Summaryj6-glucan receptors, with ligand specificity for yeast and fungal carbohydrate polymers, have been studied as phagocytic receptors of human monocytes . To characterize their structure, binding studies were carried out with human U937 cells and a rabbit IgG anti-Id that recognizes epitopes on monocyte O-glucan receptors . Unstimulated U937 cells specifically bound large amounts of the anti-ld, but almost none of the control anti-isotype. At saturation, the number of anti-Id molecules bound per U937 cell was 2.6 x 106 with an apparent K, of 1 .9 x 10 7 M -'.Immunoprecipitates from detergent lysates of surface-radioiodinated U937 cells contained only two membrane proteins with antigenic specificity for the anti-Id, one having a mol wt of 180 kD and the other 160 kD. Both proteins were disulfide-linked and presented, after reduction, as five polypeptides of 95, 88, 60, 27, and 20 kD. Detergent lysates of unlabeled U937 cells, purified by affinity chromatography on anti-ld-Sepharose, yielded the same two nonreduced proteins and five reduction products in slab gels stained with Coomassie blue. In Western blots probed with the anti-Id, the most immunoreactive nonreduced and reduced affinity-purified products were the 160 and 20 kD molecules, respectively. Immunoblots of two-dimensional gels showed the 180 and 160 kD proteins to express a common epitope through disulfide linkage to the 20 kD polypeptide. By immunoblot analysis, U937 cell glucan-binding proteins from detergent lysates contained two cell proteins antigenic for the anti-Id that were indistinguishable from affinity-purified molecules in size and subunit composition . Studies of affinity-purified proteins from detergent lysed human monocytes were characterized by immunoblot analysis and found to be identical to U937 cell R-glucan receptors. They consisted of two disulfide-linked proteins, with mol wt of 180 and 160 kD, and had in common a 20 kD polypeptide with the anti-Id epitope.glucan receptors (1) were first identified on human monocytes as phagocytic receptors which initiate phagocytosis of particulate activators of the human alternative complement pathway in the absence of opsonins (2) . Subsequent studies principally with zymosan and glucan particles, have shown that human alveolar macrophages (3), neutrophils (4-6), eosinophils (7), and murine macrophages (8, 9) possess phagocytic receptors ofcomparable ligand specificity for the (3-glucans commonly present in yeasts and fungi (10). Pathogens such as candida and aspergilli contain "yeast" glucan (11), cell wall components consisting ofbranched homopolymers of O-D-glucose with 1,3-consecutive and 1,6-crosslinked chains (12) and prototypic of Saccharomyces cerevisiae (10) . Particulate yeast glucan is similar in size and glucose composition to zymosan particles (13,14), but different in carbohydrate and protein content, with glucan particles being a purer form of zymosan (12,15) .The smallest functional unit ligand for human monocyte a-glucan receptors has been isolated from purified yeast gluca...
Antigen (Ag)-antibody (Ab) aggregates prepared with several different antigens are solubilized by fresh serum at 37 degrees C (complex-release activity of serum or CRA). The rate of solubilization varies in different systems and is strongly influenced by the affinity of Ab for the Ag in the immune precipitate. With a given Ag-Ab precipitate, the maximum amount of complex that can be solubilized by individual sera is independent of the initial concentration of complexes and cannot be increased by prolonged incubation. CRA occurs in the absence of C2 and C4, but not in the absence of C3 and factor B of the properdin pathway. Addition of C2 to C2-deficient serum or C4 to C4-deficient serum enhances CRA. Solubilization does not involve extensive degradation of the complexed antibody, as might be detected by acrylamide gel electrophoresis of released antibody after reduction and alkylation to separate H and L chains. Immune precipitates can also be solubilized by incubation with monovalent fragments (Fab or Fab') of antibodies against determinants of the Ab molecules in the immune precipitate. In contrast, F(ab')2 fragments decrease the solubility of the immune precipitates. In view of these findings, we propose that CRA is mediated by the binding of functionally monovalent C fragments (C3 and C4) onto Ab molecules in the precipitates.
Human monocytes possess a receptor for ingestion of particulate activators of the human alternative complement pathway that functions in the absence of plasma proteins and is distinct from the receptors for Fc-IgG and the major cleavage fragment of the third component of complement (C3b). Incubation of monolayers of monocytes with 1.1 x 106 to 2.2 x 107 glucan particles per ml initiated a phagocytic response comparable to that obtained with zymosan particles, of which (3-glucan is a constituent along with mannan. Maximal quantities of 4.93 ± 3.43 ng of leukotriene B4 (LTB4) and 0.43 ± 0.23 ng of leukotriene C4 (LTC4) (mean + SD, n = 3) were released by 106 monocytes stimulated with 1.1 x 107 glucan particles per ml. Preincubation of monocytes with 50 pg of soluble ,B-glucan per ml reduced subsequent monocyte ingestion of 5 x 106 zymosan particles per ml and 2.2 x 106 glucan particles per ml by 52% and 55%, respectively, and diminished release of LTB4 by monocytes stimulated with 2 X 108 zymosan particles per ml and 8.6 x 106 glucan particles per ml by 73% and 61%, respectively. Preincubation with 1 mg of soluble mannan per ml had little effect on monocyte phagocytosis or LTB4 generation in response to either zymosan or glucan particles, and neither soluble P-glucan nor mannan stimulated generation of LTB4 or LTC4. The effect of pretreatment of monocytes with soluble /-glucan was time dependent, with the maximal effect being evident within 20 min of pretreatment, and was specific for zymosan or glucan particles in that the LTB4 and LTC4 release induced by 2.5 ,M calcium ionophore A23187 was unaffected. That both phagocytosis and leukotriene generation are inhibited by soluble 3glucan but not by mannan at a rate compatible with the phagocytic process of monocyte monolayers indicates ligand specificity for a -glucan receptor. As the 3glucan receptor recognizes particulate activators of the alternative complement pathway, the nonimmune response to a single stimulus induces complement activation, phagocytosis, and leukotriene generation.In the absence of plasma proteins, human peripheral blood monocytes phagocytose zymosan particles and other particulate activators of the human alternative complement pathway through a specific trypsin-sensitive receptor that is distinct from that for Fc-IgG or the major cleavage fragment of the third component of complement (C3b) (1-3). Perturbation of this phagocytic receptor with zymosan particles stimulates monocytes to metabolize endogenous arachidonic acid to substantial quantities of leukotriene B4 (LTB4) and leukotriene C4 (LTC4), whereas comparable perturbation of monocyte phagocytic receptors for Fc-IgG does not (4). The capacity of monocyte monolayers to generate and release leukotrienes in response to zymosan particles is decreased in a dose-dependent fashion by the same low concentrations of trypsin (4) that reduce monocyte ingestion of the particulate activators, zymosan (1), rabbit erythrocytes (1), and desialated sheep erythrocytes (2), whereas the ionophore A2...
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