Macrophages, dust, and pulmonary diseases.
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Adaptation, Biological↗
Biomedical subjects
D H Bowden
Publications and source records attributed to D H Bowden.
Crocidolite-induced pulmonary fibrosis in mice. Cytokinetic and biochemical studies.
The responses of pulmonary alveolar and bronchial cells to asbestos exposure were studied by relating the cytokinetic changes of injury and repair to the inflammatory process and subsequent fibroblastic activity. The lesions were induced by intratracheal instillation of 1 mg crocidolite asbestos in mice, which were killed up to 20 weeks thereafter; 3H-thymidine was injected 1 hour before death. A rapid inflammatory response with elevated polymorphonuclear leukocytes and lysosomal enzyme release was largely over by 2 weeks, but the increase in alveolar macrophages was maintained. Focal necrosis of bronchial epithelial cells was repaired by cell regeneration, whereby new epithelial cells overgrew luminal exudates to incorporate long asbestos fibers into the peribronchial interstitium, where macrophagic granulomas formed. Increased collagen levels were largely due to stimulation of peribronchial fibroblasts. A lesser reaction of epithelial damage, Type 2 cell proliferation, and fibroblast stimulation also occurred in the alveolar walls. The results suggest that macrophage-fibroblast interactions associated with enhanced fibrosis occur readily in the peribronchial interstitium following injury and repair of epithelial cells by long asbestos fibers.
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Bronchiolar and alveolar lesions in the pathogenesis of crocidolite-induced pulmonary fibrosis in mice.
Asbestosis is generally considered to result in restrictive pulmonary disease associated with interstitial fibrosis. Recently, however, attention has focused upon bronchiolar lesions and concomitant obstruction to air flow. The responses of conducting airways and alveoli were studied over a 20 week period following instillation of crocidolite to mice. The location of the lesions and the sequential inflammatory changes were studied by bronchoalveolar lavage, light and electron microscopy; regenerative activity was monitored by autoradiographs. Within 48 h there was multifocal necrosis of bronchiolar epithelium, maximal at bifurcations where longer fibres tend to adhere. Subsequently, intralumenal exudates were overgrown by epithelium and incorporated into the bronchiolar connective tissue where active peribronchiolar granulomas persisted until 20 weeks. Alveolar lesions were located predominantly in peribronchiolar air sacs and at the junctions of bronchioles and alveolar ducts. Focal acute injury of type 1 cells and transepithelial passage facilitated transport of short asbestos fibres to the interstitium where they were phagocytozed by macrophages. Regenerative activity was prompt with active division of type 2 epithelial cells. Biochemically, collagen increased after 4 weeks, when fibrosis was predominantly located in the bronchiolar lumens and in peribronchiolar connective tissue with lesser amounts in the centrilobular alveolar interstitium. The results suggest that the longer fibres induce bronchiolar injury and a more severe fibrotic pattern similar to the recently described changes seen in human asbestosis.
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Pathways of cellular efflux and particulate clearance after carbon instillation to the lung.
The pulmonary response to the deposition of carbon particles was investigated to determine the routes of cellular efflux and the mechanisms of particulate clearance. At intervals to 6 months after the intratracheal instillation of 4 mg carbon, the lungs of mice were fixed in situ by perfusion without lavage. Within a few hours, migration of granulocytes into the bronchioles was observed; large numbers of PMNs were not seen in alveoli until 24 h. Associated with the PMN efflux there was transient oedema with no evidence of pulmonary cell necrosis. The number of free macrophages increased in response to the carbon and mononuclear cell migration into alveoli and bronchioles was also observed. The bronchiolar component of the cellular efflux indicates that PMNs and macrophages recovered from the lung by lavage are not entirely of alveolar origin. Whereas most particles, either free or in phagocytic cells, were cleared by the bronchial tree, some transepithelial passage of free particles to the interstitium was observed. Some carbon was found in hilar lymph nodes but overall lymphatic clearance was low.
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The role of cell injury and the continuing inflammatory response in the generation of silicotic pulmonary fibrosis.
