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Biomedical subjects

W D Ratnoff

Publications and source records attributed to W D Ratnoff.

At least 19 recordsLinked to original sources
PubMed1998

Detection of a soluble form of B7-1 (CD80) in synovial fluid from patients with arthritis using monoclonal antibodies against distinct epitopes of human B7-1.

The costimulatory molecule B7-1 (CD80) has been shown to be an important component for T cell immune responses. We have generated several monoclonal antibodies (PSRM-1, -2, -3, -6, and -7) against B7-1 using a human glycosylphosphatidylinositol-anchored B7-1 (GPI-B7-1) as an antigen. These monoclonal antibodies are able to detect B7-1 by flow cytometry, ELISA, and Western blotting. One antibody in particular, PSRM-3, blocks the CD28/CTLA-4 interaction with B7-1 and consequently blocks costimulation of T cells. The other PSRM monoclonal antibodies did not compete with PSRM-3 for recognition of B7-1 and also failed to block B7-1 interaction with CTLA-4 and CD28, indicating that these antibodies bind to different epitopes. PSRM-3 and -7 detect phosphatidylinositol-specific phospholipase C-released soluble GPI-B7-1 in a sandwich ELISA. We used this sandwich ELISA to assay for the presence of a soluble form of B7-1 in synovial fluids of arthritis patients. By sandwich ELISA, B7-1 was detected in the synovial fluid of 5/11 patients with rheumatoid arthritis, 5/5 patients with osteoarthritis, and 2/6 patients with other forms, including crystalline-induced arthritis. The presence of soluble B7-1 was confirmed by immunoprecipitation using PSRM-3-coupled Sepharose beads. The source and function of soluble B7-1 are unknown at present; it is possible, however, that the soluble form of B7-1 molecule may play a local immunoregulatory role which may suppress or induce inflammation depending upon whether it interacts with the T cell costimulatory CD28 molecule or the negative signaling CTLA-4 molecule.

Abatacept↗
PubMed1996

Inherited deficiencies of complement in rheumatic diseases.

Inherited complement deficiencies are rare but occur with increased frequency in systemic lupus erythematosus. This selective review summarizes the fundamentals of complement activation and biology and focuses on recent findings. system lupus erythematosus is highly associated with homozygous C1q deficiencies and with C4A null allotypes, and homozygous C2 and C3 deficiencies are less uniformly associated with autoimmune disease. This hierarchy of disease associations suggests the importance of an intact early classic activation pathway in preventing systemic lupus erythematosus. Assays of complement activation products may have greater sensitivity and specificity for systemic lupus erythematosus disease activity and lupus flares.

Autoimmune Diseases↗
PubMed1995

Immunohistochemical localization of C9 neoantigen and the terminal complement inhibitory protein CD59 in human endometrium.

PROBLEM: Human endometrium expresses complement components, receptors, and regulatory proteins, many of which appear to be expressed in a hormone-dependent manner. Whether terminal complement components are also present in the endometrium is unknown. CD59, a broadly expressed protein that blocks association of C9 with C8 in the membrane attack complex, is localized in reproductive tissue to human spermatozoa, seminal plasma, amniotic fluid, and placenta. The present study examines human endometrium for the presence of CD59 and terminal complement proteins. METHOD: Endometrial biopsies were obtained from six normal women from various phases of the menstrual cycle and analyzed by immunohistochemistry, using MEM-43 anti-human CD59 and anti-human SC5b-9 murine monoclonal antibodies and the immunoperoxidase technique. RESULTS: Both CD59 protein and SC5b-9 (C9 neoantigen) were demonstrated to be present in endometrial glandular epithelium throughout the menstrual cycle. No specific staining was demonstrated in the stromal compartment. CONCLUSION: CD59 protein and terminal complement proteins are expressed in glandular epithelial cells of normal human endometrium, in both proliferative and luteal phases, suggesting that expression is not hormonally dependent. These analyses further support the presence of a functionally active complement system in normal human endometrium.

