Number Of Pulmonary Alveolar Macrophages Research Articles

Chronic Heat Stress Weakened the Innate Immunity and Increased the Virulence of Highly Pathogenic Avian Influenza Virus H5N1 in Mice

Chronic heat stress (CHS) can negatively affect immune response in animals. In this study we assessed the effects of CHS on host innate immunity and avian influenza virus H5N1 infection in mice. Mice were divided into two groups: CHS and thermally neutral (TN). The CHS treatment group exhibited reduced local immunity in the respiratory tract, including the number of pulmonary alveolar macrophages and lesions in the nasal mucosa, trachea, and lungs. Meanwhile, CHS retarded dendritic cells (DCs) maturation and reduced the mRNA levels of IL-6 and IFN-β significantly (P < .05). After the CHS treatment, mice were infected with H5N1 virus. The mortality rate and viral load in the lungs of CHS group were higher than those of TN group. The results suggest that the CHS treatment could suppress local immunity in the respiratory tract and innate host immunity in mice significantly and moderately increased the virulence in H5N1-infected mice.

Pathogenetic process of lung tumors induced by inhalation exposures of rats to plutonium dioxide aerosols.

Sequential examinations were done on the pulmonary cytokinetics and pulmonary lesions in rats after inhalation exposure to (239)PuO(2) aerosols to investigate the pathogenesis of lung tumors. Total cell yields of lavaged bronchoalveolar cells as well as the estimated numbers of pulmonary alveolar macrophages were significantly reduced from 1 to 3 months after exposure but recovered thereafter to the control levels. The proportions of multinucleated or micronucleated pulmonary alveolar macrophages increased significantly in lavaged cells from 1 month, and the increase was sustained up to 18 months after exposure. Both tumor necrosis factor and nitric oxide were shown to be differentially released from stimulated cultures of pulmonary alveolar macrophages during the period from 6 to 18 months after exposure. The labeling indices of alveolar and bronchiolar epithelial cells treated with 5-bromo-2'-deoxyuridine increased significantly in lungs from 3 months and were sustained up to 18 months after exposure. Histopathological examinations revealed that after the early inflammation, hyperplasia and metaplasia of the lining of the bronchioloalveolar epithelium were predominant from 3 to 6 months, while adenomatous or adenocarcinomatous lesions appeared and developed from 12 months after exposure. The appearance of primary lung tumors, almost all of which were adenomas and adenocarcinomas, was found in the dose range of 1 to 2 Gy from 12 months after exposures. These results indicate that the pathogenetic process initiated by early cellular damage and alterations associated with inflammation is followed by the proliferative and metaplastic lesions of pulmonary epithelium, leading to the appearance and development of pulmonary neoplasms from 1 year after the inhalation exposures in rats that received a minimum lung dose of more than 1 Gy.

Effect of Kerosene and Its Soot on the Chrysotile-Mediated Toxicity to the Rat Alveolar Macrophages

In order to examine the pulmonary toxicity of kerosene oil and its combustion product (soot) in asbestos-exposed rats, various biochemical and chemical parameters were assayed. Treatment of rats with a single intratracheal dose of chrysotile asbestos (5 mg) and kerosene (50 μl) or its soot (5 mg) in combination led to an increased number of pulmonary alveolar macrophages (PAM), elevated levels of hydrogen peroxide, and thiobarbituric acid-reacting substances, alterations in the activities of primary (glutathione peroxidase and catalase) and secondary (glutathione reductase and glucose-6-phosphate dehydrogenase) endogenous antioxidant enzymes, and depletion in the levels of glutathione in PAM compared to the chrysotile, kerosene, or soot alone. These changes may indicate the generation of oxidative stress in the macrophages. The resulting oxidative stress may be subsequently critical in collapsing the cellular membrane, which may change the cell membrane permeability and may also damage the phagolysosomal membrane, thereby releasing the membrane bound enzymes as indicated by an increased leakage of intracellular acid phosphatase and lactate dehydrogenase. The injury to macrophages may trigger events that lead to lung fibrosis and/or malignancies in the exposed animals. This study may be helpful in understanding the etiology of certain clinical and pathological disorders in the population exposed simultaneously to both asbestos and kerosene or its combustion products.

