Cardiopulmonary function of dogs with plutonium-induced chronic lung injury

Beagle dogs had signs of restrictive lung disease 1 to 5 years after exposure by inhalation to 239PuO2 aerosols. The 239PuO2 aerosols were monodisperse with activity median aerodynamic diameters of 0.75, 1.5, or 3.0 microns. The plutonium particles produced protracted alpha irradiation of the lungs. Ten dogs had specific initial pulmonary burdens (IPB) of 330 to 4,100 kBq of 239PuO2/kg of body mass. The average onset time of clinical signs of lung injury was 3 years after exposure; the average time from the onset of signs until cardiorespiratory function evaluation was 5.5 years. A second group of 10 dogs had IPB of 110 to 2000 kBq of 239Pu/kg of body mass but no signs of lung injury. A third group of 10 dogs, not exposed to 239Pu, were matched for age and sex. Cardiopulmonary function tests were performed. Only the dogs in group I with signs of lung injury had a mild respiratory function disorder consisting of smaller lung volumes, reduced compliance, increased respiratory frequency and minute volume, and reduced carbon monoxide diffusing capacity. Cardiac function of all three groups was similar. These findings indicate that alpha irradiation of the lungs of man could produce restrictive lung disease at long times after initial exposure.     

The development and interlobar distribution of plutonium-induced pulmonary fibrosis in mice

Six-week-old mice were exposed by inhalation to an aerosol of 239PuO2 (activity median aerodynamic diameter 2.2 microns) to establish mean alveolar depositions at 2 days after exposure of 4, 40, and 930 Bq of 239Pu. Animals were killed serially after 3, 6, 12, and 18 months at which times the development of the pulmonary fibrotic lesion was assessed by both biochemical and histopathological techniques. Individual measurements of both fresh and dry weights, protein, DNA, and hydroxyproline were made on whole lung and also on each of the five constituent lobes. Early and sustained increases in lung mass, lung protein, and total lung collagen were found, together with a depression of the total cellularity of the lung at 6 and 12 months after exposure. Although at later times compensatory hypertrophy of less affected areas distorted the relationship, systematic trends in the severity of responses between lobes were found. These trends were related to the initial lobar concentrations of 239Pu.     

The cytotoxic activity of natural killer cells exposed to the joint and separate actions on the body of 239Pu, hexachlorobutadiene and tributyl phosphate

Wistar rats were subjected to a single exposure to 239Pu nitrate through inhalation with the subsequent procedure of imitation of inhalation or without it (the amount of 239Pu deposited in the lungs in 24 hr was 8.9 +/- 1.9 kBq/lung) and inhalation of hexachlorobutadiene and tributyl phosphate within one month in a combination with the radionuclide or without it. There was a 1.5-fold increase, above additive, in the harmful effect of 239Pu and chemical agents on the function of natural killers as observed 15--30 days after the combined exposure as compared to individual inhalation. On days 120 to 240 cell cytotoxic activity in rats of all groups was normalized to reach or to exceed the intact control.     

Activation of alveolar macrophages after plutonium oxide inhalation in rats: involvement in the early inflammatory response

Alveolar macrophages play an important role in the distribution, clearance and inflammatory reactions after particle inhalation, which may influence long-term events such as fibrosis and tumorigenesis. The objectives of the present study were to investigate the early inflammatory events after plutonium oxide inhalation in rats and involvement of alveolar macrophages. Lung changes were studied from 3 days to 3 months after inhalation of PuO2 of different isotopic compositions (70% or 97% 239Pu) and initial lung deposits (range 2.1 to 43.4 kBq/rat). Analyses of bronchoalveolar lavages showed early increases in the numbers of granulocytes, lymphocytes and multinucleated macrophages. The activation of macrophages was evaluated ex vivo by measurement of inflammatory mediator levels in culture supernatants. TNF-alpha and chemokine MCP-1, MIP-2 and CINC-1 production was elevated from 7 days after inhalation and remained so up to 3 months. In contrast, IL-1beta, IL-6 and IL-10 production was unchanged. At 6 weeks, pulmonary macrophage numbers and activation state were increased as observed from an immunohistochemistry study of lung sections with anti-ED1. Similarly, histological analyses of lung sections also showed evidence of inflammatory responses. In conclusion, our results indicate early inflammatory changes in the lungs of PuO2-contaminated animals and the involvement of macrophages in this process. A dose-effect relationship was observed between the amount of radionuclide inhaled or retained at the time of analysis and inflammatory mediator production by alveolar macrophages 14 days after exposure. For similar initial lung deposits, the inflammatory manifestation appears higher for 97% 239Pu than for 70% 239Pu.     

