Membrane complement receptors specific for bound fragments of C3.
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Complement Activation↗
Biomedical subjects
G D Ross
Publications and source records attributed to G D Ross.
Membrane complement receptor type three (CR3) has lectin-like properties analogous to bovine conglutinin as functions as a receptor for zymosan and rabbit erythrocytes as well as a receptor for iC3b.
Human leukocyte complement receptor type three (CR3) was shown to be lectin-like and to resemble bovine serum conglutinin (K) in that it bound to both iC3b and unopsonized yeast (Saccharomyces cerevisiae), and was inhibited by EDTA or N-acetyl-D-glucosamine (NADG). CR3 and K also bound to zymosan (Z), a yeast cell wall extract that contains primarily polysaccharide and no detectable protein. However, structural differences and the absence of K on bovine phagocytes indicated that CR3 was not the human homologue of bovine K. Phagocytic and respiratory responses to unopsonized Z were CR3 dependent because they were inhibited by monoclonal antibodies specific for the alpha-chain of CR3 and did not occur with phagocytes from patients with a genetic deficiency of CR3. The binding of CR3 to Z did not require opsonization of the Z with neutrophil-secreted C3, as Z binding and responses were not inhibited by Fab anti-C3. In addition, CR3-dependent binding of yeast occurred with neutrophils from which protein secretion was blocked by fixation with paraformaldehyde. Rabbit erythrocytes (RaE) also bound weakly to neutrophil CR3 and triggered ingestion. Anti-CR3 not only blocked the binding and ingestion of RaE but also blocked selectively the ingestion of RaEC3b without affecting the strong binding mediated by CR1. Even though sheep E and sheep EC3b were not ingested by neutrophils, a weak binding of CR3 to sheep E was suggested by the finding of 20 to 40% inhibition of sheep EAIgG ingestion by anti-CR3. Such inhibition was only observed in buffers that allowed activity of the CR3 binding site and not in buffers containing either EDTA or NADG. An apparently contradictory finding was that the weak CR3-dependent binding of Z triggered neutrophil ingestion and a superoxide burst, whereas the avid CR3-dependent binding of sheep EC3bi did not induce significant ingestion or a respiratory burst. Blocking studies with monoclonal antibodies specific for different epitopes of the alpha-chain of CR3 suggested that this might result from the presence of two distinct binding sites in CR3: one site for fixed iC3b that did not trigger functions, and a second function-triggering site for Z that did not bind to fixed iC3b.
Animals↗
Characterization of patients with an increased susceptibility to bacterial infections and a genetic deficiency of leukocyte membrane complement receptor type 3 and the related membrane antigen LFA-1.
Three children from two unrelated families had a history of recurrent bacterial infections, and their neutrophils were shown to have deficient phagocytic and respiratory responses and possible deficiencies in chemotaxis or adherence. Their neutrophils were strikingly deficient in the ability to ingest or give a respiratory burst in response to unopsonized bakers' yeast or zymosan (Z). Tests for neutrophil and monocyte CR1 (C3b/iC3b receptor) and CR3 (iC3b receptor) demonstrated rosettes with both EC3b and EC3bi. However, EC3bi were bound only to CR1, and not to CR3, because EC3bi rosettes were inhibited completely by anti-CR1. Neutrophils, monocytes, and natural killer (NK) cells also did not fluorescence stain with monoclonal antibodies specific for the alpha-chain of CR3 (anti-Mac-1, anti-Mol, OKM1, and MN-41). Quantitation of C receptors with 125I monoclonal anti-CR1 and anti-CR3 indicated that neutrophils from each patient expressed normal amounts of CR1 per cell but less than 10% of the normal amount of CR3. Examination of neutrophils by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated that a normal glycoprotein of approximately 165,000 daltons was missing. Immunoblotting of these gels indicated that the missing band was the alpha-chain of CR3. Subsequent analysis of all three patients' cells also demonstrated a deficiency of LFA-1 alpha-chain and the common beta-chain that is shared by the CR3/LFA-1/p150,95 membrane antigen family. The deficiency of LFA-1 probably explained the absent NK cell function, as normal NK cell activity is inhibited by anti-LFA-1 but not by anti-CR3. The reduced phagocytic and respiratory responses to Z were probably due to CR3 deficiency, because treatment of normal neutrophils with anti-CR3, but not anti-FLA-1, inhibits responses to Z by 80% to 90%. Ingestion of Staphylococcus epidermidis by normal neutrophils was shown to be partially inhibited by monoclonal antibodies to the alpha-chain of either CR3 or LFA-1, and monoclonal antibody to the common beta-chain inhibited ingestion by 75%. Thus, both CR3 and LFA-1 may have previously unrecognized functions as phagocyte receptors for bacteria. The absence of this type of nonimmune recognition of bacteria by these children's neutrophils may be one of the reasons for their increased susceptibility to bacterial infections.