INCREASE OF ELASTOLYTIC ACTIVITY WITHOUT ANY CHANGE OF EFFECTIVE 1-ANTITRYPSIN AND CHANGE OF OXIDE REDUCTASES IN RAT BRONCHOALVEOLAR LAVAGE FLUID AS MECHANISMS OF MINERAL DUST-INDUCED EMPHYSEMA

The lung is susceptible to injury leading to airflow obstruction and fibrosis from inhaled mineral dust, including crystalline silica. It has been established that there is a significant correlation between mineral dust exposure and the presence of emphysema at autopsy after occupational exposure. The mechanisms of the opposing processes of emphysema and fibrosis are not known. To investigate the mechanisms involved in these processes, we studied the role of elastolytic activity and its inhibitor, and oxide reductase in crystalline silica-treated rats. Wistar rats were treated with a single transtracheal instillation of crystalline silica (50 mg/kg). The control animals were treated with saline in the same manner. Elastolytic activity, 1-antitrypsin, superoxide dismutase (SOD), total glutathione (GSH), and cytokines in bronchoalveolar lavage (BAL) fluid were quantified during a 75- wk experimental period. Although the numbers of pulmonary alveolar macrophages (PAM) did not change significantly, the number of transmigrated neutrophils markedly increased until wk 18 in the silica group. Protein and elastolytic activity increased markedly from wk 3 to wk 24 without any change of 1-antitrypsin. A decrease in SOD activity and an increase of GSH, concomitant with an increase in neutrophil numbers, were observed in the silica group. Although total TGF- 1 only increased slightly between days 2 and 3, showing a low level during the next 25 days, neutrophil chemokines (cytokineinduced neutrophil chemoattractant and growth-regulated gene products) showed a high level during the 25-day experimental period. Superoxide anion generation and phagocytosis in BAL PAMs were suppressed from wk 8 in the silica group. These results suggest that the persistent increase of elastolytic activity deriving from activated PAMs and infiltrated neutrophils without any change of 1-protease inhibitor in addition to changes of oxide reductase may play pivotal roles in the development of mineral dust-induced emphysema.

Specific biological effects of an anti-rat PMN antiserum intraperitoneally infected into f344/n rats.

Neutropenia can be produced with antimitotic chemicals, but this method lacks specificity. An alternative is to use antibody-dependent cytotoxicity to produce neutropenia; however, this method has not been completely evaluated with respect to efficacy, specificity, and potential collateral damage, especially to constituents of bone marrow. This study used in vitro and in vivo methods to evaluate specific biological effects of a commercially available rabbit anti-rat neutrophil (PMN) antiserum in F344/N rats. The viability of rat pulmonary alveolar macrophages (PAMs), PMNs, and lymphocytes in vitro was quantified using a trypan blue dye exclusion test. Amounts of antiserum in vitro that rendered PMNs 100% nonviable did not decrease the viability or phagocytic ability of the PAMs and did not decrease the viability of the lymphocytes. Intraperitoneal (IP) injection of the antiserum into rats resulted in complete depletion of the PMNs and about a 50% depletion of the lymphocytes in circulating blood within 24 hours. The numbers of both cell types remained lowered for 5 days, but returned to control values by Day 6 after the IP injection. The antiserum had no effect on the numbers of PAMs or lymphocytes in the pulmonary alveolar airspaces, as determined by quantifying the numbers of these cell types in bronchoalveolar lavage fluid (BALF). The numbers of PMNs in BALF, however, decreased on Days 3 and 4 after IP injection of antiserum, but were not different from control values by Day 5. The viability of the PAMs in BALF of treated rats was not different from control values at any time point. There were no morphological indications that the injected antiserum damaged lung tissue or stem cells in bone marrow. Results demonstrate that the anti-rat PMN antiserum administered IP to F344/N rats depletes circulating PMNs and partially depletes lymphocytes for a period of about 6 days without adversely affecting the precursors of red or white blood cells in bone marrow. We concluded that the antiserum is a relatively specific way to temporarily render rats neutropenic without damaging precursor cells in bone marrow.

