The origin of alveolar macrophages in mouse radiation chimeras.

This investigation attempted to determine whether the primary source of alveolar macrophages is pulmonary or hematopoietic. We have utilized an antigenic marker to identify cells of hematopoietic origin. Mouse chimeras were produced by irradiating C57B6/AF(1) mice (900 R) and then injecting them intravenously with B10D2/AF(1) bone marrow. The donor animal has an antigenic specificity on the H-2 locus, not shared by the recipient. Alveolar macrophages were obtained by repeated lung washings with physiologic saline at 37 degrees C. Cytotoxic tests were done on bone marrow and alveolar macrophages using anti-31 mouse antibody, absorbed rabbit serum as complement, and trypan blue exclusion as a test for viability. Animals were studied at 7, 14, 21, 28, and 35-50 days and 4, 5, 8, and 11 months after irradiation and bone marrow replacement. By 21 days after irradiation, 90% of the animals had greater than 80% replacement of marrow with donor tissue; and white blood cell and alveolar macrophage counts approached normal. At this time and at later intervals the per cent of donor cells in the lung free cell population was not significantly different from the per cent of donor cells in the bone marrow. Similarly, after aerosol particulate exposure, the percentage of marrow cells and alveolar macrophages of donor origin were not significantly different. This immunologic approach suggests that alveolar macrophages in radiation chimeras are entirely of hematopoietic origin.

Sex-related differences in tumor progression associated with altered lymphocyte circulation.

Male and female ACI rats were inoculated with the syngeneic H-4-II-E hepatoma, and the natural history of the tumor, histopathology, and lymphocyte migration were studied. The tumor formed a.s.c. mass in all 16 males and in 22 of the 26 females given injections. In the males, tumors progressed, and all animals died with the mean survival time of 54 days. Complete tumor regression was observed in all but two females. In the females, there was prominent lymphocytic infiltration of the tumor, while males had no cellular reaction at the tumor site. The regional lymph nodes in males usually contained metastases and were nonreactive. The female lymph nodes did not contain metastases but contained many lymphocytes within the peripheral sinus and sinusoids. Six male-female pairs were castrated before tumor inoculation. Castration had no effect on the natural history or the etiology of the tumor. Comparing seven normal control male-female littermate pairs, there were no differences in lymphocyte accumulation in the lymph nodes 22 hr following injection of 51Cr-labeled syngeneic lymphocytes. In seven tumor-bearing male-female littermate pairs, there was a significant decrease in lymphocyte migration to the lymph nodes (p less than 0.001) in tumor-bearing males as compared to that in both their female littermates and control males. Depressed lymphocyte circulation in the males was associated with rapid progression of tumors resulting in the death of the animals. Unimpaired lymphocyte mobilization in the tumor-bearing females was associated with complete regression in most animals.

Oxidative metabolism in the alveolar macrophage: analysis by flow cytometry.