The induction of liver tumors by 239Pu citrate or 239PuO2 particles in the Chinese hamster

The influence of radiation dose distribution on the frequency of 239Pu-induced liver tumors was evaluated in the Chinese hamster. Different concentrations of 239Pu citrate 239PuO2 particles of known sizes were injected intravenously via the jugular vein. About 60% of the injected 239Pu citrate was deposited in the liver and 40% in the bone. The 239Pu citrate was rather uniformly distributed throughout the liver parenchyma. Injected plutonium oxide particles were taken up by the reticuloendothelial system with 90% of the body burden deposited in the liver. The 239PuO2 particles were localized in the Kupffer cells and produced nonuniform dose distributions that were dependent on particle size. There was an activity- and dose-dependent increase in the incidence of total liver parenchymal cell tumors following injection with either plutonium particles or citrate. For animals that received 14.0-, 2.7-, 0.3-, and 0.04-Gy dose to liver from 239Pu citrate the cumulative tumor incidence was 39, 32, 5, and 0%, respectively. Animals that were injected with the 0.24 micron 239PuO2 particles had doses of 42.0, 7.2, and 0.8 Gy to the liver and tumor incidences of 34, 26, and 5%, respectively. Plutonium citrate also produced hemangiosarcomas of the liver and tumors in bone and bone marrow. The latent period for liver tumor appearance in animals exposed to 239Pu citrate or 239PuO2 particles increased as the injected activity decreased. For animals injected with a similar total activity (7.4 Bq/g), the lifetime cumulative liver tumor incidence was similar for animals exposed to either 239Pu citrate (32%) or 239PuO2 (26%). There was little effect of particle size on liver tumor incidence. These data indicate that, in Chinese hamster liver, local radiation dose distribution is less important in altering tumor incidence than injected activity or average dose. However, the more uniform irradiation from 239Pu citrate administration was more effective in cancer production than the nonuniform irradiation from 239PuO2 particles.     

Repeated inhalation exposure of rats to aerosols of 144CeO2. I. Lung, liver, and skeletal dosimetry.

To develop a better understanding of the influence of cumulative radiation dose and dose rate to the lungs on the biological responses to inhaled radionuclides, several studies are in progress at this institute in which laboratory animals have been exposed once or repeatedly to aerosols of insoluble particles containing 144Ce or 239Pu. In the study reported here, F344 rats were exposed repeatedly to aerosols of 144CeO2 beginning at 94 days of age to reestablish desired lung burdens of 1.9, 9.2, 46, or 230 kBq of 144Ce every 60 days for 1 year (seven exposures). Other 94-day-old rats were exposed once to achieve similar desired initial lung burdens of 144Ce. Older rats were exposed once to achieve desired initial lung burdens of 46 or 230 kBq when 500 days of age, the age of the repeatedly exposed rats when exposed for the last time. Control rats were either unexposed, sham-exposed once or repeatedly, or exposed once or repeatedly to stable CeO2. Approximately equal numbers of male and female rats were used. The cumulative beta-radiation doses to the lungs, liver, and skeleton of rats exposed repeatedly were similar to those of rats with similar total lung burdens of 144Ce from a single inhalation exposure. The average beta-radiation dose rate to the lungs of the rats exposed repeatedly was about one-fifth of that in rats with similar total lung burdens after a single exposure.     

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.     

Structural chromosomal damages in myelokaryocytes exposed to plutonium-239 dioxide

Plutonium 239 dioxide, with particle sizes of 1-2 micron, injected to Wistar rats intravenously in doses of 92.5, 46.3, and 23.2 kBq/kg body mass, increased the frequency of chromosome aberrations in myelokaryocytes by 3.7, 2.3 and 1.7 times, respectively, compared to spontaneous levels.     

Functional state of the thyroid gland in sheep at a remote period after single administration of Sr-90, Pu-239 and their mixture

Five years after single intravenous injection of a mixture of 239Pu and 90Sr to semifine-wool sheep (7.4 kBq/kg + 185 kBq/kg) the iodine-fixing and hormone secreting functions of the thyroid gland were inhibited; where 90Sr alone was injected in the above dose inhibited was the hormone-secreting function only. Since 239Pu alone did not cause such alterations, the observed remote effect was attributed to the effect of the incorporated 90Sr.     