Acute, Subacute, and Subchronic Inhalation Toxicity Studies of Respirable Polymeric Methylene Diphenyl Diisocyanate (Polymeric MDI) Aerosol in Rats

Acute, Subacute, and Subchronic Inhalation Toxicity Studies of Respirable Polymeric Methylene Diphenyl Diisocyanate (Polymeric MDI) Aerosol in Rats. Reuzel, P. G. J., Kuper, C. F., Feron, V. J., Appelman, L. M., and Löser, E. (1994). Fundam. Appl. Toxicol. 22, 186-194.Short-term inhalation toxicity studies with respirable polymeric methylene diphenyl diisocyanate (polymeric MDI) aerosol were performed in rats. The 4-hr LC50 was found to be 490 mg polymeric MDI/m3 (95.5% < 4.3 μm). Exposure of (4-week-old) rats to 0, 2.2, 4.9, or 13.6 mg polymeric MDI/m3 (95% < 5 μm) for 2 weeks resulted in mortality, severe growth retardation, and elevated lung weights at 13.6 mg/m3; at 4.9 mg/m3 slight growth retardation and slightly elevated lung weights were observed. A 13-week study with 6-week-old rats exposed to 0.35, 1.4, or 7.2 mg polymeric MDI/m3 (95% < 5 μm) revealed transient growth retardation and a slightly increased number of pulmonary alveolar macrophages occasionally accompanied by increased numbers of mononuclear cells and fibroblast in alveolar septa only at 7.2 mg/m3. In a second 2-week study with 4- or 6-week-old rats exposed to 14.1 mg polymeric MDI/m3 (95% < 5 μm), 4-week-old rats died earlier and in greater numbers than 6-week-old rat. In a second 13-week study with 6-week-old rats, using exposure concentrations of 0, 4.1, 8.4, and 12.3 mg polymeric MDI/m3 (95% < 5 μm) and including a 4-week recovery period, 12.3 mg/m3 induced mortality, growth retardation, severe respiratory distress, increased lung weights, degeneration and hyperplasia of the nasal epithelium, accumulations of macrophages in the lungs and mediastinal lymph nodes, and focal inflammatory changes in the lungs. Rats exposed to 8.4 mg/m3 showed respiratory distress, lower body weights in males, increased lung weights, and similar, but much less severe, histopathological changes in the respiratory tract and mediastinal lymph nodes. Most of the histopathological changes seen at the higher concentrations were also seen at 4.1 mg/m3 but to a very minor degree and in a few rats only. At the end of the 4-week posttreatment period the microscopical changes in nose, lungs, and mediastinal lymph nodes were still present but generally to a much less degree than at the end of the exposure period. It was concluded that the dose-effect curve for repeated exposures of rats to respirable polymeric MDI is very steep, and that the "no-observed-adverse-effect level" of polymeric MDI was 1.4 mg/m3, the actual no-adverse-effect level being lower than but most probably very close to 4.1 mg/m3.

