C3H 10T Research Articles

A Modified Lethal and Potentially Lethal Model for Ultra-High Dose Rate FLASH RT

Recently, ultra-high dose rate (> 40Gy/s) radiotherapy (FLASH RT) has gained widespread interest among researchers and clinicians owing to its remarkable normal tissue sparing as well as efficient tumor control compared to conventional radiotherapy (CONV RT) with dose rates around 0.03 Gy/s. As radiobiology modeling has been the foundation of the contemporary clinical practice in radiation oncology, the urgent need for robust mathematical modeling of this paradigm-shifting treatment, FLASH RT, is difficult to overstate. This work proposes a phenomenological model as a potential candidate to explain the observed phenomena. In this study, we combined the oxygen depletion hypothesis and Curtis’ Lethal and Potentially Lethal (LPL) model. Following the oxygen depletion hypothesis on FLASH-induced normal tissue sparing, we introduced a flipped sigmoid shape function to explicitly model the time-dependent component of the radiation-induced lethal and potentially lethal lesion production rates when cells experience oxygen depletion. We conducted both numerical and analytic analyses after we selected a Logistic-type function for demonstration. Parameters from Curtis' LPL model fitting of the C3H 10T 1/2 mouse cell survival data, a generic oxygen enhancement ratio, and two newly introduced parameters describing the time scale and transition width for radiation-induced oxygen depletion were employed in our model. Both numerical and analytical tools were used to obtain the cell survival probability prediction for our modified LPL model. We demonstrated that our modified LPL model would generate asymptotical cell survival probability that decreases exponentially as the radiation dose increases in FLASH RT. Also, the numerical study indicated that our modified LPL model could produce significantly reduced normal tissue cell killings in FLASH RT. For tumor cells, since their cell environment is typically hypoxic, the mechanism mentioned above will have limited impact, and hence tumor cell death will remain at the level similar to that from CONV RT with the same total dose. We also provided an approximated analytical solution of our model’s cell radiation survival prediction for future FLASH RT study. Our modified LPL model predicts that, for normal tissue cells, the difference in the logarithm of surviving fraction between FLASH RT and CONV RT is asymptotically proportional to the radiation dose, and the cell’s oxygen enhancement ratio minus one. This modified LPL model has the potential to explain the prominent normal tissue sparing effect of FLASH RT, with interpretable parameters. Future studies are necessary to quantitatively validate the model predictions in systematically conducted FLASH RT experiments.

Quantitation of Compositional Changes in the Non-Erythroid Membrane Skeleton due to External Forces

The spectrin based membrane skeleton plays important roles in the mechanical and biological functions of living cells. Unlike the membrane skeleton of red blood cells, its non-erythroid counterpart has seen very little attention. Yet, it is assumed that also in non-erythrocytes the membrane skeleton is mostly composed of heterotetramers of α-II and β-II spectrin. Because of α-spectrins’ potential role as mechanotransducers, we investigate the effect of mechanical stimulation on the spectrin-ankyrin B membrane skeleton of NIH 3T3 fibroblasts and murine C3H 10T½ cells. Using quantitative fluorescence microscopy and biochemical techniques, the absolute changes in cellular spectrin and ankyrin B content were determined. Before stimulation the spectrins and ankyrin B make up more than 10% of cytosolic and membrane proteins. Thus, they constitute a much more significant part of the non-erythroid cytoskeleton in these cells than previously assumed. Interestingly, both cell types contain all four major isoforms of spectrin. However, while α-II spectrin is the dominant α spectrin in 3T3, in 10T½ cells it is the ‘erythroid’ α-I spectrin. Also, at least half of the α spectrins do not have β-spectrin counterparts and thus cannot be part of the classical heterotetramere structure. Mechanical stimulation decreases the overall spectrin amount by up to 60% with α-spectrins exhibiting the largest loss. In contrast, the number of ankyrin B copies increases by almost 30%. Furthermore, the fraction of polyubiquitinated spectrin actually increases, which in part could account for the reduction in α-spectrins and is in line with their proposed mechanically enhanced self-ubiquitination activity.

