Adherent Alveolar Macrophages Research Articles

Inhibition of Human Alveolar Macrophage Production of Leukotriene B4 by Acute In Vitro and In Vivo Exposure to Tobacco Smoke

The number of neutrophils in the lungs of cigarette smokers is increased. This could be a consequence of the chemotactic mediator synthesis by alveolar macrophages (AM). In order to evaluate the possible role of leukotriene B4 (LTB4) in this condition, we studied the formation of LTB4 by nonsmokers' AM exposed in vitro and in vivo to cigarette smoke and by smokers' AM. In the absence of stimulus or upon stimulation, nonsmokers' AM exposed in vitro to tobacco smoke formed less LTB4 than did nonexposed AM, e.g., the cells incubated with arachidonic acid and ionophore produced, respectively, 241 +/- 132 and 425 +/- 106 pmol LTB4/10(6) cells (mean +/- SEM) (P less than 0.01). In other experiments, smokers' AM were incubated in absence of stimulus and produced less LTB4 than did nonsmokers' AM; during a 3-h incubation, smokers' and nonsmokers' adherent AM released, respectively, 3 +/- 2 and 40 +/- 28 pmol LTB4/10(6) cells (P less than 0.05). Similarly stimulated smokers' AM produced less LTB4 than did nonsmokers' AM, e.g., the cells incubated with arachidonic acid and ionophore formed, respectively, 225 +/- 41 and 573 +/- 150 pmol LTB4/10(6) cells (P less than 0.05). In a group including mild smokers and nonsmokers, in vivo exposure to the smoke of 4 cigarettes produced a decrease in the release of LTB4 by AM, e.g., in the presence of arachidonic acid and ionophore, nonexposed and exposed AM produced, respectively, 198 +/- 38 and 143 +/- 38 pmol LTB4/10(6) cells (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Eicosanoids produced during interactions between Pseudomonas aeruginosa and alveolar macrophages are species‐dependent

Eicosanoid production during phagocytosis of pyogenic bacteria by rabbit alveolar macrophages was studied as a model of early events in the pathogenesis of pneumonia. Adherent alveolar macrophages, prelabelled with [3H]-arachidonic acid (AA), were incubated with live, opsonized Staphylococcus aureus or Pseudomonas aeruginosa (bacteria:macrophage ratio of 50:1) at 37 degrees C for 90 min. Supernatant eicosanoids were extracted and separated by reverse phase high performance liquid chromatography (RP-HPLC). While the amounts of labelled PGE2, TXB2, and PGD2 produced in response to the two organisms were equal, the amount of PGF2 alpha elicited by S. aureus amounted to three times that released during macrophage challenge with P. aeruginosa. Overall, preferential release of cyclooxygenase products occurred during phagocytosis of S. aureus. In contrast, eicosanoids identified presumptively as oxygenated metabolites of AA predominated in cultures challenged with opsonized P. aeruginosa. Live, non-opsonized P. aeruginosa elicited the same profile of eicosanoids, but in reduced amounts. Inhibitor studies indicated that these AA derivatives were not synthesized via the macrophage lipoxygenase pathway. Their production was dependent on the viability of P. aeruginosa. Macrophages challenged with opsonized, heat-killed P. aeruginosa resulted in production of an eicosanoid profile similar to that elicited by S. aureus. Secondary metabolism by P. aeruginosa of eicosanoids released from the macrophage did not contribute to the unique profile produced during the interaction of this organism with labelled macrophages. Our data indicate that during binding to macrophages, the primary human pathogen, P. aeruginosa, specifically modulates the profile of eicosanoids produced. This effect on inflammatory mediators may be of biological significance in the pathogenesis of pneumonia.

Different patterns of arachidonate metabolism in autologous human blood monocytes and alveolar macrophages.

