Plutonium microdistribution in the lungs of Mayak workers.

The degree of nonuniform distribution of plutonium in the human lung has not been determined; thus current dosimetric models do not account for nonuniform irradiation. A better scientific basis is needed for assessing the risk of developing radiation-induced disease from inhaled alpha-particle-emitting radionuclides. We measured the distribution of plutonium activity in the lung by autoradiography and related the activity to specific compartments of the lung. The study materials were lung specimens from deceased workers employed by the Mayak Production Association. The approach to analyzing these lung samples used contemporary stereological sampling and analysis techniques together with quantitative alpha-particle autoradiography. For the first time, plutonium distribution has been quantified in the human lung. The distribution of long-term retained plutonium is nonuniform, and a significant portion of plutonium was retained in pulmonary scars. In addition, a large fraction of plutonium was present in the parenchyma, where it was retained much longer than was estimated previously. The sequestration of plutonium particles in scars would greatly reduce the radiation exposure of the critical target cells and tissues for lung cancer. Thus the prolonged retention of plutonium in lung scars may not increase the dose or risk for lung cancer.

Florida Red Tide: Inhalation Toxicity of Karenia brevis Extract in Rats.

Brevetoxins are neurotoxins produced by the marine dinoflagellate Karenia brevis. Histopathologic examination of marine mammals dying following repeated exposure of brevetoxins during red tide events suggests that the respiratory tract, nervous, hematopoietic, and immune systems are potential targets for toxicity in repeatedly exposed individuals. The purpose of this experiment was to evaluate the effects of repeated inhalation of K. brevis extract on these potential target systems in rats. Male Sprague-Dawley rats were exposed four hours/day, five days/week for up to four weeks to target concentrations of 200 and 1000 µg/L K. brevis extract (approximately 50 and 200 µg/L brevetoxin-like compounds; positive neurotoxicity in a fish bioassay). Control rats were sham exposed to air. Immunohistochemical staining of pulmonary macrophages indicated deposition of brevetoxin-like compound within the lung. However, exposure resulted in no clinical signs of toxicity or behavioral changes. There were no adverse effects on hematology or serum chemistry. No histopathological changes were observed in the nose, lung, liver, kidneys, lymph nodes, spleen, or brain of exposed rats. Immune suppression was suggested by reduced responses of spleen cells in the IgM-specific antibody-forming plaque cell response assay and reduced responses of lymphocytes to mitogen stimulation in vitro. Differences between responses observed in rats in this study and those observed in manatees may be a function of dose or species differences in sensitivity.

Distribution of plutonium particles in the lungs of Mayak workers.

Lung tissues from workers at the Mayak Production Association were examined to determine the distribution of plutonium (Pu) activity in various lung compartments. Stereological sampling methods and autoradiography were used. Pu particles were identified by microscopic examination of autoradiographs and localised in one of six normal anatomic sites and two sites of fibrosis (parenchymal, non-parenchymal). Particle activity was determined by counting the number of tracks emanating from the particles. Over 50% of the Pu activity was localised in sites of fibrosis, which had significantly higher than average activity for the lung. Over 40% of the activity was in lung parenchyma. Activity in the bronchovascular interstitium was significantly lower than average. These results support the hypothesis that Pu activity is not uniformly distributed in the lung, with long-term retained particles concentrated in scars of the lung. The results may significantly affect estimates of dose from inhaled Pu.

Modifying the ICRP 66 dosimetry model based on results obtained from Mayak plutonium workers.

Results obtained in a study of the microscopic distribution of plutonium in the lungs of deceased Pu workers from the Mayak Production Association showed that the long-term retention of Pu was greater than predicted by the current ICRP 66 respiratory tract dosimetry model (HRTM). These data were therefore applied to the HRTM by modifying selected parameters, namely the transfer rate of Pu from the transformed state compartment and the fraction of Pu that transfers to the bound state compartment. Invoking the latter compartment into the modelling allowed a better representation of the long-term Pu retention as well as providing a convenient means of describing the workplace-specific characteristics of the different Pu aerosols found in the Mayak plant. In particular, the present model describes a significantly greater long-term retention of Pu nitrate aerosols in the lung compared with the Type M default.