Antigens, Surface↗
Family studies of erythrocyte complement receptor type 1 levels: reduced levels in patients with SLE are acquired, not inherited.
It has been claimed that patients with systemic lupus erythematosus (SLE) have an inherited deficiency of erythrocyte complement receptor type 1 (CR1, with ligand binding specificity for C3b, iC3b and C4b). CR1 functions as the only cofactor for factor I-mediated cleavage of iC3b to C3c and C3dg. The activity of this receptor on red cells may be an important mechanism for handling immune complexes which have bound C3b or iC3b. Radioligand binding studies were performed using a monoclonal antibody to CR1, E11, to enumerate these receptors accurately. The results confirmed that patients with SLE have a reduced number of CR1 molecules per red cell, but showed no reduction in CR1 levels amongst their consanguineous relatives. Study of 13 normal families suggested the presence of heritable factors controlling the numbers of erythrocyte CR1 molecules; in particular there was a correlation between mean parental CR1 numbers and CR1 numbers in their children. However, amongst 17 families of 19 patients with SLE, four families were identified in which genotypically 'high CR1' SLE patients had persistently low phenotypes. This is not compatible with the hypothesis that the reduction in erythrocyte CR1 numbers in these patients is inherited.
Antibodies, Monoclonal↗
Disease-associated loss of erythrocyte complement receptors (CR1, C3b receptors) in patients with systemic lupus erythematosus and other diseases involving autoantibodies and/or complement activation.
Although surface membrane density of complement receptor type one (CR1) on erythrocytes (E) is probably an inherited trait among normal individuals, recent evidence from our laboratories suggests that the reduced number of CR1 per E observed in patients with systemic lupus erythematosus (SLE) results from acquired as well as genetic factors. In the present investigation, the number of CR1 per E was quantitated with 125I-monoclonal anti-CR1 and was found to vary inversely with disease activity in patients with SLE who were followed serially for as long as 14 mo. Although evidence for E surface-bound immune complexes or fixed C3b/iC3b was not obtained, periods of disease activity and low amounts of CR1 per E correlated with the presence of 100 to 800 molecules per E of fixed C3dg fragments (less than 100 C3dg per E in normal subjects). Reduced CR1 and excess fixed C3dg on E also were observed in patients with other disorders associated with complement activation, including chronic cold agglutinin disease, autoimmune hemolytic anemia, paroxysmal nocturnal hemoglobinuria (PNH), Sjögren's syndrome, and mycoplasma pneumonia. A significant negative correlation (r = -0.498) between CR1/E and fixed C3dg/E was demonstrable in 255 individual assays evaluated by regression analysis. CR1 decreased and fixed C3dg increased during active disease; the converse was obtained during remission. In patients with active SLE, both serum complement activity and E CR1 decreased, whereas fixed C3dg fragments increased. By piecewise linear regression analysis, the appearance of 100 to 400 C3dg molecules on patients' E corresponded to a 27 to 60%, reduction in the number of CR1 per E (p less than 0.0002), confirming that fixation of C3 to E was correlated with a loss of CR1. In patients with PNH, low values for CR1 were observed on moderately complement-sensitive PNH type II E in association with increased fixed C3 fragments; however, the markedly complement-sensitive PNH type III E had essentially normal amounts of CR1 and bore little fixed C3. The addition of soluble DNA/anti-DNA immune complexes to normal blood generated levels of fixed C3dg fragments on E comparable to those observed on E from patients with SLE. Kinetic experiments indicated that C3b was fixed to E during the process of immune complex binding and release from E CR1, and that this fixed C3b was subsequently degraded rapidly to fixed iC3b and more slowly to fixed C3dg without the loss of CR1 that occurs in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)
Anemia, Hemolytic↗
Identification of an anti-monocyte monoclonal antibody that is specific for membrane complement receptor type one (CR1).