The pathogenesis of silicosis involves interaction between pulmonary macrophages and fibroblasts. The consequences of direct injury to pulmonary cells and the role of inflammatory cells other than the macrophage have received little attention. These were studied over a 20 week period after instilling silica to mice by correlating the changing inflammatory response, as revealed by bronchoalveolar lavage and lung sections, with the cellular location of silica particles and the development and resolution of granulomatous lesions. Within 24 h, a massive concentration of particles and PMN was seen in centrilobular locations with acute focal necrosis of type 1 epithelial cells. Rapid epithelial repair occurred but PMN were recovered from the lung up to 20 weeks. In the alveoli, silica was ingested by PMN and AM, resulting in the death of some cells; free particles crossed the epithelium and were found predominantly in peribronchial macrophages. Silicotic granulomas formed within a week and consisted mainly of fibroblasts macrophages and some PMN. It is suggested that the necrosis of type 1 epithelium and the continuing efflux with serial destruction of PMN may be important factors in the generation of silicotic fibrosis.
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The alveolar macrophage.
The pulmonary macrophagic system is critical to the defense of the lung, keeping the alveoli clean and sterile and responding on demand with an adaptive outpouring of new cells into the air sacs. Under basal conditions alveolar macrophages, in common with other mononuclear phagocytes, are derived from the bone marrow. A population of macrophage precursors within the pulmonary interstitium provides a reserve pool capable of proliferation and delivery of phagocytes in response to unusually heavy loads of inhaled particles. This reserve system also produces macrophages when monocytic precursors in the bone marrow are depleted by diseases such as leukemia. The alveolar macrophage is destined to ingest particulate matter and to be eliminated along the mucociliary pathway; clearance by lymphatics is of minor importance and macrophages probably do not recross the alveolar epithelium to reach the pulmonary interstitial compartment. Although the protective role of the macrophage is dominant, this cell may participate, directly or indirectly, in the genesis of two major groups of chronic pulmonary disease, interstitial fibrosis and emphysema. Such inappropriate responses involve interactions with fibroblastic cells and tissue injury initiated by proteases secreted by the macrophage.
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Unraveling pulmonary fibrosis: the bleomycin model.
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Animals↗
Role of polymorphonuclear leukocytes in silica-induced pulmonary fibrosis.
Silicosis is usually attributed to fibroblast stimulation by secretion of damaged alveolar macrophages (AMs), but the role of polymorphonuclear leukocytes (PMNs) and of continuing cell injury in the pathogenesis has not been fully studied. Mice given intratracheal injections of 2 mg of silica received 3H-thymidine 1 hour before death at intervals to 20 weeks. Cellular populations and lysosomal content of lavage fluids were correlated with morphology, DNA synthesis, and collagen content of the lung. The initial response involved rapid PMN and AM recruitment to the alveoli. Some free particles crossed Type 1 epithelial cells, and silica was found in interstitial macrophages. Focal Type 1 cell damage was rapidly repaired by Type 2 cell proliferation. Although PMN numbers dropped after a few days, they never reached control levels and rose again after 8 weeks; the number of AMs fell to control values from 2 to 8 weeks, then increased again. Glucosaminidase and glucuronidase levels in the lavage fluid were much higher than control levels throughout the study. Increased DNA synthesis by interstitial cells occurred from 2 days to 20 weeks; increased collagen synthesis was found from 4 weeks onward. The continuing inflammatory response of the lung to silica suggests may contribute to fibroblastic stimulation.
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Response of pulmonary macrophages to unilateral instillation of carbon.
Production and delivery of macrophages to the alveoli are dependent upon chemotactic and mitogenic stimuli. To determine whether these mechanisms occur uniformly throughout the lungs or are localized to the areas of particulate deposition, the authors instilled 4 mg of carbon in one lung of a rat, the other lung serving as a control. Following split-lung lavage, little change was seen in the number of cells lavaged from nontreated lungs, whereas the yield of macrophages from the black lungs rose from 32 X 10(4) to 400 X 10(4) after 2 days; control values were not attained until 3 weeks. Labeling indices for cell nuclei in the white lungs were slightly elevated, whereas in the black lungs the value rose from 0.4 to 1.0-1.5% for 3 weeks before falling to normal. The increased labeling was limited to interstitial cells. The results provide further support for the dual origin of alveolar macrophages and indicate the importance of local generation of chemotactic and mitogenic factors in stimulating the cellular response and in directing these cells to the sites of maximal deposition of particles.
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Cell turnover in the lung.