Mitochondrial DNA deletions in the peripheral blood of workers at the Mayak PA who were exposed to long-term combined effects of external γ- and internal α-radiation

The levels of large deletions in the mitochondrial DNA of workers at the Mayak Production Association (Mayak PA) who were exposed to external and combined occupational (external γ- and internal α-rays) radiation during the course of their duties were investigated. Peripheral blood-derived DNA samples were provided by the Radiobiological Human Tissue Repository of the Southern Urals Biophysics Institute (Russia). The samples were analyzed using long-extension PCR. The number of large-scale deletions in the mitochondrial DNA of workers who, in addition to external γ-radiation, were exposed to extra doses of irradiation due to incorporated ²³⁹Pu with a Pu body burden of 0.77–4.32 kBq, was 2.5-times lower compared to that of individuals who received only external γ-radiation. No significant gender-associated effects on the number of mitochondrial DNA deletions were detected among age-matched individuals.

     

A primary pulmonary sarcoma in a rhesus monkey after inhalation of plutonium dioxide

A pulmonary fibrosarcoma of bronchial origin was discovered in a Rhesus monkey that died of pulmonary fibrosis 9 years after inhalation of plutonium-239 dioxide and with a radiation dose to lung of 1400 rad (14 Gy). It grew around the major bronchus of the right cardiac lung lobe and extended into the bronchial lumen and into surrounding pulmonary parenchyma. It also readily invaded muscular pulmonary arteries, resulting in infarction and scarring in the right cardiac lobe. Despite this aggressive growth, the tumor did not metastasize. The primary cause of death was severe pulmonary fibrosis involving the alveolar septa and and perivascular and peribronchial interstitium. Bullous or pericitrical emphysema was prominent. The initial lung burden of plutonium in this monkey was 270 nCi (10 kBq) which is equivalent to approximately 500 times the maximum permissible lung burden for man on a radioactivity per unit body weight basis. The time-dose relationship for survival is consistent with that of dogs and baboons that inhaled plutonium dioxide and died with lung tumors.     

State of cellular immunity effectors of the lung after inhalation of 239Pu polymer nitrate]

Wistar rats, who inhaled 239Pu (IV) at a dose that did not reduce the average life expectancy (the initial level in the lung was 0.77 kBq) did not exhibit any essential changes in the quality and functional activity of the alveolar macrophages. Revealed were the injuries to cells of bifurcation lymph nodes, the severity of which was a function of dose absorbed in the lung.     

The antibody-producing cell count of rats exposed to polymeric 239Pu

In experiments with Wistar rats a study was made of the content of antibody-forming cells (AFC) in the spleen at remote times (3 to 12 months) after intravenous injection of 239Pu(IV) in doses of 166, 55, and 18 kBq/kg body mass. The doses absorbed in the central and peripheral immunity organs were defined. Pronounced spleen hypoplasia and profound inhibition of humoral immunity were displayed 1 year after the injection of a small amount of the radionuclide. AFC deficiency in animals was amounted to 11-32 per cent throughout the entire period of observation.     

mFISH analysis of chromosome aberrations in workers occupationally exposed to mixed radiation

We performed a study on the presence of chromosome aberrations in a cohort of plutonium workers of the Mayak production association (PA) with a mean age of 73.3 ± 7.2 years to see whether by multi-color fluorescence in situ hybridization (mFISH) translocation analysis can discriminate individuals who underwent occupational exposure with internal and/or external exposure to ionizing radiation 40 years ago. All Mayak PA workers were occupationally exposed to chronic internal alpha-radiation due to incorporated plutonium-239 and/or to external gamma-rays. First, we obtained the translocation yield in control individuals by mFISH to chromosome spreads of age-matched individuals and obtained background values that are similar to previously published values of an international study (Sigurdson et al. in Mutat Res 652:112–121, 2008). Workers who had absorbed a total dose of >0.5 Gy external gamma-rays to the red bone marrow (RBM) displayed a significantly higher frequency of stable chromosome aberrations relative to a group of workers exposed to <0.5 Gy gamma-rays total absorbed RBM dose. Thus, the translocation frequency may be considered to be a biological marker of external radiation exposure even years after the exposure. In a group of workers who were internally exposed and had incorporated plutonium-239 at a body burden >1.48 kBq, mFISH revealed a considerable number of cells with complex chromosomal rearrangements. Linear associations were observed for translocation yield with the absorbed RBM dose from external gamma-rays as well as for complex chromosomal rearrangements with the plutonium-239 body burden.     