A monoclonal antibody (E11) was produced by immunization of mice with intact human cells of monocyte lineage. Despite the finding that E11 did not inhibit rosettes with C3b-coated sheep erythrocytes (EC3b), several lines of evidence indicated that E11 was specific for complement receptor type one (CR1). All monocytes, neutrophils, lymphocytes and erythrocytes that reacted with E11 formed EC3b rosettes. The E11 antigen on these cells was shown to be a molecule of 222 +/- 10 kDa. Treatment of lymphocytes, monocytes, and neutrophils with E11 followed by fluorescein-coupled F(ab')2 anti-mouse-IgG at 37 degrees C in buffer lacking sodium azide, led to capping or apparent endocytosis of the E11 antigen and a diminution in CR1 activity of 88%, 59% and 25%, respectively. This same treatment had no detectable effect on monocyte or neutrophil CR3 activity (EC3bi rosettes). Furthermore, with E11-capped lymphocytes, the residual EC3b rosetting was capped directly over the E11-fluorescence cap, whereas EC3d,g rosetting (CR2 specific) was undiminished and distributed evenly around the circumference of cells containing E11-fluorescence caps. Finally, the binding of E11 to cells was inhibited by the prior treatment of these cells with a well characterized rabbit polyclonal anti-CR1. These data indicated that E11 was specific for a site in CR1 that was distal from the C3b-binding site, so that E11 was unable to block CR1 activity. E11 proved to be useful for identifying CR1 on various cells in tissue sections, and for quantitating CR1 on erythrocytes and neutrophils. Erythrocytes and neutrophils from normal individuals were found to bind an average of 610 and 4.6 X 10(4) 125I-labeled E11 molecules per cell. When E11 was visualized in tissues by immunoperoxidase staining, the cells that apparently contained the greatest amounts of CR1 were dendritic reticulum cells and kidney podocytes. The E11 reactive dentritic reticulum cells were characteristic of both follicular and diffuse follicular center cell tumors. Lymphocytes from patients with chronic lymphocytic leukemia (CLL) characteristically expressed little E11, confirming earlier studies that CLL cells lacked CR1 activity detected by EAC1-3b rosette formation. Because normal B cells have been shown to express CR1 at a very early stage of maturation, the absence of CR1 on CLL cells is discordant with the immature nature of CLL cells defined by immunoglobulin expression.
Animals↗
Phagocytosis by human monocyte-derived macrophages. Independent function of receptors for C3b (CR1) and iC3b (CR3).
Human monocytes and monocyte-derived macrophages were examined for their ability to bind and ingest C3-coated sheep erythrocytes (E). Greater than 90% of monocytes and macrophages formed rosettes with EC3b and EC3bi prepared with 15,000-20,000 molecules of C3 per E. Binding of EC3b to the monocyte or macrophage surface was inhibited by rabbit anti-C3b receptor (CR1), but was not inhibited by two different monoclonal anti-C3bi receptor (CR3) antibodies. EC3bi rosette formation was inhibited by monoclonal anti-CR3, but not by anti-CR1. Monocytes and macrophages did not form rosettes with similarly prepared EC3d,g or EC3d. Macrophages cultured for 7 days, but not freshly isolated monocytes, phagocytosed both EC3b and EC3bi. This ability was a consequence of macrophage maturation, as no external stimuli were present during in vitro culture. Experiments directed to determine if EC3b was converted to EC3bi before ingestion suggested that macrophage CR1 and CR3 mediated phagocytosis independently. No evidence was obtained that during the phagocytosis assay, macrophage factor I converted EC3b to EC3bi. The number of E bound or ingested by monocytes and macrophages was dependent on the number of molecules of C3b or iC3b bound per E. Monocytes and macrophages did not require the presence of either Ca++ or Mg++ for rosette formation with EC3b, whereas both divalent cations were required for optimum rosette formation with EC3bi. The presence of divalent cations was required for macrophage phagocytosis of EC3b and EC3bi. For ingestion of EC3b, Mg++ alone was sufficient, and the addition of Ca++ did not increase the number of EC3b ingested. For ingestion of EC3bi, both Ca++ and Mg++ were required for optimal phagocytosis, and their effect was concentration dependent and additive.