Modulation of silica-induced lung injury by reducing lung non-protein sulfhydryls with buthionine sulfoximine

A role for non-protein sulfhydryl moieties (NPSH) (e.g., glutathione) in silica (SI)-induced cellular inflammation and fibrosis was examined in C57B1/6 mice depleted of lung NPSH by buthionine sulfoximine (BSO). Lung NPSH levels in the BSO-treated groups were reduced to approx. 50% of the non-BSO-treated animals. In BSO-treated Si-injected (2 mg/mouse) animals, the number of pulmonary alveolar macrophages (PAM) lavaged from the lungs was significantly increased on day 1 and decreased on day 7. Moreover, BSO-treated Si-exposed mice evidenced significantly more lavage protein and albumin on days 1 and 3, respectively, than non-BSO-treated Si-exposed mice. Si-induced collagen deposition, however, was decreased by 18% in the BSO-treated animals. These data suggest that lung NPSH lessens the potential of silica to elicit acute lung injury.

Cytotoxicity of Fibrogenic Asbestos and Silica on Murine Pulmonary Alveolar Macrophage Cology-Forming Cells

AbstractToxicity of fibrogenic asbestos and silica particles on the in vitro colony-forming capacity of murine pulmonary alveolar macrophages (PAM) was tested to elucidate the possibility that the proliferation or the maintenance of the PAM population could be affected by inhaled substances. Intratracheal instillation of crocidolite asbestos or Min-u-sil silica particles into mice induced significant and persistent reduction of alveolar macrophage colony-forming cells (AM-CFC) during the period of 1 to 6 months after treatments as compared to untreated control or groups of mice administered with nonfibrogenic TiO2 particles. Total numbers of PAM recovered by bron-choalveolar lavage, however, were not significantly different from the controls after asbestos or silica instillation. Instead, inflammatory exudation of polymorphonuclear leukocytes, lymphocytes, and monocytes significantly increased in the groups of asbestos- or silica-instilled animals, but not in TiO2-instilled mice. These data suggest that i...

DETERMINATION OF THE TOXICITY OF CYCLOTRIPHOSPHAZENE HYDRAULIC FLUID BY 21-DAY REPEATED INHALATION AND DERMAL EXPOSURE

Cyclotriphosphazene (CTP) ester is one of a series of compounds developed for use as a fire-resistant hydraulic fluid. The most significant routes of industrial exposure to hydraulic fluids are dermal, because of spills or leaks, and aerosol inhalation from pressurized system leaks. This study was designed to evaluate the toxic effects associated with repeated or continuous exposure to CTP by both dermal and inhalation routes. Male and female Fischer 344 (F-344) rats were exposed for 3 weeks to air alone, or to 0.25, 0.50, or 1.00 mg CTP/L. No deaths or signs of toxic stress occurred during the exposure period. A depression in mean body weight gain and increases in numbers of pulmonary alveolar macrophages and renal hyaline droplets were noted in both genders. Male and female New Zealand White (NZW) rabbits were treated dermally for 3 weeks with mineral oil, or 0.25, 0.50, or 1.00 g CTP/kg. No toxic effects were noted in either gender of rabbits.

Determination of the Toxicity of Cyclotriphosphazene Hydraulic Fluid by 21-Day Repeated Inhalation and Dermal Exposure

Cyclotriphosphazene (CTP) ester is one of a series of compounds developed for use as a fire-resistant hydraulic fluid. The most significant routes of industrial exposure to hydraulic fluids are dermal, because of spills or leaks, and aerosol inhalation from pressurized system leaks. This study was designed to evaluate the toxic effects associated with repeated or continuous exposure to CTP by both dermal and inhalation routes. Male and female Fischer 344 (F-344) rats were exposed for 3 weeks to air alone, or to 0.25, 0.50, or 1.00 mg CTP/L. No deaths or signs of toxic stress occurred during the exposure period. A depression in mean body weight gain and increases in numbers of pulmonary alveolar macrophages and renal hyaline droplets were noted in both genders. Male and female New Zealand White (NZW) rabbits were treated dermally for 3 weeks with mineral oil, or 0.25, 0.50, or 1.00 g CTP/kg. No toxic effects were noted in either gender of rabbits.