Frequency of structural chromosome aberrations in the hepatocytes of rats exposed to polymeric 239Pu

Intravenous injection of polymeric 239Pu(IV) nitrate (166.5, 55.5 and 18.5 kBq/kg body mass) to Wistar rats was shown to produce biphase changes in the frequency of hepatocyte chromosome aberrations. The increase in the structural damages to chromosomes at later times of observation was a pronounced function of radiation dose. The absence of such a dependence at early times was evidently due to the elimination of damaged liver parenchyma cells.     

Biochemical changes in rat tissues as affected by the combined action of external gamma-radiation and 239Pu in different doses

Under the combined effect of external gamma-radiation (25.8 and 103.2 mC/kg) and 239Pu (18.5 and 92.5 kBq/kg body weight) the changes in the nucleic acids of rat thymus tissues depended upon the intensity of gamma- and alpha-radiation. The damage to the thymus produced by the combined effect was weaker than that of the effect of the two types of radiation delivered separately. In bone tissue, summation of the oppositely directed changes in alkaline phosphatase activity was registered after the effect of gamma-radiation and 239Pu delivered separately.     

Effects of the single or repeated inhalation exposure of Syrian hamsters to aerosols of 239PuO2

Male Syrian hamsters were scheduled to be exposed by inhalation approximately every 60 days for 1 year (7 exposures) to aerosols of 239PuO2 beginning at 84 days of age. Other hamsters were exposed once when 84 or 320 days of age. Plutonium-239 deposited in the lungs by the repeated inhalation exposures was cleared from the lungs at a rate similar to that following a single inhalation exposure. The incidence of radiation pneumonitis, bronchiolar epithelial hyperplasia, and alveolar squamous metaplasia were the only lesions that were related to radiation dose. Only two primary lung tumours were found among the hamsters exposed to 239PuO2. No primary lung tumours were found in the control hamsters. It was concluded that the incidence of lung tumours was not increased by the protraction of the alpha radiation dose to the lungs from repeated inhalation exposure.     

Effects of protraction of the alpha dose to the lungs of mice by repeated inhalation exposure to aerosols of 239PuO2

To determine the long-term biological effects of protracted alpha irradiation of the lung, 84-day-old C57BL/6J mice were repeatedly exposed by inhalation to aerosols of 239PuO2 every other month for up to six exposures in 10 months to reestablish lung burdens of 20, 90, or 460 Bq. Other mice were exposed only once when either 84 or 460 days of age to achieve desired initial lung burdens of 20, 90, 460, or 2300 Bq. Suitable control groups were maintained. Groups of mice with similar cumulative alpha doses to the lung had 3.4 to 4.4 times greater incidence of pulmonary tumors (adenomas and adenocarcinomas) when the dose to the lung was protracted by the repeated inhalation exposures compared to mice that received a single inhalation exposure. Excess pulmonary tumors per unit dose to the lung were also greater in groups of repeatedly exposed mice compared to those exposed only once. Repeatedly exposed mice also died earlier with pulmonary tumors than did those exposed once. It appears that protraction of an alpha dose to lungs increases the carcinogenic risk of inhaled 239PuO2 in mice.     

Radiosensitivity and proliferative activity of vertebral CFU-S surviving in mice injected with oncogenic activities of 239Pu or 241Am

Survival, radiosensitivity and capability to produce differentiated progeny were followed in CFU-S from lumbar vertebrae of mice injected with 198.6 kBq 239Pu/kg or 208.6 kBq 241Am/kg. The CFU-S assay and 59Fe uptake into spleen colonies were used. The number of CFU-S from treated mice was significantly lower than in controls. Higher radiosensitivity of CFU-S from 239Pu- or 241Am-treated mice was demonstrated using additional exposure to 0.5 Gy X-rays 1, 24, 48, 72 hrs after cell transplantation and expressed more precisely by survival curves obtained 1 hr after the marrow cell injection. The effect of 239Pu on CFU-S was characterized by Do 0.58 Gy (n = 0.91) and that of 241Am by Do 0.64 Gy (n = 0.91); corresponding control values were Do 0.89 Gy, n = 1.11. Lower iron utilization due not only to the decreased CFU-S numbers, but also to the defective production of erythroid cells per one CFU-S was found. Complexity of radiation effect on hemopoietic stem cells was demonstrated by the present study.