Radiation Standards are currently driven by data derived using human epidemiology approaches. These methods lack the sensitivity needed to measure a change in cancer frequency following low levels of radiation exposures such as those associated with occupational or environmental exposures. Since it is not possible to measure changes in cancer frequency at low doses, the linear-no-threshold model (LNTH) is used to extrapolate cancer risks from high radiation dose levels to the risks from exposures at levels of concern. This LNTH extrapolation does not consider many biological processes that may limit the expression and repair of damage, The LNTH is used because it is thought to be conservative, but the hypothesis may or may not reflect biological reality. in the past, it has been very difficult to get information on the biological changes induced by low levels of radiation. However, with recent rapid advances in technology and cellular and molecular biology, and with improvement in detection capability, it is now possible to measure damage at level of exposure that were not possible in the past. It is important that these techniques and methods be applied to understanding biological effects at low levels of exposure so that our radiation standards are based on the best scientific data available. To meet this challenge, the DOE has initiated a research program focused on biological responses to low doses of low-LET radiation. This research may have major impact on changing some current radiation paradigms. These changes will be beneficial in construction of extrapolation models needed to estimate low-level radiation risk. This paper will briefly review the research being conducted, describe an example of the type of research progress that has been made and review the potential for the results of this research to alter current paradigms in radiation biology. Such paradigm changes have the potential to alter basic models of radiation induced cancer and could impact radiation standards.

PURPOSE:This review is to evaluate the use of biomarkers as an indication of past exposure to radiation or other environmental insults, individual sensitivity and risk for the development of late occurring disease.OVERVIEW:Biomarkers can be subdivided depending on their applications. Markers of exposure and dose can be used to reconstruct and predict past accidental or occupational exposures when limited or no physical measurements were available. Markers of risk or susceptibility can help identify sensitivity individuals that are at increased risk for development of spontaneous disease and may help predict the increased risk in sensitive individuals associated with environmental or therapeutic radiation exposures. Markers of disease represent the initial cellular or molecular changes that occur during disease development. Each of these types of biomarkers serves a unique purpose.OUTLINE:This paper concentrates on biomarkers of dose and exposure and provides a brief review of biomarkers of sensitivity and disease. The review of biomarkers of dose and exposure will demonstrate the usefulness of biomarkers in evaluation of physical factors associated with radiation exposure, such as LET, doserate and dose distribution. It will also evaluate the use of biomarkers to establish relationships that exist between exposure parameters such as energy deposition, environmental concentration of radioactive materials, alpha traversals and dose. In addition, the importance of biological factors on the magnitude of the biomarker response will be reviewed. Some of the factors evaluated will be the influence of species, tissue, cell types and genetic background. The review will demonstrate that markers of sensitivity and disease often have little usefulness in dose-reconstruction and, by the same token, many markers of dose or exposure may not be applicable for prediction of sensitivity or risk.

Because radon and its progeny (referred to collectively here as radon) emit alpha particles with a wide range of energies, as well as beta particles and gamma-rays, it is important to quantitate the relationship between initial damage induced by radon and that by acute low-LET radiation. We have evaluated dose-response relationships for induction of micronuclei both in vivo and in vitro following exposure to radon or 60Co. To determine if isolation procedures altered the cells' responsiveness to 60Co gamma-ray exposures, animals were exposed before cell isolation, or cells were isolated and then exposed. The data were described by linear dose-response functions and were not significantly different when the radiation exposure was in vivo or in vitro (respectively micronuclei/1000 binucleated cells = 1.6 +/- 6.5 + 62 +/- 2.7 D; micronuclei/1000 binucleated cells = 15.4 +/- 26.0 + 54.6 +/- 11.4 D, where D is in Gy). Primary rat lung fibroblasts (RLF) or Chinese hamster ovary (CHO-K1) cells were exposed in vitro to either radon or 60Co gamma-rays. Radon was 10.9 +/- 2.6 and 12.5 +/- 2.4 times as effective per Gy of radiation dose in producing micronuclei as was 60Co in RLF and CHO-K1 cells respectively. To determine the relative biological effectiveness of in vivo radon exposure, animals were exposed to either radon or 60Co, and lung fibroblasts were isolated and evaluated for radiation-induced micronuclei. In vivo radon exposure was 10.6 +/- 1.0 times as effective as acute whole-body 60Co exposure in producing micronuclei in lung fibroblasts. Different cell lines and exposure conditions resulted in similar effectiveness factors. Such ratios help evaluate the biological damage, hazard and risk associated with radon inhalation.