We evaluated the reagents dichlorofluorescin (DCFH) and hydroethidine (HE) for use in flow cytometric analysis of the respiratory burst of alveolar macrophages and monocytes. DCFH and HE are non-fluorescent precursors which can be oxidized intracellularly to the fluorescent compounds dichlorofluorescein and ethidium. Alveolar macrophages (AMs) loaded with either DCFH or HE were analyzed after phorbol myristate acetate (PMA) stimulation. The results, expressed as fmol/cell oxidation product (DCF or ethidium) after fluorometric standardization of the flow cytometer, show that both DCFH (273 +/- 48, mean increase over control +/- SE, fmol/cell, N = 9) and HE (416 +/- 54, N = 11) detected the substantial respiratory burst of hamster AMs. Similar results were obtained with normal human AMs. By using multiparameter analyses, the oxidative response of AMs ingesting opsonized fluorescent latex beads was measured in subpopulations ingesting increasing numbers of particles. A graded increase in oxidation of both DCFH and HE was found in response to increasing phagocytosis. Ingestion of fluoresceinated staphylyococcal bacteria caused similar changes in HE-loaded AMs. Inhibition of respiration with antimycin showed that approximately 95% of the increased oxidative metabolism of hamster AMs ingesting opsonized beads or bacteria was mitochondrial. The remaining 5% (10-40 fmol/cell) is membrane-derived oxidative activity quantitatively similar to that measured in assays of extracellular release of H2O2. Monocytes loaded with either DCFH or HE showed substantial increases in fluorescence after PMA stimulation (mean % increase over control +/- SE at 30 min: 464 +/- 104, DCFH, 505 +/- 156, HE). While DCHF is known to measure H2O2, HE is less well characterized. Exposure of cells to an extracellular source of both superoxide anion (O2-) and H2O2, xanthine oxidase-xanthine, resulted in marked oxidation of intracellular HE. Addition of both superoxide dismutase and catalase blocked this oxidation, indicating that HE can detect both O2- and H2O2. These agents can be useful probes for precise analysis of oxidative metabolism during phagocytosis in AMs and other mononuclear phagocytes.

Sulfite stimulates NADPH oxidase of human neutrophils to produce active oxygen radicals via protein kinase C and Ca2+/calmodulin pathways.

The effect of sulfite on the oxidative metabolism of human neutrophils was studied in vitro. Superoxide anion production of PMN was determined using superoxide dismutase-inhibitable lucigenin-dependent CL. The addition of sulfite in concentrations of 0.01 mM-1 mM results in an up to 6-fold increase in CL of nonstimulated neutrophils at 37 degrees C and pH 7. Neutrophils stimulated with zymosan or PMA have an additional 2-fold stimulation when sulfite is added. Higher sulfite concentrations (2 mM-10 mM) decrease the CL of both nonstimulated and stimulated cells. The activity of NADPH oxidase, responsible for O2.- production, is significantly increased in neutrophils incubated with 1 mM sulfite. Neutrophils from patients with chronic granulomatous disease, which are cytochrome b558 negative or have p47phox deficiency, exhibit no significant NADPH oxidase activity and show no increase in CL by sulfite. Inhibitors of protein kinase C, H7, and calphostin C, as well as inhibitors of Ca(2+)- and calmodulin-dependent processes, W7, and R 24 571, completely inhibited the increased CL of sulfite-treated neutrophils. These findings indicate that sulfite in low concentrations stimulates neutrophils to produce superoxide anions by activation of NADPH oxidase through a signal transduction pathway involving protein kinase C and Ca2+/calmodulin.

Factors associated with correct antemortem diagnosis of major pulmonary embolism.

Difficulties persist in the accurate clinical diagnosis of major pulmonary embolism despite the availability of lung scans and pulmonary angiography. To evaluate factors associated with the correct antemortem diagnosis of pulmonary embolism, we reviewed all 1,455 autopsy reports at the Peter Bent Brigham Hospital from 1973 to 1977. Of 54 patients identified with anatomically major pulmonary embolism at autopsy, 16 (30 percent) had correct antemortem diagnosis. Accuracy was far greater in postoperative patients (64 percent) (p = 0.02) and in patients with autopsy-proved venous thrombosis (55 percent) (p = 0.005). Lung scanning (82 percent) (p = 0.0002) and pulmonary angiography (80 percent) (p = 0.05) during the 10 days prior to death were also associated with an increased tendency to correct clinical diagnosis of pulmonary embolism. In contrast, among 21 patients with autopsy-proved major pulmonary embolism who also had pneumonia, no pulmonary embolism was diagnosed before death (p = 0.0001). Furthermore, among patients 70 years of age or older, only 10 percent with pulmonary embolism at postmortem examination had a correct diagnosis prior to death (p = 0.02). In patients with pneumonia or in elderly patients, an increased awareness of the possibility of pulmonary embolism and more frequent use of lung scanning and pulmonary angiography may increase the accurate clinical diagnosis of pulmonary embolism.

