Biomedical subjects

D Frankenberg

Publications and source records attributed to D Frankenberg.

At least 19 recordsLinked to original sources

PubMed2006

The PTB microbeam: a versatile instrument for radiobiological research.

The PTB microbeam is routinely used for the irradiation of living cells using protons (1-20 MeV) and alpha particles (1-28 MeV). The beam diameter is approximately 2 microm (fwhm), achieved by focussing, resulting in an excellent energy resolution and practically no scattered particles. Recently, an electrostatic beam scanner was added to the facility which allows targeting of each cell within 1 ms. This and other improvements led to an increase in the experimental speed of the system to a maximum of 50,000 cells per hour including all experimental steps. To improve the versatility of the facility further, a module for automatic quantification of immunocytochemical staining was implemented. This allows the analysis of protein activation, taking into account the positional information of the irradiation run.

Cell Culture Techniques↗

PubMed2006

Neoplastic transformation of a human hybrid cell line by alpha particles in relation to mammography X rays.

The hybrid cell line CGL1 is the only cell line which allows quantitation of neoplastic transformation in human cells. Hybrid cells were exposed to 3.4 MeV alpha particles or to mammography X rays (29 kV(p)) and both survival and neoplastic cell transformation were assayed. For comparison, previously published data obtained with 200 kV(p) X rays are also included. Alpha particles were significantly more efficient than 29 kV(p) X rays in cell killing. Surprisingly, the efficiency of mammography X rays for the induction of neoplastic cell transformation was close to that observed for alpha particles.

Alpha Particles↗

PubMed2002

Mutation induction and neoplastic transformation in human and human-hamster hybrid cells: dependence on photon energy and modulation in the low-dose range.

Mutation induction in the HPRT gene of human fibroblasts after irradiation with mammography-like 29 kVp or 200 kVp x-rays shows radiohypersensitivity for doses smaller than approximately 0.5 Gy. Similarly, mutation induction in the CD 59 gene on human chromosome 11 in A(L) cells shows radiohypersensitivity for doses smaller than approximately 0.5 Gy after exposure to 200 kVp x-rays, but not after irradiation with low-filtered 30 kVp x-rays. The RBE values of 29 and 30 kVp x-rays relative to 200 kVp x-rays are strongly dose dependent. For neoplastic transformation of human hybrid (CGL1) cells after irradiation with 29 or 200 kVp x-rays or 60Co gamma rays a linear-quadratic dose relationship was observed with RBE values of approximately four and eight for mammography relative to 200 kVp x-rays and 60Co gamma rays, respectively.

Animals↗

PubMed2002

Enhanced mutation and neoplastic transformation in human cells by 29 kVp relative to 200 kVp X rays indicating a strong dependence of RBE on photon energy.

The fundamental assumption implicit in the use of the atomic bomb survivor data to derive risk estimates for occupational and medical exposures is that the gamma rays of Hiroshima and Nagasaki are considered as equal efficiencies to other low LET radiations up to an LET of 10 keV.micron-1. For breast cancer induction, neoplastic cell transformation, mutation, reciprocal translocations and dicentrics in human lymphocytes, a strong and very similar dependence of the RBE values on photon energy or on LET is observed. Experimental data on mutation induction and neoplastic cell transformation in human cells show that 29 kVp X rays are by a factor of 4 and 3.4, respectively, more effective compared with 200 kVp X rays. These data are in excellent agreement with the data in the literature.

Cell Transformation, Neoplastic↗

PubMed2002

Enhanced neoplastic transformation by mammography X rays relative to 200 kVp X rays: indication for a strong dependence on photon energy of the RBE(M) for various end points.