Calcium↗
Generation of three different fragments of bound C3 with purified factor I or serum. II. Location of binding sites in the C3 fragments for factors B and H, complement receptors, and bovine conglutinin.
The many different recognized functions of C3 are dependent upon the ability of the activated C3 molecule both to bind covalently to protein and carbohydrate surfaces and to provide binding sites for as many as eleven different proteins. The location of the binding sites for six of these different proteins (factors B and H, complement receptors CR(1), CR(2) and CR(3) and conglutinin) was examined in the naturally occurring C3-fragments generated by C3 activation (C3b) and degradation by Factor I (iC3b, C3c, C3d,g) and trypsin (C3d). Evidence was obtained for at least four distinct binding sites in C3 for these six different C3 ligands. One binding site for B was detectable only in C3b, whereas a second binding site for H and CR(1) was detectable in both C3b and iC3b. The affinity of the binding site for H and CR(1) was charge dependent and considerably reduced in iC3b as compared to C3b. H binding to iC3b-coated sheep erythrocytes (EC3bi) was measurable only in low ionic strength buffer (4 mS). The finding that C3c-coated microspheres bound to CR(1), indicated that this second binding site was still intact in the C3c fragment. However, H binding to C3c was not examined. A third binding site in C3 for CR(2) was exposed in the d region by factor I cleavage of C3b into iC3b, and the activity of this site was unaffected by the further I cleavage of iC3b into C3d,g. Removal of the 8,000-dalton C3g fragment from C3d,g with trypsin forming C3d, resulted in reduced CR2 activity. However, because saturating amounts of monoclonal anti-C3g did not block the CR(2)-binding activity of EC3d,g, it appears unlikely that the g region of C3d,g or iC3b forms a part of the CR(2)-binding site. In addition, detergent-solubilized EC3d (C3d-OR) inhibited the CR(2)-binding activity of EC3d,g. Monocytes and neutrophils, that had been previously thought to lack CR(2) because of their inability to form EC3d rosettes, did bind EC3d,g containing greater than 5 x 10(4) C3d,g molecules per E. The finding that monocyte and neutrophil rosettes with EC3d,g were inhibited by C3d-OR, suggested that these phagocytic cells might indeed express very low numbers of CR(2), and that these CR(2) were detectable with EC3d,g and not with EC3d because C3d,g had a higher affinity for CR2 than did C3d. A fourth C3 binding site for CR(3) and conglutinin (K) was restricted to the iC3b fragment. Because of simultaneous attachment of iC3b to phagocyte CR3 and CR(3), the characteristics of iC3b binding to CR3 could only be examined with phagocytes on which the CR(1) had been blocked with anti-CR(1). Inhibition studies with EDTA and N-acetyl-D-glucosamine demonstrated a requirement for both calcium cations and carbohydrate in the binding of EC3bi to CR3 and to K. However, CR(3) differed from K in that magnesium cations were required in addition to calcium for maximum CR(3) binding activity, and NADG produced less inhibition of CR(3) activity than of K activity.
Animals↗
Studies of the Epstein Barr virus receptor found on Raji cells. II. A comparison of lymphocyte binding sites for Epstein Barr virus and C3d.