Antigens↗
PubMed1992

Effect of glycoinositolphospholipid anchor lipid groups on functional properties of decay-accelerating factor protein in cells.

The inositol ring in the glycoinositolphospholipid (GPI) anchor of human decay-accelerating factor (DAF) is unmodified in nucleated cells, whereas it is fatty acid acylated in erythrocytes (Ehu). To assess the effect of this and of the glycerol sn-2-associated acyl substituent on the abilities of DAF to cell membrane incorporate and function, 1) endogenous (physiologically anchored) DAF proteins bearing three- and two-"footed" GPI anchors were purified from Ehu and HeLa cells and 2) synthetic DAF variants bearing alternative one- "footed" anchors (retaining either the sn-1 glycerol- or inositol-associated lipid) were prepared by alkaline hydroxylamine treatment and phosphatidylinositol-specific phospholipase D digestion of Ehu DAF, respectively. The different DAF species were added to antibody-sensitized sheep erythrocytes (EshA) and their abilities to insert into the plasma membranes of the cells and control subsequent complement activation on their surfaces were compared. DAF proteins bearing all four GPI anchor structures adhered to the Esh hemolytic intermediates and inhibited expression of C3 convertase (C4b2a) activity. However, mixing of DAF-treated EshA with untreated EshAC142 and stripping of cell-associated DAF proteins with vesicles showed that only the physiologically anchored proteins remained stably associated with the lipid bilayer and functioned intrinsically. Both three- and two-"footed" Ehu and HeLa DAF proteins exhibited comparable ability to incorporate and function in the intermediates as well as to accumulate to levels 1000-fold higher/cell in Schistosoma mansoni schistosomula. These findings indicate that 1) an intact inositolphospholipid-containing GPI anchor is necessary for stable membrane integration and intrinsic function, 2) endogenous GPI anchors (with either unsubstituted and acylated inositol) incorporate and function with comparable efficiency, and 3) the transfer of either endogenous DAF form can account for the previously described circumvented uptake of human C3b by blood stage schistosomula.

Blood Proteins↗
PubMed1992

Structural properties of the glycoplasmanylinositol anchor phospholipid of the complement membrane attack complex inhibitor CD59.

CD59, the membrane regulator of autologous C5b-9 channel formation, exhibits variable sensitivity to cleavage by phosphatidylinositol-specific phospholipase C (PI-PLC), an enzyme that releases glyco-inositolphospholipid (GPI)-anchored proteins from cell surfaces. To determine whether the GPI-anchor phospholipid of CD59 is similar to that of decay-accelerating factor (DAF) and whether variation in its structure underlies its variable enzyme susceptibility, the GPI anchors of the two proteins expressed on erythrocytes, polymorphonuclear and mononuclear leucocytes were compared in situ and after purification. Flow cytometric analyses of PI-PLC-treated cells showed parallel cell type specific release of both proteins as a function of enzyme concentration. Non-denaturing PAGE analyses of alkaline/hydroxylamine-treated proteins (affinity-purified from [125I]-surface-labelled cells) provided evidence for (i) comparable proportions of GPI-anchor acylation, and (ii) alkali-resistant rather than alkali-sensitive lipid substituents in erythrocytes. These findings argue that the differential C5b-9 sensitivity that distinguishes paroxysmal nocturnal haemoglobinuria II and III erythrocytes does not derive from expression of CD59 molecules with alternative GPI-anchor phospholipid structures.

Antigens, CD↗
PubMed1990

Structural basis for variations in the sensitivity of human decay accelerating factor to phosphatidylinositol-specific phospholipase C cleavage.

