Hyper-radiosensitivity Research Articles

The Response of Primary Keratinocytes and Fibroblasts from Cancer Patients to Multiple Low-Dose Irradiations

In our previous study, using the micronucleus (MN) assay, a hyper-radiosensitivity (HRS)-like phenomenon was observed after single low doses for fibroblasts from two and keratinocytes from four of the 40 patients studied. In this paper, we report the response of primary keratinocytes from 23 and fibroblasts from 21 of these cancer patients to multiple low-dose irradiations and answer the question regarding whether the patients with an HRS-like response after single low doses also demonstrate chromosomal hypersensitivity after multiple low doses. The cells were irradiated with three doses of 0.25 Gy separated by 4-h intervals, and MN induction was compared with that after the same total dose given as a single fraction of 0.75 Gy. Similarly, the effect of three doses of 0.5 Gy was compared with that of a single dose of 1.5 Gy. For fibroblasts from two and keratinocytes from four patients who demonstrated a single-dose HRS-like response, a significant inverse effect of fractionation (greater MN induction after three doses of 0.25 Gy than after a single dose of 0.75 Gy) was observed, which suggests a repeated hypersensitive response after each dose of 0.25 Gy. Such an effect was not seen for the cells from 19 patients who were single-dose HRS-like negative. In conclusion, an inverse fractionation effect for MN induction that was observed in fibroblasts from two and keratinocytes from four patients after three doses of 0.25 Gy (but not 3 x 0.5 Gy) reflects the chromosomal hyper-radiosensitivity seen in the same patients in response to single low doses.

Transition in Survival From Low-Dose Hyper-Radiosensitivity to Increased Radioresistance Is Independent of Activation of ATM SER1981 Activity

The molecular basis of low-dose hyper-radiosensitivity (HRS) is only partially understood. The aim of this study was to define the roles of ataxia telangiectasia mutated (ATM) activity and the downstream ATM-dependent G(2)-phase cell cycle checkpoint in overcoming HRS and triggering radiation resistance. Survival was measured using a high-resolution clonogenic assay. ATM Ser1981 activation was measured by Western blotting. The role of ATM was determined in survival experiments after molecular (siRNA) and chemical (0.4 mM caffeine) inhibition and chemical (20 microg/mL chloroquine, 15 microM genistein) activation 4-6 h before irradiation. Checkpoint responsiveness was assessed in eight cell lines of differing HRS status using flow cytometry to quantify the progression of irradiated (0-2 Gy) G(2)-phase cells entering mitosis, using histone H3 phosphorylation analysis. The dose-response pattern of ATM activation was concordant with the transition from HRS to radioresistance. However, ATM activation did not play a primary role in initiating increased radioresistance. Rather, a relationship was discovered between the function of the downstream ATM-dependent early G(2)-phase checkpoint and the prevalence and overcoming of HRS. Four cell lines that exhibited HRS failed to show low-dose (<0.3-Gy) checkpoint function. In contrast, four HRS-negative cell lines exhibited immediate cell cycle arrest for the entire 0-2-Gy dose range. Overcoming HRS is reliant on the function of the early G(2)-phase checkpoint. These data suggest that clinical exploitation of HRS could be achieved by combining radiotherapy with chemotherapeutic agents that modulate this cell cycle checkpoint.

Low-Dose Radiation Response of Primary Keratinocytes and Fibroblasts from Patients with Cervix Cancer

The aim of the present study was to examine, using the micronucleus (MN) assay, the low-dose radiation response of normal skin cells from cancer patients and to determine whether the hyper-radiosensitivity (HRS)-like phenomenon occurs in cells of these patients. Primary skin fibroblasts and keratinocytes derived from 40 patients with cervix cancer were studied. After in vitro gamma irradiation with single doses ranging from 0.05 to 4 Gy, MN induction was assessed. For each patient, the linear-quadratic (LQ) model and the induced repair (IR) model were fitted over the whole data set. In fits of the IR model, an HRS-like response after low doses (seen as the deviation over the LQ curve) was demonstrated for the fibroblasts of two patients and for the keratinocytes of four other patients. The alpha(s)/alpha(r) ratio for the six patients ranged from 2.7 to 15.4, whereas the values of the parameter d(c) ranged from 0.13 to 0.36 Gy. No relationship was observed between chromosomal radiosensitivity of fibroblasts and keratinocytes derived from the same donor in the low-dose (0.1-0.25 Gy) region. In conclusion, the fact that low-dose chromosomal hypersensitivity was observed for cells of only six of the patients studied suggests that it is not a common finding in human normal cells and can represent an individual characteristic.