Development of surface membrane characteristics of "premedullary" macrophages in organ cultures of embryonic rat and hamster lungs.

A replicating population of non-monocyte-derived free cells appears in organ-cultured embryonic rat lungs, indistinguishable from alveolar macrophages by classical criteria such as ultrastructure, lysosomal enzyme cytochemistry, and phagocytic behavior. We demonstrate similar events in cultured embryonic hamster lungs and development of macrophage-associated properties on the plasmalemma of these cells in both species. Immunoperoxidase localizations were obtained using monoclonal antibodies against alveolar macrophage antigen (HAM1) in hamsters, and rat macrophage antigen (ED1) and leukocyte-common antigen (OX1) in rats. Fc and C3b receptors were identified in both species by immune rosetting. HAM1 staining, perinuclear in rare cells at explantation, gains definitive surface localization 3-4 days later as cells prepare to emerge through the pleura. ED1 and OX1 cytoplasmic staining first occurs after 24 hr, increases as macrophages multiply and congregate beneath the pleura, and translocates to the plasmalemma of emerged cells. Some glass-adherent cells from lung explants have Fc receptors. The proportion rises sharply for 24 hr and equals fully emerged cells (90-95%) by days 3-4. At first phagocytosis is slow to follow Fc receptor binding, but ingestion time decreases to 3-10 min as macrophages mature. A minority of emerged macrophages bind complement-opsonized erythrocytes, which are rarely taken up. These properties are shared by alveolar macrophages of adults.

Inhaled concentrated ambient particles are associated with hematologic and bronchoalveolar lavage changes in canines.

Pulmonary inflammatory and hematologic responses of canines were studied after exposure to concentrated ambient particles (CAPs) using the Harvard ambient particle concentrator (HAPC). For pulmonary inflammatory studies, normal dogs were exposed in pairs to either CAPs or filtered air (paired studies) for 6 hr/day on 3 consecutive days. For hematologic studies, dogs were exposed for 6 hr/day for 3 consecutive days with one receiving CAPs while the other was simultaneously exposed to filtered air; crossover of exposure took place the following week (crossover studies). Physicochemical characterization of CAPs exposure samples included measurements of particle mass, size distribution, and composition. No statistical differences in biologic responses were found when all CAPs and all sham exposures were compared. However, the variability in biologic response was considerably higher with CAPs exposure. Subsequent exploratory graphical analyses and mixed linear regression analyses suggested associations between CAPs constituents and biologic responses. Factor analysis was applied to the compositional data from paired and crossover experiments to determine elements consistently associated with each other in CAPs samples. In paired experiments, four factors were identified; in crossover studies, a total of six factors were observed. Bronchoalveolar lavage (BAL) and hematologic data were regressed on the factor scores. Increased BAL neutrophil percentage, total peripheral white blood cell (WBC) counts, circulating neutrophils, and circulating lymphocytes were associated with increases in the aluminum/silicon factor. Increased circulating neutrophils and increased BAL macrophages were associated with the vanadium/nickel factor. Increased BAL neutrophils were associated with the bromine/lead factor when only the compositional data from the third day of CAPs exposure were used. Significant decreases in red blood cell counts and hemoglobin levels were correlated with the sulfur factor. BAL or hematologic parameters were not associated with increases in total CAPs mass concentration. These data suggest that CAPs inhalation is associated with subtle alterations in pulmonary and systemic cell profiles, and specific components of CAPs may be responsible for these biologic responses.

Pulmonary vascular disease and hypertension after valve surgery for mitral stenosis.

