The role of macrophages in particle translocation from lungs to lymph nodes.

Red fluorescent and green fluorescent microspheres were instilled into separate but adjacent areas of dog lung lobes. After 7 days, the tracheobronchial lymph nodes that drained both of the instilled areas contained many macrophages with all red or all green microspheres but rarely both. This indicates that the particles did not translocate passively and that lung macrophages phagocytized the microspheres in the lung and carried them to the tracheobronchial lymph nodes. In addition, two populations of pulmonary alveolar macrophages (PAM's), one that had phagocytized red microspheres in vivo and one that had phagocytized green microspheres, were lavaged from the lungs of dogs, mixed into one population, and instilled back into a previously unexposed lung lobe of the same dogs. As in the first experiment, the tracheobronchial lymph nodes that drained the instilled area contained numerous macrophages with either all red or all green microspheres. This suggested that the instilled PAM's had migrated to the tracheobronchial lymph nodes. Thus, lung macrophages, including PAM's probably play a critical role in the induction of lung immunity and in protection from disease by determining particle translocation.

Single inhalation exposure to 90SrCl2 in the beagle dog: late biological effects.

Late-occurring biologic effects were studied in beagle dogs that were given graded levels of 90SrCl2 via single brief inhalation exposures and were subsequently observed for their life-span. Due to the soluble chemical form of the aerosol, 90Sr was rapidly translocated from lung and deposited in bone where it was subsequently retained for a long period of time. Radiation-induced lesions were confined to the bone, bone marrow, and adjacent soft tissue. Forty-five primary bone tumors occurred in 31 of 66 exposed dogs. Metastasis occurred from 21 tumors, with the lung being the most frequent site of metastasis (76%). Twenty-seven tumors were classified as different subtypes of osteosarcoma, 14 as hemangiosarcomas, 3 as fibrosarcomas, and 1 as a myxosarcoma. Four carcinomas arising from soft tissues adjacent to bone were also considered to be 90Sr induced. In contrast to bone tumors arising in beagles chronically exposed to 90Sr through ingestion, histologic lesions of radiation osteodystrophy were minimal in this study, indicating that these lesions are not a necessary precursor of osteosarcoma development. The incidences of hemangiosarcomas (31%) and telangiectatic osteosarcomas (11%) in addition to osteosarcomas suggest that the cell of origin for all of these neoplasms is a multipotent mesenchymal cell with the potential for various morphologic expressions dependent on local environmental factors.

A relevant dose of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone is extensively metabolized and rapidly absorbed in the canine tracheal mucosa.

Lung cancer is largely a site-of-entry disease caused by inhaled carcinogenic agents, especially tobacco smoke. Two major groups of procarcinogens, tobacco-specific nitrosamines and polycyclic aromatic hydrocarbons, are putative agents, but their relative contributions are disputed. An important indicator of relative potency for these compounds is the dose to the target epithelial cells. Although we have reported the dose of polycyclic aromatic hydrocarbons to the canine tracheal epithelium [Gerde et al., Carcinogenesis (Lond.), 18: 1825-1832, 1997; Gerde et al., Carcinogenesis (Lond.), in press, 1998], the purpose of the current study was to characterize the absorption and metabolism of low levels of one tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), in the canine trachea. One hundred ng of tritiated NNK were instilled in the distal trachea of the dog. Blood was repeatedly sampled from the azygous vein and both sides of the systemic circulation from 15 s to 30 min after instillation. Tissues were then removed and analyzed for the tritiated NNK and its metabolites. Autoradiography was used to determine the depth distribution of tritium in the tracheal mucosa. Most NNK appeared rapidly in the blood draining the airway mucosa, but there was also a slow clearance phase. During absorption, NNK was distributed within the entire depth of the mucosa to the tracheal cartilage; however, a portion was conspicuously bound to the mucin component of the mucous lining layer. Reversible binding to mucin may be largely responsible for the slow clearance phase. Despite the rapid absorption of most of the tritium, NNK was nonetheless extensively metabolized in the tracheal mucosa. Systemic metabolism was also rapid: within 18 min of instillation, the NNK parent compound had disappeared from the systemic circulation, and 45 min after instillation, no NNK was found in the trachea or any distal tissue. Although the rapid absorption and distribution of NNK and its metabolites ensured widespread and extensive distal binding in all tissues, first-pass metabolism and activation of NNK in the airway mucosa were sufficiently rapid to cause levels of binding at the site of absorption to be approximately 20-fold those of distal tissues. NNK may thus act as a site-of-entry carcinogen. This observation may be important in estimating the contribution of NNK to lung cancer relative to other carcinogens and for explaining increased incidences of oral cancers in users of snuff and chewing tobacco in which NNK is present in high concentrations.