Rodent cell transformation assays—A brief historical perspective

In vitro cell transformation is a process characterized by a series of progressive distinctive events that often emulate manifestations occurring in vivo and which are associated with neoplasia. Attendant cellular and sub-cellular alterations include, among others: cellular immortality, phenotypic changes, aneuploidy, genetic variability, cellular disarray, anchorage-independent growth, and tumorigenicity in vivo. Early chemically induced neoplastic transformation studies involved the use of normal diploid (Syrian) hamster embryo (SHE) cells and monitored the formation of morphologically altered colonies. Later investigations employed primarily two established mouse cell lines, i.e. the BALB/c 3T3 A31 cell line and the C3H 10T 1/2 cell line, and monitored the induction of morphologically aberrant foci. In either case, such transformed cellular clusters (colonies and foci) could induce tumors upon inoculation in vivo. Some subsequent noteworthy advancements using these systems included pH adjustments, metabolic supplementation, amplification of expression of formerly latent transformed foci, concurrent detection of mutagenesis and transformation, and use of a Bhas 42 cell line (v-Ha-ras transfected BALB/c 3T3 cells) to detect both tumor initiators and promoters. Over time, such transformation assay systems have been found useful in academic, industry and regulatory laboratories, generally for research purposes, but also occasionally as screening tools for potential chemical carcinogens. Nevertheless, to date, use of these assays for decision-making purposes in the regulatory arena remains elusive and will require comprehensive validation to gain universal acceptance.

Increased Frequency of Spontaneous Neoplastic Transformation in Progeny of Bystander Cells from Cultures Exposed to Densely Ionizing Radiation

An increased risk of carcinogenesis caused by exposure to space radiation during prolonged space travel is a limiting factor for human space exploration. Typically, astronauts are exposed to low fluences of ionizing particles that target only a few cells in a tissue at any one time. The propagation of stressful effects from irradiated to neighboring bystander cells and their transmission to progeny cells would be of importance in estimates of the health risks of exposure to space radiation. With relevance to the risk of carcinogenesis, we investigated, in model C3H 10T½ mouse embryo fibroblasts (MEFs), modulation of the spontaneous frequency of neoplastic transformation in the progeny of bystander MEFs that had been in co-culture 10 population doublings earlier with MEFs exposed to moderate doses of densely ionizing iron ions (1 GeV/nucleon) or sparsely ionizing protons (1 GeV). An increase (P<0.05) in neoplastic transformation frequency, likely mediated by intercellular communication through gap junctions, was observed in the progeny of bystander cells that had been in co-culture with cells irradiated with iron ions, but not with protons.

Mild hyperthermia as a potential mechanism to locally enhance cell growth kinetics

The effect of mild hyperthermia on growth kinetics of two human glioma and one mouse fibroblast cell line was evaluated over a 12 h (short-term) or 7 day (long-term) period. All cell lines showed growth enhancement at 38°C, although the effect in C3H 10T ½ mouse fibroblasts was more pronounced and had a more rapid onset than in U87MG or LN71 glioma cells. At 39°C, growth of C3H 10T ½ cells was slightly reduced and glioma cell lines similar to that of their respective 37°C controls. At 40°C, C3H 10T ½ cells showed a more rapid and dramatic growth reduction than glioma cell lines and accumulated in both G0/G1 and G2 checkpoint compartments. In contrast, U87MG cells accumulated only in G2 and LN71 showed no checkpoint accumulation. The findings indicate that cell lines are differentially responsive to small temperature elevations. If similar differences exist between normal and diseased human tissues, local application of mild hyperthermia might offer a noninvasive and cost-effective method to achieve local enhancement of drugs that target proliferating tissue.

Melengestrol acetate enhances adipogenic gene expression in cultured muscle-derived cells