Effects of subchronic inhalation exposure of rats to emissions from a diesel engine burning soybean oil-derived biodiesel fuel.

There is increasing interest in diesel fuels derived from plant oils or animal fats ("biodiesel"), but little information on the toxicity of biodiesel emissions other than bacterial mutagenicity. F344 rats were exposed by inhalation 6 h/day, 5 days/wk for 13 wk to 1 of 3 dilutions of emissions from a diesel engine burning 100% soybean oil-derived fuel, or to clean air as controls. Whole emissions were diluted to nominal NO(x) concentrations of 5, 25, or 50 ppm, corresponding to approximately 0.04, 0.2, and 0.5 mg particles/m(3), respectively. Biologically significant, exposure-related effects were limited to the lung, were greater in females than in males, and were observed primarily at the highest exposure level. There was a dose-related increase in the numbers of alveolar macrophages and the numbers of particles in the macrophages, as expected from repeated exposure, but no neutrophil response even at the highest exposure level. The macrophage response was reduced 28 days after cessation of the exposure. Among the high-level females, the group mean lung weight/body weight ratio was increased, and minimal, multifocal bronchiolar metaplasia of alveolar ducts was observed in 4 of 30 rats. Lung weights were not significantly increased, and metaplasia of the alveolar ducts was not observed in males. An increase in particle-laden macrophages was the only exposure-related finding in lungs at the intermediate and low levels, with fewer macrophages and fewer particles per macrophage at the low level. Alveolar histiocytosis was observed in a few rats in both exposed and control groups. There were statistically significant, but minor and not consistently exposure-related, differences in body weight, nonpulmonary organ weights, serum chemistry, and glial fibrillary acidic protein in the brain. There were no significant exposure-related effects on survival, clinical signs, feed consumption, ocular toxicity, hematology, neurohistology, micronuclei in bone marrow, sister chromatid exchanges in peripheral blood lymphocytes, fertility, reproductive toxicity, or teratology. This study demonstrated modest adverse effects at the highest exposure level, and none other than the expected physiological macrophage response to repeated particle exposure at the intermediate level.

Assessing the uniformity of plutonium alpha radiation dose in human lung: the Mayak experience.

Radiation-induced lung cancer risk is currently estimated based on epidemiological data from populations exposed either to relatively uniform, low-LET radiation, or from uranium miners who inhaled radon and its progeny. Inhaled alpha-emitting radionuclides (e.g. Pu and Am) produce distinctive dose patterns that may not be adequately modelled at present. Thus the distribution of Pu is being measured in formalin-fixed autopsy lung tissue from former workers at the Mayak Production Association, and which is maintained in a tissue archive at SUBI. Lungs are sampled using contemporary stereological techniques and Pu particle activities and locations are determined using quantitative autoradiography and morphological identification of lung structures. To date, > 80% of Pu particles have been observed in parenchymal lung tissues with higher concentrations being found in scar tissue. Concentrations of Pu particles in conducting airways are uniformly low, thus indicating that long-term-retained Pu particles are non-uniformly distributed in human lung, mostly in the parenchyma.

Influence of exposure concentration or dose on the distribution of particulate material in rat and human lungs.

Differences among species in the anatomic sites of particle retention could influence responses to inhaled particles. In this study, we used morphometric techniques to examine the influence of exposure concentration on particle retention in histologic sections from rats and humans. The rats had been exposed for 24 months to diesel exhaust at 0.35, 3.5, or 7.0 mg soot/m(3). The human subjects were nonsmokers who did not work as miners, nonsmoking coal miners who worked under the current standard of 2 mg dust/m(3) for 10-20 years (mean = 14 years), and nonsmoking coal miners who worked under the former standard of < 10 mg dust/m(3) for 33-50 years (mean = 40 years). The distribution of retained particles within the lung compartments was markedly different between species. In all three groups of rats, 82-85% of the retained particulate material was located in the alveolar and alveolar duct lumens, primarily in macrophages. In humans, 57, 68, and 91% of the retained particulate material was located in the interstitium of the lung in the non-miners, coal miners under the current standard, and coal miners under the former standard, respectively. These results show that chronically inhaled diesel soot is retained predominantly in the airspaces of rats over a wide range of exposures, whereas in humans, chronically inhaled particulate material is retained primarily in the interstitium. In humans, the percentage of particles in the interstitium is increased with increased dose (exposure concentration, years of exposure, and/or lung burden). This difference in distribution may bring different lung cells into contact with the retained particles or particle-containing macrophages in rats and humans and may account for differences in species response to inhaled particles.