As peripheral blood monocytes (PBM) differentiate into tissue macrophages, they undergo a variety of functional changes. One such difference which has been described is an enhanced metabolism of arachidonic acid (AA) via the 5-lipoxygenase (5-LO) pathway in alveolar macrophages (AM) as compared to PBM. In order to elucidate a possible mechanism for this difference, we compared the metabolism of endogenously released AA mobilized by agonists and of exogenously supplied fatty acid in adherent autologous PBM and AM obtained from six normal subjects. Exogenous AA was metabolized to larger amounts of both cyclooxygenase (CO) and 5-LO products by PBM as compared with AM. Although the two cell types released similar amounts of endogenous AA in response to ionophore A23187, marked differences in the pattern of its metabolism were observed. In PBM, a large proportion of released AA remained unmetabolized, and that which was metabolized was converted predominantly to CO products. In contrast, arachidonate released by AM was efficiently metabolized, predominantly via the 5-LO pathway. Similar results were obtained when cells were stimulated with the particulate zymosan, with PBM synthesizing mainly CO and AM, mainly 5-LO eicosanoids. In addition, culture of PBM for up to 5 days in an aerobic environment did not alter their response to A23187 stimulation. These results suggest that the lesser 5-LO metabolism by PBM than AM is not explained by lesser phospholipase or 5-LO activities, but rather a compartmentalization of the endogenous AA deacylated by phospholipase and the 5-LO enzyme in the PBM. The acquisition of the capacity to metabolize endogenous AA to large quantities of 5-LO products as mononuclear phagocytes differentiate in the lung may equip them with the ability to mount an inflammatory response in the alveolar space.

Reduced responsiveness of bronchoalveolar cells from sensitised guinea-pigs to the cyclic AMP—Stimulating effect of prostaglandin E 2 and β-adrenoceptor agonists

Prostaglandin E2 and the beta-adrenoceptor agonist salbutamol were less effective in increasing the intracellular cAMP content of bronchoalveolar cells and adherent alveolar macrophages from ovalbumin-sensitised as compared to control guinea-pigs. This refractoriness to the cyclic AMP-stimulating effects of PGE2 and salbutamol induced by the sensitisation procedure may be relevant to the altered bronchopulmonary responsiveness in asthma.

Immobilization of alveolar macrophages for measurement of in vitro dissolution of aerosol particles.

The dissolution of aerosol 57CO3O4 particles was investigated by exposing baboon alveolar macrophages (AM) to the particles and immobilizing the AM in alginate macrobeads crosslinked with Ca2+ ions. AM were obtained from bronchopulmonary lavage and, after immobilization, were stored in culture medium enriched with Ca2+. Dissolution of 57Co3O4 particles by these AM was measured for up to 16 days by gamma counting of filtered media containing beads. For these particles, the daily dissolution rate was 0.242 per cent versus 0.072, 0.089 and 0.091 per cent for beads containing particles only, particles combined with AM enzymes, and non-phagocytic cells, respectively. The rate obtained with immobilized AM is very close to the rates previously found for adherent AM in culture. Moreover, AM that had phagocytosed PuO2 particles before immobilization by the same method gave a mean daily dissolution rate close to that obtained in vivo with baboons. We also demonstrated, by dissolving the alginate network, that immobilization did not affect AM viability. The present method of immobilization seems very promising compared to conventional methods because it facilitates manipulation, allows the use of cells at a high density in a small volume, and gives results for particles with a very low dissolution rate.

Chemiluminescent responses of macrophages exposed to biomaterials: a biocompatibility screening test.

Chemiluminescence (CL) response of alveolar macrophages (AM) exposed to either phagocytosable or nonphagocytosable particles of polymethylmethacrylate (PMMA) or titanium in vitro was monitored. The results were correlated to known in vivo bone tissue compatibility of the materials. Moreover, the quantity and morphology of adherent AM on titanium and PMMA disks were assessed using scanning electron microscopy (SEM). Our results with PMMA indicated that a high CL response to phagocytosable as well as nonphagocytosable particles of the biomaterial in question, correlated with poor bone tissue compatibility. By contrast, a low CL response to phagocytosable particles and no response to nonphagocytosable particles correlated with a satisfactory bone tissue compatibility, as in the case of titanium. SEM showed 48% more AM adherent to PMMA than titanium per unit area. Also, the percentage of cells with membrane damage was three times higher on PMMA than on titanium. It appears that the CL response of AM exposed to biomaterials in vitro thus can be used as an easy to do screening test for potential orthopaedic implant materials before in vivo tests are considered.

Hydrogen peroxide release by alveolar macrophages from sarcoid patients and by alveolar macrophages from normals after exposure to recombinant interferons alpha A, beta, and gamma and 1,25-dihydroxyvitamin D3.