Melengestrol acetate (MGA) has been used in the United States for nearly 40 yr to enhance feedlot heifer performance, yet unequivocal studies have not been conducted to discover the mechanism of action. Our hypothesis was that MGA may induce various populations of muscle-derived cells (MDC) to the adipogenic pathway in both a bovine and murine cell culture model. To determine this, MDC were digested from the semimembranosus muscle tissue of six 14-mo-old crossbred steers. The addition of insulin, oleic acid, and ciglitizone (IOC) with cultured bovine MDC resulted in morphological differences compared with control cultures. Multilocular lipid droplets stained with Oil Red O were seen not only in single MDC, but also in fused myotubes. An increase (P < 0.05) in relative PPARgamma messenger RNA (mRNA) levels was measured in MDC incubated with IOC. However, myogenin mRNA levels in MDC incubated with IOC were repressed (P < 0.05) compared with nontreated MDC. Cultures of MDC treated with 10 microM insulin, 10 microM oleic acid, 10 microM ciglitizone, 10 nM estradiol-17beta (E2), and 10 nM MGA resulted in cultures with highly distributed lipid droplets not only in single cells, but also in the multinucleated myotubes. Relative C/EBPbeta and PPARgamma mRNA levels in total RNA isolated from MDC treated with MGA increased (P < 0.05) compared with control cultures. Estradiol treatment had no effect (P > 0.05) on these mRNA levels. The addition of both E2 and MGA to MDC increased (P < 0.05) C/EBPbeta mRNA levels and tended (P = 0.06) to increase the PPARgamma mRNA level. There was no difference (P > 0.10) in relative myogenin mRNA among the control, E(2), and MGA treatments. Relative C/EBPbeta, PPARgamma, and myogenin mRNA levels were investigated in murine C2C12, C3H 10T 1/2, and 3T3-L1 cells. Treatment of cultures with 10 nM MGA increased C/EBPbeta levels (P < 0.05) in C2C12 myoblasts and tended (P = 0.08) to increase C/EBPbeta levels in 3T3-L1 preadipocytes. These data indicate that populations of cells are present in postnatal skeletal muscle that, under the appropriate stimuli in a culture model, express adipogenic genes and accumulate lipids. In addition, the synthetic progestogen MGA appeared to upregulate the genes necessary for conversion to the adipogenic pathway.

Risks Associated with Low Doses and Low Dose Rates of Ionizing Radiation: Why Linearity May Be (Almost) the Best We Can Do

Deterministic and stochastic effects associated with high-dose ionizing radiation (x-ray) exposure have been known for almost as long as ionizing radiation itself (1–3). At lower doses, radiation risks are primarily stochastic effects, in particular, somatic effects (cancer) rather than the deterministic effects characteristic of higher-dose exposure (4–6). In contrast to deterministic effects, for stochastic effects, scientific committees generally assume that at sufficiently low doses there is a positive linear component to the dose response—that is, that there is no threshold (4–6). This does not preclude there being higher-order (eg, quadratic) powers of dose in the dose response that may be of importance at higher doses. It is on this basis that models linear (or linear-quadratic) in dose are often used to extrapolate the experience of the Japanese atomic bomb survivors (who were typically exposed at a high dose rate to moderate doses [average, 0.1 Sv]) to estimate risks from low doses and low dose rates (4–6). Most population-based cancer risk estimates are based primarily on the Japanese atomic bomb survivor Life Span Study (LSS) cohort data (4–6). However, evidence of excess risks comes from a large number of other studies as well. In the parallel editorial (7), evidence is presented for possible real (or at least “practical”) thresholds or “hormetic” (beneficial) effects of low doses of ionizing radiation. As we summarize here, and in contrast to the arguments of Tubiana et al (7), we judge that there is little epidemiologic or biologic evidence for these for cancer. The arguments are of three forms: (a) assessment of the degree of curvature in the cancer dose response within the Japanese atomic bomb survivors and other exposed groups (in particular, departure from linear or linear-quadratic curvature), (b) consistency of risks between the Japanese and other moderate- and low-dose cohorts, and (c) assessment of biologic data on mechanisms. Most of the information on radiation-induced cancer risk comes from (a) the Japanese atomic bomb survivors, (b) medically exposed populations, (c) occupationally exposed groups, and (d) environmentally exposed groups (6). In the higher-dose radiation therapy studies, where doses received are very much higher than in the LSS, sometimes in the range at which cell sterilization occurs, excess cancer risks per unit dose tend to be less than in comparable subsets of the LSS (8,9). However, as we show, risks in moderate- and low-dose medically and occupationally exposed groups are generally consistent with those in the LSS.

ROLE OF SMAD AND WNT SIGNALING PATHWAYS IN THE FIBROTIC DIFFERENTIATION OF C3H 10T (1/2) MULTIPOTENT CELLS INDUCED BY MYOSTATIN.