The toxicity of insoluble cerium-144 inhaled by beagle dogs: non-neoplastic effects.

The biological effects of inhaled beta-particle-emitting radionuclides are not well known. The non-neoplastic diseases induced by an inhaled, relatively insoluble form of cerium-144 ((144)Ce) were studied in beagle dogs exposed to graded activity levels of (144)Ce in fused aluminosilicate particles by a single, brief inhalation exposure and observed for their life span. The initial lung burdens (ILBs) achieved ranged from 0.000093-7.6 MBq (144)Ce/kg body weight. The (144)Ce was retained in the lung with an effective half-life of about 190 days. Significant (144)Ce was translocated to the tracheobronchial lymph nodes, and the concentration exceeded that of the lung at about 400 days after inhalation exposure. Significant radiation doses were delivered to the lung and tracheobronchial lymph nodes and to the heart adjacent to the tracheobronchial lymph nodes. Radiation pneumonitis was the predominant non-neoplastic disease. The dose response for radiation pneumonitis indicated that an ILB of 1.4 MBq/kg would cause death from radiation pneumonitis in 50% of the exposed dogs. This ILB resulted in a pulmonary dose to death of about 350 Gy. The tracheobronchial lymph nodes developed lesions in dogs with ILBs lower than those causing radiation pneumonitis. The overall results of this study, however, showed that (144)Ce, inhaled in an insoluble form, did not cause any unique or inexplicable biological effects in dogs or cause effects at unusually low doses that might call current radiation protection guidelines into question.

1,3-butadiene: cancer, mutations, and adducts. Part I: Carcinogenicity of 1,2,3,4-diepoxybutane.

Reports in the literature suggest that one reason for the greater sensitivity of mice to the carcinogenicity of 1,3-butadiene (BD) is that exposed mice metabolize much more of the BD to 1,2,3,4-diepoxybutane (BDO2) than do exposed rats. The purpose of this study was to determine the tumorigenicity of BDO2 in rats and in mice exposed to the same concentration of the agent. Female B6C3F1 mice and Sprague-Dawley rats, 10 to 11 weeks old, 56 per group, were exposed by inhalation to 0, 2.5, or 5.0 ppm BDO2, 6 hours/day, 5 days/week for 6 weeks. Preliminary dosimetry studies in rodents exposed for 6 hours to 12 ppm BDO2 indicated that blood levels would be expected to be approximately 100 and 200 pmol/g at the two exposure concentrations in the rat and twice those levels in the mouse. During the 6-week exposure, the mice at the high exposure level showed signs of labored breathing during the last week, and four mice died. In the others, however, the respiratory symptoms disappeared after exposure ended. Rats showed no clinical signs of toxicity during exposure but developed labored breathing after the end of the exposure leading to the death of 13 rats within 3 months. At the end of the exposure, some animals (8 per group) were evaluated for the acute toxicity resulting from the BDO2 exposure. The remaining exposed rats and mice were held for 18 months for observation of tumor development. At the end of the exposure, rats had no biologically significant alteration in standard hematological parameters, but mice had a dose-dependent increase in neutrophils and decrease in lymphocytes. In both species the significant histopathologic lesions were in the nose, concentrated around the main airflow pathway. Necrosis, inflammation, and squamous metaplasia of the nasal mucosa, as well as atrophy of the turbinates, were all present at the end of exposure to 5.0 ppm. Within 6 months, necrosis and inflammation subsided, but squamous metaplasia remained in the mice. In rats that died after exposure, squamous metaplasia was seen in areas of earlier inflammation and, in other rats, extended beyond those areas with time. The metaplasia was severe enough to restrict and occlude the nasopharyngeal duct. Later, keratinizing squamous cell carcinomas developed from the metaplastic foci in rats but not mice. At the end of 18 months, the only significant increase in neoplasia in the exposed rats was a dose-dependent increase in neoplasms of the nasal mucosa (0/47, 12/48, and 21/48 for the control, 2.5 ppm, and 5.0 ppm exposures, respectively). Neoplasia of the nasal mucosa did not increase significantly in the mice; neoplastic lesions in the mice were observed in reproductive organs, lymph nodes, bone, liver, Harderian gland, pancreas, and lung. The only significant increase in neoplasms in a single organ in the mice was in the Harderian gland (0/40, 2/42, and 5/36 for the control, 2.5 ppm, and 5.0 ppm exposures, respectively). This tumor accounts for the apparent trend toward an increase in total neoplastic lesions in mice as a function of dose (10/40, 7/42, and 16/36 for control, 2.5 ppm, and 5.0 ppm exposures, respectively). These findings indicate that the metabolite of BD, BDO2, is carcinogenic in the respiratory tract of rats. An increase in respiratory tract tumors was not observed in similarly exposed mice despite the fact that preliminary studies indicated mice should have received twice the dose to tissue compared with the rats. High cytosolic activity of detoxication enzymes in the mouse may account, in part, for the differences in response.