A comparison was made between the binding sites of two receptors that are believed to be closely associated on human B lymphocytes: complement receptor type two (CR2) that is specific for C3d fragments, and the receptor (EBVR) for Epstein Barr virus (EBV). Isolated fluid-phase CR2 bound to C3d on erythrocytes (EC3d) and inhibited both B cell-EC3d rosettes and the agglutination of EC3d by anti-C3d, it failed to inhibit either the binding or superinfection of B cells by EBV. By contrast, isolated fluid-phase EBVR inhibited EBV B cell binding activity and superinfection but had no CR2 activity. In addition, radiolabeled CR2 bound to EC3d and anti-CR2-Sepharose, whereas radiolabeled EBVR did not. Purified fluid-phase C3d fragments inhibited EC3d rosette formation with CR2+/EBVR+ cells but did not inhibit EBV binding. However, EBV binding to B cells did inhibit EC3d rosette formation. Clones of human/mouse somatic cell hybrids made from CR2+/EBVR+ human B lymphoblastoid cell and CR2-/EBVR- mouse myeloma cell parents expressed either EBVR or CR2 but only rarely expressed both EBVR and CR2. This suggested that the genes for EBVR and CR2 were located on two different human chromosomes. Thus it was concluded that CR2 is probably not the binding site for EBV.
Animals↗
Characterization of the lymphocyte membrane receptor for factor H (beta 1H-globulin) with an antibody to anti-factor H idiotype.
Antibody to the binding site (idiotype) of anti-factor H was shown to have specificity for both B lymphocyte membrane H receptors and C3b. Goat F(ab')(2) anti-human H was purified by absorption and elution from H agarose and used for rabbit immunization to produce anti-anti-H (aaH). After absorption with nonimmune goat IgG, (125)I-labeled aaH bound to B lymphocytes and to sheep erythrocytes coated with C3b (EC3b) but did not bind to T lymphocytes or to EC3d. All B cell- and C3b-specific activities of the aaH were removed and subsequently recovered by absorption and elution of the antibody from either C3-agarose or goat-anti-H-agarose. This indicated that the aaH probably recognized a single common antigenic structure that was shared by anti-H, C3b, and the membranes of B cells. Affinity-purified aaH resembled H in that it bound to B cells, blocked the uptake of H onto B cell H receptors, and triggered B cells to release endogenous factor I (C3b inactivator). In addition, aaH functioned with factor I as either a cofactor for cleavage of fluid-phase C3b or a potentiator for cleavage of bound C3b. This same spectrum of C3 binding functions could not be demonstrated with either sheep anti-C3b or rabbit-anti-C3c. Analysis by sodium dodecyl sulfate- polyacrylamide get electrophoresis of the [(3)H]leucine intrinsically labeled B cell proteins reactive with the purified aaH revealed proteins of 100,000 M(r) and 50,000 M(r) without reduction, and after complete reduction of disulfide bonds, a single protein band of 50,000 M(r). This same protein molecular weight profile was also demonstrated with labeled B cell proteins that were absorbed and eluted from H-agarose. Thus, aaH is apparently specific for both B cell H receptors and C3b. However, because parallel analysis of C3b confirmed its known 115,000- and 75,000-M(r) polypeptide chain structure, the H receptor is probably not C3b and shares only the structure of the H binding site with C3b.
Animals↗
Identification of a C3bi-specific membrane complement receptor that is expressed on lymphocytes, monocytes, neutrophils, and erythrocytes.