Learning ability in aged beagle dogs is preserved by behavioral enrichment and dietary fortification: a two-year longitudinal study.

The effectiveness of two interventions, dietary fortification with antioxidants and a program of behavioral enrichment, was assessed in a longitudinal study of cognitive aging in beagle dogs. A baseline protocol of cognitive testing was used to select four cognitively equivalent groups: control food-control experience (C-C), control food-enriched experience (C-E), antioxidant fortified food-control experience (A-C), and antioxidant fortified food-enriched experience(A-E). We also included two groups of young behaviorally enriched dogs, one receiving the control food and the other the fortified food. Discrimination learning and reversal was assessed after one year of treatment with a size discrimination task, and again after two years with a black/white discrimination task. The four aged groups were comparable at baseline. At one and two years, the aged combined treatment group showed more accurate learning than the other aged groups. Discrimination learning was significantly improved by behavioral enrichment. Reversal learning was improved by both behavioral enrichment and dietary fortification. By contrast, the fortified food had no effect on the young dogs. These results suggest that behavioral enrichment or dietary fortification with antioxidants over a long-duration can slow age-dependent cognitive decline, and that the two treatments together are more effective than either alone in older dogs.

Comparison of cell-mediated and humoral immunity in the dog lung after localized lung immunization.

This study evaluated cell-mediated immunity (CMI) and antibody production serially in control and immunized lung lobes of beagle dogs. A fiberoptic bronchoscope was used to immunize selected airways in the left cardiac lung lobe with 10(10) sheep red blood cells (SRBC); the right cardiac lobe received saline as control. The immune responses produced by this localized lung immunization were evaluated in cells and fluids obtained from blood and serial bronchial washings of the immunized and control lung lobes from 5 to 21 days after immunization. The antigen-specific production of procoagulant activity (LPCA) and inhibition of migration of alveolar macrophages by SRBC antigen were used to measure CMI in lavage cells from immunized and control lung lobes. The level of specific IgM and IgG antibody in lavage fluid from the control and immunized lung lobes was evaluated with the enzyme-linked immunosorbent assay (ELISA). The results of this study showed a significantly greater percentage of neutrophils and lymphocytes in the lavage fluid from the immunized lung lobes than from the control lung lobes. The increased number of lymphocytes in the immunized lung lobes showed a positive correlation with increased antibody concentrations. In contrast to the antibody response, CMI as measured by LPCA and MIF assays was positive, with nearly equal responses in control and immunized lung lobes. Even though there were only a few lymphocytes in the control lung lobes, there were apparently enough specific immune lymphocytes present that produced mediators after antigen stimulation to result in positive CMI responses.

The effect of local antibody responses on in vivo and in vitro phagocytosis by pulmonary alveolar macrophages.

This study used both in vitro and in vivo techniques to determine if local antigen deposition in the lung has a localized effect on immune phagocytosis by pulmonary alveolar macrophages (PAM). Using a fiberoptic bronchoscope, dogs were immunized in the left cardiac and left diaphragmatic lobes with sheep red blood cells (sRBC). The right cardiac and right diaphragmatic lobes of the same animals received saline as controls. Unimmunized dogs served as additional controls. On days 2, 6, 9, 13, and 16 after immunization, the left and right diaphragmatic lobes were lavaged, and the cells and fluids were analyzed in vitro. Opsonizing antibody in lavage fluids was first detectable at 6 days, peaked at 9-13 days, and was significantly higher in the immunized lobe than in the control lobe. Phagocytosis of sRBC caused by cytophilic antibody on PAM also peaked at 9 to 13 days. Significantly more cytophilic antibody activity was detected on day 9 in the immunized lobes, than in the control lobes. In vivo phagocytosis of sRBC was evaluated in the alveoli of immunized and control lobes of immunized dogs and a control lobe of unimmunized dogs. Phagocytosis of sRBC by PAM in the immunized lobes was about four times greater than that of the control lobes and about 40 times greater than that of a control lobe of an unimmunized dog. These results indicate that the local deposition of a particulate antigen in the lung had a localized effect on immune phagocytosis. These data suggest that the accumulation of antibody-secreting cells in the alveolus may play a critical role in pulmonary defense mechanisms.