Region-specific age at onset of beta-amyloid in dogs.

Cortical patterns of beta-amyloid (Abeta) deposition were evaluated in 40 beagle dogs ranging in age from 2 to 18 years. Abeta deposition in the prefrontal, occipital, parietal and entorhinal cortices was visualized by using an antibody against Abeta1-42. A logistic regression was used to estimate differences in age-at-onset and rate of deposition of Abeta as a function of brain region. The earliest and most consistent site of Abeta deposition with age was in the prefrontal cortex. Entorhinal Abeta deposition was not consistently observed until the age of 14 years, but was present in a subset of dogs under the age of 14 years. These regional vulnerabilities to Abeta accumulation are similar to those seen in the aging human. By using parameters derived from regression analyses, it may be possible to predict the presence of Abeta within specific brain regions in individual dogs. We propose that these models will be a useful tool to evaluate interventions that delay the age of onset or slow the rate of accumulation of Abeta in the dog.

Animal models of emphysema and their relevance to studies of particle-induced disease.

Emphysema is a pulmonary disease that may be exacerbated by inhaled particles. Over the years, many animal models of emphysema have been developed that may be useful in studying the effects of inhaled particles on humans with emphysema. Models have been described in many species, and many approaches have been described for inducing emphysema. Emphysema in humans is a parenchymal component of chronic obstructive pulmonary disease and frequently coexists in a complex with disease of the airways such as bronchitis. Animal models of emphysema usually recapitulate only one or a few aspects of this complex disease. Thus, the emphysema model must be selected carefully in order to answer specific questions about the interactive effects of particles and emphysema.

Carcinogenicity of inhaled butadiene diepoxide in female B6C3F1 mice and Sprague-Dawley rats.