Genotoxic damage induced by radon and its progeny was investigated using the micronucleus assay in deep-lung fibroblasts to compare the response induced in vitro with that induced from inhalation of radon and its progeny in vivo. Male Wistar rats were exposed to 0, 115, 213 and 323 working-level months (WLM) of radon and its progeny by inhalation. After sacrifice, the cells were isolated and grown in culture, and the frequency of micronuclei was determined. A linear increase in the frequency of micronuclei was measured as a function of exposure [micronuclei/1000 binucleated cells = (29 +/- 9) + (0.47 +/- 0.04) WLM]. To compare exposure in WLM to dose in mGy, and to study how cell proliferation influences the way inhalation of radon and its progeny induces micronuclei, lung fibroblasts were isolated and exposed in vitro to graded doses from radon and its progeny after either 16 or 96 h in tissue culture. Cell cycle stage at the time of exposure was determined using flow cytometry. Primary lung fibroblasts exposed as either nondividing or dividing cells showed dose-dependent increases in micronuclei [micronuclei/1000 binucleated cells = (33 +/- 40) + (593 +/- 68)D and micronuclei/1000 binucleated cells = (27 +/- 69) + (757 +/- 88)D, respectively, where D is dose in Gy]. Results showed no significant influence (P = 0.20) of cell proliferation at the time of exposure on the frequency of micronuclei induced by radon and its progeny. Comparing dose-response relationships for nondividing cells to the exposure response for cells exposed by inhalation of radon and its progeny, it was estimated that a 1-WLM exposure in vivo caused the same amount of cytogenetic damage as produced by 0.79 mGy in vitro. In vivo/in vitro research using the micronucleus assay in lung fibroblasts serves as a powerful tool to estimate effective dose to cells in the respiratory tract after inhalation of radon and its progeny. Such studies form the basis for understanding the relationship between exposure, dose and biological damage.

Chromosome aberration frequency has been widely used to estimate dose in radiation accidents when no physical dosimetry was available. Basal and secretory cells are thought to represent the major cells at risk in the upper respiratory tract for induction of cancer. Using an in vivo/in vitro approach, we have evaluated chromosome aberrations to determine radiation sensitivity of basal and secretory cells and to estimate the dose to tracheal epithelial cells following radon inhalation. Tracheal epithelial cells were isolated, sorted by flow cytometry into populations of basal and secretory cells, grown in serum-free medium, and exposed to graded doses of x rays. Chromosome aberration frequency was determined in both types of cells. A 2-Gy dose of x rays induced 0.52 {+-} 0.07 or 0.55 {+-} 0.07 aberrations/cells in the basal and secretory cells, respectively. These data suggest that the radiation sensitivity is similar for the two cell populations and, because these are the cells that divide, the measured aberration frequencies can provide an estimate of radiation dose. After evaluating the response to x rays it was essential to determine the dose-response relationship for tracheal epithelial cells exposed to alpha particles. Tracheal cells were isolated from the trachea, allowed to attach tomore » a 3-{mu}m Mylar film, exposed to a {sup 238}Pu source, and grown in culture. The frequency of chromosome aberrations was then determined. One gray of alpha exposure produced 0.55 {+-} 0.06 aberrations/cell as well as 0.11 {+-} 0.04 dicentrics plus rings/cell. This provides a benchmark against which radon-induced aberrations in the trachea cells exposed to radon can be compared. In other experiments, two groups of F-344/N rats inhaled radon and its progeny at concentrations and times to produce total exposures of either 900 or 1000 working level months (WLM).« less

Silicon carbide whiskers (SiCW) and continuous glass filaments are important components of composite materials having potentially widespread use in the automotive, aerospace, and power generation industries. We determined the in vitro activity of three well-characterized samples of silicon carbide whiskers and a continuous glass filament sample in four different cellular assays and compared this to the activities of UICC crocidolite, JM Code 100 glass microfiber, and erionite in the same assay systems. The SiCW had a diameter range of 0.32-0.75 microns and a length range of 4.5-20.1 microns. The SiCW was significantly toxic; on a mass basis, one SiCW sample was more toxic than crocidolite; however, JM Code 100 glass microfiber, which is not toxic in vivo (i.e., it does not cause fibrogenesis or carcinogenesis when inhaled), was also more toxic than crocidolite. The glass filament sample was the least cytotoxic of all the samples tested. On a fiber number basis, all three SiCW samples were more toxic than crocidolite. The results of our study showed that SiCW exhibits significant in vitro biological reactivity. Thus, despite the caution that must be exercised in extrpolating the results of in vitro studies to conclusions about in vivo health effects, SiCW should be considered toxic until further toxicological data are available.