On the possible increase in local tumour control probability for gliomas exhibiting low dose hyper-radiosensitivity using a pulsed schedule

Using modelling, we have developed a treatment strategy for gliomas exhibiting low dose hyper-radiosensitivity (HRS) that employs both a reduced dose-rate and pulsed treatment dose delivery. The model exploits the low dose hypersensitivity observed in some glioma cell lines at low radiation doses. We show, based on in vitro data, that a pulsed delivery of external beam radiation therapy could yield significant increases in local control. We therefore propose a pulsed delivery scheme for the treatment of gliomas in which the daily treatment fraction is delivered using 0.20 Gy pulses, separated by three minutes for a time-averaged dose-rate of 0.0667 Gy/min. The dose per pulse of 0.2 Gy is near or below the transition dose observed in vitro for four of the five glioma cell lines we have studied. Using five established glioma cell lines our modelling demonstrates that our pulsed delivery scheme yields a substantial increase in tumour control probability (TCP).

Low-Dose Hyper-radiosensitivity is not Caused by a Failure to Recognize DNA Double-Strand Breaks

One of the earliest cellular responses to radiation-induced DNA damage is the phosphorylation of the histone variant H2AX (gamma-H2AX). gamma-H2AX facilitates the local concentration and focus formation of numerous repair-related proteins within the vicinity of DNA DSBs. Previously, we have shown that low-dose hyper-radiosensitivity (HRS), the excessive sensitivity of mammalian cells to very low doses of ionizing radiation, is a response specific to G(2)-phase cells and is attributed to evasion of an ATM-dependent G(2)-phase cell cycle checkpoint. To further define the mechanism of low-dose hyper-radiosensitivity, we investigated the relationship between the recognition of radiation-induced DNA double-strand breaks as defined by gamma-H2AX staining and the incidence of HRS in three pairs of isogenic cell lines with known differences in radiosensitivity and DNA repair functionality (disparate RAS, ATM or DNA-PKcs status). Marked differences between the six cell lines in cell survival were observed after high-dose exposures (>1 Gy) reflective of the DNA repair capabilities of the individual six cell lines. In contrast, the absence of functional ATM or DNA-PK activity did not affect cell survival outcome below 0.2 Gy, supporting the concept that HRS is a measure of radiation sensitivity in the absence of fully functional repair. No relationship was evident between the initial numbers of DNA DSBs scored immediately after either low- or high-dose radiation exposure with cell survival for any of the cell lines, indicating that the prevalence of HRS is not related to recognition of DNA DSBs. However, residual DNA DSB damage as indicated by the persistence of gamma-H2AX foci 4 h after exposure was significantly correlated with cell survival after exposure to 2 Gy. This observation suggests that the persistence of gamma-H2AX foci could be adopted as a surrogate assay of cellular radiosensitivity to predict clinical radiation responsiveness.

A modelling study of the potential influence of low dose hypersensitivity on radiation treatment planning