Previous studies suggest that the greater sensitivity of mice, compared to rats, to the carcinogenicity of 1,3-butadiene (BD) is linked to higher rates of BD metabolism to butadiene diepoxide (BDO2) by mice than rats. The purpose of this study was to determine the tumorigenicity of BDO2 in mice and rats exposed by inhalation to the same concentrations of the agent. Female B6C3F1 mice and Sprague-Dawley rats, 10-11 weeks old, 56/group, were exposed to 0, 2.5, or 5.0 ppm BDO2, 6 h/day, 5 days/week for 6 weeks. At the end of the BDO2 exposure, 8 animals/group were evaluated for toxicity. The remainder of the exposed rats and mice were held for up to 18 months for observation of tumor development. At the end of the exposure, rats had no biologically significant alteration in standard hematological parameters, but mice had a dose-dependent increase in neutrophils and decrease in lymphocytes. Most of the significant lesions in both species were in the nose, concentrated around the main airflow pathway. Necrosis, inflammation, and squamous metaplasia of the nasal mucosa, as well as atrophy of the turbinates, were all present in animals exposed to 5.0 ppm. In mice, necrosis and inflammation subsided within 6 months, but squamous metaplasia remained. In rats that died after exposure, squamous metaplasia was seen in areas of earlier inflammation and extended beyond those areas with time. The metaplasia was severe enough to restrict and occlude the nasopharyngeal duct. Later, keratinizing squamous-cell carcinomas developed from metaplastic foci in rats, but these were not seen in mice. At the end of 18 months, the only significant increase in neoplasia in the exposed rats was a dose-dependent increase in neoplasms of the nasal mucosa (0/47, 12/48, and 21/48 for the control, 2.5 ppm, and 5.0 ppm exposures, respectively). Neoplasia of the nasal mucosa did not increase significantly in the mice. Neoplastic lesions in the mice were observed in reproductive organs, lymph nodes, bone, liver, Harderian gland, pancreas, and lung, but the only significant increase in neoplasms in a single organ in the mice was in the Harderian gland (0/40, 2/42, and 5/36 for the control, 2.5 ppm, and 5.0 ppm exposures, respectively). This tumor accounts for the apparent trend toward an increase in total neoplastic lesions in mice as a function of dose (10/40, 7/42, and 16/36 for control, 2.5 ppm, and 5.0 ppm, respectively). These findings indicate that the metabolite of BD, BDO2, is carcinogenic in the upper respiratory tract of rats. An increase in upper respiratory tract tumors was not observed in similarly exposed mice, despite the fact that preliminary studies indicated mice should have received twice the dose to tissue than did the rats. Higher cytosolic activity of detoxication enzymes has been reported in the liver and lung cells of the mouse compared to the rat, and this may account, in part, for the differences in response. The transport of externally administered BDO2, into the cell and through the cytoplasm, might allow detoxication of the molecule before it reaches critical sites on the DNA. The results indicate that the site of formation of the BDO2 is important for tumor induction.

Comparative stochastic effects of inhaled alpha- and beta-particle-emitting radionuclides in beagle dogs.

The stochastic effects of inhaled, insoluble particles of alpha- or beta-particle-emitting radionuclides were compared in dogs. Male and female beagle dogs were exposed briefly by nasal inhalation to relatively insoluble aerosols of (239)PuO(2) or (144)Ce in fused aluminosilicate particles (FAP) and observed for cancer for their lifetimes. The initial lung burden and retention of each radionuclide was determined by whole-body counting of the emissions from (144)Ce-(144)Pr- or (169)Yb-labeled (239)PuO(2). Lung doses were calculated for each dog from these data. The lung doses ranged from 0.21 to 1200 Gy for (144)Ce FAP and 1.6 to 58 Gy for (239)PuO(2). Dogs with doses to the lung of about 60 Gy or greater from (144)Ce or about 2 Gy or greater from (239)PuO(2) had an increased incidence of lung carcinomas. In dogs exposed to (144)Ce FAP, three organs were targets for neoplasia: lung, tracheobronchial lymph nodes, and heart. The insoluble FAP carried to the lymph nodes draining the lung delivered high radiation doses to the nodes and adjacent heart, resulting in hemangiosarcomas of these organs. In the lung, high radiation doses induced hemangiosarcomas and carcinosarcomas. At lower doses, carcinomas of various histological patterns were induced in the lung. In dogs exposed to (239)PuO(2), the lung was the sole target organ for neoplasia. Nearly all of these neoplasms were carcinomas of various histological patterns. These results indicated that relatively low doses of alpha-particle radiation can induce pulmonary cancers, but relatively large doses of beta-particle radiation are required. In addition, inhaled beta-particle emitters can also induce cancers in lung-associated lymph nodes and heart at these larger absorbed radiation doses.

Comparative deterministic effects of inhaled, insoluble alpha- and beta-particle-emitting radionuclides in dogs.

