targeting cells individually with counted particles.

Charged-particle microbeams provide a unique opportunity to control precisely, the dose to individual cells and the localization of dose within the cell. The Gray Laboratory is now routinely operating a charged-particle microbeam capable of delivering targeted and counted particles to individual cells, at a dose-rate sufficient to permit a number of single-cell assays of radiation damage to be implemented. By this means, it is possible to study a number of important radiobiological processes in ways that cannot be achieved using conventional methods. This report describes the rationale, development and current capabilities of the Gray Laboratory microbeam.

DNA↗
PubMed1997

A charged-particle microbeam: II. A single-particle micro-collimation and detection system.

The use of a charged-particle microbeam provides a unique opportunity to control precisely, the number of particles traversing individual cells and the localization of dose within the cell. The accuracy of 'aiming' and of delivering a precise number of particles crucially depends on the design and implementation of the collimation and detection system. This report describes the methods available for collimating and detecting energetic particles in the context of a radiobiological microbeam. The arrangement developed at the Gray Laboratory uses either a 'V'-groove or a thick-walled glass capillary to achieve 2-5 microns spatial resolution. The particle detection system uses an 18 microns thick transmission scintillator and photomultiplier tube to detect particles with > 99% efficiency.

Particle Accelerators↗
PubMed1997

DNA double-strand break distributions in X-ray and alpha-particle irradiated V79 cells: evidence for non-random breakage.

Many studies have shown that with increasing LET of ionizing radiation the RBE (relative biological effectiveness) for dsb (double strand breaks) induction remains around 1.0 despite the increase in the RBE for cell killing. This has been attributed to an increase in the complexity of lesions, classified as dsb with current techniques, at multiply damaged sites. This study determines the molecular weight distributions of DNA from Chinese hamster V79 cells irradiated with X-rays or 110 keV/micron alpha-particles. Two running conditions for pulsed-field gel-electrophoresis were chosen to give optimal separation of fragments either in the 225 kbp-5.7 Mbp range or the 0.3 kbp to 225 kbp range. Taking the total fraction of DNA migrating into the gel as a measure of fragmentation, the RBE for dsb induction was less than 1.0 for both molecular weight regions studied. The total yields of dsb were 8.2 x 10(-9) dsb/Gy/bp for X-rays and 7.8 x 10(-9) dsb/Gy/bp for alpha-particles, measured using a random breakage model. Analysis of the RBE of alpha-particles versus molecular weight gave a different response. In the 0.4 Mbp-5.7 Mbp region the RBE was less than 1.0; however, below 0.4 Mbp the RBE increased above 1.0. The frequency distributions of fragment sizes were found to differ from those predicted by a model assuming random breakage along the length of the DNA and the differences were greater for alpha-particles than for X-rays. An excess of fragments induced by a single-hit mechanism was found in the 8-300 kbp region and for X-rays and alpha-particles these corresponded to an extra 0.8 x 10(-9) and 3.4 x 10(-9) dsb/bp/Gy, respectively. Thus for every alpha-particle track that induces a dsb there is a 44% probability of inducing a second break within 300 kbp and for electron tracks the probability is 10%. This study shows that the distribution of damage from a high LET alpha-particle track is significantly different from that observed with low LET X-rays. In particular, it suggests that the fragmentation patterns of irradiated DNA may be related to the higher-order chromatin repeating structures found in intact cells.

Alpha Particles↗
PubMed1996

Inactivation of V79 cells by low-energy protons, deuterons and helium-3 ions.

Previous work by ourselves and by others has demonstrated that protons with a linear energy transfer (LET) about 30 keVmum(-1)are more effective at killing cells than doubly charged particles of the same LET. In this work we show that by using deuterons, which have about twice the range of protons with the same LET, it is possible to extend measurements of the RBE of singly charged particles to higher LET (up to 50 keVmum(-1). We report the design and use of a new arrangement for irradiating V79 mammalian cells. Cell survival measurements have been made using protons in the energy range 1.0-3.7 MeV, deuterons in the energy range 0.9-3.4MeV and 3He2+ ions in the energy range 3.4-6.9 MeV. This corresponds to volume-averaged LET (within the cell nucleus) between 10 and 28 keVmum(-1) for protons, 18-50 keVmum(-1) for deuterons, and 59-106 keVmum(-1) for helium ions. Our results show no difference in the effectiveness of protons and deuterons matched for LET. However, for LET above about 30 keVmum(-1) singly charged particles are more effective at inactivating cells than doubly-charged particles of the same LET and that this difference can be understood in terms of the radial dose distribution around the primary ion track.

Animals↗
PubMed1994

Effect of radiation quality on lesion complexity in cellular DNA.