Human decay-accelerating factor (DAF) proteins expressed on E and nucleated cells differ in their susceptibility to phosphatidylinositol (PI)-specific phospholipase C (PLC) cleavage/release. To investigate the mechanism of this difference, the glycoinositol-phospholipid anchoring moieties of E DAF, and of HeLa cell, polymorphonuclear cell, and lymphocyte DAF were structurally compared. Labeling of PI-PLC-resistant E DAF with 3-(trifluoromethyl)-3-(m-[125I]-iodophenyl)-diazirine ([125I]TID) and TLC analysis of nitrous acid deamination anchor fragments showed a predominant phospholipid species with less polar migration than the 125I-TID-labeled PI. Gas chromatographic analyses of methanolyzed E protein revealed 2.20 +/- 0.16 mol of fatty acids [16:0, 18:0, 18:1, 20:4, 22:4, and 22:5 (0.76, 0.36, 0.25, 0.15, 0.40, 0.28 mol, respectively)] and 0.86 +/- 0.05 mol of inositol per mol of N-terminal Asp. Gas chromatography-mass spectroscopy demonstrated principally myo-inositol but also variable amounts of the chiro-isomer. Nondenaturing polyacrylamide gel electrophoresis of 14C-radiomethylated E protein revealed that pretreatment with hydroxylamine, a reagent which removes ester-linked lipids, rendered it PI-PLC susceptible. In contrast, parallel analyses of 35S-cys-labeled PI-PLC-sensitive HeLa DAF protein revealed only minor amounts of the hydroxylamine-sensitive phospholipid species. Similar results were obtained with 125I-surface-labeled DAF from polymorphonuclear cells, as well as from unstimulated peripheral blood and anti-CD3-activated lymphocytes. These findings demonstrate that, rather than PI, the E DAF anchor contains an inositol alkylacylglycerol-phospholipid which is heterogeneous with respect to acyl groups and inositol isomers, that an ester-linked substitution in this inositolphospholipid underlies the resistance of E DAF protein to PI-PLC cleavage/release, and that this structural modification is cell-specific.

CD55 Antigens↗
PubMed1989

Late appearance of phospholipid platelet-activating factor and leukotriene B4 in human skin after repeated antigen challenge.

Inflammatory mediators were assessed in supernatants of chamber fluids from eight ragweed- or grass-sensitive subjects during antigen-induced cutaneous inflammatory responses. Platelet activating factor (PAF) accumulated at concentrations of 1 pm to 90 mumol/L in six of eight subjects beginning at 3 hours and continuing for 9 hours after antigen challenge. Leukotriene B4 (LTB4) was detectable at cutaneous sites of antigen challenge in five of five subjects throughout the 9-hour period at levels from 1 to 36 nmol, a range of 38% to 80% of which were omega-oxidation metabolites. Histamine levels peaked in the first hour at 106 +/- 18 ng/ml and decreased to a plateau of 11 to 13 ng/ml at 3 to 9 hours after antigen challenge. No PAF and only very low levels of LTB4 (0.1 to 1.3 nmol) and of histamine (less than 2 ng/ml) were detected at buffer-control sites during the 9 hours of study. Continuous antigen exposure thus results in the persistent release of histamine and LTB4 and the late appearance of PAF, all of which may contribute to the chronicity of allergic disorders and may have a bearing on the IgE-mediated, late-phase cutaneous response.

Adult↗
PubMed1989

Identification of leukotriene B4 as the neutrophil chemotactic factor released by antigen challenge from passively sensitized guinea pig lungs.

Neutrophils are prominent in some IgE-mediated allergic reactions and may contribute to the pathophysiology of immediate hypersensitivity. Antigen challenge of fragments of guinea pig lung tissue that were passively sensitized with IgE or IgG antibody evoked the release of neutrophil chemotactic activity (NCA) in parallel with histamine. The NCA released from lung tissue by both IgG- and IgE-dependent stimulation coeluted from a column of Sephacryl S-300 with synthetic leukotriene B4 (LTB4). The NCA in eluates from the Sephacryl S-300 column contained LTB4, as determined by high-performance liquid chromatography and specific radioimmunoassay, in quantities that accounted for the observed chemoattractant activity in the eluates. Furthermore, the NCA of supernatants from antigen-challenged lung fragments was reduced by a mean of 80% after absorption with a monoclonal antibody to LTB4. LTB4 thus constitutes the major functional constituent of NCA released after anaphylactic challenge of IgE- and IgG-sensitized guinea pig lung tissue.