Surface morphology and morphometry of rat alveolar macrophages after ozone exposure

As the ultrastructural data on the effects of ozone on pulmonary alveolar macrophages (PAM) are lacking, transmission (TEM) and scanning (SEM) electron microscopy were performed on rat PAM present in alveolar lavages following exposure to ozone. Rats were continuously exposed for 7 d to ozone concentrations ranging from 0.25 to 1.50 mg/m3 for 7 d followed by a 5-d recovery period. Additionally, morphometry on lung sections was performed to quantitate PAM. In a second experiment rats were continuously exposed to 1.50 mg O3/m3 for 1, 3, 5, or 7 d. To study the influence of concurrent ozone exposure and lung infection, due to Listeria monocytogenes, rats were exposed for 7 d to 1.50 mg O3/m3 after a Listeria infection. The surface area of lavaged control PAM was uniformly covered with ruffles as shown by SEM and TEM. Exposure to 0.5 mg ozone/m3 for 7 d resulted in cells partly covered with microvilli and blebs in addition to normal ruffles. The number of large size PAM increased with an increase in ozone concentration. After 1 d of exposure, normal-appearing as well as many small macrophages with ruffles and scattered lymphocytes were seen. Lavage samples taken after 5 or 7 d of exposure showed an identical cell composition to that taken after 3 d of exposure. After Listeria infection alone, lavage samples consisted of mainly lymphocytes and some macrophages. Small quantitative changes, such as an increase in the number of polymorphonuclear neutrophils and large-size PAM, occurred in lavages after ozone exposure and infection with L. monocytogenes. Morphometric examination of lung sections revealed a concentration-related increase in the number of PAM, even in animals exposed to 0.25 mg ozone/m3 for 7 d. Centriacinar regions were more severely affected than other regions of lung tissue. By 5 d after termination of exposure to ozone, the number of lysozyme-positive alveolar cells was still significantly increased in centriacinar areas of the lung. The results indicate that ozone exposure causes major changes in the number, size, and surface morphology of PAM in rat lung. Furthermore, the results presented here suggest that changes in alveolar macrophage function are reflected by morphological changes.

Response of rat alveolar macrophages to ozone: quantitative assessment of population size, morphology, and proliferation following acute exposure.

The purpose of this study was to evaluate the in vivo effects of an acute exposure to low levels of ozone on rat pulmonary alveolar macrophages (PAM). Fisher 344 rats exposed to 0.0, 0.12, 0.8, or 1.5 ppm O3 for 6 h were killed immediately after and 3, 18, 42, or 66 h after ozone exposure and their lungs were lavaged. Compared to sham-exposed (control) rats, exposure to 0.12 ppm O3 had no measurable effect on the total number, labeling index (LI), mitotic index (MI), or morphology of rat alveolar macrophages. The number of neutrophils was significantly (p less than or equal to 0.001) greater than in controls at 3, 18, and 42 h after exposure to 1.5 ppm O3 and 42 h after exposure to 0.8 ppm O3. The number of PAM was approximately twice that of controls 42 and 66 h after exposure to 0.8 and 1.5 ppm O3. There was a significant (p less than or equal to 0.001) increase in PAM MI 42 and 66 h after exposure to 1.5 ppm O3 and 42 h after 0.8 ppm O3. The increase in the number of PAM in mitosis was preceded by an increase in PAM LI. The PAM LI was significantly (p less than or equal to 0.001) greater than controls 18 and 42 h after exposure but returned to near normal levels by 66 h after exposure. There was a transient decrease in the mean nuclear/cytoplasmic ratio of PAM from rats exposed to 1.5 ppm O3 18 and 42 h after exposure due to an increase in the mean PAM cytoplasmic area. Comparison of the PAM population doubling time (Dt) and cell cycle time (Ct) suggest that PAM proliferation played a significant role in the observed increase in PAM following exposure to 0.8 and 1.5 ppm O3. These results highlight the dynamic response of PAM to an acute exposure to ozone and suggest that the proliferative response of pulmonary alveolar macrophages may be a useful indicator of pulmonary damage following inhalation of an irritant oxidant.