The reduction of pulmonary hypertension that occurs within 24 hours of valve replacement for mitral stenosis is well documented, but patients who die after surgery have not been adequately studied. Clinical and autopsy data for 16 patients who died following mitral valve replacement were reviewed. The emphasis was on preoperative and postoperative pulmonary arterial pressure and pulmonary vascular disease, including arterial, venous, and capillary changes. Morphologic features were graded and summed to obtain an additive histologic assessment (AHA). Patients were divided into three groups: 1) those who had uneventful operations and early postoperative periods but died prior to discharge; 2) those who had postoperative difficulty, with identifiable acute anatomic causes of death; and 3) those who had postoperative difficulty, with no apparent acute anatomic cause of death. In group 1 (n = 4) the preoperative pulmonary arterial pressure was 43 +/- 17 mm Hg, and AHA ranged from 0 to 4; in group 2 (n = 5) the preoperative pulmonary arterial pressure was 60 +/- 15 mm Hg, but AHA ranged only from 2 to 5. In group 3 (n = 7) the preoperative pulmonary arterial pressure was 59 +/- 12 mm Hg; AHA ranged from 6 to 9, significantly higher than that of the other groups (P less than 0.005). Three patients from group 3 had elevated pulmonary arterial pressure (60, 52, and 50 mm Hg three, six, and 15 days after surgery, respectively). Two additional patients had right heart failure with normally contracting left ventricles terminally. It is concluded that some patients with mitral stenosis who die after surgery with persistently elevated pulmonary arterial pressure have sufficiently severe pulmonary vascular disease to account for their persistent pulmonary hypertension and death.

Pathologic patterns of Serratia marcescens pneumonia.

To characterize the pulmonary lesions caused by Serratia marcescens, the authors reviewed all autopsy-culture-proven cases of S. marcescens pneumonia occurring at their hospital between 1968 and mid-1980. In 16, S. marcescens was the only organism cultured from the lungs during life or at autopsy. This report describes primarily these pure infections. Two histopathologic reactions were seen. Nine non-neutropenic patients had acute, hemorrhagic bronchopneumonia, seven with microabscesses and two with larger cavities. In seven, distinctive vasculitis was apparent in vessels larger than 75 microns in diameter; intramural gram-negative rods were identified in two. Seven immunosuppressed patients had diffuse neutropenic pneumonitis resembling diffuse alveolar damage, with extensive intra-alveolar fibrinous exudates and pulmonary hemorrhage. In two patients, bacteria without cellular reaction were present. In patients with prolonged infections, focal areas of intra-alveolar organization and bronchiolitis obliterans accompanied both patterns. Since the incidence of nosocomial S. marcescens infection is increasing and since pneumonia caused by this organism is recognizable histologically, autopsy cultures positive for S. marcescens should not be disregarded.

Imaging of lymphoid structures with indium-111-labeled lymphocytes.

The ability of indium-111 (111In)-oxine-labeled syngeneic lymphocytes to migrate normally and their suitability for imaging both normal lymphoid structures and those with metastatic disease were assessed. Sixty-two ACI rats were studied, 34 of which received injections in the left foot pad with 1 X 10(6) syngeneic H-4-II-E hepatoma cells nine to 44 days before imaging. Most animals were bearing palpable tumors when imaged. In 53 experiments, 1-6 X 10(8) 111In-oxine-labeled lymphocytes with a labeling concentration of 5-80 microCi/10(8) cells were injected intravenously. Gamma camera images were obtained 22 hours later. After the last image, the animals were killed. The lymph nodes, liver, spleen, lungs, and left femur were dissected, and the recovered radioactivity was determined in a gamma well counter. Lymph nodes could be partially or completely visualized in 70% of the animals (15 tumor-bearing and 16 normal out of 44 technically satisfactory experiments). Large metastatic nodes were seen clearly. Lymphocytes labeled with 111In-oxine exhibited a normal migration from blood to lymph nodes.

Transmission electron microscopy of intracellular particles of polyethylene from joint replacement prostheses: size distribution and cellular response.