Understanding the critical lesions induced by ionizing radiation in DNA and their relationship to cellular effects is an important challenge in radiation biology. Much evidence has suggested that DNA double-strand breaks (dsb) are important lesions. Establishing a cause and effect relationship between initial levels of DNA dsb, their repair rate or the level of residual unrepaired breaks, and cellular effects has proved difficult in mammalian cells. Several studies have measured yields of DNA dsb after irradiation with radiations of differing linear energy transfer (LET). In general the RBEs for dsb induction (20-100 keV/microns) have been lower than the RBEs measured for cell survival and in many cases are around 1.0. Several studies have shown differences in the rejoining of dsb with less dsb rejoined after high-LET irradiation in comparison with low-LET radiation. These results suggest that there may be differences in the types of lesions induced by different radiations and scored as DNA dsb using current techniques. Track structure modelling studies have suggested that some lesions induced will be clustered at the sites of energy depositions and that uniquely large energy deposition events are produced by high-LET radiations. Assays need to be developed to measure complex lesions in both model DNA and cellular systems. Different levels of complexity need to be considered such as clustering of radicals close to DNA, localized areas of DNA damage (1-20 bp) and lesions which may be induced over larger distances. Studies using new and existing assays of DNA damage, coupled with irradiation at various LETs, are directed at understanding the role of lesion complexity in relation to cellular effects.

DNA↗
PubMed1994

Action spectra for single- and double-strand break induction in plasmid DNA: studies using synchrotron radiation.

Ionizing radiations deposit a wide range of energies in and around DNA and this leads to a corresponding spectrum of complexity of the lesions induced. The relationships between the amount of energy deposited and the yields and types of damage induced are important in modelling the physical and chemical stages of radiation effect and linking them to biological outcome. To study these relationships experimentally, plasmids were mounted as a monolayer and exposed in vacuum to near-monoenergetic photons from the Daresbury Synchrotron. After irradiation, the DNA was washed off and assayed for single-(ssb) and double-strand breaks (dsb) using agarose gel electrophoresis. Dose-effect relationships for ssb and dsb induction were obtained at various energies in the range 8-25 eV. The initial responses in the low-dose region allowed damage yields to be estimated. However, a common feature is that the responses showed energy-dependent plateaus at higher doses as if a fraction of the DNA were shielded. Various measures were taken both to minimize and to correct for this effect. The data appear to show that the yields of ssb and dsb increase only slowly with photon energies > 10 eV, with a suggestion of similar threshold energies for both lesions. In the energy range covered, the yield of ssb is 12-20-fold greater than that of dsb. The data indicate that ssb and dsb may have a common precursor in this system. Earlier work with low-energy electrons showed that at 25 eV ssb were induced but no dsb were detected.

DNA↗
PubMed1994

Quantification of radiation-induced hydroxyl radicals within nucleohistones using a molecular fluorescent probe.

We present a method that specifically records .OH formation within histones and possibly at other sites in irradiated nucleohistone. The approach uses the radiation-induced fluorescence emissions from a chromatin-conjugated .OH detector, SECCA (a succinylated derivative of coumarin), that is converted to a fluorescent derivative, 7-hydroxy-SECCA (7-OH-SECCA), after interaction with .OH in neutral aqueous solutions. It is shown that (a) the fluorescent product 7-OH-SECCA cannot be generated by direct radiation effects after gamma or neutron irradiation of SECCA; (b) when SECCA-labeled histone is complexed with DNA to form nucleohistone, the physical properties of the modified nucleohistone are similar to those of unlabeled nucleoprotein; and (c) after irradiation of SECCA-labeled nucleohistone, a linear induction of the fluorescence signal is observed within the radiation doses examined (0.3-30 Gy). Since the sample remains available for further studies after registration of the optical signal, the current approach should permit the investigator to correlate in a single sample the localization and frequency of .OH formation with the results of other assays.

Animals↗
PubMed1993

Measurement of DNA damage by electrons with energies between 25 and 4000 eV.

All ionizing radiations deposit energy stochastically along their tracks. The resulting distribution of energies deposited in a small target such as the DNA helix leads to a corresponding spectrum in the severity of damage produced. So far, most information about the probable spectra of DNA lesion complexity has come from Monte Carlo studies which endeavour to model the relationship between the energy deposited in DNA and the damage induced. The aim of this paper is to establish methods of determining this relationship by irradiating pBR322 plasmid DNA using low energy electrons with energies comparable with the minimum energy thought to produce critical damage. The technique of agarose gel electrophoresis has been used to ascertain the fraction of DNA single- and double-strand breaks induced by monoenergetic electrons with energies as low as 25 eV. Our data show that the threshold electron energy for induction of single-strand breaks is < 25 eV, and for double-strand breaks between 25 and 50 eV.

DNA↗
PubMed1990

The irradiation of V79 mammalian cells by protons with energies below 2 MeV. Part II. Measurement of oxygen enhancement ratios and DNA damage.

The effectiveness of low-energy (below 2 MeV) protons at inducing DNA damage in the form of single- and double-strand breaks has been determined. Protons with mean energies of 1.90 MeV, 1.15 MeV and 0.76 MeV corresponding to track average LETs of 17 keV/microns, 24 keV/microns and 32 keV/microns, respectively, have been used and compared with 250 kVp X-rays and 3.8 MeV 238Pu alpha-particles. Although there was variation in the RBE for DNA ssb induction with LET, the RBEs for dsb induction at all three proton energies and for 3.8 MeV alpha-particles were all around 1.0. This suggests that, if DNA dsb are important in radiation-induced cell lethality, the probability of an induced dsb leading to a lethal event increases with increasing LET of radiation. Oxygen enhancement ratios were measured for both cell survival and DNA dsb induction, and in both cases a decrease in OER with increasing LET was observed.