Precise information on cell turnover in the lung is surprisingly limited and it is derived for the most part from the study of small rodents. The determination of cell cycle time is dependent upon the identification of a pure population of cells. This requirement poses a particular problem in the analysis of the mixed epithelial components of the tracheobronchial tree and with the miscellany of cells that populate the pulmonary interstitium. Cell turnover in general is related to the need for replacement of aging or injured cells. This explains the rapid epithelial turnover in the major airways when compared with the much slower replacement of cells in the more stable milieu of the alveoli. Bronchiolar cytokinetics poses a special problem in rodents, which, unlike humans, have no respiratory bronchioles. This particular problem demands continuing study with special emphasis on the kinetics of the several cell types that populate the intermediate channels linking the airways with the air sacs.
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Endothelial injury and repair in radiation-induced pulmonary fibrosis.
Cytokinetic relationships between endothelial cells and fibroblasts during lung injury and repair in mice have been studied in a morphologic, autoradiographic, and biochemical study following whole body irradiation. After 650 rads, endothelial injury accompanied by interstitial edema was seen between weeks 1 and 2. The cell labeling curve had two components: predominant endothelial labeling to 3 weeks, then a smaller rise in DNA synthesis in interstitial cells. There was focal fibrosis but little change in total hydroxyproline to 20 weeks. After 1000 rads, cell injury, still confined to the endothelium, was more severe and lasted up to 6 weeks. Increased DNA synthesis occurred in the endothelium between Weeks 2 and 8 and in interstitial cells from Week 3 to 16, when total hydroxyproline was significantly elevated and many fibrotic areas were seen in the lung. The results indicate that acute endothelial injury may be rapidly repaired with little fibroblastic stimulation, whereas severe or prolonged injury with delayed regeneration disturbs endothelial-mesenchymal relationships. This may be a key factor in promoting fibroblast proliferation and the deposition of collagen.
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Alveolar macrophage response to carbon in monocyte-depleted mice.
The predominant source of alveolar macrophages is monocytic migration with a smaller proportion arising from dividing interstitial cells. To determine how this system responds to particulate loading when depleted of monocytes, mice received 650 rad whole body irradiation followed by 4 mg carbon instilled into the trachea. Control groups included irradiated mice given no carbon and nonirradiated mice given carbon. After irradiation alone, the number of monocytes fell to a very low level for at least 3 wk, whereas the output of alveolar macrophages remained normal. In nonirradiated mice, carbon induced a tenfold increase in macrophages the first day, dropping rapidly after 1 wk. Macrophage output in irradiated animals given carbon doubled the first day, then rose to 4 times normal after 2 wk. This limited response to carbon was accompanied by increased DNA synthesis in pulmonary interstitial cells, but the number of circulating monocytes and labeling of alveolar macrophages remained very low. The results indicate that when monocytes are not available, output of alveolar macrophages is maintained by precursor cells in the pulmonary interstitium. This interstitial compartment provides an essential back-up mechanism capable of responding to increased particulate loading in both normal and monocytopenic animals.
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Effects of irradiation on macrophagic response and transport of particles across the alveolar epithelium.
Macrophagic production and particulate penetration into the lung were studied in mice depleted of monocytes by whole body irradiation. Subsequent intratracheal instillation of 4 mg carbon resulted in a much smaller adaptive increase in the number of alveolar macrophages, as compared with nonirradiated animals. The decreased macrophagic output was associated with increased passage of free carbon across the Type 1 alveolar epithelium in increased accumulation of carbon in interstitial macrophages and hilar lymph nodes of irradiated mice. The results suggest that interference with the adaptive outpouring of alveolar macrophages following a particulate load facilitates the interaction of particles with macrophages within the pulmonary interstitium, where fibrogenic factors released in response to toxic agents would have a maximum effect.
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Chemotactic and mitogenic components of the alveolar macrophage response to particles ad neutrophil chemoattractant.
The pulmonary response to instilled particulates involves initial efflux of polymorphonuclear leukocytes (PMNs) and increased production of alveolar macrophages (AMs). The relationships of the cell migration and division components of the AM production to the initial PMN response after intrapulmonary carbon or latex administration are examined. Supernatants of lung lavages taken during early PMN migration to the alveoli promote sequential migration of PMNs, migration of monocytes, and division of pulmonary interstitial cells in normal mice. Supernatants taken during the phase of increased cell division in the lung produce no such effects, implying tht factors responsible for chemotaxis and mitosis are generated rapidly and are short-lived. A possible source is the interaction of AMs with particles, since supernatants of such incubations induce an inflammatory response in vivo. Similarly, when a synthetic chemoattractant is used, efflux of PMNs is followed by macrophages arising from migration of monocytes and from division of interstitial cells. The results suggest that particulate instillation in the lung stimulates a standard inflammatory response in which rapid generation of a factor(s) chemotactic for PMNs also attracts mononuclear cells to the alveoli. The initial efflux of cells may be explained by migration from the blood, but continued demand or replacement requires mitotic activity of precursors. For the alveolar macrophages, this includes division of cells in the pulmonary interstitium.