Identification of plasma cells in lung alveoli and interstitial tissues after localized lung immunization.

Large numbers of antibody-forming cells (AFC) can be identified with the Jerne plaque assay in immunized lung lobes of the beagle dog after localized instillation of particulate antigen. Published data suggest that these AFC are secreting antibody and are responsible for increased levels of specific antibody in immunized lung lobes. If AFC in the lung are actively secreting antibody, there should be an increase in the number of mature plasma cells in lung lobes exposed to antigen. The purposes of this study were to evaluate the number of lymphoid cells present in immunized and control lung lobes and to determine if lung immunization produces a localized increase in the number of plasma cells. Sheep red blood cells (SRBC), autologous dog red blood cells, and saline were instilled into specific lung lobes of beagle dogs with the aid of a fiberoptic bronchoscope. Light and transmission electron microscopy studies of tissues from lung lobes instilled with SRBC showed perivascular infiltrates and intra-alveolar accumulations of lymphoid cells which were not present in control lung lobes. The morphology of these lymphoid cells ranged from small lymphocytes through mature plasma cells. From 5% to 15% of the cells present in the interstitial tissues and alveoli of immunized lung lobes were plasma cells. These observations suggest that lymphoid cells which entered the SRBC immunized lung can mature to plasma cells which are probably responsible for the local production of antigen-specific antibody.

Migration of neutrophils from lung to tracheobronchial lymph node.

Translocation of particulate antigen deposited in the lung to tracheobronchial lymph nodes (TBLN) is important in the induction of pulmonary immune responses. We have previously shown that alveolar macrophages can contribute to particle translocation to TBLN, but whether neutrophils can also contribute to this process is not known. To determine if neutrophils can carry particles to the TBLN, dog neutrophils were elicited by instillation of red or green fluorescent microspheres into individual lung lobes. Autologous neutrophils that had phagocytized fluorescent microspheres were then instilled into an unexposed lobe of the same dog's lung. After 24 hr, the TBLN of instilled dogs had numerous neutrophils, 99% of which contained either red or green fluorescent microspheres but not both. Use of the two different colored microspheres as labels precluded the possibility that neutrophils had phagocytized the microspheres in the TBLN. In a second experiment, dogs were depleted of peripheral blood neutrophils by injections of hydroxyurea. Hydroxyurea-treated and normal dogs were instilled with fluorescent microspheres and killed after 40 h. Hydroxyurea treatment reduced neutrophil accumulation in the lung by 79% and reduced particle translocation to the TBLN by 80%. Results of these experiments indicate that neutrophils are similar to pulmonary alveolar macrophages (PAM) in their ability to phagocytize particles in the lung and then migrate to the TBLN.

Landmark discrimination learning in the dog: effects of age, an antioxidant fortified food, and cognitive strategy.

The landmark discrimination learning test can be used to assess the ability to utilize allocentric spatial information to locate targets. The present experiments examined the role of various factors on performance of a landmark discrimination learning task in beagle dogs. Experiments 1 and 2 looked at the effects of age and food composition. Experiments 3 and 4 were aimed at characterizing the cognitive strategies used in performance on this task and in long-term retention. Cognitively equivalent groups of old and young dogs were placed into either a test group maintained on food enriched with a broad-spectrum of antioxidants and mitochondrial cofactors, or a control group maintained on a complete and balanced food formulated for adult dogs. Following a wash-in period, the dogs were tested on a series of problems, in which reward was obtained when the animal responded selectively to the object closest to a thin wooden block, which served as a landmark. In Experiment 1, dogs were first trained to respond to a landmark placed directly on top of coaster, landmark 0 (L0). In the next phase of testing, the landmark was moved at successively greater distances (1, 4 or 10 cm) away from the reward object. Learning varied as a function of age group, food group, and task. The young dogs learned all of the tasks more quickly than the old dogs. The aged dogs on the enriched food learned L0 significantly more rapidly than aged dogs on control food. A higher proportion of dogs on the enriched food learned the task, when the distance was increased to 1cm. Experiment 2 showed that accuracy decreased with increased distance between the reward object and landmark, and this effect was greater in old animals. Experiment 3 showed stability of performance, despite using a novel landmark, and new locations, indicating that dogs learned the landmark concept. Experiment 4 found age impaired long-term retention of the landmark task. These results indicate that allocentric spatial learning is impaired in an age-dependent manner in dogs, and that age also affects performance when the distance between the landmark and target is increased. In addition, these results both support a role of oxidative damage in the development of age-associated cognitive dysfunction and indicate that short-term administration of a food enriched with supplemental antioxidants and mitochondrial cofactors can partially reverse the deleterious effects of aging on cognition.