The objectives of this transmission electron microscopy study of peri-implant tissues retrieved at revision arthroplasty were to (1) determine the size distribution of intracellular polyethylene particles, and (2) assess the cellular response to phagocytosed polyethylene particles as revealed by the condition of the cellular organelles. The frequency distributions of intracellular polyethylene particle sizes for 15 cases of total hip replacement showed that more than 75% of the particles had lengths of less than 0.5 microm. More than 90% of the particles were less than 1.0 microm in size. In comparison, the frequency distribution for the particles in cellscomprising tissue retrieved from three total knee replacement prostheses showed that only 43% of the particles were less than 0.5 microm in length and 72% were less than 1 microm in size. There was no statistically significant difference in the mean particle length between the specimens from the hip and knee patients. The majority of the cells containing polyethylene were without signs of degeneration. The cytoplasmic and nuclear membranes were intact. Several electron lucent voids which once contained polyethylene particles were seen surrounded by several healthy appearing mitochondria, which displayed sharp membranes and intact cristae. There were no signs of a cytotoxic response to polyethylene at the ultrastructural level.

Contribution of osmium tetroxide to the image quality and detectability of iron in cells studied by electron spectroscopic imaging and electron energy loss spectroscopy.

The detection of elemental distributions within ultrastructural cellular components presents a number of challenges. There are many technical questions that need to be resolved including optimal fixation protocols. Another is the impact of heavy metals, such as osmium tetroxide (OsO4), on the detectability of other elements when OsO4 is used in chemical fixation protocols for biological samples. OsO4 was examined by varying its concentrations from 0% to 1% and time of fixation from 5 to 30 minutes with hamster alveolar macrophages. The morphological quality of cellular images observed and the detectability of iron using electron spectroscopic imaging (ESI) and electron energy loss spectroscopy (EELS) were evaluated. One percent OsO4 for 30 minutes in the chemical fixation protocol enhances the quality of the ESI and does not interfere with the ESI or EELS signal of iron. Positive results from both methods indicate the presence of the specific element. The loss of 59Fe during the chemical fixation procedure was also studied. Less than 10% was lost during the primary fixation step, but minimal losses occurred through dehydration, embedding, and sectioning. Careful technical assessment of the presence of an element as well as factors which might interfere with its detection is an important step in the application of any analytical microscopic technique.

Peritoneal responses to implanted fabrics used in operating rooms.

Fibers obtained from woven and nonwoven surgical fabrics were operatively instilled into the peritoneal cavities of rats to assess pathologic responses at 2, 8, 16, and 32 weeks postoperatively. Adhesions were found in 20% of the sham-operated animals and in 22% and 29% of the animals with fibers from woven and nonwoven fabrics, respectively. These differences were not significant. The adhesions were not progressive with time and were not related to increased morbidity or mortality rates. Foreign-body granulomas were found in 8% of the sham-operated animals and in 96% and 89% of the animals with fibers from woven and nonwoven fabrics, respectively. Differences between the groups with fibers were not significant. There were no morphologic differences in the granulomas of woven and nonwoven materials; they did not increase in number of enlarge with time. We conclude that responses to cellulose fibers, whether from cotton or from nonwoven materials produced from wood fibers, are virtually identical.

Anatomic localization of 24- and 96-h particle retention in canine airways.

Long-term retention of particles in airways is controversial. However, precise anatomic localization of the particles is not possible in people. In this study the anatomic location of retained particles after shallow bolus inhalation was determined in anesthetized, ventilated beagle dogs. Fifty 30-cm(3) boluses containing monodisperse 2.5-micron polystyrene particles (PSL) were delivered to a shallow lung depth of 81-129 cm(3). At 96 h before euthanasia, red fluorescent PSL were used; at 24 h, green fluorescent PSL and (99m)Tc-labeled PSL were used. Clearance of (99m)Tc-PSL was measured during the next 24 h. Sites of particle retention were determined in systematic, volume-weighted random samples of microwave-fixed lung tissue. Precise particle localization and distribution was analyzed by using gamma counting, conventional fluorescence microscopy, and confocal microscopy. Within 24 h after shallow bolus inhalation, 50-95% of the deposited (99m)Tc-PSL were cleared, but the remaining fraction was cleared slowly in all dogs, similar to previous human results. The three-dimensional deposition patterns showed particles across the entire cross-sectional plane of the lungs at the level of the carina. In these locations, 33 +/- 9.9% of the retained particles were found in small, nonrespiratory airways (0.3- to 1-mm diameter) and 49 +/- 10% of the particles in alveoli; the remaining fraction was found in larger airways. After 96 h, a similar pattern was found. These findings suggest that long-term retention in airways is at the bronchiolar level.