Background and purposeLow dose hyper-radiosensitivity (HRS) has been observed in both normal tissues and tumours. This modelling study explores the possible impact of HRS on radiation treatment planning. Patients and methodsThe interplay between volume-effect and HRS was studied in an idealized comparison of partial versus whole organ irradiation. In the further studies, CT scans of three previously scanned patients were used to estimate normal tissue complication probability (NTCP) for the kidneys after a conformal and a conventional treatment plan with and without consideration of HRS. ResultsIdealized treatment plans were compared as pairs of a conventional and a conformal plan both treating the same target volume to the same dose per fraction. Contour maps of the difference in NTCP between paired plans showed a strong dependence on the magnitude of both the volume effect and the HRS effect. For more clinically realistic treatment plans with NTCP calculated for the kidney, the balance between the sparing due to the LQ effect and the increased sensitivity due to the HRS effect was dependent on both the dose distribution and the fractionation. ConclusionsHRS may potentially affect radiotherapy treatment planning and the relative importance of HRS is larger in a tissue or organ with a pronounced volume effect. If HRS is expressed in some normal tissues or organs, this could offset much of the sparing predicted by the LQ formalism. However, in some clinical situations the NTCP calculated with correction for HRS may still be lower than the NTCP calculated from the uncorrected physical doses.

Not all 2 gray radiation prescriptions are equivalent: Cytotoxic effect depends on delivery sequences of partial fractionated doses

To test whether or not the commonly prescribed daily dose of 2 Gy (whole fraction), when delivered as various partial fraction (PF) dose sequences simulating clinical treatment fields, produces equal biologic effects. Eleven actively proliferating cell lines derived from human and animal tissues were used in this study. 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) and clonogenic assays were used to determine the radiation effects on cell proliferation and survival, respectively. The 2 Gy dose was divided into 2 or more PFs for delivery to simulate the delivery of clinical treatment fields. Most irradiation sequences contained two parts consisting of at least 1 small PF, denoted by S which was 0.5 Gy or less, and a large PF, denoted by L which was 1 Gy or more. Irradiation schemes were designed to include the following conditions: (a) the 2 Gy dose divided into combinations of an L-dose and one or more S-doses; (b) the L-dose given either before or after the S-doses; and (c) delivery of all partial fractions within a fixed total time. Significant differences in biologic effect were observed between sequences in which the L-dose was given before or after the S-doses in both the MTT and clonogenic assays. Nearly all the latter schemes, that is S-L, produced greater cytotoxic effects than the L-S schemes. These data demonstrate that the biologic effects of 2 Gy may differ in different clinical settings depending on the size and sequence of the partial fractions. The variation between cytotoxic effects is likely a result of the combination of low-dose hyper-radiosensitivity (HRS) and higher-dose increased radioresistance (IRR) effects established recently. We suggest that to ensure the optimal biologic effect of a prescribed dose of 2 Gy clinically, it is critical to consider the sequence in which the treatment fields are delivered when partial fractions of different sizes are used.

Ultrafractionation does not Improve the Results of Radiotherapy in Radioresistant Murine DDL1 Lymphoma

Low-dose hyperradiosensitivity (HRS), i.e., a relatively higher efficacy of doses < or = 0.5 Gy compared to doses > 1 Gy, has been shown in a number of tumor cell lines in vitro. Therefore ultrafractionated irradiation, i.e., application of very low doses per fraction, has been proposed to improve the effects of radiotherapy. The present study investigates ultrafractionation (UF) in radioresistant murine DDL1 T-cell lymphoma in mice. UF was performed with 0.4 Gy per fraction, three fractions per day at 7 days per week, and conventional fractionation (CF) with 1.68 Gy per fraction, one fraction per day at 5 days per week. Tumor growth delay was evaluated for 2, 4 and 6 weeks of irradiation as time that tumors needed to reach fivefold the starting volume (GD(V5)). GD(V5) was not significantly different between UF and CF. The composite median relative GD(V5) calculated for all tumors irradiated in the present study was 1.00 [95% confidence interval 0.99; 1.08] in the CF and 0.99 [0.92; 1.01] in the UF arm (p = 0.24). UF was not more efficient than CF in DDL1 tumors. Taken together with previous experiments on human A7 glioblastoma, which showed a negative effect of UF on local tumor control, the preclinical data obtained in this laboratory so far do not support the use of ultrafractionated schedules in radiotherapy.

