Geraldine M. Brown

Inflammation generating potential of long and short fibre amosite asbestos samples.

Previous studies have shown that long thin asbestos fibres are more pathogenic in in vivo and more active in in vitro assays than short fibre samples. In the present study a long fibre amosite asbestos sample and a short fibre sample prepared from it were tested for ability to cause inflammation in the peritoneal cavity of the mouse; a UICC sample intermediate in fibre size and an inert compact dust, TiO2, were also tested. The ability of the dust samples to cause inflammation, as judged by macrophage and neutrophil recruitment, was ranked in the order long fibre greater than UICC greater than short fibre greater than TiO2. Ability of amosite samples to cause inflammation was therefore related to the proportion of long fibres. The enhanced ability of long fibres to cause inflammation and cause macrophage activation is probably a key factor in the ability of long fibres to cause pulmonary fibrosis and may also be important in fibre carcinogenesis.

Kinetics of the bronchoalveolar leucocyte response in rats during exposure to equal airborne mass concentrations of quartz, chrysotile asbestos, or titanium dioxide.

The kinetics of the bronchoalveolar response was assessed in rats exposed, at equal airborne mass concentration (10 mg/m3), to titanium dioxide--a non-pathogenic dust--and the two pathogenic mineral dusts quartz and chrysotile asbestos. Rats were killed at intervals over a 75 day exposure period and groups of rats exposed for 32 and 75 days after recovery for two months. Bronchoalveolar lavage was carried out and the lavage fluid characterised for cellular content, macrophage activation, and concentrations of free total protein, lactate dehydrogenase, and N-acetyl-beta-D-glucosaminidase. Inhalation exposure to the two pathogenic dusts resulted in an increased number of leucocytes, macrophage activation, and increased levels of free enzymes and total protein. The pattern and magnitude of the responses to quartz and chrysotile differed. Chrysotile caused less inflammation than quartz, and the main cellular response peaked around the middle of the period of dust exposure whereas the highest levels of enzymes occurred towards the end. The difference in timing suggests that macrophages were not available for lavage towards the end of the exposure, owing to their playing a part possibly in deposition of granulation tissue. Quartz caused a greater cellular and enzyme response than chrysotile, particularly towards the end of the dust exposure phase. There was a noticeable progression of inflammation in the quartz exposed groups left to recover for two months, but not in the chrysotile recovery groups.

Bronchoalveolar leukocyte response in experimental silicosis: Modulation by a soluble aluminum compound

The biological properties of quartz have been related to its surface reactivity. We have addressed the role of particle surface reactivity in mediating the biological activity of quartz in mixed dusts, by treating the quartz with aluminum lactate. Intratracheal instillation of untreated quartz in rats caused a rapid, sustained alveolitis and bronchoalveolar leukocytes obtained from these animals had enhanced activity in degrading fibronectin, but reduced ability to mount a respiratory burst. Quartz pretreated with aluminum elicited a markedly reduced inflammatory response; the reduced activity of the treated quartz was also reflected in the attenuated change in the key functional parameters, oxidant production and proteolysis of fibronectin. Late intratracheal dosing with aluminum after the quartz-induced alveolitis was well established reduced the inflammatory response and abrogated the effect of quartz on the respiratory burst, but did not alter fibronectin degradation by the leukocytes. Aluminum did not affect the inflammatory response to Corynebacterium parvum and thus the effect was on the quartz particles and not on the inflammatory leukocytes. These findings have implications for the likely pulmonary responses to mixed dusts containing quartz and aluminum silicate clays.

New perspectives on basic mechanisms in lung disease. 5. Respirable industrial fibres: mechanisms of pathogenicity.