Visual-discrimination learning ability and beta-amyloid accumulation in the dog.

Young, middle-aged, and old beagle dogs were tested on several visual-discrimination tasks: reward- and object-approach learning, object discrimination and reversal, long-term retention of a reversal problem, and a size-discrimination task. Beta-amyloid accumulation in the entorhinal, prefrontal, parietal, and occipital cortices was quantified using immunohistochemical and imaging techniques at the conclusion of cognitive testing. Middle-aged and old dogs were impaired in size-discrimination learning. In each task, a subset of aged dogs was impaired relative to age-matched peers. Beta-amyloid accumulation was age-dependent. However, not all middle-aged and old dogs showed beta-amyloid accumulation in the entorhinal cortex. The error scores from dogs tested with a nonpreferred object during visual discrimination learning and from reversal learning were correlated with beta-amyloid in the prefrontal but not entorhinal cortex. Size-discrimination and reward and object-approach learning error scores were correlated with beta-amyloid accumulation in the entorhinal but not prefrontal cortex. The results of these studies support an association between cognitive test and the location and extent of beta-amyloid pathology.

Dietary enrichment counteracts age-associated cognitive dysfunction in canines.

Advanced age is accompanied by cognitive decline indicative of central nervous system dysfunction. One possibly critical causal factor is oxidative stress. Accordingly, we studied the effects of dietary antioxidants and age in a canine model of aging that parallels the key features of cognitive decline and neuropathology in humans. Old and young animals were placed on either a standard control food, or a food enriched with a broad spectrum of antioxidants and mitochondrial enzymatic cofactors. After 6 months of treatment, the animals were tested on four increasingly difficult oddity discrimination learning problems. The old animals learned more slowly than the young, making significantly more errors. However, this age-associated decline was reduced in the animals fed the enriched food, particularly on the more difficult tasks. These results indicate that maintenance on foods fortified with complex mixtures of antioxidants can partially counteract the deleterious effects of aging on cognition.

Magnetic resonance imaging of anatomic and vascular characteristics in a canine model of human aging.

Dogs exhibit both neuroanatomical and cognitive changes as a function of age that parallel those seen in aging humans. This study describes in vivo changes in neuroanatomical and cerebrovascular characteristics of the canine brain as a function of age in a group of dogs ranging from 4 to 15 years old. Dynamic contrast-enhanced magnetic resonance imaging (MRI) was used to measure the kinetics of contrast agents in the brain. Measures of vascular volume and blood-brain barrier (BBB) permeability were derived from a pharmacokinetic analysis. Cortical atrophy and ventricular enlargement were characteristic features of the aged canine brain. Vascular volume did not vary as a function of age and BBB permeability exhibited a nonsignificant increasing trend with age. However, BBB dysfunction was detected in one middle-aged dog that in addition to having unusually large ventricles, demonstrated an early onset of diffuse senile plaques at postmortem. These findings indicate that BBB dysfunction detected by magnetic resonance imaging may be useful for predicting and potentially diagnosing early pathological conditions.

Species differences in the metabolism and disposition of inhaled 1,3-butadiene and isoprene.

Species differences in sensitivity to carcinogenic effects from inhaled 1,3-butadiene might stem, at least in part, from differences in uptake, metabolism, and distribution of 1,3-butadiene. To examine this possibility, rats, mice, and monkeys were exposed to stepped concentrations of 14C-labeled 1,3-butadiene and the chemically related compound, isoprene. Respiratory data were collected during exposure and were used to determine fractional uptake. Rates and routes of excretion of retained radioactivity were also determined and blood levels of potentially toxic metabolites were measured. In some cases, the concentrations of hemoglobin adducts were determined. For rodents, the tissue distribution of metabolites was examined. Some results from these continuing studies to date are: a) mice achieve higher blood concentrations of reactive metabolites than do rats; b) blood levels of toxic metabolites are lower in monkeys than in rodents; c) uptake and retention of 1,3-butadiene is nonlinear in the range where long-term toxicity studies have been conducted; d) the efficiency of production of reactive metabolites decreases with increased inhaled concentrations of 1,3-butadiene; e) repeated exposure to 1,3-butadiene does not induce the metabolism of 1,3-butadiene in rodents; f) hemoglobin adducts of 1,3-butadiene are potential dosimeters of exposure; and g) rats inhaling isoprene produce reactive metabolites analogous to those produced during inhalation of 1,3-butadiene. The available data indicate that major differences in the biological fate of inhaled 1,3-butadiene occur among species, and these differences, at least in part, account for those in species sensitivity to the toxicity of inhaled 1,3-butadiene.

