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Title: Studies on the alveolar macrophage
Author: Diaz Amor, Patricia
ISNI:       0000 0001 3423 5850
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1980
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This thesis is concerned with (1) the identification of membrane receptors on alveolar macrophages, particularly those for histamine, and (2) some of the metabolic changes which the cell undergoes following incubation with histamine, either in free solution or when bound to particles. In preliminary experiments, in which some of the variables associated with the "rosette technique" were examined, it was shown that alveolar macrophages bind IgG-coated erythrocytes (EAg) as well as sheep cells sensitized with IgM and whole serum as a source of complement (EAm"C3"). Therefore, lung macrophages, like most other leucocytes, appear to bear "immunological receptors" for IgG and complement (presumably C3). The "rosette technique" was then employed for the identification of histamine receptors on various guinea pig leucocytes using histamine coupled as a rabbit serum albumin conjugate (H-RSA) to formalised ox red cells. It was found that the number of "histamine rosettes" varied from 60-81% for alveolar macrophages, 14-73% for peritoneal macrophages, 14-30% for blood monocytes, 27-48% for lymph node cells, 7-24% for blood lymphocytes and 0-29% for peritoneal and blood neutrophils. Virtually no histamine rosettes were formed with eosinophils or basophils. The percentage of rosette-forming target cells was directly related to the concentration of erythrocyte-bound H-RSA. Free histamine partially inhibited rosette formation by alveolar macrophages in a dose-dependent fashion from 10⁻³ to 10⁻⁵ moles.1⁻¹ , and complete inhibition was achieved by the H-RSA conjugate. In contrast, amines closely related to histamine such as L-histidine and the major histamine catabolites, imidazoleacetic acid, 1,4-methylhistamine, l-methyl-4- imidazoleacetic acid and N-acetylhistamine, had no inhibitory effect. The histamine Hl-receptor antagonists, mepyramine and chlorpheniramine, and the Hl-receptor agonist, 2-(2-aminoethyl) thiazole, inhibited rosette formation by alveolar macrophages in a dose-dependent fashion. In contrast, the H2-receptor antagonists, burimamide and metiamide, and the H2-receptor agonists, Dimaprit and 4-methylhistamine, were inactive. These- experiments suggested that (1) compared to other leucocytes, histamine receptors are particularly well expressed on the alveolar macrophage, (2) these receptors have a high degree of specificity for histamine in that other amines, closely related chemically, did not inhibit rosette formation, and (3) the binding of histamine to the alveolar macrophage membrane is HI- and not H2-receptor dependent. Experiments were then undertaken to determine whether free histamine, or histamine bound to particles, interacted with its membrane receptor to initiate the "respiratory burst" in alveolar macrophages. Superoxide anion formation (0₂⁻) and chemiluminescence, recognized features of the "respiratory burst" of phagocytic cells, were generated by alveolar macrophages following incubation with H-RSA bound to zymosan (H-RSAZ) (and also H-RSA linked to a number of other inert particles such as Sephadex and polystyrene beads). The degree of 0₂⁻ generation and light emission was comparable to that achieved with serum treated zymosan (STZ). Superoxide formation by alveolar macrophages was related to the concentration of histamine conjugate bound to the zymosan particles as well as the time of incubation. As with histamine rosettes, 0₂⁻ production generated by H-RSAZ in alveolar macrophages also appeared to be dependent on HI-, but not H2-, histamine receptors. Chemiluminescence induced by H-RSAZ was also time- and dose-dependent and totally inhibitible by superoxide dismutase indicating that the 0₂⁻ formation and the burst of light emission are closely related events in the alveolar macrophage. Histamine in free solution did not appear to promote 0₂⁻ generation or chemiluminescence in alveolar macrophages. Studies were then undertaken in an attempt to determine whether histamine influenced the release of the lysosomal enzyme, ß-glucuronidase, following incubation of alveolar macrophages with complement-coated zymosan. Histamine induced a small, but significant, increase in STZ-induced ß-glucuronidase release but had no apparent effect on the elaboration of the cytoplasmic enzyme, lactic dehydrogenase. Experiments to determine whether this was an HI- and/or H2- effect were inconclusive. H-RSAZ alone did not appear to influence ß-glucuronidase release. The general conclusion of this work is that there is a direct relationship between histamine and lung macrophages but that for the initiation of the "respiratory burst" histamine requires to be bound to particles to achieve appreciable biological effects. Since histamine is known to bind to a number of plasma proteins ("histaminopexy") these observations may be of significance both physiologically, in the regulation of oxygen-dependent killing by lung macrophages, and also in disease states where high concentrations of histamine in bronchial secretions may adversely influence the normal functions of alveolar macrophages.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available