Fibrosis (progressive scarring) is a leading cause of organ failure worldwide. Our results indicate that myostatin (Mst), the only known negative regulator of muscle mass, induces the appearance of myofibroblasts in a multipotent cell line (C3H 10T [1/2]) of mouse origin. In this study we show that Mst induces collagen deposition in this cell line, promoting fibrosis, and that this process is mediated through the Smad and Wnt signaling transduction pathways and some of their downstream components. C3H 10 T (1/2) cells were incubated for 2 days with 5′-azacytidine to induce differentiation and then treated with R-Mst (4 μg/mL) in a time course manner (0.5, 1, 2, 3, 24 hours and 3 days); TGF-β (10 ng/ml) was used as a positive control. Cells were also transfected with pcDNA-Mst and two different Mst-shRNA plasmids to overexpress and to silence Mst expression, respectively. Transcriptional regulation of the Smad and Wnt signaling components and induction of genes involved in collagen synthesis were evaluated by three different pathway focus designed microarrays, which have genes each one related to the TGF-β-BMP, osteogenesis, and Wnt signaling pathway. Results were confirmed by immunocytochemistry (ICC) for PAI-1, TGF-β1, collagen I, III and p-Smad 2/3, 3, 4, and 7. The total RNA from cells treated with R-Mst for 3 Hs and 3 days was analyzed by microarrays and qRT-PCR. In parallel, cells were fixed with 2% p-formaldehyde and analyzed by ICC. The main results of the microarray analysis at 3 hours showed that procollagen type IIα1, IVα6, IXα1,3, Smad 3, TGF-β1, Smad-7, and Fst (follistatin), an inhibitor of Mst activity, and Wntif1 were up-regulated; β-catenin, Wnt 4, ctbp1 were down-regulated, and Smad-1, 2, 5, and 6 did not change. After 3 days of incubation with R-Mst: procollagen type IIα1, IVα6, IXα1, 3, Fst, and Smad 7 were up-regulated and β-catenin, Wnt 4, and ctbp1 were down-regulated, whereas Smad 1, 2, and 5 remained the same and Smad 3 was undetectable. Again, the results for collagen I and III, p-Smad 2/3, 3, 4, and 7 were confirmed by ICC, and, in general, the results from the microarrays were confirmed by quantitative RT-PCR and RT2 PCR profiler array. In conclusion, Mst induces the expression of PAI-1, TGF-β1, and collagen deposition in C3H 10 T (1/2) cells by signaling through the Smad /Wnt signaling pathway.

The number of genes changing expression after chronic exposure to Code Division Multiple Access or Frequency DMA radiofrequency radiation does not exceed the false‐positive rate

Experiments with cultured C3H 10T 1/2 cells were performed to determine if exposure to cell phone radiofrequency (RF) radiations induce changes in gene expression. Following a 24 h exposure of 5 W/kg specific adsorption rate, RNA was extracted from the exposed and sham control cells for microarray analysis on Affymetrix U74Av2 Genechips. Cells exposed to 0.68 Gy of X-rays with a 4-h recovery were used as positive controls. The number of gene expression changes induced by RF radiation was not greater than the number of false positives expected based on a sham versus sham comparison. In contrast, the X-irradiated samples showed higher numbers of probe sets changing expression level than in the sham versus sham comparison.

Gene Expression does not Change Significantly in C3H 10T½ Cells after Exposure to 847.74 CDMA or 835.62 FDMA Radiofrequency Radiation

In vitro experiments with C3H 10T(1/2) mouse cells were performed to determine whether Frequency Division Multiple Access (FDMA) or Code Division Multiple Access (CDMA) modulated radiofrequency (RF) radiations induce changes in gene expression. After the cells were exposed to either modulation for 24 h at a specific absorption rate (SAR) of 5 W/ kg, RNA was extracted from both exposed and sham-exposed cells for gene expression analysis. As a positive control, cells were exposed to 0.68 Gy of X rays and gene expression was evaluated 4 h after exposure. Gene expression was evaluated using the Affymetrix U74Av2 GeneChip to detect changes in mRNA levels. Each exposure condition was repeated three times. The GeneChip data were analyzed using a two-tailed t test, and the expected number of false positives was estimated from t tests on 20 permutations of the six sham RF-field-exposed samples. For the X-ray-treated samples, there were more than 90 probe sets with expression changes greater than 1.3-fold beyond the number of expected false positives. Approximately one-third of these genes had previously been reported in the literature as being responsive to radiation. In contrast, for both CDMA and FDMA radiation, the number of probe sets with an expression change greater than 1.3-fold was less than or equal to the expected number of false positives. Thus the 24-h exposures to FDMA or CDMA RF radiation at 5 W/kg had no statistically significant effect on gene expression.