A comparison of the predicted risks of developing osteosarcoma for dogs exposed to 238PuO2 based on average bone dose or endosteal cell dose.

Dose-response relationships observed in laboratory animals can be used to identify possible human risk factors and may also be used in a quantitative manner when human data are not available. This paper presents an analysis of the dose dependency of osteosarcoma incidence in beagle dogs given a single inhalation exposure to a monodisperse aerosol of 238PuO2. We were particularly interested in comparing the predicted risks that were based on average bone dose with those based on endosteal cell dose and in evaluating the advantages of using a more biologically relevant cell-specific dose in risk estimation. The endosteal cell dose was calculated using the method of Marshall et al. (Health Phys. 35, 91-101, 1978), as extended to account for exposure by inhalation. The relationship between dose and time to tumor was analyzed by the proportional hazards regression model. The probability of developing osteosarcoma was strongly dependent on dose for dogs receiving low doses, but this was not true for dogs receiving high doses. The predicted risk based on endosteal cell dose was not consistently higher or lower than the risk based on average bone dose at various times after exposure, because the relationship between these two doses was not linear with respect to time. Also, as a result of the nonlinear relationship between these two doses, the risk estimated based on endosteal cell dose would not be a fixed factor of that based on the average dose. Random errors in the measured initial lung burden had a relatively large impact on the predicted risk based on endosteal cell dose, and the difference between the estimated risk of developing osteosarcoma based on endosteal cell dose and that based on average bone dose is likely to be within the error margins of the estimated risks.

Biological effects of inhaled 144CeCl3 in beagle dogs.

The biological effects of 144Ce were studied in beagle dogs that were exposed to graded activity levels of 144CeCl3 via a single, brief inhalation exposure and observed for their life span. The long-term retained body burdens ranged from 0.06 to 13 MBq/kg with a median of 1.2 MBq/kg. After a short residence time in the lung, most of the 144Ce was translocated to liver and skeleton, where the 144Ce was retained with a half-time approaching the physical half-life of 144Ce, 284 days. Significant radiation doses were delivered to the lung, 28 Gy (median) and 2.5-370 Gy (range); liver, 68 Gy (median) and 6.1-250 Gy (range); and skeleton, 21 Gy (median) and 1.9-100 Gy (range). Lesions induced by the beta-particle radiation were noted in the lung, liver, skeleton, bone marrow, and oral and nasal mucosae closely associated with bone. Early deaths (within 2.5 years) were generally related to hematological dyscrasia, radiation pneumonitis, or hepatocellular degeneration and atrophy. Neoplasms that occurred relatively early, from 2.2-6.8 years after exposure, were noted in the liver, bone, bone marrow and oral mucosa closely associated with bone. Neoplasms that occurred later, beyond 7 years after exposure, were noted in the liver, lung and nasal mucosa closely associated with bone. Increased numbers of neoplasms were not found in two other organs that had relatively high radiation doses, namely the thyroid and kidney. Only one primary bone tumor was noted, but 11 tumors of bone-associated tissues (oral and nasal mucosae and bone marrow) were found. Radiation doses and effects in tissues adjacent to bone, especially those of epithelial or marrow origin, should be considered when determining risks from internally deposited bone-seeking radionuclides, such as 144Ce. The property of 144Ce in depositing on and remaining associated with bone surfaces for long times may be an important factor in the radiation dose to bone marrow and epithelium adjacent to bone.

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.

Lung lavage therapy to lessen the biological effects of inhaled 144Ce in dogs.