This report compares the deterministic effects from an alpha-particle-emitting radionuclide, (239)PuO(2), and a beta-particle emitter, (144)Ce in fused aluminosilicate particles (FAP). The studies were conducted in beagle dogs of both genders exposed by inhalation to aerosols of the radionuclides. The initial lung burdens of (239)Pu and (144)Ce were determined by whole-body counting of the (169)Yb added to the plutonium aerosol during its preparation or the (144)Ce and its progeny (144)Pr. In addition, organ retention data were obtained from parallel serial sacrifice studies with the same aerosols. After exposure, the dogs were observed for health effects over their lifetime. The deterministic effects observed for both of these relatively insoluble aerosols were lymphopenia, fibrosis, atrophy of the lung-associated lymph nodes, and radiation pneumonitis. Due to the longer half-life of plutonium, the lymphopenia was more prolonged and the clinical course of the radiation pneumonitis more chronic than that resulting from cerium. The greater tissue penetration of the beta-particle emissions from the cerium resulted in more uniform dose distribution over the lung and the atria of the heart than from the alpha-particle emissions from plutonium.

Cigarette smoke exposure produces more evidence of emphysema in B6C3F1 mice than in F344 rats.

Cigarette smoke (CS) causes pulmonary emphysema in humans, but results of previous studies on CS-exposed laboratory animals have been equivocal and have not clearly demonstrated progression of the disease. In this study, morphometry and histopathology were used to assess emphysema in the lungs of B6C3F1 mice and Fischer-344 rats. The animals were exposed, whole-body, to CS at a concentration of 250 mg total particulate matter/m3 for 6 h/day, 5 days/week, for either 7 or 13 months. Morphometry included measurements of parenchymal air space enlargement (alveolar septa mean linear intercept [Lm], volume density of alveolar air space [VVair]), and tissue loss (volume density of alveolar septa [VVspt]). In addition, centriacinar intra-alveolar inflammatory cells were counted to assess species differences in the type of inflammatory response associated with CS exposure. In mice, many of the morphometric parameters indicating emphysema differed significantly between CS-exposed and control animals. In CS-exposed rats, only some of the parameters differed significantly from control values. The Lm in both CS-exposed mice and rats was increased at 7 and 13 months, indicating an enlargement of parenchymal air spaces, but the VVair was increased significantly only in CS-exposed mice. The VVspt was decreased at both time points in mice, but not in rats, indicating damage to the structural integrity of parenchyma. Morphologic evidence of tissue destruction in the mice included alveoli that were irregular in size and shape and alveoli with multiple foci of septal discontinuities and isolated septal fragments. Morphometric differences in the mice at 13 months were greater than at 7 months, suggesting a progression of the disease. Inflammatory lesions within the lungs of mice contained significantly more neutrophils than those lesions in rats. These results suggest that B6C3F1 mice are more susceptible than F344-rats to the induction of emphysema by this CS exposure regimen and that in mice the emphysema may be progressive. Furthermore, the type of inflammatory response may be a determining factor for species differences in susceptibility to emphysema induction by CS exposure.

Dosimetry and acute toxicity of inhaled butadiene diepoxide in rats and mice.