Expression of the Proto-oncogeneFosafter Exposure to Radiofrequency Radiation Relevant to Wireless Communications

In this study the expression levels of the proto-oncogene Fos were measured after exposure to radiofrequency (RF) radiation at two relatively high specific absorption rates (SARs) of 5 and 10 W/kg for three types of modulated signals: 847.74 MHz code division multiple access (CDMA), 835.62 MHz frequency division multiple access (FDMA), and 836.55 MHz time division multiple access (TDMA). This work was undertaken to confirm a previous report by Goswami et al. (Radiat. Res. 151, 300-309, 1999) that CDMA and FDMA radiation caused small but statistically significant increases in Fos levels as cells entered plateau phase during exposure. No effects on Myc or Jun levels were observed in that study. Therefore, in the present study, analyses were restricted to Fos expression during the transition from exponential growth to plateau phase. Fos expression was measured using the real-time polymerase chain reaction (RT-PCR) technique. Serum-stimulated C3H 10T(1/2) cells were used as a positive control for Fos expression. Possible influences of final cell number or pH variability on Fos expression were evaluated. Expression of Fos mRNA in C3H 10T(1/2) cells was not significantly different from that found after sham exposure at either SAR level for any signal modulation. Therefore, the results of Goswami et al. could not be confirmed.

Evidence That Osteogenic Progenitor Cells in the Human Tunica Albuginea May Originate from Stem Cells: Implications for Peyronie Disease1

Tissue ossification in Peyronie disease (commonly known as Peyronie's disease [PD]), a localized fibrotic lesion within the tunica albuginea (TA) of the penis, may result from osteogenic differentiation of fibroblasts, myofibroblasts, and/or adult stem cells in the TA, and may be triggered by chronic inflammation, oxidative stress, and profibrotic factors like transforming growth factor beta 1 (TGFB1). In this study, we have investigated whether cultures of cells from normal TA and PD plaques undergo osteogenesis, express markers for stem cells, and originate other cell lineages via processes modulated by TGFB1. We found that TA and PD cells in osteogenic medium (OM) expressed osteogenic markers, alkaline phosphatase, and osteopontin and underwent calcification. PD cells, but not TA cells, formed foci in soft agar that were positive for alkaline phosphatase and calcification and expressed the mRNAs for osteoblast-specific factors pleiotrophin and periostin and bone morphogenic protein 2. Both cultures expressed stem cell marker CD34 antigen but not protein tyrosine phosphatase, receptor type c. TA and PD cells expressed smooth-muscle cell markers smoothelin and transgelin. None of the cultures underwent adipogenesis in adipogenic medium. Incubation with TGFB1 increased osteogenesis and myofibroblast differentiation and reduced CD34 antigen expression in both cultures. TA and PD cells modulated the differentiation of the multipotent C3H 10T(1/2) cells in dual cultures, into osteoblasts and myofibroblasts. In conclusion, both TA and PD cultures contain cells, presumably stem cells, that undergo osteogenic and myofibroblast differentiation, and may induce these processes by paracrine interactions. This may explain progression of fibrosis in the PD plaque and its eventual calcification.

The bystander response in C3H 10T1/2 cells: the influence of cell-to-cell contact.

Although radiation-induced heritable damage in mammalian cells was thought to result from the direct interaction of radiation with DNA, it is now accepted that biological effects may occur in cells that were not themselves traversed by ionizing radiation but are close to those that were. However, little is known about the mechanism underlying such a bystander effect, although cell-to-cell communication is thought to be of importance. Previous work using the Columbia microbeam demonstrated a significant bystander effect for clonogenic survival and oncogenic transformation in C3H 10T(1/2) cells. The present study was undertaken to assess the importance of the degree of cell-to-cell contact at the time of irradiation on the magnitude of this bystander effect by varying the cell density. When 10% of cells were exposed to a range of 2-12 alpha particles, a significantly greater number of cells (P < 0.0001) were inactivated when cells were irradiated at high density (>90% in contact with neighbors) than at low density (<10% in contact). In addition, the oncogenic transformation frequency was significantly higher in high-density cultures (P < 0.0004). These results suggest that when a cell is hit by radiation, the transmission of the bystander signal through cell-to-cell contact is an important mediator of the effect, implicating the involvement of intracellular communication through gap junctions.