The fundamental assumption implicit in the use of the atomic bomb survivor data to derive risk estimates is that the gamma rays of Hiroshima and Nagasaki are considered to have biological efficiencies equal to those of other low-LET radiations up to 10 keV/microm, including mammography X rays. Microdosimetric and radiobiological data contradict this assumption. It is therefore of scientific and public interest to evaluate the efficiency of mammography X rays (25-30 kVp) to induce cancer. In this study, the efficiency of mammography X rays relative to 200 kVp X rays to induce neoplastic cell transformation was evaluated using cells of a human hybrid cell line (CGL1). For both radiations, a linear-quadratic dose-effect relationship was observed for neoplastic transformation of CGL1 cells; there was a strong linear component for the 29 kVp X rays. The RBE(M) of mammography X rays relative to 200 kVp X rays was determined to be about 4 for doses < or = 0.5 Gy. A comparison of the electron fluences for both X rays provides strong evidence that electrons with energies of < or = 15 keV can induce neoplastic transformation of CGL1 cells. Both the data available in the literature and the results of the present study strongly suggest an increase of RBE(M) for carcinogenesis in animals, neoplastic cell transformation, and clastogenic effects with decreasing photon energy or increasing LET to an RBE(M) approximately 8 for mammography X rays relative to 60Co gamma rays.

Animals↗

PubMed2000

Induction of DNA double-strand breaks in mammalian cells and yeast.

Induction of DNA double-strand breaks (dsb) and their distribution are dependent on the energy deposition pattern within the cell nucleus (physical structure) and the ultrastructure of the chromosomes and its variation by the cell cycle and gene activities (biological structure). For electron radiation very similar RBE-values are observed for mammalian and yeast cells (AlK, 1.5 keV, 15 keV/micrometer: 2.6 in mammalian cells and 2.2 in yeast; CK 0.278 keV, 23 keV/micrometer: approx. 2.5 in mammalian cells and 3.8 in yeast). In contrast, the RBE-values for the induction of dsb of 4He2+ and light ions in the LET range from about 100 keV/micrometer up to 1000 keV/micrometer are significantly higher for yeast cells compared to mammalian cells. For example, the RBE-value of alpha-particles (120 keV/micrometer) is about 1.2 for mammalian cells whereas for yeast the RBE-value is about 2.5. The yeast chromatin has less condensed fibres compared with mammalian cells. Since a single CK photoelectron can induce only one dsb, the different condensation of the mammalian and yeast chromatin has no influence. However, particles may induce more than one dsb when traversing a chromatin fibre. The probability for the induction of closely neighboured dsb is higher the more condensed the chromatin fibres are. Since small DNA fragments (50 bp up to several kbp) are lost by standard methods of lysis, the underestimation of dsb yields increases with fibre condensation, which is in accordance with the observes dsb yields in mammalian cells and yeast. In order to obtain relevant yields of dsb (and corresponding RBE-values) the measurement of all DNA fragments down to about 50 bp are needed.

DNA Damage↗

PubMed1999

Induction of DNA double-strand breaks by 1H and 4He lons in primary human skin fibroblasts in the LET range of 8 to 124 keV/microm.

Yields of DNA double-strand breaks were determined in primary human skin fibroblasts exposed to 1H and 4He ions at various linear energy transfers (LETs) and to 15 MeV electrons as the reference radiation. The values obtained for the relative biological effectiveness (RBE) were 2.03, 1.45 and 1.36 for 1H ions at LETs of 35, 23 and 7.9 keV/microm, respectively, and 1.2, 1.18, 1.38 and 1.31 for 4He ions at LETs of 124, 76, 35 and 27 keV/microm, respectively. The data were obtained using pulsed-field gel electrophoresis of DNA released from cells using the chromosomes of the yeast Saccharomyces cerevisiae as length markers and fitting the experimental mass distributions of fragmented DNA to those obtained by computer simulation of the random breakage of human chromosomes. The RBE values for induction of DSBs in mammalian cells cannot be fitted to a common RBE-LET relationship for electrons and 1H, 4He and light ions. Comparison of the RBEs for mammalian cells with the corresponding RBEs obtained for yeast cells shows similar RBEs of electrons for yeast and mammalian cells; however, for 4He and light ions in the LET range of 100 to 1000 keV/microm, the RBEs for yeast are significantly higher compared with mammalian cells. These characteristics of the RBE-LET relationships for yeast and mammalian cells are attributed to the fraction of small DNA fragments induced by particles when traversing the higher-order chromatin structures which are different to some extent in these two cell types.