Low-dose hyperradiosensitivity of human glioblastoma cell lines in vitro does not translate into improved outcome of ultrafractionated radiotherapy in vivo

Purpose: Low dose hyperradiosensitivity (HRS) has been observed in HGL21- and T98G human glioblastoma cells in vitro. The present study investigates whether these effects translate into improved outcome of ultrafractionated irradiation (UF) in vivo.Material and methods: T98G or HGL21 were transplanted on the hind leg of nude mice. Tumours were irradiated with UF (3 fractions of 0.4 Gy per day, interval 4 h, 7 days per week) or with conventional fractionation (CF; 1 fraction of 1.68 Gy per day, 5 days per week) over 2 or 4 weeks in HGL21 and 2,4 or 6 weeks in T98G. In HGL21, graded top-up doses under clamped hypoxia were applied after 4 weeks of fractionated irradiation. Additional groups of animals were irradiated with single doses under clamp hypoxic conditions with or without whole body irradiation (WBI) before tumour transplantation. Experimental endpoints were growth delay (time to 5-fold starting volume, GDV5) and local tumour control.Results: In T98G tumours median relative GDV5 was 1.2 [95%C.I. 0.96; ∞] in the CF and 0.8 [0.7; 1.02] in the UF arm (p = 0.009) indicating that ultrafractionation is less efficient than conventional fractionation. The TCD50 value of 33.5 Gy [22; 45] after UF was higher than TCD50 of 23.6 Gy [16; 31] after CF (p = 0.15). In HGL21 the median relative GDV5 was not significantly different between CF and UF. The top-up TCD50 value of 16.1 Gy [95% C.I. 9; 23 Gy] after CF was significantly lower than the corresponding value of 33.2 Gy [23; 44] after UF irradiation (p = 0.007), indicating a higher efficacy of CF compared to UF.Conclusion: The results on human T98G and HGL21 glioblastoma do not support the hypothesis that HRS in vitro translates into improved outcome of ultrafractionated irradiation in vivo.

Low-dose hyper-radiosensitivity: a consequence of ineffective cell cycle arrest of radiation-damaged G2-phase cells.

This review highlights the phenomenon of low-dose hyper- radiosensitivity (HRS), an effect in which cells die from excessive sensitivity to small single doses of ionizing radiation but become more resistant (per unit dose) to larger single doses. Established and new data pertaining to HRS are discussed with respect to its possible underlying molecular mechanisms. To explain HRS, a three-component model is proposed that consists of damage recognition, signal transduction and damage repair. The foundation of the model is a rapidly occurring dose-dependent pre-mitotic cell cycle checkpoint that is specific to cells irradiated in the G2phase. This checkpoint exhibits a dose expression profile that is identical to the cell survival pattern that characterizes HRS and is probably the key control element of low-dose radiosensitivity. This premise is strengthened by the recent observation coupling low- dose radiosensitivity of G2-phase cells directly to HRS. The putative role of known damage response factors such as ATM, PARP, H2AX, 53BP1 and HDAC4 is also included within the framework of the HRS model.

Early effects of low dose-rate radiation on cultured tumor cells.

Low-dose hyperradiosensitivity (HRS) has been found for several cell types after exposure to low doses, < 0.5 Gy, of high dose-rate (typically 50-150 Gy/h) low-LET radiation. HRS precedes the occurrence of a relative resistance for doses above 0.5-1 Gy. A critical question is whether HRS is of importance in radionuclide therapy where the dose-rate is low but the total dose might be high. An indication that cells exposed to low dose-rate can be kept hyperradiosensitive has recently been published. We have in the present study applied cells without (glioma U373MG) and with (glioma U118MG and colon carcinoma HT29) HRS and studied early effects, up to one week, during low dose-rate (LDR), 0.05-0.09 Gy/hours, exposure (total dose after one week: 11.8 +/- 1.5 Gy). The cells were grown on thin foils above a (32)P source placed in a cell culture chamber. Cell number reductions, cell-cycle disturbances, and changed numbers of apoptotic cells were analyzed after continuous LDR exposures. There seemed to be no relation with HRS when the cell number reduction was considered. The U373MG cells, lacking HRS, had the strongest cell number reduction due to a combination of a G(2) block and increased apoptosis. The U118MG and HT29 cells, both having HRS, had surprisingly low cell number reductions. U118MG had only a G(2) block but no increase in apoptosis. HT29 had both a G(2) block and an increase in apoptosis but the apoptosis change was somewhat smaller than for U373MG. Thus, there seemed to be no obvious relation between HRS and early cellular effects when the cells were analyzed after continuous LDR exposure.