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Alveolar response to injury.
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Cell Communication↗
Dose response of the pulmonary macrophagic system to various particulates and its relationship to transepithelial passage of free particles.
Alveolar macrophages are thought to arise from both marrow-derived monocytes and pulmonary interstitial cells. Macrophage kinetics are now studied under various conditions of alveolar loading using several doses of carbon (0.03 micrometers diameter), polystyrene latex (0.1 and 1.0 micrometers), and heat-killed bacteria. In serial studies we examined the number of macrophages recovered by lavage, DNA synthesis by lung cells on autoradiographs, and the passage of free particles into lung tissues by electron microscopy. The dual origin of the alveolar macrophage was confirmed for each particulate. The observed peak in macrophagic output at 1 day may be explained by monocytic egress. With greater loads, the peak value did not increase, but the continuing macrophagic production correlated with a period of interstitial cell proliferation. For all particles used, the number of new macrophages was related more closely to number of particles instilled than to the total dose by weight delivered to the lungs. With increasing number, more free particles of carbon and latex crossed the Type 1 epithelium to be phagocytized by interstitial macrophages. The results suggest that the adaptive outpouring of alveolar macrophages occurs by an acceleration of the normal biphasic pathway; when the adaptive response is prolonged, the interstitial compartment appears to be the predominant source of new cells.
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Uptake of 3H prednisolone by fetal lung explants: role of intercellular contacts in epithelial maturation.
Epithelial maturation in the developing lung is accelerated by glucocorticoids and it has been suggested that a fibroblast-derived factor may be important in regulating this response. In the present study, uptake of 3H prednisolone by cultured explants of fetal rat lung is used to correlate cellular location of the steroid with changes in epithelial--mesenchymal cell relationships at different stages of development. At days 17 and 18 of gestation, there is substantial incorporation of steroid by epithelial and interstitial cells with no morphologic evidence of accelerated maturation. At days 19 and 20, uptake of prednisolone is maximal with preferential incorporation by Type 2 cells which now contain an increased number of lamellar bodies. In the bronchioles, steroid uptake is observed in Clara cells but not in ciliated cells. Lung development results in a progressively closer anatomic relationship between alveolar epithelial and interstitial cells and at the time of maximal steroid response, day 19-20, narrow "gap junctions" are observed between these cell types. The results suggest that, although steroid is bound to several cell types from an early stage of development, the effects on epithelial maturation occur relatively late in gestation when close epithelial-mesenchymal cell contacts are made.
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Role of monocytes and interstitial cells in the generation of alveolar macrophages I. Kinetic studies of normal mice.
Following monocytic depletion, alveolar macrophages are produced by a local cell renewal system within the pulmonary interstitium. The contributions of hematopoietic and interstitial components to the output of macrophages in the normal state is not known. In the present autoradiographic study, sequential labeling and grain counts in monocytes, interstitial cells, and free alveolar macrophages are examined in mice up to 7 weeks after three injections of 3H-thymidine; the animals also received colchicine 4 hours before sacrifice. The labeling patterns of interstitial cells and macrophages are biphasic. The initial peaks with very few mitotic figures are consistent with migration of blood monocytes across the interstitium to the alveoli. In the second phase, when no labeled monocytes are present, a few heavily labeled macrophages are observed up to 7 weeks; in this period mitoses and heavy labeling are seen in a few interstitial mononuclear cells. It appears that, under normal circumstances, the predominant mechanism of macrophage production is direct passage of monocytes across the interstitium to the alveoli. This may be rapid at the thin walled capillaries and take up to 2 days where the interstitium is thicker. A smaller proportion of cells appears to arise by division of resident interstitial cells with subsequent migration to the alveoli. The sojourn of free macrophages in the alveolus under basal conditions is 7 days.
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