The physicochemical properties of particles influence their in vivo toxicity following deposition in the respiratory tract. To evaluate the relative contributions of mass and surface area to particle-induced toxicity, rat pulmonary alveolar macrophages (PAM) were exposed to four types of particles in vitro. We used three beryllium metal samples: relatively large (Be-II) and relatively small (Be-V) sized fractions of beryllium metal obtained from an aerosol cyclone, and a beryllium metal aerosol generated by laser vaporization of bulk beryllium metal in an argon atmosphere (Be-L). We also used glass beads (GB) as a negative control particle. End points examined included cell viability, determined by trypan blue dye exclusion, and changes in phagocytic ability, measured by counting the number of sheep red blood cells internalized by the PAM. Phagocytic ability was inhibited by exposure to beryllium particles at concentrations that did not cause appreciable cell death. Results describing effects based on the mass concentration of particles in culture medium were transformed by the amount of specific surface area of the particles to permit the expression of toxicity relative to the amount of particle surface per unit volume of culture medium. On a mass basis, the order of particle-related cytotoxicity was Be-L greater than Be-V greater than Be-II greater than GB, and for inhibition of phagocytosis, the order was Be-L approximately Be-V greater than Be-II greater than GB. When analyzed on a specific surface area basis, the cytotoxicity of the different materials became more similar in a fashion that was largely predicted by the amount of surface of the particles administered. However, because differences in specific surface area among the beryllium particle samples did not entirely predict cytotoxicity, we concluded that factors in addition to specific surface area influenced the expression of toxic effects in cultures of PAM exposed to beryllium metal.

Beryllium and nickel are toxic following deposition in the respiratory tract. Toxicity may be related to the bioavailability of these metals, which is in turn influenced by in vivo dissolution. This study was conducted to investigate the role of differing physical and chemical properties of beryllium and nickel compounds on their cytotoxicity to cell systems in vitro. We hypothesized that the two major factors influencing cytotoxicity are chemical form and specific surface area. Various forms of beryllium and nickel compounds were administered to cell cultures. The toxicity observed for particulate compounds was compared on the basis of particle mass and particle surface area. We found that for a given compound, toxicity was associated with the surface area of the compound available for interaction with cells.

Genetic toxicology studies were conducted on organic dyes and mixtures used in colored smoke munitions. The dyes studied included Solvent Red 1; two different batches (Lot 1 and Lot 2) of Disperse Red 11; terephthalic acid; and a mixture of 25 parts Solvent Red 1, 5 parts Disperse Red 11, and 16 parts terephthalic acid. The dyes were evaluated for their ability to produce mutations in Salmonella bacterial strains and in Chinese hamster ovary (CHO) cells. The dyes were also tested in CHO cells to determine cytotoxicity and the induction of sister chromatid exchanges and chromosome aberration. None of the dyes were genotoxic in the standard Ames assay using bacterial strain TA1535 or TA100 with or without the addition of S‐9 or in TA98 and TA1538 without S‐9. With S‐9, Disperse Red 11 (Lot 2) showed significant mutagenic activity in TA98 and TA1538 which increased as a function of S‐9 concentration. However, the maximum level of mutagenic activity detected was low (3.8 revertants/μg). The azo dye Solvent Red 1 was also negative in a pre‐incubation assay designed to reduce azo compounds to free amines. Solvent Red 1 was cytotoxic to mammalian cells, caused a significant increase in SCE, but was not mutagenic or clastogenic. Disperse Red 11 (Lot 1 and Lot 2) were not cytotoxic or clastogenic but produced an increase in cell cycle time and SCE frequency. Only Disperse Red 11 (Lot 2) increased mutations in the CHO/hypoxanthine‐guanine phosphoribosyltransferase (HGPRT) assay. The mutagenic activity of the dye mixture was not significant, suggesting no synergistic interaction between the dyes. These studies demonstrated that none of the dyes was clastogenic and that a contaminant in Disperse Red 11 (Lot 2) may be responsible for the weak mutagenic activity in both mammalian and bacterial cell systems.