Background Fibrous materials have much to commend them in industrial applications, where they can offer, in varying degree, reinforcement, thermal and electrical insulation, flexibility, and strength. In the first part of this century the fibrous material that was available, cheaply and in bulk, that could provide these properties to industry was the asbestos group of silicate minerals (table 1). The fibrous, crystalline, asbestiform habit, however, also means that during mining and industrial use fibres are released into the air. Some of these are in the respirable size range and may be deposited in the bronchiolar-alveolar region. Subsequently we have come to realise that asbestos inhalation is associated with several different lung diseases-namely, asbestosis (interstitial fibrosis), carcinoma of the lung, mesothelioma, and small airways diseaseand the association is now well documented. This information on the harmful effects of asbestos has led to bans in some countries and considerable litigation and claims for damages throughout the developed world. Regulatory and legal pressures on asbestos have contributed to an increase in the use of alternative materials, both natural and manmade, including some fibrous materials. More importantly, advances in materials science have identified ar, ever widening range of applications in which the fibrous nature of a material offers positive advantages. Thus in addition to their traditional use in thermal insulation new fibre composites have found new uses, such as metal reinforcement, to which asbestos was never suited (table 2). The finding that in at least some biological assays some non-asbestos fibres could cause the same types of effect as asbestos has raised the question of whether all fibres have the same pathogenic potential as asbestos. A major approach in present research on fibres is therefore to compare any fibre under study with asbestos and to gauge the potential for

Degradation of connective tissue components by lung derived leucocytes in vitro: role of proteases and oxidants

Inflammatory leucocytes are implicated in connective tissue damage during chronic inflammatory lung disease. In an investigation of the role of leucocytes in connective tissue derangements in the lung, inflammatory leucocytes were generated in rat lungs by intratracheal instillation of inflammatory agents and retrieved by bronchoalveolar lavage. The proteolytic activities of control macrophages and of two inflammatory cell populations were compared; iodinated collagen, laminin, and fibronectin matrices were used. The inflammatory cells caused consistently and substantially more degradation of the matrices than the controls on a per cell basis. The oxidant scavengers superoxide dismutase and catalase did not inhibit matrix degradation, but alpha 1 protease inhibitor and alpha 2 macroglobulin were inhibitory. It is concluded that matrix damage in this assay is enhanced by inflammatory cells and is mediated principally by serine protease activity.

Inflammatory responses in lungs of rats inhaling coalmine dust: enhanced proteolysis of fibronectin by bronchoalveolar leukocytes.

Chronic exposure to coalmine dust is associated with the accumulation of inflammatory leukocytes in the bronchoalveolar region of the lung and, in the long term, with fibrosis and emphysema of the lung parenchyma. Degradation of connective tissue by inflammatory leukocytes has been implicated in the parenchymal damage that precedes the development of fibrotic or emphysematous lesions in the lung. The ability of inflammatory leukocytes obtained by bronchoalveolar lavage from rats inhaling coalmine dust to degrade fibronectin in vitro was assessed. The animals were exposed to an airborne mass concentration of dust similar to the maximum permissible level in United Kingdom collieries. The bronchoalveolar lavage cell population showed changes with duration of dust exposure; there were increases in the total number of leukocytes and in the percentage of polymorphonuclear leukocytes, and the macrophage component of the lavage became increasingly activated, as assessed by the ability of these cells to spread on glass. In addition, degradation of a radiolabelled fibronectin matrix by the coalmine dust exposed bronchoalveolar leukocytes increased with duration of dust exposure. Thus exposure to airborne coalmine dust causes an influx of inflammatory leukocytes to the alveolar region. These cells have enhanced ability to degrade fibronectin and this may be important in subsequent disease development.

Activated human peripheral blood neutrophils produce epithelial injury and fibronectin breakdown in vitro

The ability of peripheral blood neutrophils to injure elements of the alveolar septum was assessed. Experiments consisted of activating previously isolated neutrophils (PMNs) with a soluble (phorbol myristate acetate, PMA) and a particulate (zymosan) trigger, and measuring detachment of51Cr-labeled epithelial cells. In addition, the mechanisms of epithelial injury were investigated by including anti-proteinase and antioxidants in the system. Untriggered PMNs produced only slight detachment injury to epithelial cells at two effector-target cell ratios, this effect being dramatically increased after triggering with PMA; zymosan caused triggering but less than was produced by PMA. Alpha-1-protease inhibitor produced a decrease in detachment when both PMA- and zymosan-triggered cells were the effectors; superoxide dismutase did not significantly reduce detachment. The ability of triggered PMNs to cause proteolysis of fibronectin, as measured in a radiolabeled fibronectin matrix degradation assay, was related to their triggerability in the detachment assay, zymosan being largely ineffective in triggering enhanced proteoiysis. These findings suggest that fibronectin is important in maintaining the integrity of the alveolar epithelial surfaces. Furthermore, in inflamed alveoli, activated PMNs can release proteinases, which cause degradation of this matrix component leading to compromise of the alveolar epithelial barrier.