The bystander effect in experimental systems and compatibility withradon-induced lung cancer in humans

Bystander effects following exposure to α-particles have been observedin C3H 10T 1/2cells and in other experimental systems, and imply that linearly extrapolatinglow-dose risks from high-dose data might materially underestimate risk. Theratio of lung cancer risk among persons exposed to low and high doses ofradon daughters is 2.4–4.0, with an upper 95% confidence limit (CL) ofabout 14. Assuming that the bystander effect observed in the C3H 10T 1/2data applies to human lung cells in vivo, the epidemiological dataimply that the number of neighbouring cells that can contribute to thebystander effect is between 0 and 1, with an upper 95% CL of about7. As a consequence, the bystander effect observed in the C3H 10T 1/2 systemprobably does not play a large part in the process of radon-induced lung carcinogenesisin humans. Other experimental data relating to the bystander effect after α-particleexposure are surveyed; some of these data are more compatible with theepidemiological data.

The effect of 835.62 MHz FDMA or 847.74 MHz CDMA modulated radiofrequency radiation on the induction of micronuclei in C3H 10T(1/2) cells.

To determine if radiofrequency (RF) radiation induces the formation of micronuclei, C3H 10T(1/2) cells were exposed to 835.62 MHz frequency division multiple access (FDMA) or 847.74 MHz code division multiple access (CDMA) modulated RF radiation. After the exposure to RF radiation, the micronucleus assay was performed by the cytokinesis block method using cytochalasin B treatment. The micronuclei appearing after mitosis were scored in binucleated cells using acridine orange staining. The frequency of micronuclei was scored both as the percentage of binucleated cells with micronuclei and as the number of micronuclei per 100 binucleated cells. Treatment of cells with cytochalasin B at a concentration of 2 microg/ml for 22 h was found to yield the maximum number of binucleated cells in C3H 10T(1/2) cells. The method used for the micronucleus assay in the present study detected a highly significant dose response for both indices of micronucleus production in the dose range of 0.1-1.2 Gy and it was sensitive enough to detect a significant (P > 0.05) increase in micronuclei after doses of 0.3 Gy in exponentially growing cells and after 0.9 Gy in plateau-phase cells. Exponentially growing cells or plateau-phase cells were exposed to CDMA (3.2 or 4.8 W/kg) or FDMA (3.2 or 5.1 W/kg) RF radiation for 3, 8, 16 or 24 h. In three repeat experiments, no exposure condition was found by analysis of variance to result in a significant increase relative to sham-exposed cells either in the percentage of binucleated cells with micronuclei or in the number of micronuclei per 100 binucleated cells. In this study, data from cells exposed to different RF signals at two SARs were compared to a common sham-exposed sample. We used the Dunnett's test, which is specifically designed for this purpose, and found no significant exposure-related differences for either plateau-phase cells or exponentially growing cells. Thus the results of this study are not consistent with the possibility that these RF radiations induce micronuclei.

The radiation-induced bystander effect for clonogenic survival.

It has long been accepted that the radiation-induced heritable effects in mammalian cells are the result of direct DNA damage. Recent evidence, however, suggests that when a cell population is exposed to a low dose of alpha particles, biological effects occur in a larger proportion of cells than are estimated to have been traversed by alpha particles. Experiments involving the Columbia University microbeam, which allows a known fraction of cells to be traversed by a defined number of alpha particles, have demonstrated a bystander effect for clonogenic survival and oncogenic transformation in C3H 10T(1/2) cells. When 1 to 16 alpha particles were passed through the nuclei of 10% of a C3H 10T(1/2) cell population, more cells were unable to form colonies than were actually traversed by alpha particles. Both hit and non-hit cells contributed to the outcome of the experiments. The present work was undertaken to assess the bystander effect of radiation in only non-hit cells. For this purpose, Chinese hamster V79 cells transfected with hygromycin- or neomycin-resistance genes were used. V79 cells stably transfected with a hygromycin resistance gene and stained with a nuclear dye were irradiated with the charged-particle microbeam in the presence of neomycin-resistant cells. The biological effect was studied in the neomycin-resistant V79 cells after selective removal of the hit cells with geneticin treatment.

