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R F Todd

Publications and source records attributed to R F Todd.

At least 73 records · Page 4Linked to original sources
PubMed1993

Lectin-like inhibition of immune complex receptor-mediated stimulation of neutrophils. Effects on cytosolic calcium release and superoxide production.

We have tested the role of lectin-like interactions, with particular emphasis on CR3, in insoluble immune complex (IC)-mediated activation of human polymorphonuclear leukocytes (PMN). The ability of IC to trigger intracellular Ca2+ release and O2- production by normal PMN, saccharide-treated cells, and CR3-deficient PMN (leukocyte adhesion deficiency, LAD, patients) were tested. When indo-1-labeled normal PMN were stimulated with IC in Ca(2+)-free buffer, intracellular Ca2+ rose from approximately 100 nM to approximately 230 nM. However, when LAD PMN were tested, a small rise in intracellular Ca2+ was observed. Because previous studies have shown that certain saccharides inhibit CR3-Fc gamma RIII co-capping, we tested a panel of saccharides to determine their ability to influence IC-mediated intracellular Ca2+ release. When normal PMN were exposed to 0.15 M N-acetyl-D-glucosamine (NADG), D-mannose, or alpha-methyl-mannoside, the Ca2+ response to IC was significantly reduced. However, addition of 0.15 M glucose, raffinose, sucrose, galactose, fructose, or sorbitol did not significantly affect the Ca2+ response, suggesting that the response was specific for certain saccharides. To test the physiologic consequences of these Ca2+ signals, we have examined the ability of saccharides to influence O2- production by normal PMN and the ability of LAD PMN to produce O2- upon triggering by IC. Normal PMN stimulated with IC generated 4.3 +/- 1.7 nmol/10(6) cells/30 min of O2-. In contrast, O2- production was inhibited by 0 to 20% by glucose, galactose, sucrose, sorbitol, fructose, and raffinose and > or = 50% by NADG and mannose. LAD PMN, which display diminished Ca2+ signals, were found to produce O2- at 47 +/- 6% of control levels. NADG and mannose dose-response studies indicated that they cooperatively block O2-.

Adult↗
PubMed1993

Cocapping of the leukoadhesin molecules complement receptor type 3 and lymphocyte function-associated antigen-1 with Fc gamma receptor III on human neutrophils. Possible role of lectin-like interactions.

We have tested the possible physical interactions between the iC3b receptor (CR3), lymphocyte function-associated Ag-1, and class III Fc gamma receptor (Fc gamma RIII) at neutrophil surfaces. Cells were labeled using fluorochrome-conjugated Fab or F(ab')2 fragments of antireceptor mAb. Labeled receptors were capped using second-step F(ab')2 fragments of goat anti-mouse Fab antiserum. After 20 min at 37 degrees C, 68% of the cells capped the anti-CR3 plus second-step complex. Capping was time, temperature, and cytochalasin B sensitive. When capped cells were probed with Fab' or F(ab')2 fragments of anti-Fc gamma RIII labeled with a distinct fluorochrome, 41% of the cells cocapped Fc gamma RIII. Indistinguishable results were obtained when potential antibody combining sites within caps were blocked with a large excess of Fab or F(ab')2 fragments. When Fc gamma RIII was capped, 49% of the cells cocapped CR3. Similarly, LFA-1 cocapped with both CR3 and Fc gamma RIII. Importantly, other membrane components including HLA class I, Mo5, CD13, CR type 1, and IL-8 receptors and N-4-nitrobenzo-2-oxa-1, 3-diszole L-alpha-dimyristoyl phosphatidylethanolamine did not cocap with CR3. However, the positive control Con A did cocap with CR3 and Fc gamma RIII. We next evaluated the effect of saccharides on CR3-Fc gamma RIII cocapping and found that 0.15 M N-acetyl-D-glucosamine (NADG), alpha-methyl-D-mannoside, and D-mannose significantly inhibited cocapping by 70, 58, and 48%, respectively. No inhibition was obtained using glucose, galactose, N-acetyl-neuraminic acid, fucose, sorbitol, fructose, or sucrose. Similarly, Fc gamma RIII-lymphocyte function-associated-1 cocapping was inhibited by NADG. However, the cocapping of CR3 with lymphocyte function-associated-1 or Con A were not affected by 0.15 M NADG, which suggests that NADG inhibition of leukoadhesin-Fc gamma RIII cocapping is not due to a general effect of NADG on capping. Inasmuch as Fc gamma RIII is a glycophospholipid-linked membrane protein, we speculate that it interacts with CR3 and/or lymphocyte function-associated-1 via lectin-like interactions.