Butadiene diepoxide (BDO2), a metabolite of 1,3-butadiene (BD) and potent mutagen, is suspected to be a proximate carcinogen in the multisite tumorigenesis in B6C3F1 mice exposed to BD. Rats, in contrast to mice, do not form much BDO2 when exposed to BD, and they do not form cancers after exposure to the low levels of BD at which mice develop lung and heart tumors. Tests were planned to determine the direct carcinogenic potential of BDO2 in similarly exposed rats and mice, to see if they would develop tumors of the lung (the most sensitive target organ in BD-exposed mice) or other target tissues. The objective of the current series of studies was to assess the acute toxicity and dosimetry to blood and lung of BDO2 administered by various routes to B6C3F1 mice and Sprague-Dawley rats. The studies were needed to aid in the design of the carcinogenesis study. Initial studies using intraperitoneal injection of BDO2 were designed to determine the rate at which each of the species cleared the compound from the body; the clearance was equally fast in both species. A second study was designed to determine if the highly reactive BDO2, when deposited in the lung, would enter the bloodstream from the lung; intratracheally instilled BDO2 did enter the bloodstream, indicating that exposure via the lungs would result in BDO2 reaching other organs of the body. In a third study, rats and mice were exposed by inhalation for 6 h to 12 ppm BDO2 to determine blood and lung levels of the compound. Concentrations of BDO2 in the lung immediately after the exposure were 2 to 3 times higher than in the blood in both species (approximately 500 and 1000 pmol/g blood in the rat and mouse, respectively). As expected, mice received a higher dose/g tissue than did rats, consistent with the higher minute volume/kg body weight of the mice. The inhalation dosimetry study was followed by a histopathology study to determine the acute toxicity to rodents following a single, 6-h exposure to 18 ppm BDO2. No clinical signs of toxicity were observed; lesions were confined to the olfactory epithelium where areas of necrosis were observed. Analysis of bronchoalveolar lavage fluid did not indicate pulmonary inflammation. Based on these findings, an attempt was made to expose rats and mice repeatedly (for 7 days) to 10 and 20 ppm BDO2, but these exposure concentrations proved too toxic, due to inflammation of the nasal mucosa and occlusion of the nasal airway, a lesion that cannot be tolerated by obligate nose breathers. Finally, the toxicity of rats and mice exposed 6 h/day for 5 days to 0, 2.5, or 5.0 ppm BDO2 was determined. The repeated exposures caused no clinical signs of toxicity, nor were any lesions observed in the respiratory tract or other major organs. Therefore, the final design selected for the carcinogenesis study comprised exposing the rats and mice for 6 h/day, 5 days/week for 6 weeks to 0, 2.5, or 5.0 ppm BDO2.

Chromatin clearance in C57Bl/10 mice: interaction with heparan sulphate proteoglycans and receptors on Kupffer cells.

Chromatin is an important autoantigen in the pathogenesis of systemic lupus erythematosus (SLE) as an immunogen and as a part of nephritogenic immune complexes. Earlier studies focused on clearance of DNA. However, DNA released into the circulation from dying cells is found associated with histones in nucleosomes. The liver is the major organ involved in clearance of chromatin from the circulation of mice. Heparan sulphate proteoglycans (HSPG) have been implicated in the clearance of various charged molecules. Receptor-mediated clearance of ssDNA by the liver has also been reported. Because chromatin contains positively charged histones in addition to DNA, we wished to determine if HSPG and/or DNA receptors are involved in chromatin clearance. The rate of clearance of H1-stripped chromatin from the bloodstream of C57Bl/10 mice was markedly decreased by prior treatment of mice with Heparinase I. Clearance was also inhibited by heparin, heparan sulphate, and DNA, but not by colominic acid. DNA was the most effective inhibitor of clearance and released chromatin from sites of clearance. Depletion of Kupffer cells and splenic macrophages using liposome-encapsulated Clodronate (dichloromethylene bisphosphonate) markedly inhibited chromatin clearance. These data suggest that chromatin clearance is mediated by charge interactions with cell surface HSPG and by DNA receptors. Clearance and degradation of chromatin require functional macrophages in the liver and spleen.

Toxicity of inhaled 91YCl3 in dogs.

This study was conducted in dogs to determine the toxicity of inhaled 91YCl3, which is of interest because 91Y is a fission-product radionuclide that is abundant in a reactor inventory after sustained operation. Yttrium-91 has a short half-life, 59 days, and decays with the emission of beta particles and low-yield gamma rays. The study was conducted in 58 beagle dogs with equal numbers of males and females. Forty-six dogs inhaled the 91YCl3 aerosol, while 12 served as controls. Four exposure levels were used. To determine the long-term retained burden (LTRB) of 91Y, each dog was periodically whole-body counted and its excreta were analyzed radiochemically. Over time, the 91Y transferred from the lung primarily to the skeleton and liver. The dogs were observed over their life spans for biological effects. Fatal hematological dyscrasia occurred from 12 to 33 days after exposure in the dogs with the highest LTRBs. Bone-associated tumors of the nasal and oral mucosae occurred in 5 dogs from 2000 to 5800 days after they inhaled the 91YCl3 aerosols. Five dogs died with malignant lung tumors and 2 dogs with malignant liver tumors. The results of this study were compared to those from similar studies in beagles that inhaled 90SrCl2 or 144CeCl3 or were injected with 137CsCl. The comparison showed that the biological effects in each study were clearly dependent on the cumulative doses to critical organs.