Anaphylaxis↗
PubMed1988

Sulfidopeptide-leukotriene peptidases in pulmonary edema fluid from patients with the adult respiratory distress syndrome.

The human pulmonary edema fluid concentrations of LTC4 and of LTD4 and LTE4, derived peptidolytically from LTC4, were assessed by radioimmunoassays of the mediators resolved by reverse-phase high-performance liquid chromatography. The mean pulmonary edema fluid concentration (+/- SD) of LTD4 of 19.2 +/- 25.6 nM for 12 patients with the adult respiratory distress syndrome and of LTE4 of 192 +/- 309 nM for 10 of the patients were significantly higher (P less than 0.005 and P less than 0.05) than those of 2.2 +/- 2.4 and 11.0 +/- 18.2 nM, respectively, for 10 patients with cardiogenic pulmonary edema, whereas the lower mean concentrations of LTC4 were not significantly different for the two groups. Pulmonary edema fluid from five patients with adult respiratory distress syndrome, one with cardiogenic pulmonary edema, and one with an indeterminate syndrome contained similar concentrations of peptidoleukotriene peptidases. The LTC4 and LTD4 peptidolytic activities in ARDS fluids were 81 and 142 kD, respectively, by gel filtration. The extents of peptidolysis of [3]LTC4 and [3]LTD4 by 100 microliter of pulmonary edema fluid attained respective mean maximum levels of 74.5 +/- 2.9% (N = 5) and 37.7 +/- 10.2% (N = 4) after 30 min at 37 degrees C and were inhibited by serine-borate and by cysteine, respectively. The predominance of LTD4 and LTE4 over LTC4 in states of altered pulmonary vascular pressure and permeability thus is attributable to two distinct peptidases.

Adolescent↗
PubMed1988

Distinctive patterns of release of neuroendocrine peptides after nasal challenge of allergic subjects with ryegrass antigen.

The concentrations of the neuropeptides substance P, somatostatin, and calcitonin gene-related peptide in human nasal secretions were quantified by radioimmunoassays, concurrently with that of histamine, in the course of nasal challenge of allergic and control subjects with ryegrass antigen to examine contributions of neuromediation of the tissue response. Each of the neuropeptides and histamine were detected in nasal lavage fluid prior to challenge. In allergic patients, but not normal controls, antigen evoked significant increases of 3-fold in histamine at 15-60 min, 1.5- to 4-fold in calcitonin gene-related peptide at 15 min-24 hr, and more than 2-fold in somatostatin at 6 hr, without altering the concentration of substance P in nasal lavage fluid. The identity of the neuropeptides was confirmed chromatographically. Thus calcitonin gene-related peptide may mediate nasal congestion directly and somatostatin may be one of the factors regulating the late involvement of basophils and mast cells in allergic rhinitis.

Administration, Intranasal↗
PubMed1988

Decay-accelerating factor protects human tumor cells from complement-mediated cytotoxicity in vitro.