Inflammation in the lungs of rats after deposition of dust collected from the air of wool mills: the role of epithelial injury and complement activation

In a previous study assessing respiratory symptoms in individuals employed in wool textile mills in the north of England relations between symptoms of chronic bronchitis, breathlessness and wheeze, and rhinitis and current exposure to airborne mass concentration of dust were shown. As preliminary steps in defining the potential hazard associated with dust from the air of wool mills the ability of inspirable dust, collected from the air of wool textile mills, to cause inflammation when injected into the lungs of rats was determined. Dusts were collected from the beginning of wool processing (opening) in one factory and from the middle (combing) and late (backwinding) stages of the process in two other factories. Ability of the dusts to cause inflammation was assessed by instillation into the lungs of rats followed by bronchoalveolar lavage. All the dusts caused some inflammation which peaked on day 1 and did not persist beyond one week. A distinctive aggregation response of mononuclear cells in the lavage, however, had a different time course, peaking at day 7. An attempt was made to determine how the wool mill dusts caused inflammation and experiments showed that the dusts themselves had no inherent chemotactic activity but that they did have a pronounced ability to generate chemotaxins in serum and so could activate complement in lung fluid. In addition, dust collected from ledges in the mills had the ability to injure epithelial cells in vitro which could also contribute to inflammation. A role for endotoxin in the inflammatory activity of the dusts was not discounted and a leachate of the dust had the ability to cause inflammation when injected into the lungs of rats. Wool mill dust is likely to be a complex mixture of materials and these experiments represent a preliminary approach to understanding the biological activity of the whole unfractionated dust and further studies are in progress to define more accurately the toxic material(s) in the dust.

Persistent biological reactivity of quartz in the lung: raised protease burden compared with a non-pathogenic mineral dust and microbial particles.

This study assessed the potential harmfulness of particles in the lung by measuring their ability to elicit and maintain an inflammatory response and to damage lung tissue. It compared the inflammogenicity of two nondurable, biological particulates (Corynebacterium parvum and zymosan) with a pathogenic mineral dust (quartz) and a nonpathogenic dust (titanium dioxide) by dosing rats via the intratracheal route and measuring the consequent alveolitis. The magnitude and duration of the inflammatory response were assessed by measuring the total number of leucocytes and the percentage of neutrophils obtained by bronchoalveolar lavage. Two key functional parameters of the lavaged leucocytes--ability to degrade fibronectin and production of plasminogen activator--were also measured. A marked inflammatory response had occurred by one day after instillation, characterised by increases in total leucocyte numbers and percentage of neutrophils in the bronchoalveolar lavages, with all four test materials. In all but the quartz exposed animals, the inflammation subsided rapidly thereafter, approaching control levels by 15 days after injection; in the quartz exposed animals the alveolitis persisted for up to 30 days. All of the inflammogens generated chemotaxins in rat serum in vitro and so, by analogy, might also be expected to generate chemotactic activity in alveolar lining fluid which could contribute to the generation of an inflammatory response. The cellular inflammatory response was accompanied by a concomitant increase in the proteolytic activity of the bronchoalveolar lavage leucocytes but production of plasminogen activator remained unchanged. In vitro exposure to the inflammogens had no effect on the proteolytic activity against fibronectin or on the plasminogen activator activity of bronchoalveolar leucocytes.

Assessment of mineral dust cytotoxicity toward rat alveolar macrophages using a 51Cr release assay

An assay was developed to assess the cytotoxicity of mineral dust by measuring release of 51Cr from prelabeled rat alveolar macrophages. Optimal conditions for the assay are described, the most notable being use of 2% albumin instead of fetal calf serum. The assay demonstrated loss of label into the supernatant when prelabeled macrophages were cultured with the two pathogenic mineral dusts, quartz and chrysotile asbestos. In contrast the inert mineral dust titanium dioxide had very little effect on 51Cr release by rat alveolar macrophages.