DNA↗

PubMed1999

Chromosome aberrations induced in human lymphocytes by 3.45 MeV alpha particles analyzed by premature chromosome condensation.

Premature chromosome condensation (PCC) experiments using human lymphocytes with centromere staining have shown that after exposure to 3.45 MeV alpha-particle radiation, the full number of dicentric chromosomes appears when the cell fusion protocol is applied immediately after irradiation. In this case, the time available for repair and misrepair of DNA damage is only about 30 min. The number of dicentrics does not change with a further increase in the time available for chromatin rearrangement. This fast response confirms the expectation based on our previous experiments using PCC with 150 kV X rays in which the alpha component of the yield of dicentrics was found to appear when the cell fusion protocol was applied immediately after irradiation, whereas the beta component was delayed by several hours. The time constant for rejoining of the excess acentric chromosome fragments is found to be donor-specific and not to differ for alpha particles and X rays, but alpha-particle radiation leaves a larger fraction of the excess acentric fragments unrejoined. The RBEs of the 3.45 MeV alpha-particle radiation compared to 150 kV X rays, evaluated for the alpha component for the yield of dicentrics and for the yield of unrepaired acentric fragments, have almost equal values of about 4. This is consistent with data in the literature on chromosome aberrations observed in metaphase that show the equality of the RBE values for production of dicentrics and acentric fragments. Our experimental results concerning the fast kinetics of the alpha component of the yield of exchange-type chromosome aberrations are not consistent with Lea's pairwise lesion interaction model, and they support the proposed alternative mechanism of lesion-nonlesion interaction between chromatin regions carrying clustered DNA damage and intact chromatin regions.

Alpha Particles↗

PubMed1998

Radiation rendered more cytotoxic by fludarabine monophosphate in a human oropharynx carcinoma cell-line than in fetal lung fibroblasts.

PURPOSE: Fludarabine monophosphate (fludarabine-P) is a relatively new drug in the treatment of different haematological diseases. The mechanism of action also implies a possible role of this drug as a radiosensitizer. Up to now no in vitro investigations dealing with radiosensitizing effects of fludarabine-P in carcinoma cell lines and fibroblasts have been published. The aim of our studies was to analyse the cytotoxic and radiosensitizing effects of different dosages and application schedules of fludarabine-P in a human squamous carcinoma cell line of the oropharynx (ZMK-1) and of fetal lung fibroblasts (MRC-5) in vitro. Possible mechanisms of interaction of fludarabine-P and radiation were investigated. METHODS: ZMK-1 and MRC-5 cells were cultured under standard conditions with different concentrations of fludarabine-P in combination with escalating doses of radiation. Cytotoxic effects were measured by colony-forming assays. Induction and rejoining of radiation-induced DNA double-strand breaks after incubation with fludarabine-P were measured using constant-field gel electrophoresis. Incubation times for rejoining varied from 0 h to 24 h. RESULTS: Fludarabine-P showed a radiosensitizing activity in ZMK-1 tumour cells and MRC-5 fibroblasts. The observed effects depended on the concentration and the incubation time. The largest effect was demonstrable for an incubation of 5 days, which started shortly before irradiation, whereas an incubation solely before irradiation did not have a clear effect on the cellular survival. The sensitizer enhancement ratio, at the 10% survival level, in the ZMK-1 cells was 2.2 in comparison to 1.6 in MRC-5 cells. The analysis of the interaction of fludarabine-P and ionising radiation by means of the isobologram approach, revealed an overadditive effect in the tumour cell line and an additive effect in the lung fibroblasts. Fludarabine-P did not modify the rejoining of radiation-induced DNA double-strand breaks in either cell line. CONCLUSIONS: We conclude that fludarabine-P in clinically attainable doses is a strong radiosensitizer in ZMK-1 cells and has a lower activity in the MRC-5 fibroblasts in vitro. The radiosensitization of fludarabine-P seems to be over additive in the malignant cells and additive in normal fetal fibroblasts. This would indicate that fludarabine-P might enhance the therapeutic ratio of radiation. Further investigations are warranted to identify the potential of this drug as a radiosensitizer in vivo and to elucidate the mechanism of interaction of the drug and radiation.