The disialoganglioside GD2 is expressed on a wide spectrum of human tumor types, including neuroblastomas and melanomas. Upon binding of 3F8, a murine monoclonal antibody (MAb) specific for GD2, neuroblastomas and some melanomas are sensitive to killing by human complement, whereas some melanomas are not. To investigate the mechanism underlying these differences in complement mediated cytotoxicity, complement-insensitive melanoma cell lines were compared with respect to expression of the decay-accelerating factor (DAF), a membrane regulatory protein that protects blood cells from autologous complement attack. While DAF was undetectable among neuroblastomas, it was present in complement-insensitive melanomas. When the function of DAF was blocked by anti-DAF MAb, C3 uptake and complement-mediated lysis of the insensitive melanoma lines were markedly enhanced. F(ab')2 fragments were as effective in enhancing lysis as intact anti-DAF MAb. The DAF-negative and DAF-positive melanoma cell lines were comparably resistant to passive lysis by cobra venom factor-treated serum. The data suggest that in some tumors, DAF activity accounts for their resistance to complement-mediated killing. The ability to render these cells complement-sensitive by blocking DAF function may have implications for immunotherapy.

Antibodies, Monoclonal↗
PubMed1986

Decreased expression of the C3b/C4b receptor (CR1) and the C3d receptor (CR2) on B lymphocytes and of CR1 on neutrophils of patients with systemic lupus erythematosus.

Decreased numbers of complement receptor type 1 (CR1) have been observed on erythrocytes of patients with systemic lupus erythematosus (SLE) and on glomerular podocytes of patients having proliferative nephritis of SLE. In the present study, the analysis of the cellular expression of CR1 has been extended to include leukocytes. In addition, expression by B lymphocytes of the C3d receptor (CR2), which also serves as the receptor for the Epstein-Barr virus, was assessed. Receptor expression was measured by 2-color fluorescent flow cytometry of peripheral blood B cells, identified by the presence of the B1 antigen, that had also been stained with anti-CR1 or anti-CR2. B cells from 17 patients with SLE exhibited a mean relative fluorescence for CR1 that was 61% of that found in 17 normal individuals (P less than 0.001). The expression of CR2 by the patients' B cells (n = 14) was 62% of that of the B cells from normal subjects (n = 17) (P less than 0.001). The expression of CR1 correlated with that of CR2 among patients (r = 0.63; P less than 0.01) but not with the expression of CR2 among normal individuals (r = 0.36; P greater than 0.1). The mean CR1 content of the patients' neutrophils was only 59% of the normal mean (P less than 0.001). Thus, abnormalities of complement receptor expression occur on the leukocytes of patients with SLE. These deficiencies may be secondary to interaction of the cells with the products of complement activation, or, in some individuals, the deficiencies may be familial.

Antibodies, Antinuclear↗
PubMed1980

Activation of the alternative complement pathway by Histoplasma capsulatum.

The present study was performed to determine whether Histoplasma capsulatum (yeast phase) is able to activate the alternative complement pathway. H. capsulatum yeasts were shown to consume C3 in C4-deficient guinea pig serum. Immunoelectrophoretic conversion of alternative pathway component factor B confirmed that C3 consumption was mediated by activation of the alternative pathway. These results demonstrate that H. capsulatum is able to activate the alternative complement pathway.

Animals↗
PubMed1980

The rabbit properdin system: I. Identification of a new factor and purification of rabbit properdin.

Pure preparations of rabbit properdin were obtained from rabbit serum by ion-exchange chromatography. These preparations functioned as properdin when they were measured with the zymosan assay or with a serum reagent selectively depleted of properdin by a specific immunoabsorbent. Properdin in these preparations was in its activated state. A new serum factor was required to measure properdin activity when purified preparations of rabbit properdin were tested with the zymosan assay. This factor was designated as ZBP, or zymosan-binding protein. ZBP appeared to be distinct from known components of the alternative complement pathway and the classical complement system, and it did not appear to be an immunoglobulin.

Animals↗
PubMed1980

The familial occurrence of polycythemia vera: report of a father and son, with consideration of the possible etiologic role of exposure to organic solvents, including tetrachloroethylene.

The occurrence of polycythemia vera in a father and son, both of whom had intermittent exposure to organic solvents, including tetrachloroethylene and Stoddard solvent, is reported. Only three other well substantiated familial occurrences of polycythemia vera, none encompassing successive generations, were found among many reported instances.

Adult↗