Adult↗
PubMed1993

Role of beta 1, beta 2 integrins and ICAM-1 in lung injury after deposition of IgG and IgA immune complexes.

After intrapulmonary deposition of IgG or IgA immune complexes, injury has been recently shown to be CD18-dependent in both cases and E-selection-dependent only in the former case. In our studies further evaluation of the requirements for beta 1 and beta 2 integrins and intercellular adhesion molecule-1 (ICAM-1) has been undertaken. In the IgG immune complex model, which is neutrophil dependent, anti-CD11a reduced injury (as measured by changes in permeability and hemorrhage) by 61 and 43%, respectively, whereas a newly developed anti-CD11b produced minimal protection (16 and 19%, respectively). Treatment of rats with increasing doses (1.5- and 3.0-fold) of antibody to rat CD11b failed to demonstrate additional protective effects in this model of injury. Anti-ICAM-1 reduced the parameters of injury by 61 and 78%, respectively, while anti-VLA-4 reduced the injury parameters by 40 and 35%, respectively. There were reductions in lung content of myeloperoxidase, roughly corresponding to the protective effects of the interventions. In the IgA immune complex model of injury, in which lung macrophages appear to be the effector cells, anti-CD11a reduced the injury parameters (permeability and hemorrhage) by 36 and 33%, respectively, whereas anti-CD11b reduced the parameters of injury by 63 and 67%, respectively. In this model, anti-ICAM-1 reduced the parameters of injury by 61 and 56%, respectively, while anti-VLA-4 reduced the parameters by 77 and 62%, respectively. The cell content of bronchoalveolar lavage fluids revealed changes that have been shown to reflect protective interventions in both models of immune complex-induced injury. These findings suggest that, in IgG and IgA immune complex models of lung injury, both VLA-4 and ICAM-1 are required, although lymphocyte function-associated Ag-1 is the predominant beta 2 integrin requirement in the IgG immune complex-induced model of injury and Mac-1 is the predominant requirement for IgA immune complex-induced lung injury. Thus, engagement in the lung of adhesion molecules in a manner leading to injury depends on the nature of the inflammatory stimulus and the type of phagocytic cells involved in the development of injury.

Animals↗
PubMed1993

Role of leukocyte adhesion molecules in complement-induced lung injury.

The bolus i.v. infusion of cobra venom factor into rats results in acute lung injury that is neutrophil-dependent, oxygen radical-mediated, and requires CD18. In our studies a more precise definition regarding the role of beta-integrins and requirements for cytokines was obtained by the use of blocking antibodies. Lung injury was quantitated by changes in permeability (leakage of 125I-BSA) and hemorrhage (extravasation of 51Cr-RBC). In animals treated with anti-CD11a, the permeability and hemorrhage parameters were reduced by 30 and 29%, respectively. Treatment with anti-CD11b resulted in reductions in permeability and hemorrhage by 53 and 48%, respectively, whereas anti-intercellular adhesion molecule-1 reduced the parameters of injury by 60 and 75%, respectively. Not surprisingly, treatment with antibodies to very late Ag-4, TNF-alpha and IL-1 failed to show any protective effects, which contrasts to the requirements for these molecules in lung injury after deposition of IgG immune complexes. Protective interventions were associated with a reduction in lung content of myeloperoxidase. These studies indicate that, in the cobra venom factor model of acute lung injury in rats, engagement of Mac-1, lymphocyte function-associated Ag-1, and intercellular adhesion molecule-1 are essential, whereas, in contrast to other models of neutrophil mediated lung injury, cytokines (TNF-alpha and IL-1) and very late Ag-4 are not required for the full development of injury.