The Bystander Effect in C3H 10T Cells and Radon-Induced Lung Cancer

Little, M. P. and Wakeford, R. The Bystander Effect in C3H 10T(1/2) Cells and Radon-Induced Lung Cancer. Radiat. Res. 156, 695-699 (2001).Bystander effects, whereby cells that are not directly exposed to radiation exhibit adverse biological effects, have been observed in a number of experimental systems, including C3H 10T(1/2) cells exposed to alpha-particle radiation. The bystander effect implies that risks from exposure to low doses of radiation obtained by linear extrapolation from data for high-dose exposures might be substantial underestimates. The best estimate of the ratio of the lung cancer risk among persons exposed to low (residential) doses of radon daughters to that among persons (underground miners) exposed to high doses of radon daughters is in the range of 2.4-4.0, with an upper 95% confidence limit of about 14. Assuming that the bystander effect observed in the in vitro C3H 10T(1/2) cell system applies to human lung cells in vivo, these epidemiological data imply that the central estimate of the number of neighboring cells that can contribute to the bystander effect is between 0 and 1, with an upper 95% confidence limit of about 7. As a consequence, the bystander effect observed in the experimental C3H 10T(1/2) cell system probably does not play a large part in the process of radon-induced lung carcinogenesis in humans.

Radiofrequency electromagnetic fields do not alter the cell cycle progression of C3H 10T and U87MG cells.

The effects of exposure to radiofrequency electromagnetic fields (RF EMFs) on cell cycle progression of mouse fibroblasts C3H 10T(1/2) and human glioma U87MG cells were determined by the flow cytometric bromodeoxyuridine pulse-chase method. Cells were exposed to a frequency-modulated continuous wave at 835.62 MHz or a code division multiple access RF EMF centered on 847.74 MHz at an average specific absorption rate of 0.6 W/kg. Five cell cycle parameters, including the transit of cells through G(1), G(2) and S phase and the probability of cell division, were examined immediately after the cells were placed in the fields or after they had been kept in the fields for up to 100 h. The only significant change observed in the study was that associated with C3H 10T(1/2) cell cultures moving into plateau phase toward the later times in the long-exposure experiment. No changes in the cell cycle parameters were observed in cells exposed to either mode of RF EMFs when compared to sham-exposed cells in either of the cell lines studied during the entire experimental period. The results show that exposure to RF EMFs, at the frequencies and power tested, does not have any effect on cell progression in vitro.

Measurement of DNA damage in mammalian cells exposed in vitro to radiofrequency fields at SARs of 3-5 W/kg.

In the present study, we determined whether exposure of mammalian cells to 3.2-5.1 W/kg specific absorption rate (SAR) radiofrequency fields could induce DNA damage in murine C3H 10T(1/2) fibroblasts. Cell cultures were exposed to 847.74 MHz code-division multiple access (CDMA) and 835.62 frequency-division multiple access (FDMA) modulated radiations in radial transmission line (RTL) irradiators in which the temperature was regulated to 37.0 +/- 0.3 degrees C. Using the alkaline comet assay to measure DNA damage, we found no statistically significant differences in either comet moment or comet length between sham-exposed cells and those exposed for 2, 4 or 24 h to CDMA or FDMA radiations in either exponentially growing or plateau-phase cells. Further, a 4-h incubation after the 2-h exposure resulted in no significant changes in comet moment or comet length. Our results show that exposure of cultured C3H 10T(1/2) cells at 37 degrees C CDMA or FDMA at SAR values of up to 5.1 W/kg did not induce measurable DNA damage.

Adaptive Response and the Bystander Effect Induced by Radiation in C3H 10T½ Cells in Culture

This paper discusses two phenomena of importance at low doses that have an impact on the shape of the dose-response relationship. First, there is the bystander effect, the term used to describe the biological effects observed in cells that are not themselves traversed by a charged particle, but are neighbors of cells that are; this exaggerates the effect of small doses of radiation. Second, there is the adaptive response, whereby exposure to a low level of DNA stress renders cells resistant to a subsequent exposure; this reduces the effect of low doses of radiation. The present work was undertaken to assess the relative importance of the adaptive response and the bystander effect induced by radiation in C3H 10T(1/2) cells in culture. When the single-cell microbeam delivered from 1 to 12 alpha particles through the nuclei of 10% of C3H 10T(1/2) cells, more cells were inactivated than were actually traversed by alpha particles. The magnitude of this bystander effect increased with the number of particles per cell. An adaptive dose of 2 cGy of gamma rays, delivered 6 h beforehand, canceled out about half of the bystander effect produced by the alpha particles.