An association between the radiation-induced arrest of G2-phase cells and low-dose hyper-radiosensitivity: a plausible underlying mechanism?

The survival of asynchronous and highly enriched G1-, S- and G2-phase populations of Chinese hamster V79 cells was measured after irradiation with 60Co gamma rays (0.1-10 Gy) using a precise flow cytometry-based clonogenic survival assay. The high-dose survival responses demonstrated a conventional relationship, with G2-phase cells being the most radiosensitive and S-phase cells the most radioresistant. Below 1 Gy, distinct low-dose hyper-radiosensitivity (HRS) responses were observed for the asynchronous and G2-phase enriched cell populations, with no evidence of HRS in the G1- and S-phase populations. Modeling supports the conclusion that HRS in asynchronous V79 populations is explained entirely by the HRS response of G2-phase cells. An association was discovered between the occurrence of HRS and the induction of a novel G2-phase arrest checkpoint that is specific for cells that are in the G2 phase of the cell cycle at the time of irradiation. Human T98G cells and hamster V79 cells, which both exhibit HRS in asynchronous cultures, failed to arrest the entry into mitosis of damaged G2-phase cells at doses less than 30 cGy, as determined by the flow cytometric assessment of the phosphorylation of histone H3, an established indicator of mitosis. In contrast, human U373 cells that do not show HRS induced this G2-phase checkpoint in a dose-independent manner. These data suggest that HRS may be a consequence of radiation-damaged G2-phase cells prematurely entering mitosis.

Low-dose reduction in transformation frequency compared to unirradiated controls: the role of hyper-radiosensitivity to cell death.

Calculations based on plausible parameters taken from the existing experimental database, and new measurements on the cell cycle dependence of low-dose hyper-radiosensitivity (HRS) of non-tumorigenic HeLa x skin fibroblast human hybrid cells, provide the first experimental evidence that the selective killing of a transformation-sensitive G(2)/M-phase subpopulation as a consequence of low-dose HRS could account in part for the observed reduction of induced transformation frequencies at low doses to values below that observed spontaneously. However, it is clear that other mechanisms associated with classical adaptive response, such as induced DNA repair, are also likely to be involved.

Effects of cell cycle phase on low-dose hyper-radiosensitivity

To examine the low-dose radiation response of human glioma cell lines separated into different cell-cycle phases and to determine if low-dose hyper-radiosensitivity (HRS) differs in populations defined by cell-cycle position. To assess whether predictions of the outcome of multiple low-dose regimens should take account of cell-cycle effects. The clonogenic survival of G1, G2 and S phase cells was measured after exposure to single doses of X-rays in two human glioma cell lines. One cell line (T98G) showed marked HRS when asynchronous cells were irradiated, while the other (U373) did not. Separation of populations and high-resolution cell counting was achieved using a fluorescence activated cell sorter. Sorted cell populations were irradiated with 240 kVp X-rays to doses between 0.05 and 5Gy. The resulting cell-survival versus dose data were comparatively fitted using the linear-quadratic and induced-repair models in order to assess the degree of HRS. In both cell lines the low-dose response was altered when different populations were irradiated. In T98G cells, all populations showed HRS, but this was most marked in G2 phase cells. In U373 cells, no HRS was found in G1 or S phase cells, but HRS was demonstrable in G2 phase cells. HRS was expressed by the whole cell population of T98G cells but the size of the effect varied with cell-cycle phase and was most marked in the G2 population. In U373 cells, the effect could only be demonstrated in G2 cells. This implies that HRS is primarily a response of G2 phase cells and that this response dominates that seen in asynchronous populations. Actively proliferating cell populations may therefore demonstrate a greater increase in radiosensitivity to very low radiation doses compared with quiescent populations.