Animals↗
PubMed1993

Receptor-receptor interactions of complement receptor type 3 in neutrophil membranes.

The leukocyte integrin CR3 (CD11b/CD18) is known to participate in a variety of cell functions. Recent studies have indicated that CR3 may communicate with other plasma membrane receptors to carry out several cell functions. In this review we discuss these potential receptor-receptor interactions of CR3 and present a unifying model of CR3's diverse functions.

Animals↗
PubMed1993

Requirements for leukocyte adhesion molecules in nephrotoxic nephritis.

Requirements for leukocyte adhesion molecules as well as cytokines have been determined in the rat model of acute nephrotoxic nephritis. Proteinuria (at 24 h) and neutrophil accumulation in renal glomeruli (at 6 h) have been used as the endpoints. For full accumulation in glomeruli of neutrophils as well as full development of proteinuria, requirements have been demonstrated for TNF alpha, (but not IL-1), CD11b (but not CD11a), very late arising-4 (CD49d/CD29), and intercellular adhesion molecule-1 but not endothelial leukocyte adhesion molecule-1 (E-selectin). By immunohistochemical approaches, infusion of antibody to glomerular basement membrane induced glomerular upregulation of intercellular adhesion molecule-1, endothelial leukocyte adhesion molecule-1, and vascular adhesion molecule-1. Treatment of rats with anti-TNF alpha or soluble recombinant human TNF receptor-1 blocked this expression. Renal arterial infusion of TNF alpha induced glomerular expression of all three endothelial adhesion molecules, but infusion of IL-1 beta did not. These data suggest that, in neutrophil and complement-dependent anti-glomerular basement membrane-induced acute nephritis in rats, there are selective requirements for cytokines, beta 1 and beta 2 integrins, and endothelial adhesion molecules. These requirements contrast with those found in other vascular beds in which complement and neutrophil-induced vascular injury has been induced by deposition of immune complexes.

Acute Disease↗
PubMed1993

Defect of a complement receptor 3 epitope in a patient with systemic lupus erythematosus.

Complement receptor 3 (CR3) is expressed on cells of the reticuloendothelial system and involved in the clearance of immune complexes. In this article a patient with a deficiency of the C3bi binding site of this receptor is described. Clinically this patient exhibited predominantly cutaneous manifestations of a systemic lupus erythematosus with an immune vasculitis and panniculitis. The deficiency of the CR3 epitope was demonstrated using flow cytometry. The functional relevance of this defect was demonstrated in a rosetting assay with C3bi-loaded erythrocytes. C3bi binding was found to be significantly decreased. Furthermore, there was an impairment of phagocytosis of opsonized Escherichia coli. The CR3 defect is not due to an autoantibody but is assumed to have a genetic basis. These data suggest that the defect of the CR3 may be involved in the pathogenesis of the immune vasculitis in this patient.

Adult↗
PubMed1993

Role of beta 2 integrins and ICAM-1 in lung injury following ischemia-reperfusion of rat hind limbs.

Ischemia/reperfusion involving the hind limbs of rats results in both local injury to skeletal muscle as well as injury to lungs, as measured by increased vascular permeability (125I-labeled bovine serum albumin leakage) and hemorrhage (extravasation of 51Cr-labeled rat erythrocytes). In the current study, we have focused on events in lungs occurring during reperfusion of hind limbs. Analysis of blood neutrophils obtained 4 hours after reperfusion has indicated up-regulation of CD11b and CD18 but not CD11a. Plasma from the same animals demonstrate the ability to induce similar effects in normal blood neutrophils, indicative of the presence of a neutrophil-activating agent in plasma. During reperfusion, lung injury, which develops progressively over a 4-hour period, has been shown to be neutrophil-dependent and requires CD11a/CD18 and CD11b/CD18 as well as intercellular adhesion molecule-1. These data suggest that ischemia and reperfusion injury of rat lower extremities causes systemic changes that result in neutrophil-dependent lung injury that is beta 2 integrin- (leukocyte function antigen-1, Mac-1) and intercellular adhesion molecule-1-dependent.