We measured H2O2 release by human alveolar macrophages (AM) from normals and sarcoid patients in suspension immediately after bronchoalveolar lavage in the presence and absence of the triggering agent, phorbol myristate acetate (PMA). AM from 11 sarcoid patients produced a mean (+/- SE) of 21.7 +/- 2.3 and 5.9 +/- 3.4 nmol H2O2/10(6) macrophages in the presence and absence of PMA, respectively. By contrast, AM from normals (n = 6) produced 9.8 +/- 1.7 and 1.6 +/- 0.7 nmol H2O2/10(6) macrophages with and without PMA, respectively. Macrophage activation, as monitored by H2O2 production, did not correlate with the angiotensin-converting enzyme levels, the result of gallium-67 scans, or the percent of lymphocytes in the bronchoalveolar lavage. To determine whether AM from normals could be stimulated to increase their H2O2 production to the level seen in patients with sarcoid, we measured H2O2 released by adherent AM after incubation in each of four potential activating agents: recombinant interferons alpha A, beta, gamma (rIFN alpha A, rIFN beta, and rIFN gamma, respectively), and 1,25-dihydroxyvitamin D3. H2O2 release in the range seen in sarcoid patients could be induced in PMA-triggered AM from normals by rIFN gamma in a time- (t1/2 approximately 1 d) and dose-dependent fashion (threefold increase, EC50 5 antiviral U/ml) and by rIFN alpha A and rIFN beta at higher concentrations, but not by 1,25-dihydroxyvitamin D3.

Enhanced interleukin 1 production by alveolar macrophages and increase in Ia-positive lung cells in silica-exposed rats.

Inhalation exposure to silica dust enhanced interleukin 1 (IL-1) production by alveolar macrophages (AM), which is attributable to an increase in Ia-positive lung cells. While the proportion of Ia-positive cells in lavaged bronchoalveolar cells (BAC) was much lower (0-3%) in unexposed control rats, about a third of the rats that inhaled silica showed higher proportions (8.0-18.5%); these were designated "Ia-high" exposed animals. The number of total cells, Ia-positive cells and lymphocytes in BAC was significantly increased (P less than 0.05, P less than 0.001, and P less than 0.001, respectively) in these "Ia-high" exposed animals, compared to the control animals. Adherent AM populations obtained from BAC preparations also contained significantly higher (P less than 0.001) proportions of Ia-positive cells in the "Ia-high" exposed animals. When these adherent AM cultures were stimulated with lipopolysaccharide, IL-1 activity of the culture supernatants was enhanced and was significantly higher (P less than 0.001) in the "Ia-high" exposed rats, compared to the control animals. These results indicate that silica-exposure can induce populational changes in lung cells and also activation of AM associated with the increase in Ia-positive cells.

Quantitative evaluation of opsonin-independent phagocytosis by alveolar macrophages in monolayer using polystyrene microspheres

Macrophages can bind and engulf a variety of particles in the absence of specific opsonins. Polystyrene-type microspheres are often employed to quantitative opsonin-independent phagocytic activities of macrophages in vitro. Reliable measurement of this cell function, however, requires the ability of the investigator to distinguish between particles that are merely attached to the cell surface and those that are actually internalized. We have developed a simple, rapid, and reproducible method for quantitating phagocytosis using polystyrene microspheres and adherent alveolar macrophages. Basically, particles associated with macrophages after a given incubation time are microscopically quantitated on a cell-by-cell basis before and after toluene dissolution of external particles. Particle/macrophage values obtained after toluene treatment exclusively index phagocytosis.

Human alveolar macrophage inhibition of lung fibroblast growth. A prostaglandin-dependent process.

The processes controlling fibrosis in the human lung are poorly understood. Mononuclear cells may be important regulators of fibroblast function, and the alveolar macrophage (AM) is the major mononuclear phagocyte of the human lung. To further understand AM-fibroblast interactions in the lung, supernatants from unstimulated and stimulated adherent human AM were obtained and their effect on the log phase growth of human lung fibroblasts was characterized. Alveolar macrophage supernatants inhibited fibroblast growth in a dose-dependent fashion. This inhibition was the result of a heat-stable (56 degrees C, 1 h), nondialyzable (molecular weight cutoff, 12,000 daltons), soluble factor(s), which was preferentially elaborated by smaller and denser macrophage subpopulations. The inhibitory capacity of an AM supernatant was directly related to its capacity to stimulate fibroblast prostaglandin production. Blocking fibroblast prostaglandin production reversed the inhibition of fibroblast growth caused by AM supernatants. Thus, AM inhibition of fibroblast growth may be mediated in part by fibroblast prostaglandin production. Alveolar macrophage inhibition of fibroblast growth may be important in preventing fibrosis in normal persons and/or in inhibiting the fibrotic response seen with pulmonary inflammation.