The Induction of Nuclear Abnormalities in the Alveolar Macrophages of Mouse Lung After Inhalation of &lt;sup&gt;239&lt;/sup&gt;PuO&lt;sub&gt;2&lt;/sub&gt;

Abstract Effects of inhaled 239PuO2 on the free-cell population of the lungs of CBA/H mice have been studied using broncho-alveolar lavage following initial alveolar depositions (IADs) of 209 to 2800 Bq. The numbers of pulmonary alveolar macrophages (PAM) were quantified using a radioactive tracer technique. The number of PAM declined soon after exposure; the extent of the depression and the time taken to reach the nadir were both dose-dependent. Numbers eventually returned to normal but, with high IADs, only after 3 months. The number of binucleate PAM and those with micronuclei increased following relatively low IADs in a dose-dependent manner. Micronucleate PAM were the most sensitive indicator of cellular damage as a result of 239PuO2 inhalation, providing a short-term assessment of radiation damage to lung cells at doses known to produce lung tumours. The results also provide further evidence that mitosis in PAM, or in their precursor cells, does occur within the lung.

Proliferation of pulmonary alveolar macrophages during postnatal development of rabbit lungs.

The purpose of this study was to determine if proliferation of pulmonary alveolar macrophages (PAM) played a significant role in establishing the PAM population of the lung during postnatal development. New Zealand albino rabbits were killed at 0.5, 1 through 5, 7, and 14 days and 4 months after birth and their lungs were lavaged. Cell yield in the lavage was determined by hemocytometer counts, and the percentage of PAM in mitosis (mitotic index) was determined from cytocentrifuge preparations. The total number of PAM increased from 1.5 X 10(6) at 1 day to 8.38 X 10(6) at 14 days after birth. The mitotic index (MI) was 0.6% at 0.5 days after birth, increased to 1.6% at 1 day, and remained elevated through 5 days. By 14 days, the MI declined to 0.2%. The cell cycle time (Ct) of the PAM population was calculated from the MI and ranged from 1.8 to 2.4 days during Days 1 through 5 of life. Direct measurements of the doubling time (Dt) of PAM in the lavage revealed that the PAM population doubled twice over this same time period. Because Ct was equal to Dt during Days 1 through 5, we conclude that proliferation of PAM was the primary mechanism by which the PAM population increased during the immediate postnatal development of the rabbit lung. No evidence was obtained indicating that migratory monocytes or interstitial macrophages were involved with this process of population expansion. This study adds to the growing literature demonstrating that the intraalveolar proliferation of "free" PAM is the major local source of PAM in the lung.

Alveolar macrophage phagocytic kinetics following pulmonary parainfluenza-3 virus infection.

Qualitative and quantitative evaluations of the cellular components of bronchoalveolar washings of calves with experimental parainfluenza-3 virus pneumonitis and control calves were made. Calves were exposed to 10(9) TCID50 of PI-3 by intranasal aerosol exposure and bronchoalveolar cells obtained 7 days after infection by volume-controlled bronchopulmonary lavage. Transient tachypnea and pyrexia occurred in all infected calves, and virus was recoverable at 7 days from nasal swabs and lung tissue. Pulmonary lesions were typical of viral pneumonitis, characterized by patchy alveolitis and bronchiolitis with accumulations of cells and inflammatory debris. The mean total lavage cell yield was elevated in the virus-infected calves, and the percentage of neutrophils was elevated (P less than 0.05). Increased numbers of pulmonary alveolar macrophages (PAM) were also recovered but the difference was not significant. Linear regression equations showed that a decreased proportion of PAM from virus-infected animals were phagocytic. The mean initial phagocytic rates of macrophages from calves with viral pneumonitis were significantly decreased (P less than 0.05) over controls. This difference was concentration dependent and required a phagocytic stimulus in excess of 12.5 X 10(6) beads/ml. Studies of phagocytic kinetics showed that PAM from calves with viral pneumonitis had a lower Vmax than PAM from control calves, but that Km values were comparable. No differences in PAM beta-glucuronidase and acid phosphatase activity were observed. These results indicate depressed phagocytic function in PI-3 virus-inflamed lungs relative to controls. In concert with virus-induced airway lesions, such in vivo depression of PAM phagocytic functions would be expected to depress pulmonary particulate clearance and lung defense mechanisms.