Animals↗
PubMed1992

cDNA for Mo3, a monocyte activation antigen, encodes the human receptor for urokinase plasminogen activator.

We have cloned the cDNA for Mo3, an activation Ag expressed by human monocytes and myelomonocytic cell lines after stimulation by PMA, LPS, muramyl dipeptide, certain cytokines, and cAMP agonists. We have previously shown that Mo3 expression in vivo is associated predominantly with macrophages in inflammatory sites. Mo3 is a highly glycosylated protein of about 50 kDa in monocytes and U-937 cells and is anchored to the plasma membrane by glycosyl-phosphatidylinositol linkage. We purified Mo3 protein by cleavage from the U-937 cell surface with phosphatidylinositol-specific phospholipase C, followed by affinity chromatography using a mAb. An internal peptide sequence was determined and used to design oligonucleotide probes for screening an expression cDNA library. Nucleotide sequencing indicated that the complete coding sequence encodes 335 amino acids, including a predicted signal peptide of 22 residues and a hydrophobic C-terminal portion that is probably cleaved during formation of the GPI linkage. The resulting mature protein of about 290 amino acids is consistent with the 29-kDa molecular mass of deglycosylated Mo3. A Northern blot of RNA from U-937 cells revealed a 1.5-kb band that was induced by PMA treatment. Mo3 cDNA was transfected into Cos cells and surface expression of Mo3 was detected by ELISA using various anti-Mo3 mAb. We performed a computer search of the National Biomedical Research Foundation database and found that Mo3 is identical to the human receptor for the urokinase plasminogen activator (uPA-R). Purified soluble Mo3, as well as anti-Mo3 antibodies, were able to block uPA binding to its receptor on U-937 cells, indicating that Mo3 is indeed uPA-R. The use of these anti-Mo3 antibodies may be helpful in assessing the role of uPA-R in processes such as inflammation and tumor invasion.

Amino Acid Sequence↗
PubMed1992

Surface-bound immune complexes trigger transmembrane proximity between complement receptor type 3 and the neutrophil's cortical microfilaments.

To characterize the transmembrane associations participating in antibody-dependent binding, we have used resonance energy transfer (r.e.t.) microscopy to assess the molecular proximity of complement receptor type 3 (CR3) and Fc gamma R type II (Fc gamma RII) and type III (Fc gamma RIII) with microfilaments during neutrophil adherence to untreated surfaces, surfaces coated with BSA, surfaces coated with BSA/anti-BSA F(ab')2 complexes, or surfaces coated with BSA/anti-BSA rabbit IgG immune complexes. Receptors were labeled with fluorescein-conjugated antireceptor Fab fragments, whereas microfilaments were labeled with rhodamine-phalloidin. CR3-to-microfilament r.e.t. was dramatically increased in neutrophils adherent to IgG immune complex-coated surfaces but not untreated or control surfaces. However, the low level of r.e.t. between donor-labeled anti-Fc gamma RII Fab fragments and rhodamine-phalloidin was not affected by any condition including surface-bound immune complexes. Significant r.e.t. levels between Fc gamma RIII and microfilaments were not found under any condition. We suggest that CR3 plays an important role in tethering surface-attached immune complexes to the neutrophil's cytoskeleton.

Actin Cytoskeleton↗
PubMed1992

Roles of beta 2 integrins of rat neutrophils in complement- and oxygen radical-mediated acute inflammatory injury.