Cells expressing a membrane C receptor (CR(3)) specific for C3b-inactivator- cleaved C3b (C3bi) were identified by rosette assay with C3bi-coated sheep erythrocytes (EC3bi) or C3bi-coated fluorescent microspheres (C3bi-ms). C3bi- ms, probably because of their smaller size, bound to a higher proportion of cells than did EC3bi. C3bi-ms bound to greater than 90 percent of mature neutrophils, 85 percent of monocytes, 92 percent of erythrocytes, and 12 percent of peripheral blood lymphocytes. Binding of C3bi-ms to neutrophils, monocytes, and erythrocytes was inhibited by fluid-phase C3bi, Fab anti-C3c, or Fab anti-C3d but was not inhibited by F(ab')(2) anti-CR(1) (C3b receptor) or F(ab')(2) anti-CR(2) (C3d receptor) nor by fluid-phase C3b, C3c, or C3d. This indicated that monocytes, neutrophils, and erythrocytes expressed C3bi receptors (CR(3)) that were separate and distinct from CR(1) and CR(2) and specific for a site in the C3 molecule that was only exposed subsequently to cleavage of C3b by C3b inactivator and that was either destroyed, covered, or liberated by cleavage of C3bi into C3c and C3d fragments. Lymphocytes differed from these other cell types in that they expressed CR2 in addition to CRa. Lymphocyte C3bi-ms rosettes were inhibited from 50 to 84 percent by F(ab')(2)-anti-CR(2) or fluid-phase C3d, whereas C3d-ms rosettes were inhibited completely by F(ab')(2) anti-CR(2), fluid-phase C3bi, or fluid- phase C3d. Thus, with lymphocytes, C3bi was bound to CR(3), and in addition was bound to CR(2) by way of the intact d region of the C3bi molecule. In studies of the acquisition of C receptors occurring during myeloid cell maturation, the ability to rosette with C3bi-coated particles was detected readily with immature low-density cells, whereas this ability was nearly undetectable with high density mature polymorphonuclear cells. This absence of C3bi binding to polymorphs was not due to a loss of the CR(3) but instead was due to the maturation-linked acquisition of the abiity to secrete elastase that cleaved reagent particle-bound C3bi into CR(3)-unreactive C3d. Neither neutrophils nor monocytes bound C3d-coated particles at any stage of maturation. Assay of CR(3) with mature neutrophils required inhibition of neutrophil elastase with either soybean trypsin inhibitor or anti-elastase antibodies, and the amounts of these elastase inhibitors required to allow EC3bi rosette formation increased with neutrophil maturation. Because lymphocytes bound C3bi to CR(2) as well as to CR(3), specific assay of lymphocyte CR(3) required saturation of membrane CR(2) with Fab' anti-CR(2) before assay for rosettes with C3bi-ms. Only 3.5 percent of anti-CR(2)- treated peripheral blood lymphocytes bound C3bi-ms. Therefore, among normal blood lymphocytes the majority of the 12 percent C3bi-ms-binding cells expressed only CR(2) (8.5 percent), and the small proportion of C3bi-ms- binding cells that expressed CR(3) (3.5 percent) represented a distinct subset from the CR2(+) cells. Double-label assay indicated that 3.0 percent out of 3.5 percent of these CR(3)-bearing lymphocytes were B cells because they expressed membrane immunoglobulins. Of the remaining CR(3)(+) cells, 0.2 percent expressed either Leu-1 or 3A1 T cell antigens, and 0.6 percent expressed the OKM-1 monocyte-null lymphocyte determinant.
Cell Membrane↗
Generation of three different fragments of bound C3 with purified factor I or serum. I. Requirements for factor H vs CR1 cofactor activity.
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Animals↗
Structure and function of membrane complement receptors. Summary.
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Complement Activating Enzymes↗
Assay of membrane complement receptors (CR1 and CR2) with C3b- and C3d-coated fluorescent microspheres.
A sensitive and specific fluorescence assay for membrane complement (C) receptors (CR1 and CR2) was developed with purified C3b and C3d fragments coupled to fluorescent microspheres (0.9 mu diameter). C3-microspheres (C3-ms) bound to cells with low numbers of receptors that were undetectable by other assay techniques. Inhibition studies with anti-CR1 and anti-CR2 demonstrated that C3b-ms and C3d-ms bound exclusively to CR1 and CR2, respectively. Preparation of the C3-ms required only small amounts of partially purified C3 and no immunoglobulin or other C components. Once formed, the C3-ms were stable for up to 4 mo at 4 degrees C.
Binding, Competitive↗
Isolation of lymphocyte membrane complement receptor type two (the C3d receptor) and preparation of receptor-specific antibody.