DNA synthesis in pulmonary alveolar macrophages and type II cells: effects of ozone exposure and treatment with alpha-difluoromethylornithine.

An increase in the number of pulmonary alveolar macrophages (AM) can be induced by a number of toxic insults to the lung, including ozone, an important photochemical oxidant air pollutant. This increase could arise from an influx of monocytes from the vascular or interstitial compartments, or from proliferation of AM in situ. While proliferation of alveolar type II cells after oxidant exposure has been well documented, it is not clear whether AM are also capable of this response. Rats were exposed to air or to 0.12, 0.25, or 0.50 ppm ozone for 1, 2, 3, 7, or 14 d, 20 h/d. The labeling index in both AM and type II cells increased about 10-fold after 2 d of exposure to 0.25 and 0.50 ppm of ozone, but returned to control levels by the end of 1 wk of exposure. These changes closely paralleled the temporal and dose-response characteristics of changes in total lung DNA synthesis. alpha-Difluoromethylornithine (DFMO) administered to rats during a 2-d exposure to 0.50 ppm ozone did not inhibit the ozone-induced increase in labeling index in AM or type II cells, although evidence of inhibition of lung ornithine decarboxylase activity was obtained, and the ozone-induced increase in total lung DNA synthesis was inhibited by 23%. These results suggest that, like type II cells, AM are capable of entering the cell cycle and synthesizing new DNA in situ in response to short-term exposure to environmentally relevant doses of ozone, and that the ozone-induced stimulation of DNA synthesis in these cell types was refractory to inhibition by DFMO.

Preliminary studies of the interaction between 239PuO2 and cigarette smoke in the mouse lung.

Our current experiments were designed to show whether 12 months' exposure to cigarette smoke enhances the incidence of lung tumours in mice that had previously inhaled 239PuO2. These periods of smoke exposure are almost complete. After death their lungs will be cleared and any nodules found will be sectioned for histopathology. This paper reports the results of two preliminary experiments conducted earlier. The first study showed that mice could tolerate the proposed smoking regime for 3 months, with no sign of ill health in any animal throughout. The major difference found was a reduced growth rate in both smoke- and sham-exposed mice relative to that of cage controls. After 3 months of treatment, histopathology and morphometry of lung sections found only slight smoke-induced changes. These included a reduced proportion of alveolar space and an increased number of pulmonary alveolar macrophages (PAM) per unit area. Bronchopulmonary lavage showed that the PAM from smoke-exposed mice were larger than those from sham-exposed or control mice and that an increased proportion of cells were binucleate. All mice in the second study were initially exposed to 239PuO2, then subsequently divided into three treatment groups as above. Cigarette smoke exposure was shown to inhibit the removal of 239Pu from the lung whilst sham exposure had no effect. Smoke exposure also produced an increase and sham exposure a decrease in lung weights relative to those of cage controls. The latter was probably as a result of their lower growth rate. In our current experiments it is likely that the group receiving 239PuO2, then smoke, will receive a higher radiation dose to lung than those receiving 239PuO2 only. Any increased tumour incidence found will be considered in conjunction with this evidence.