The roles of beta 2 integrin molecules in neutrophil accumulation and tissue injury have been examined by the use of antibodies that are reactive with human CD11b and CD18 and cross-react with the homologous epitopes on rat neutrophils. Adherence to rat pulmonary artery endothelial cells by human neutrophils and endothelial cell killing by phorbol ester-activated human neutrophils required CD11b, CD11c, and CD18. Companion adherence studies between rat neutrophils and endothelial cells revealed a requirement for both CD11b and CD18. Neither anti-CD11b nor anti-CD18 depressed in vitro responses (O2- generation and chemotactic migration) of rat neutrophils. The accumulation of neutrophils in glycogen-induced peritoneal exudates was diminished substantially in rats treated with either anti-CD18 or anti-CD11b. In oxidant-mediated acute lung injury induced by rapid intravascular infusion of cobra venom factor, treatment of rats with either anti-CD18 or anti-CD11b significantly attenuated injury as assessed by increases in vascular permeability and hemorrhage. These protective effects correlated morphologically with diminished adhesion of neutrophils to interstitial intrapulmonary capillary endothelial cells. In studies of immune complex (BSA-anti-BSA)-induced alveolitis and dermal vasculitis, anti-CD18 had protective effects at all doses of anti-BSA employed. The protective effects of anti-CD18 correlated with diminished neutrophil accumulation in tissues at lower doses of anti-BSA. Although anti-CD11b was not effective under the same experimental conditions, intratracheal administration of this antibody conveyed protection against immune complex-induced lung injury, suggesting that both CD11b and CD18 are required for the full expression of injury. The current studies also demonstrated that when surface-bound IgG immune complexes were treated with fresh rat serum, the increment in O2- and TNF alpha generated by alveolar macrophages was suppressed by anti-CD18, but not by anti-CD11b, suggesting a heretofore unrecognized role for CD18 in the O2- and TNF-alpha responses of alveolar macrophages. Thus, neutrophil beta 2 integrins play a requisite role for the full expression of complement-dependent and oxygen radical-mediated injury of the lung and dermal vasculature.

Animals↗
PubMed1992

Mesangial cell killing by leukocytes: role of leukocyte oxidants and proteolytic enzymes.

Mesangial cells from human and rat kidney were examined for sensitivity to killing by neutrophils. Cells from both species were sensitive to killing by phorbol myristate acetate-stimulated neutrophils. Catalase was highly protective while superoxide dismutase was less protective and a number of protease inhibitors were not protective. Strong protection was also observed with the iron chelators, deferoxamine and phenanthroline, and with the hydroxyl radical scavengers, dimethylthiourea and 5,5-dimethyl-1-pyrroline N-oxide. Pretreatment of the mesangial cells with deferoxamine followed by washing also provided protection. Mesangial cells were also killed by reagent hydrogen peroxide (H2O2) but were much less sensitive to injury by direct application of proteolytic enzymes. The ability of H2O2 to injure mesangial cells was prevented by pre-incubation of the H2O2 with human leukocyte myeloperoxidase. These data suggest that killing is due primarily to the generation of H2O2 by the stimulated neutrophils and its further reduction in an iron-catalyzed reaction. The hydroxyl radical may be the reduction product that actually mediates lethal injury but lack of scavenger specificity prevents definitively concluding this. Mesangial cell killing by activated neutrophils could be significantly inhibited by monoclonal antibodies to CD11/CD18 molecules, suggesting that close contact between the target and effector cells is required for cytotoxicity. Although qualitatively similar to endothelial cells, the mesangial cells appeared to be quantitatively more oxidant sensitive than previously examined human and rat endothelial cells. Taken together, these data show that mesangial cells from rat and human are sensitive to leukocyte-induced injury and that injury results via an oxidant pathway.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Expression of monocyte activation antigen Mo3 on the surface of peripheral blood monocytes from patients with multiple sclerosis.