Carcinogenic responses of transgenic heterozygous p53 knockout mice to inhaled 239PuO2 or metallic beryllium.

The transgenic heterozygous p53+/- knockout mouse has been a model for assessing the tumorigenicity of selected carcinogens administered by noninhalation routes of exposure. The sensitivity of the model for predicting cancer by inhaled chemicals has not been examined. This study addresses this issue by acutely exposing p53+/- mice of both sexes by nose-only inhalation to either air (controls), or to 1 of 2 levels of 239PuO2 (500 or 100 Bq 239Pu) or beryllium (Be) metal (60 or 15 micrograms). Additional wild-type p53+/+ mice were exposed by inhalation to either 500 Bq of 239PuO2 or 60 micrograms of Be metal. These carcinogens were selected because they operate by differing mechanisms and because of their use in other pulmonary carcinogenesis studies in our laboratory. Four or 5 of the 15 mice per sex from each group were sacrificed 6 mo after exposure, and only 2 pulmonary neoplasms were observed. The remainder of the mice were held for life-span observation and euthanasia as they became moribund. Survival of the p53+/- knockout mice was reduced compared to the p53+/+ wild-type mice. No lung neoplasms were observed in p53+/- mice exposed to air alone. Eleven of the p53+/- mice inhaling 239PuO2 developed pulmonary neoplasms. Seven p53+/+ mice exposed to 239PuO2 also developed pulmonary neoplasms, but the latency period for pulmonary neoplasia was significantly shorter in the p53+/ mice. Four pulmonary neoplasms were observed in p53+/- mice exposed to the higher dose of Be, whereas none were observed in the wild-type mice or in the heterozygous mice exposed to the lower dose of Be. Thus, both p53+/- and p53+/+ mice were susceptible to 239Pu-induced carcinogenesis, whereas the p53+/- but not the p53+/+ mice were susceptible to Be-induced carcinogenesis. However, only 2 pulmonary neoplasms (1 in each of the 239PuO2 exposure groups) were observed in the 59 p53+/ mice that were sacrificed or euthanatized within 9 mo after exposure, indicating that the p53+/- knockout mouse might not be appropriate for a 6-mo model of carcinogenesis for these inhaled carcinogens.

Statistical modeling of carcinogenic risks in dogs that inhaled 238PuO2.

Combined analyses of data on 260 life-span beagle dogs that inhaled 238PuO2 at the Inhalation Toxicology Research Institute (ITRI) and at Pacific Northwest National Laboratory (PNNL) were conducted. The hazard functions (age-specific risks) for incidence of lung, bone and liver tumors were modeled as a function of cumulative radiation dose, and estimates of lifetime risks based on the combined data were developed. For lung tumors, linear-quadratic functions provided an adequate fit to the data from both laboratories, and linear functions provided an adequate fit when analyses were restricted to doses less than 20 Gy. The estimated risk coefficients for these functions were significantly larger when based on ITRI data compared to PNNL data, and dosimetry biases are a possible explanation for this difference. There was also evidence that the bone tumor response functions differed for the two laboratories, although these differences occurred primarily at high doses. These functions were clearly nonlinear (even when restricted to average skeletal doses less than 1 Gy), and evidence of radiation-induced bone tumors was found for doses less than 0.5 Gy in both laboratories. Liver tumor risks were similar for the two laboratories, and linear functions provided an adequate fit to these data. Lifetime risk estimates for lung and bone tumors derived from these data had wide confidence intervals, but were consistent with estimates currently used in radiation protection. The dog-based lifetime liver tumor risk estimate was an order of magnitude larger than that used in radiation protection, but the latter also carries large uncertainties. The application of common statistical methodology to data from two studies has allowed the identification of differences in these studies and has provided a basis for common risk estimates based on both data sets.