Macrophage Depletion of Mouse Lung Following Inhalation of 239 PuO 2

Changes in the free-cell population of the lungs of two strains of mice (SAS/4 and CBA/H) were studied up to 4 months after inhalation exposure to a sized fraction of 239PuO2 particles (1.5 micron AMAD) to give initial alveolar depositions (IADs) ranging from 17 to 810 Bq. A sample of the free-cell population of the lung was recovered by bronchoalveolar lavage, and a radiometric method was used to estimate the total number of pulmonary alveolar macrophages (PAM) in the lung. The response of the lung to 239PuO2 was characterized by an initial, dose-dependent depression in the total number of PAM following an IAD as low as 50 Bq. At IADs greater than 150 Bq, the initial depression continued for longer, merging into a chronic phase in which the PAM were larger and were accompanied by a minor infiltration of leukocytes. These findings were confirmed by histology, which also revealed focal accumulations of Type II pneumocytes. The results indicate that inhaled alpha-emitting particles are effective at producing a depletion in the alveolar macrophage population at relatively low IADs and that chronic effects on the cells can be produced by higher concentrations.

Abnormal phagolysosome fusion in pulmonary alveolar macrophages of rats exposed chronically to cigarette smoke.

Cigarette smoking is strongly associated with functional and morphologic changes in pulmonary alveolar macrophages (PAM). Phagocytic activity is a primary function of PAM, and although the ingestion of particles appears to be normal in the PAM of cigarette smokers, published data suggest that antimicrobial activity of these cells might be diminished. Because phagolysosome fusion (PLF) is an important aspect of the phagocytic process subsequent to the ingestion phase, PLF was evaluated in PAM lavaged from the lungs of male Fisher 344 rats that had been exposed for 8 wk to whole cigarette smoke, to the gas phase of cigarette smoke, or to air as a sham control. Using the acridine orange assay with viable yeast as the phagocytic challenge, we found no difference in the numbers of PAM from each group that had phagocytic activity after a 2-h challenge. However, PLF expressed as the number of orange-fluorescing phagosomes per total number of yeast-containing phagosomes was 35 +/- 1% (X +/- SE) for the group exposed to whole smoke, 53 +/- 2% for the gas phase group, and 65 +/- 1% for the control group. These differences are highly significant, with p less than 0.001 for the whole smoke versus control and p less than 0.01 for the gas phase versus control. Tight phagosomal membranes suggest normal PLF, whereas loose membranes suggest that PLF is inhibited. When the structure of yeast-containing phagosomes was examined and PLF was calculated with the number of phagosomes with tight membranes substituted for the number of orange-fluorescing phagosomes, PLF in smoke-exposed PAM was found to be significantly (p less than 0.01) different from that in sham-exposed control PAM.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitation of leukocytes in bronchoalveolar lavage samples from rats after intravascular injection of endotoxin.

Although it has been demonstrated in several animal species that neutrophils aggregate in the pulmonary microvasculature after intravenous infusion of chemotactic factors or substances that activate the systemic complement system, studies in rabbits have revealed that affected neutrophils do not migrate out of capillaries into air spaces unless the infusion processes are combined with manipulative procedures involving the airways, such as intubation or instillation of an anesthetic agent. In this study, we report that after intravenous infusion of endotoxin (Escherichia coli) into Sprague-Dawley rats, in the absence of airway manipulation, the absolute number of neutrophils in bronchoalveolar lavage (BAL) samples increased significantly 24 and 48 h after injection. This represented nearly a 24-h delay from the time that maximal numbers of neutrophils were seen in lung tissue and approximately an 18-h delay from the time that maximal numbers appeared in peripheral blood. We also found that after infusion of endotoxin the number of pulmonary alveolar macrophages (PAM) recoverable in BAL samples fell dramatically within 2 h and remained suppressed for at least 12 h. We conclude that: (1) neutrophils are capable of migration into air spaces after aggregation in pulmonary capillaries in the absence of airway manipulative procedures or instillation of chemoattractants into air spaces and (2) endotoxemia affected surface parameters of PAM in vivo that determine recoverability by lavage.