Exposure of human peripheral blood monocytes (PBM) to phorbol esters, bacterial products, and cyclic adenosine monophosphate agonists is known to stimulate expression of a plasma membrane antigen ([Ag]; Mo3). Mo3 is recognized by two monoclonal antibodies, Mo3e (IgM), and Mo3f (IgG). Surface Mo3 is barely detectable by indirect immunofluorescence flow cytometry in nonstimulated monocytes. Mo3-positive monocytes have been found in inflammatory tissues, but increased surface expression of Mo3 in PBM has not been seen in any patient group. We report that PBM from patients with chronic progressive MS (CPMS) express increased Mo3. PBM from patients with other neurologic diseases and healthy controls express little measurable Mo3. No difference was seen in class II major histocompatibility complex Ag expression and in Mo2 (CD14) expression. Exposure of PBM to lipopolysaccharide (10 mg/ml) enhanced Mo3 expression in both MS patients and controls. Mo3 expression on CPMS PBM was not dependent on culture conditions. Taken together, our observations suggest that monocytes from patients with MS are stimulated in vivo to express activation Ag Mo3, but that Mo3-positive monocytes need not be upregulated for HLA-DR.

Cell Separation↗
PubMed1992

Modulation and function of intercellular adhesion molecule-1 (CD54) on human retinal pigment epithelial cells.

As part of the blood-retina barrier, the neuroectodermally-derived retinal pigment epithelial (RPE) monolayer is strategically positioned to interact with circulating leukocytes and regulate their access to the retina. We, therefore, studied whether human RPE cells express intercellular adhesion molecule-1 (ICAM-1), a specialized cell surface glycoprotein that binds the leukocyte function antigen-1 receptor present on all leukocytes. Using specific monoclonal antibody to ICAM-1, immunohistochemical staining of freshly-isolated primary and fourth passaged human RPE cells resulted in delicate reaction product that increased dramatically upon exposure to human recombinant (r) interferon-gamma (rIFN-gamma), interleukin-1-beta (rIL-1 beta), or tumor necrosis factor-alpha (rTNF-alpha). Fluorescence-activated cell sorting analysis demonstrated 2-fold increases in constitutive RPE ICAM-1 expression within 6 hours of exposure to physiologic concentrations of rIFN-gamma, rIL-1 beta, or rTNF-alpha. In standardized leukocyte adherence assays, cultured RPE cells showed avid binding of neutrophils that increased significantly after stimulation with rIFN-gamma, rIL-1 beta, or rTNF-alpha (p less than 0.001). In parallel assays, monoclonal antibody to either ICAM-1 on RPE cells, or subunits of leukocyte function antigen-1 receptors on leukocytes significantly blocked leukocyte binding to unstimulated (p less than 0.001) or rIFN-gamma-stimulated RPE cells (p less than 0.001). To demonstrate RPE ICAM-1 expression in intact human tissue, fresh uveoretinal explants were exposed to rIFN-gamma, rIL-1 beta, or rTNF-alpha and stained using mAb to ICAM-1. Tissue sections of cytokine-stimulated explants revealed dramatic increases in RPE ICAM-1 immunoreactivity over the low levels observed in unstimulated uveoretinal tissue. Our results indicate that: (a) ICAM-1 is expressed at low levels on unstimulated RPE cells, (b) RPE ICAM-1 may be augmented by inflammatory cytokines, and (c) RPE ICAM-1 is a functional receptor mediating leukocyte binding. ICAM-1 on RPE cells at the blood-retina barrier may regulate leukocytic infiltration in ocular diseases in which leukocytes are important pathogenetically and may be important to the generation of ocular immune responses.

Cell Adhesion↗
PubMed1992

Differential expression of human corneal and perilimbal ICAM-1 by inflammatory cytokines.