Effects of cell cycle phase on low-dose hyper-radiosensitivity

Purpose : To examine the low-dose radiation response of human glioma cell lines separated into different cell-cycle phases and to determine if low-dose hyper-radiosensitivity (HRS) differs in populations defined by cell-cycle position. To assess whether predictions of the outcome of multiple low-dose regimens should take account of cell-cycle effects. Materials and methods : The clonogenic survival of G1, G2 and S phase cells was measured after exposure to single doses of X-rays in two human glioma cell lines. One cell line (T98G) showed marked HRS when asynchronous cells were irradiated, while the other (U373) did not. Separation of populations and high-resolution cell counting was achieved using a fluorescence activated cell sorter. Sorted cell populations were irradiated with 240 kVp X-rays to doses between 0.05 and 5Gy. The resulting cell-survival versus dose data were comparatively fitted using the linear-quadratic and induced-repair models in order to assess the degree of HRS. Results : In both cell lines the low-dose response was altered when different populations were irradiated. In T98G cells, all populations showed HRS, but this was most marked in G2 phase cells. In U373 cells, no HRS was found in G1 or S phase cells, but HRS was demonstrable in G2 phase cells. Conclusions : HRS was expressed by the whole cell population of T98G cells but the size of the effect varied with cell-cycle phase and was most marked in the G2 population. In U373 cells, the effect could only be demonstrated in G2 cells. This implies that HRS is primarily a response of G2 phase cells and that this response dominates that seen in asynchronous populations. Actively proliferating cell populations may therefore demonstrate a greater increase in radiosensitivity to very low radiation doses compared with quiescent populations.

Radiosensitivity of V79 cells after alpha particle radiation at low doses.

Low dose hyper-radiosensitivity (HRS) of V79 cells was demonstrated after irradiation with gamma rays and 4He2+ ions of various linear energy transfer (LET) values (58.9, 79.3 and 101.7 keV.micron-1). In parallel, the cytogenetic analysis showed an LET dependence of aberrations at a dose of 1 Gy, while the observed chromatid fragments appeared to vary with the number of 4He2+ ions traversing the cell nucleus. The results of both studies are correlated so as to achieve a better understanding of the so-called induced radioresistance. The cell mechanism of radioresistance appears to be induced after a certain amount of energy is deposited in the cell nucleus. This amount depends both on the radiation quality as well as the number of particles traversing the cell, inducing chromosome alterations and chromatid damage.

No benefits of ultrafractionation in two head-and-neck cancer cell lines with different inherent radiosensitivity

Purpose: To assess if ultrafractionation is applicable in the context of an unknown hyperradiosensitivity (HRS) status, we studied the survival and repair capacity of two tumor cell lines after irradiation with two different dose/fractionation schedules that can be used in a clinical setting.Methods and Materials: Squamous cell carcinoma cell lines SCC-3 (radioresistant) and SCC-6 (radiosensitive) were used. Survival was studied by clonogenic assay after multiple fractions of 0.5 Gy (2 fractions/day, 6-h interval) and 2 Gy (1 fraction/day) for a total dose of 8 Gy of γ-rays. The capacity to repair single-strand and double-strand breaks (SSB, DSB) was assessed by comet assay. The messenger RNA (mRNA) levels of DNA-dependent protein kinase (PK) components were analyzed by RNase protection and real-time polymerase chain reaction (PCR).Results: In both cell lines, no apparent difference was noted between the two fractionation protocols. In particular for SCC-3, the mean surviving fraction tended to be lower after 2 Gy than after 0.5 Gy fractions. In SCC-3 and SCC-6 no significant difference was observed in the repair capacity of SSB and DSB after exposure to single doses of 0.5 Gy or 2 Gy. After exposure to the same single doses, the mRNA levels of DNA-PK catalytic subunit (PKcs), Ku 70, and Ku 80 were similar.Conclusions: Our data do not support the concept of ultrafractionation, at least when using fractions of 0.5 Gy in the cell lines studied. This suggests that methods for testing HRS status in individual tumors need to be developed before the relevance of ultrafractionation can be investigated in the clinic.