To evaluate the therapeutic effects of removal of an internally deposited radionuclide on long-term biological effects, lung lavage was used to treat dogs that had inhaled 144Ce in a relatively insoluble form, in fused aluminosilicate particles. Either 10 lung lavages were performed between Days 2 and 56 after exposure or 20 lung lavages were performed between Days 2 and 84 after exposure. Approximately one-half of the 144Ce was removed by the lavages, resulting in a corresponding reduction in the total absorbed beta dose to lung. The mean survival time of the treated dogs was 1270 days compared to 370 days for untreated dogs whose initial pulmonary burdens of 144Ce were similar. Treated dogs died late from cancers of the lung or liver, whereas the untreated dogs died at much earlier times from radiation pneumonitis. Dogs treated with lung lavage but not exposed to 144Ce had a mean survival of 4770 days. We concluded that removal of 144Ce from the lung by lavage resulted in increased survival time and in a change in the biological effects from inhaled 144Ce from early-occurring inflammatory disease to late-occurring effects, principally cancer. In addition, the biological effects occurring in the treated dogs could be better predicted from the total absorbed beta dose in the lung and the dose rate after treatment rather than from the original dose rate to the lung. Therefore, we concluded that prompt treatment to remove radioactive materials could be of significant benefit to persons accidentally exposed to high levels of airborne, relatively insoluble, radioactive particles.

Influence of dose rate on survival time for 239PuO2-induced radiation pneumonitis or pulmonary fibrosis in dogs.

Beagle dogs were exposed once or repeatedly to 0.75-microns-diameter monodisperse aerosols of 239PuO2 by pernasal inhalation. The dogs that were exposed once received alveolar depositions (+/- standard deviation) of 3.9 +/- 1.9 kBq/kg body mass and accumulated doses of 23 +/- 8 Gy to the lung before death at 5.4 +/- 1.7 years after exposure. Dogs exposed repeatedly received a total alveolar deposition of 5.3 +/- 0.9 kBq/kg body mass during 7 to 10 semiannual exposures and accumulated doses of 22 +/- 5 Gy to the lung before death at 4.9 +/- 0.7 years after first exposure. Clearance of the plutonium from the lung in the dogs exposed repeatedly was slower than in the dogs exposed once. All dogs in the repeated-exposure study and all but one dog in the single-exposure study died from radiation effects. Pulmonary fibrosis accounted for 72% of the radiation-related deaths in the single-exposure study and 87% in the repeated-exposure study. The remaining dogs died with pulmonary cancer. Based on total cumulative radiation dose, the times after exposure to death from radiation pneumonitis and pulmonary fibrosis were not significantly different for single and repeated exposures. Thus dose rate does not appear to be an important factor in predicting death from radiation pneumonitis or pulmonary fibrosis for dogs inhaling 239PuO2.

The biological effects of radium-224 injected into dogs.

A life-span study was conducted in 128 beagle dogs to determine the biological effects of intravenously injected 224Ra chloride. The 224Ra chloride was prepared by the same method used for intravenous injections in humans who were treated for ankylosing spondylitis and tuberculosis. Thus the results obtained from dogs can be compared directly to the population of treated humans, both for the elucidation of the effect of exposure rate and for comparison with other radionuclides for which data for humans are unavailable. Using equal numbers of males and females, the dogs were injected with one of four levels of 224Ra resulting in initial body burdens of approximately 13, 40, 120 or 350 kBq of 224Ra kg-1 body mass. A control group of dogs was injected with diluent only. All dogs were divided further into three groups for which the amount of injected 224Ra (half-life of 3.62 days) or diluent was given in a single injection or divided equally into 10 or 50 weekly injections. As a result of these three injection schedules, the accumulation of dose from the injected 224Ra was distributed over approximately 1, 3 or 12 months. Each injection schedule included four different injection levels resulting in average absorbed alpha-particle doses to bone of 0.1, 0.3, 1 and 3 Gy, respectively. The primary early effect observed was a hematological dyscrasia in the dogs receiving either of the two highest injection levels. The effect was most severe in the dogs receiving a single injection of 224Ra and resulted in the death of three dogs injected at the highest level. The late-occurring biological effects were tumors. Bone tumors were the most common followed by tumors in the nasal mucosa. The occurrence of bone tumors was highest in the dogs given the highest dose in 50 injections. The age-specific incidence rate for mammary tumors was increased in all three injection groups. The results of this study revealed two important exposure-rate effects. Hematological dyscrasia was amplified by delivery of relatively high doses at a high exposure rate. In contrast, bone tumors were amplified by delivery of relatively high doses at a lower exposure rate (i.e. dose delivered over 1 year rather than 1-3 months). There was a dose-response relationship for the induction of nasal mucosal tumors and mammary tumors. These findings in dogs are similar to those in humans injected with 224Ra, except for the nasal tumors. The calculated risk of developing a bone tumor was about 40 times higher in dogs than reported for humans.

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.