The mechanisms that regulate corneal infiltration by circulating leukocytes in inflammatory diseases are poorly understood. In this study, we investigated the effects of pro-inflammatory cytokines on corneal endothelial (CE) and stromal (CS) expression of intercellular adhesion molecule-1 (ICAM-1), a specialized cell surface glycoprotein that binds the leukocyte function antigen-1 (LFA-1) receptor present on all leukocytes and enhances immune responses. Using specific monoclonal antibody (mAb) to ICAM-1, immunohistochemical staining of intact human corneas resulted in discrete, granular reaction product in CE and CS cells as well as perilimbal vascular endothelium that increased dramatically when exposed to human recombinant interleukin-1-beta (rIL-1 beta), tumor necrosis factor-alpha (rTNF-alpha), and interferon-gamma (rIFN-gamma). Immunoreactive ICAM-1 in CE and CS cells was differentially increased by each of these cytokines. In contrast, immunoreactive endothelial-leukocyte adhesion molecule-1 and vascular cell adhesion molecule-1 were not detected in any CE or CS cells of unstimulated or cytokine-stimulated corneas. In standardized leukocyte adherence assays, neutrophil binding to CE surfaces of whole corneas increased significantly upon exposure to rIL-1 beta, rTNF-alpha, or rIFN-gamma (P less than 0.001). In parallel assays, mAb to ICAM-1 on CE cells or subunits of LFA-1 receptors on leukocytes, but not control mAb, significantly blocked leukocyte binding to unstimulated (P less than 0.01) or rIFN-gamma-stimulated corneas (P less than 0.001). Our results indicate that: (1) ICAM-1 is expressed at low levels on unstimulated CE cells, CS cells, and perilimbal vascular endothelium; (2) ICAM-1 may be augmented differentially in corneal and perilimbal tissue by pro-inflammatory cytokines; and (3) ICAM-1 is a functional ligand mediating corneal-leukocyte binding. Differential expression of ICAM-1 within corneal tissue may regulate keratitic precipitate formation, leukocyte trafficking and accumulation, and localized generation of immune responses.

Cell Adhesion Molecules↗
PubMed1991

Regulation of transendothelial neutrophil migration by endogenous interleukin-8.

Movement of neutrophils from the bloodstream to inflamed tissue depends on the activation of both the neutrophil and the endothelial cell. Endothelial cells lining the postcapillary venule respond to proinflammatory mediators by expressing adhesion molecules and synthesizing a variety of neutrophil-activating factors. Endothelial cell production of a 77-amino acid variant of interleukin-8 (IL-8) was found to be a requirement for the invasion of neutrophils through a vessel wall model. IL-8 secreted by cytokine- or lipopolysaccharide-stimulated endothelial cells induced the rapid shedding of neutrophil lectin adhesion molecule-1, the up-regulation of leukocyte beta 2 integrins, and the attachment and transmigration of the neutrophils. Thus, endogenous endothelial IL-8 regulates transvenular traffic during acute inflammatory responses.

Antigens, CD↗
PubMed1991

Purification, biochemical composition, and biosynthesis of the Mo3 activation antigen expressed on the plasma membrane of human mononuclear phagocytes.

Mo3 is an activation Ag expressed on the surface of human mononuclear phagocytes stimulated in vitro or in vivo by various activating factors. Mo3 is obtained by immunoprecipitation with anti-Mo3 mAb from lysates of PMA-stimulated U-937 cells. The Ag is a heterogeneous glycoprotein with a molecular mass range of 42 to 66 kDa (nonreducing conditions) containing N-linked carbohydrate chains. When the cells are treated with phosphatidylinositol-specific phospholipase C, greater than 60% of total precipitable gp42-66 Ag is released in the supernatant. This phosphatidylinositol-specific phospholipase C-sensitive linkage to the plasma membrane has provided a means for the one-step purification of Mo3 by immunoaffinity chromatography. The eluted soluble Mo3 (sMo3) was greater than 90% pure as documented by the appearance of a single major protein peak on reverse phase HPLC and SDS-PAGE. The average yield was 12.1 micrograms/10(8) cells. Sufficient quantities of sMo3 have been purified to permit the determination of amino acid and carbohydrate composition. Complex N-linked carbohydrates make up nearly 50% of the glycoprotein content and contribute to its heterogeneity. An anti-Mo3 polyclonal antiserum generated from sMo3 was used to immunoprecipitate Mo3 and its precursor from biosynthetically labeled, PMA-stimulated U-937 cells or LPS-stimulated monocytes. These 35S-methionine "pulse-chase" experiments demonstrated the existence of a 40- to 42-kDa endo-beta-N-acetylglucosaminidase-sensitive precursor, which over a period of 4 to 5 h gave rise to an endo-beta-N-acetylglucosaminidase-resistant, but N-glycanase-sensitive 42- to 66-kDa mature form.

Amino Acids↗