Carcinoma, Squamous Cell↗

PubMed1998

A review of dsb induction data for varying quality radiations.

PURPOSE: This short review summarizes the data obtained with various techniques for measuring the yields of double strand breaks (dsb) produced by particle radiations of differing linear energy transfer (LET) in order to obtain relative biological effectiveness (RBE) values. RESULTS AND CONCLUSIONS: Studies aimed at understanding the interactions of different types of radiation with cellular DNA have monitored the yields of DNA dsb versus radiation quality. Several techniques have been used to measure dsb yields in mammalian cells, and these include: neutral sedimentation gradients, filter elution and more recently pulsed field gel electrophoresis techniques (PFGE). Recent developments in PFGE have allowed the measurement of both the yields and the distribution of breaks within the genome, which go part of the way to explaining the RBE values close to 1.0 previously measured using other approaches with various radiation qualities. It is clear that future studies to determine the effectiveness of radiations of differing LET must use techniques that determine both yields and distributions of dsb, and assays need to be developed to allow these measurements at biologically relevant doses.

Alkalies↗

PubMed1998

Transformation of C3H 10T1/2 cells by low doses of ionising radiation: a collaborative study by six European laboratories strongly supporting a linear dose-response relationship.

For the assessment of radiation risk at low doses, it is presumed that the shape of the low-dose-response curve in humans for cancer induction is linear. Epidemiological data alone are unlikely to ever have the statistical power needed to confirm this assumption. Another approach is to use oncogenic transformation in vitro as a surrogate for carcinogenesis in vivo. In mid-1990, six European laboratories initiated such an approach using C3H 10T1/2 mouse cells. Rigid standardisation procedures were established followed by collaborative measurements of transformation down to absorbed doses of 0.25 Gy of x-radiation resulting in a total of 759 transformed foci. The results clearly support a linear dose-response relationship for cell transformation in vitro with no evidence for a threshold dose or for an enhanced, supralinear response at doses approximately 200-300 mGy. For radiological protection this represents a large dose, and the limitations of this approach are apparent. Only by understanding the fundamental mechanisms involved in radiation carcinogenesis will further knowledge concerning the effects of low doses become available. These results will, however, help validate new biologically based models of radiation cancer risk thus providing increased confidence in the estimation of cancer risk at low doses.

Animals↗

PubMed1996

Two types of double-strand breaks in electron and photon tracks and their relation to exchange-type chromosome aberrations.

Yields of DNA double-strand breaks (dsb), i.e. the average number of dsb, N, per relative molar mass, M(r), and dose, D, produced by electrons and photons in the energy range 50 eV-1 MeV were calculated. The experimental data of dsb induction by ultrasoft x-rays and by photons agree well with the calculated yields of dsb as a function of photon energy. The dsb are classified into simple and complex ones. Energy transfers of less than about 200 eV producing at least two ionizations generate mainly simple dsb, while low-energy electrons with an initial energy between 200 and 500 eV induce preferentially complex dsb. Assuming that dsb is the main DNA lesion leading to exchange-type chromosome aberrations (etca), three different mechanisms have to be considered: 1) complex dsb on its own; 2) interaction between two dsb induced by the same primary particle; and 3) interaction between two dsb induced by different primary particles. Mechanisms 1) and 2) produce a linear term, whereas mechanism 3) leads to a quadratic term for the yield of etca. The sum of contributions 1) and 2) to the yield of dicentrics describes fairly well the non-trivial structure of the experimental data. The results suggest that interaction between complex dsb does not contribute significantly to the formation of dicentrics via mechanism 3).