Evidence for the involvement of DNA-dependent protein kinase in the phenomena of low dose hyper-radiosensitivity and increased radioresistance

Purpose : To investigate the role of DNA-dependent protein kinase (DNA-PK) in the phenomena of low dose hyper-radiosensitivity (HRS) and increased radioresistance (IRR) using the genetically related M059 cell lines of disparate PRKDC status. Materials and methods : Clonogenic survival was measured for the three cell lines following low doses of X-irradiation using a flow-activated cell sorting (FACS) plating technique. The presence of PRKDC, G22p1 and Xrcc5 proteins was determined by Western blotting and a kinase assay used to measure DNA-PK complex activity. Results : The survival responses for the three cell lines over the 0-0.3Gy dose range were comparable, but differences in radiosensitivity were evident at doses >0.4Gy. M059K and M059J/Fus1 cells (both PRKDC competent) exhibited marked HRS/IRR responses, albeit to different extents. M059J cells (PRKDC incompetent) were extremely radiosensitive exhibiting a linear survival curve with no evidence of IRR. The presence of IRR was coincident with the presence of PRKDC protein and functional DNA-PK activity. Conclusions : HRS is a response that is independent of DNA-PK activity. In contrast, IRR showed a dependence on the presence of PRKDC protein and functional DNA-PK activity. These data support a role for DNA-PK activity in the IRR response.

Cell cycle and growth response of CHO cells to X-irradiation: Threshold-free repair at low doses

Purpose: To test the hypothesis of a threshold for induced repair of DNA damage (IR) and, secondarily, of hyperradiosensitivity (HRS) to low-dose X-irradiation. Methods and Materials: Exponentially growing Chinese hamster ovary cells (CHO) were X-irradiated with doses from 0.2 to 8 Gy. Survival data were established by conventional colony-forming assay and flow-cytometric population counting. The early cell cycle response to radiation was studied based on DNA-profiles and bromodeoxyuridine pulse-labeling experiments. Results: Colony-forming data were consistent with HRS. However, these data were of low statistic significance. Population counting provided highly reproducible survival curves that were in perfect accord with the linear-quadratic (LQ) model. The dominant cell cycle reaction was a dose-dependent delay of G2 M and late S-phase. Conclusion: There was no evidence for a threshold of IR and for low-dose HRS in X-irradiated CHO cells. It is suggested that DNA damage repair activity is constitutively expressed during S-phase and is additionally induced in a dose-dependent and threshold-free manner in late S-phase and G2. The resulting survival is precisely described by the LQ model.

Low-dose hypersensitivity: current status and possible mechanisms

Purpose: To retain cell viability, mammalian cells can increase damage repair in response to excessive radiation-induced injury. The adaptive response to small radiation doses is an example of this induced resistance and has been studied for many years, particularly in human lymphocytes. This review focuses on another manifestation of actively increased resistance that is of potential interest for developing improved radiotherapy, specifically the phenomenon in which cells die from excessive sensitivity to small single doses of ionizing radiation but remain more resistant (per unit dose) to larger single doses. In this paper, we propose possible mechanisms to explain this phenomenon based on our data accumulated over the last decade and a review of the literature. Conclusion: Typically, most cell lines exhibit hyper-radiosensitivity (HRS) to very low radiation doses (<10 cGy) that is not predicted by back-extrapolating the cell survival response from higher doses. As the dose is increased above about 30 cGy, there is increased radioresistance (IRR) until at doses beyond about 1 Gy, radioresistance is maximal, and the cell survival follows the usual downward-bending curve with increasing dose. The precise operational and activational mechanism of the process is still unclear, but we propose two hypotheses. The greater amount of injury produced by larger doses either ( 1) is above a putative damage-sensing threshold for triggering faster or more efficient DNA repair or ( 2) causes changes in DNA structure or organization that facilitates constitutive repair. In both scenarios, this enhanced repair ability is decreased again on a similar time scale to the rate of removal of DNA damage.