Functional characterization of recombinant rat macrophage inflammatory protein-1 alpha and mRNA expression in pulmonary inflammation.

Chemokines are important inflammatory mediators that function by activating and recruiting leukocytes to an inflamed tissue. We have recently cDNA cloned the rat chemokine macrophage inflammatory protein-1 alpha (MIP-1 alpha) (1). In the present study, we characterize the biological function of recombinant MIP-1 alpha protein and describe expression of its mRNA both in vitro and in a rat model of lung inflammation. In vitro rat rMIP-1 alpha protein was chemotactic for both polymorphonuclear leukocytes (PMNs) and macrophages with maximal activity at 50 nM for both cell types. In in vivo studies, we found that intratracheal instillation of 1 and 5 micrograms of rMIP-1 alpha resulted in a significant (P < 0.05) influx of cells, primarily monocytes/macrophages, into the airspace of the lungs after 6 h. Mean numbers of lavagable PMNs were not elevated significantly (P < 0.05) for either dose of MIP-1 alpha. As a model of inflammation, rats were intratracheally instilled with 0.1 mg/kg bacterial lipopolysaccharide (LPS). Bronchoalveolar lavage (BAL) was performed 3 h later. Instillation of LPS resulted in an acute neutrophilia, but no significant change in lavagable macrophages. BAL cells from control animals (saline instilled) displayed no basal mRNA expression of either MIP-1 alpha or MIP-2 (positive control). In contrast, both MIP-1 alpha and MIP-2 mRNA levels increased markedly in BAL cells from rats instilled with LPS. The rat alveolar macrophage cell line (NR8383) also showed increased MIP-1 alpha mRNA levels in response to LPS (10 micrograms/ml) with a maximal increase after 6-8 h. The induction of MIP-1 alpha mRNA expression by LPS in NR8383 cells was attenuated by cotreatment with the antioxidants N-acetylcysteine and dimethylsulfoxide, suggesting that the induction of MIP-1 alpha mRNA by LPS is mediated via the generation of reactive oxygen species. We conclude that MIP-1 alpha is a potent chemoattractant for macrophages in vivo, and its mRNA expression in macrophages and BAL cells in response to inflammatory stimuli suggests a fundamental role in acute pulmonary inflammation.

Mechanisms of morbidity and mortality from exposure to ambient air particles.