Chromosome Aberrations↗

PubMed1995

Influence of non-homology between recombining DNA sequences on double-strand break repair in Saccharomyces cerevisiae.

In this paper we study the influence of non-homology between plasmid and chromosomal DNA on the efficiency of recombinational repair of plasmid double-strand breaks and gaps in yeast. For this purpose we used different combinations of plasmids and yeast strains carrying various deletions within the yeast LYS2 gene. A 400 bp deletion in plasmid DNA had no effect on recombinational plasmid repair. However, a 400 bp deletion in chromosomal DNA dramatically reduced the efficiency of this repair mechanism, but recombinational repair of plasmids linearized by a double-strand break with cohesive ends still remained the dominant repair process. We have also studied the competition between recombination and ligation in the repair of linearized plasmids. Our experimental evidence suggests that recombinational repair is attempted but aborted if only one recombinogenic end with homology to chromosomal DNA is present in plasmid DNA. This situation results in a decreased probability of non-recombinational (i.e. ligation) repair of linearized plasmid DNA.

Chromosomes, Fungal↗

PubMed1995

Molecular mechanism of potentially lethal damage repair. I. Enhanced fidelity of DNA double-strand break rejoining under conditions allowing potentially lethal damage repair.

This study contributes to the elucidation of the molecular mechanism underlying potentially lethal damage (PLD) repair. Repair of DNA double-strand breaks (dsbs) is involved in PLD repair in yeast, i.e. in the enhanced survival of cells due to post-irradiation treatment under non-growth conditions before plating cells on nutrient agar (growth conditions). However, dsbs are rejoined when cells are kept either in non-growth or growth medium. One possibility to explain the enhanced survival of cells after post-irradiation treatment in non-growth medium might be an enhanced fidelity of dsb rejoining under non-growth relative to growth conditions. We have addressed this problem by using a plasmid-mediated assay. Into one of the two selectable plasmid markers a single dsb was introduced by a restriction enzyme. The cut plasmid was transfected into an appropriate yeast mutant. Transformants that had correctly rejoined the dsb were selected on the basis of restoration of the function of the cut gene. The yeast mutant was allowed to rejoin the cut plasmid under either non-growth or growth conditions. The results show that the fidelity of dsb rejoining is higher in cells kept under non-growth relative to growth conditions.

DNA Repair↗

PubMed1995

Different repair kinetics for short and long DNA double-strand gaps in Saccharomyces cervisiae.

The kinetics of recombinational repair of plasmid DNA double-strand breaks (dsb) and gaps (dsg) of different sizes and ends were studied. For this purpose we used the mutant rad54-3 of the yeast Saccharomyces cerevisiae, which is temperature dependent with respect to genetic recombination and rejoining of dsb/dsg, allowing us to stop these processes by shifting cells to the restrictive temperature. We found that the kinetics of repair of cohesive-ended dsb and small gaps (up to 400 bp) are similar and characterized by two phases separated by a plateau. In contrast, large gap (1.4 kbp) repair proceeds with different kinetics exhibiting only the second phase. We also investigated the repair kinetics of 400 bp gaps introduced into plasmid DNA with and without homology to chromosomal DNA allowing recombinational repair and non-recombinational repair (ligation), respectively. We found that gaps introduced in plasmid sequences homologous to chromosomal DNA are rapidly repaired by recombination. In contrast, recircularization of the gapped plasmid by ligation is as slow and inefficient as ligation of a cohesive-ended dsb. The kinetics of repair of gapped plasmids may be explained by assuming a constitutive level of enzymes responsible for the first phase of recombinational repair, while inducible enzymes, which become available at the end of the plateau, carry out the second phase of repair.