A glycoprotein binding complement component C3d was isolated from media used for culture of Raji human lymphoblastoid cells. Analysis by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and gas/liquid chromatography indicated that the C3d-binding glycoprotein consisted of a single polypeptide chain with extensive intrachain disulfide bonds, a molecular weight of 72,000, and several different bound carbohydrates. Several lines of evidence indicated that this medium-derived C3d-binding protein originated from membrane complement receptor type two (CR2, the C3d receptor), presumably shed during membrane turnover. The C3d-binding protein bound to sheep erythrocytes coated with C3d (EC3d) but not to sheep erythrocytes coated with C3b (EC3b). Antisera, prepared by immunization with the purified C3d-binding glycoprotein, inhibited lymphocyte rosette formation with EC3d but not with EC3b. Analysis by sodium dodecyl sulfate gel electrophoresis of the radiolabeled and solubilized lymphocyte antigens reactive with the anti-C3d-binding protein sera revealed a single-chain cell-surface protein of molecular weight 72,000 that was apparently identical to the isolated C3d-binding protein. Parallel assay of lymphocytes for Cr2 by direct immunofluorescence with F(ab')2 anti-C3d-binding protein (anti-CR2) and rosette formation with EC3d indicated that both assays had the same specificity and nearly the same sensitivity. With both systems CR2 expression was limited to B cells, and was undetectable on T cells, monocytes, or neutrophils.
Animals↗
Characteristics of isolated erythrocyte complement receptor type one (CR1, C4b-C3b receptor) and CR1-specific antibodies.
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Animals↗
Release of endogenous C3b inactivator from lymphocytes in response to triggering membrane receptors for beta 1H globulin.
Human bone marrow-derived lymphocytes and cells from B lymphoblastoid lines were shown to have specific membrane receptors for beta 1H globin. Lymphocytes responded to the presence of beta 1H by releasing endogenously-synthesized C3b-inactivator. Very little spontaneous release of C3b-inactivator occurred in the absence of beta 1H. beta 1H-treated lymphocytes that either lacked complement receptor type one (CR1, the C4b-C3b receptor) or had their CR1 blocked with Fab'-anti-CR1 formed rosettes with C3b-coated sheep erythrocytes (EC3b) by adherence to complement receptor type two (CR2, the C3d-C3bi receptor). The mechanism of the beta 1H-induced EC3b rosette formation was shown to involve the release of lymphocyte C3b-inactivator that cleaved bound C3b into C3bi forming EC3bi. This lymphocytes-generated EC3bi then bound to CR2, forming rosettes. beta 1H-induced EC3b rosettes were completely inhibited by the presence of either anti-C3b-inactivator, F(ab')2-anti-CR2, Fab-anti-C3c, or Fab-anti-C3d, but were unaffected by the presence of fluid-phase concentrations of beta 1H up to 5.5 mg/ml or Fab'-anti-CR1. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography demonstrated that cell-free supernates of beta 1H-treated lymphocytes cleaved 3H-labeled C3b on EC3b into C3bi. Inhibition studies with chelating agents and sodium azide suggested that the release of C3b inactivator might involve a calcium and energy-dependent transport of this enzyme across the membranes of beta 1H-triggered cells. Because plasma beta 1H and C3b-inactivator are known to have important functions in the distinction of alternative pathway-activating substances from normal tissue, it is possible that this beta 1H receptor-C3b-inactivatory releasing system in lymphocytes may have an analogous function.
B-Lymphocytes↗
Analysis of the different types of leukocyte membrane complement receptors and their interaction with the complement system.
The specificity, distribution, and structure of 8 different types of leukocytes membrane complement (C) receptors (CR1, CR2, CR3, and receptors for C1q, beta 1H, C3e, C3a, and C5a) are discussed. Recent data are reviewed on the synthesis of C components by macrophages and B lymphocytes, and how these components may function in the activation of these two cell type by the C system. Commonly used C receptor assay procedures are evaluated in terms of both specificity and sensitivity. Specific assay procedures are recommended for measuring CR1 (C4b-C3b receptor), CR2 (C3d receptor), CR3 (C3bi receptor), and the beta 1H receptor. Assays include both rosette and fluorescence procedures for detection of C receptors on either mouse or human leukocytes. Primary systems have been selected for optimal sensitivity and specificity, and where possible, acceptable alternative systems that are less sensitive or specific are suggested for laboratories lacking facilities for C purification.
Animals↗