Animals↗
PubMed1989

The irradiation of V79 mammalian cells by protons with energies below 2 MeV. Part I: Experimental arrangement and measurements of cell survival.

The relative biological effectiveness (RBE) has been determined for protons with mean energies of 1.9, 1.15 and 0.76 MeV, from measurements of the survival of V79 Chinese hamster cells. The cells are supported as a monolayer and are swept through a beam of scattered protons produced using a 4 MeV Van de Graaff accelerator. An estimation of the dose and unrestricted linear energy transfer (LET) variation within the sensitive volume of the cells is given for the three proton energies. The RBEs for cell survival (relative to 250 kVp X-rays) at the 10 per cent survival level are 1.6, 1.9 and 3.36 for protons with track-average LETs of 17, 24 and 32 keV microns-1 respectively, and the data suggest that protons are most effective at about 40-50 keV microns-1. It is shown that the proton RBEs can be reconciled with those of other light ions if plotted against z*2/beta 2 (where z* is the effective charge and beta is the relative velocity) rather than against LET.

Animals↗
PubMed1989

Measurement and analysis of supralinearity in LiF TLD-100 irradiated by 1.5 keV x-rays.

Measurements of the response of lithium fluoride TLD-100 to 1.5 keV ultrasoft x-rays, 240 keV x-rays and 137Cs gamma-rays at doses below 70 Gy are presented and the implications for the track interaction model are discussed. This model proposes that thermoluminescence is due to the combination electrons and holes formed along secondary charged particle tracks. As the dose is increased, the average distance between particle tracks is reduced and there is a greater probability of combination between neighbouring tracks; this mechanism is used to explain the supralinear response of lithium fluoride to certain radiations. Measurements using ultrasoft x-rays are interesting in this context because the energy from each photon is deposited in a small volume, therefore the probability of combination between neighbouring events should only be significant at high doses. The results, however, show that lithium fluoride exhibits a significant supralinear response to 1.5 keV x-rays after only a few tens of grays and suggests that the track interaction model alone cannot explain the phenomenon of supralinearity.

Cesium Radioisotopes↗
PubMed1989

Measurements of neutron energy using a recoil-proton telescope and a high-pressure ionization chamber.

Two very different techniques for measuring the energy of neutrons in the energy range 0.1-10 MeV are presented and compared. A recoil-proton spectrometer is used to determine the energy spectra of neutrons produced by the d(4)-Be and p(4)-Be reactions down to the low-energy threshold of 0.7 MeV. The same radiation fields are also measured with a recently developed method using a high-pressure ionization chamber that can be used to determine the mean energy of the neutrons in a mixed neutron-gamma radiation field provided the gamma-ray absorbed dose fraction is determined independently. An intercomparison of the two methods shows that the high-pressure ionization chamber compares well and supplements the established recoil-proton spectrometer technique. The almost isotropic response of the chamber has enabled measurements to be made of the variation of mean neutron energy with depth in water for the two radiation fields.

Fast Neutrons↗
PubMed1989

Measurement of DNA damage and cell killing in Chinese hamster V79 cells irradiated with aluminum characteristic ultrasoft X rays.

Chinese hamster V79 cells were irradiated with 1.487 keV aluminum characteristic X rays produced using a cold-cathode discharge tube. Under aerobic conditions a relative biological effectiveness (RBE) of 2.18 for cell killing in comparison to 250-kVp X rays was measured using cells grown in suspension and irradiated on membrane filters. DNA damage in the form of single-strand (ssb) and double-strand breaks (dsb) was measured using the filter elution technique. The aerobic RBEs are 1.64 for dsb induction and 0.49 for ssb induction, consistent with the view that dsb are more closely related to cell kill than ssb. A reduced oxygen enhancement ratio (OER) for cell killing was measured for Al-K X rays, but the OER for dsb induction was similar to that measured for 250-kVp X rays. A curvilinear relationship between dsb induction and dose is observed, similar to that seen for 250-kVp X rays. This agrees with the concept that ultrasoft X rays produce critical lesions similar to hard X rays but with a greater efficiency per unit dose.

Animals↗
PubMed1988

Interaction between X-ray and alpha-particle damage in V79 cells.

V79 cells have been exposed to X-rays or 238Pu alpha-particles or to X-rays following priming alpha-particle doses of 0.5, 2 or 2.5 Gy. The survival curve for exposure to alpha-particles was exponential with a D0 of 0.89 Gy. Following exposure to priming alpha-particle doses the resulting X-ray survival curves had the same slope as the single dose X-ray curve, but a reduced shoulder. For alpha-particle priming doses of 0.5 and 2 Gy this reduction was the same as for the same X-ray doses. 2.5 Gy alpha-particles reduced the subsequent X-ray curve Dq to almost zero. alpha-particles do cause damage capable of interacting with X-ray damage.

Alpha Particles↗