DNA Repair↗

PubMed1995

0.3 keV carbon K ultrasoft X-rays are four times more effective than gamma-rays when inducing oncogenic cell transformation at low doses.

Oncogenic transformation and inactivation were investigated in C3H10T1/2 mouse embryo fibroblasts exposed to proton-induced 0.28 keV carbon K (CK)-characteristic X-rays and 60Co gamma-rays as reference radiation at high dose-rate (2-3 and 0.7 Gy/min respectively). Both oncogenic cell transformation and cell inactivation followed a linear-quadratic relationship with dose. At low doses where the linear component dominates CK ultrasoft X-rays were more effective, by a factor of 4, at inducing oncogenic cell transformation and cell inactivation compared with 60Co gamma-rays. For both endpoints the RBE of CK ultrasoft X-rays gradually decreased with increasing dose mainly due to the greater quadratic component for 60Co gamma-rays compared with CK ultrasoft X-rays. Our experimental data are in agreement with the hypothesis that single DNA double-strand breaks (dsbs), which are induced by 0.28-keV ultrasoft CK X-rays, may lead to oncogenic cell transformation. With increasing absorbed dose, i.e. with decreasing mean distance between dsbs induced by 0.28-keV ultrasoft X-rays, oncogenic cell transformation and cell inactivation may also be induced by interaction between those dsbs.

Animals↗

PubMed1994

Evidence against the "oxygen-in-the-track" hypothesis as an explanation for the radiobiological low oxygen enhancement ratio at high linear energy transfer radiation.

Oxygen sensitizes cells toward the effect of ionizing radiation. This sensitization, quantified by the oxygen enhancement ratio (OER), decreases with increasing ionization density or linear energy transfer (LET) of the radiation applied. One explanation for the decreased OER at high LET offers the "oxygen-in-the-track" hypothesis. It claims that oxygen is produced in the track of densely ionizing particles providing an oxic microenvironment around the relevant cellular target molecules, even if cells are exposed under anoxic atmospheric conditions. Experimental evidence is presented against this hypothesis. It is based on the different kinetic pattern of DNA double-strand-break rejoining observed in yeast cells exposed under oxic or anoxic conditions to 3.5 MeV alpha-particles.

DNA↗

PubMed1994

Repair of DNA double-strand breaks and its effect on RBE.

DNA double-strand breaks (DSB) are induced linearly with absorbed dose both for sparsely and densely ionizing radiations. By enzymatic repair the linear relationship between the number of DSB and absorbed dose is converted into a non linear one. Furthermore, the RBE-values of high LET radiations for residual DSB increase with increasing amount of DSB repair especially in the low dose range. Unrepaired and/or misrepaired DSB are supposed to be responsible for chromosomal aberrations, cell killing, oncogenic cell transformation and gene mutation. At low doses, for these endpoints much higher RBE-values than those for initial DSB are observed. However, with increasing doses the RBE-values for these endpoints approach those for initial DSB. These observations are likely to be interpreted using the following two parameters of the energy deposition structure: 1. The distribution of clusters with respect to their size at the nm-scale and to the number of ionizations per cluster (cluster distribution). 2. The distribution of distances between clusters of definite size and with definite number of ionizations (distance distribution of clusters). For the induction of DSB solely the ionization density in clusters of nm-dimensions (i.e. the cluster distribution) is important. For unrepaired or misrepaired DSB (responsible for chromosome aberrations, cell killing, oncogenic cell transformation and gene mutation) both the cluster distribution and the distance distribution of clusters are relevant. At low doses the distance distribution of clusters along a single particle track determines the RBE-value. However, with increasing dose the distribution of clusters produced by all particles traversing the cell nucleus becomes increasingly determinant. Here, solely the cluster distribution is important as it is the case for the induction of DSB.

Alpha Particles↗