The studies reported here assessed pathophysiologic mechanisms that result from exposure to concentrated ambient particles (CAPs) in animals with and without cardiopulmonary compromise. These studies were carried out to determine the biologic plausibility of epidemiologic observations of increases in particulate air pollution associated with increases in human morbidity and mortality. Dogs were exposed two at a time to CAPs or filtered air via tracheostomy for six hours per day on three consecutive days. The electrocardiogram (ECG) and breathing pattern were recorded continuously, and indicators of inflammation were also assessed. In one experimental design, normal dogs were exposed in pairs to CAPs and subsequently to filtered air or to filtered air and subsequently CAPs (the double CAPs/double sham design). Comparisons were made between the CAPs measurements and each dog's own sham responses. In another design, one dog was exposed to CAPs while the chambermate received a sham exposure; these experiments were followed by crossover of the protocol the subsequent week (the crossover design). Comparisons were made between the CAPs exposure and both the chambermate's sham and each dog's own sham responses. The crossover experiments were conducted in normal animals and in animals who had undergone balloon occlusion of the left anterior descending (LAD) coronary artery to induce myocardial compromise. The effects of CAPs in animals with induced chronic bronchitis were part of the original specific aims; because these studies were not fully pursued, the results are presented only in Appendix A. In normal dogs, analyses of all double CAPs and crossover studies revealed low frequency (LF) and high frequency (HF) powers for heart rate variability (HRV) that were significantly higher for CAPs exposure compared to sham exposure. Variation in day-to-day exposure concentrations, aerosol composition, and pathophysiologic responses were also found. The crossover design, continuous measures of aerosol mass, and biologic responses were incorporated in the development of a statistical model that allowed isolation of changes associated with CAPs from changes due to animal variations. Comparison of individual exposures with this model revealed a range from no response in any measured parameter to statistically significant changes in cardiac autonomic balance, pulmonary air flow, and breathing pattern. On days in which dogs showed statistically significant changes in responses, the findings were consistent in both cardiac and respiratory parameters. Days associated with significant increases in LF and HF HRV, LF/ HF HRV ratio, and heart rate standard deviation (HR SD) were also associated with decreases in average heart rate. These same days had decreases in respiratory frequency, tidal volume, minute volume, and peak flows with corresponding increases in respiratory cycle times and enhanced pause (Pauenh), a measure of bronchoconstriction. These cardiac and respiratory changes suggest an effect mediated via both the sympathetic nervous system and the vagus nerve. Alternatively, days associated with increased heart rate had decreases in the HR SD; decreases or no change in HF and LF HRV; increases in respiratory flows and volumes; and decreases in breathing cycle times, all suggesting only sympathetic nervous system mediation. When all data from the crossover design experiments were assessed with this model, the heart rate and respiratory rate were significantly decreased in relation to both cumulative and actual exposure and the LF HRV, LF/HF HRV ratio, HR SD, and all other respiratory parameters were significantly increased (p < 0.0001 for all). When cardiac data were grouped by days in which the air mass trajectory came from the north or northwest (versus west, south, east, or northeast), significant increases in HR SD and HF HRV and significant decreases in average heart rate were associated with the northwest trajectory. (ABSTRACT TRUNCATED)

Ultrafine particles in human lung macrophages.

As knowledge about size dependency of particle toxicity continues to grow, attention has been focused on ultrafine particles (i.e., < 0.1 microm in diameter). In recent studies with rats, investigators learned that ultrafine particles likely have greater pulmonary toxicity than larger particles, and it is possible that exposure to, and accumulation of, these particles in the human lung may be associated with adverse respiratory health effects. As part of an ongoing study, the authors performed bronchoalveolar lavage in 14 healthy current nonsmokers to investigate the extent to which ultrafine particles were present in lung macrophages. In addition, 10 of the 14 subjects performed pulmonary function tests. Eleven of the 14 subjects were utility workers, and 3 were nonmaintenance employees of a university. The authors used a Zeiss CEM902 electron microscope to study macrophages isolated from bronchoalveolar lavage fluid. Morphometric quantification revealed ultrafine particles in lung macrophages of all 14 volunteers; the average number of ultrafine particles/microm3 cytoplasm per cell (UFavg) ranged from 34 to 231 (mean = 95, standard deviation = 54). Regression analysis showed that the UFavg was associated inversely with percent predicted forced expiratory volume in 1 second (FEV1.0) (beta = -1.2 percent predicted FEV1.0/10 ultrafine particles x microm3 cytoplasm per cell [standard error = 0.45, p = .031). The demonstration of ultrafine particles in all 14 subjects, independent of occupational exposure, suggests that there is environmental exposure to ultrafine particles. The negative association between the number of ultrafine particles and ventilatory function demonstrates a need for further investigation into the pulmonary health effects of ultrafine particles.