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Biomedical subjects

A Nicholson-Weller

Publications and source records attributed to A Nicholson-Weller.

At least 37 records · Page 2Linked to original sources
PubMed1994

Terminal complement proteins C5b-9 release basic fibroblast growth factor and platelet-derived growth factor from endothelial cells.

Interactions between endothelium and vascular smooth muscle cells play a major role in the biology of the blood vessel wall. Growth factors released from endothelial cells control in part the normal and pathological proliferation of vascular smooth muscle cells. Endothelial deposits of C5b-9 proteins, the membrane attack complex of complement (MAC), have been found in a variety of pathological tissues in which cell proliferation is an early characteristic abnormality, including atherosclerosis. We have explored a possible bridging role for terminal complement C5b-9 proteins in eliciting focal signals for cell proliferation by releasing growth factors from endothelial cells. We found that both bovine aortic and human umbilical vein cells respond to the MAC by releasing basic fibroblast growth factor and platelet-derived growth factor. These mitogens stimulate DNA synthesis in Swiss 3T3, vascular smooth muscle, and glomerular mesangial cells. Based on these findings, we propose that complement-induced release of mitogens from endothelial cells is a novel pathogenic mechanism for proliferative disorders.

Animals↗
PubMed1994

Structure and function of decay accelerating factor CD55.

Studies of decay-accelerating factor (DAF) function and structure are reviewed. DAF was first recognized as a species restricting factor operating at the level of C3/C5 activation. Cloning of the gene indicates that DAF has four short consensus repeats of the type characteristic of the regulators of complement activation gene cluster family. The third short consensus repeat is responsible for DAF's complement regulatory activity and signaling. DAF, like other glycophosphatidylinositol (GPI) anchored proteins, is associated with tyrosine kinases, and these kinases are probably the signaling devices. The details of how DAF's GPI anchor in the outer leaflet of plasma membrane connects with the tyrosine kinases on the inner leaflet are not known. Although DAF does not have an essential role in controlling hemolysis of erythrocytes, it does have important role in regulating the deposition of C3 on nucleated cells. The therapeutic potential of DAF is discussed.

Animals↗
PubMed1993

Molecular cloning and characterization of human eosinophil Charcot-Leyden crystal protein (lysophospholipase). Similarities to IgE binding proteins and the S-type animal lectin superfamily.

We have isolated and sequenced a 598-bp full length cDNA clone for the human Charcot-Leyden crystal (CLC) protein (eosinophil lysophospholipase), the unique and prominent constituent of human eosinophils and basophils that forms the hexagonal bipyramidal crystals classically observed in tissues and secretions from sites of eosinophil-associated inflammation. A 426-bp open reading frame encoded a 142-amino acid polypeptide with a predicted molecular mass of 16.5 kDa and isoelectric point of 7.28. The deduced amino acid sequence of CLC protein showed 20 to 30% similarity over regions of approximately 100 amino acids with the carboxyl-terminal domains of four IgE-binding proteins, including the 31-kDa human and rat IgE-binding proteins, the 35-kDa mouse carbohydrate binding protein (CBP35), Mac-2, the murine macrophage cell surface protein that is identical to CBP35, and the human homologue of Mac-2. These proteins are members of a superfamily of beta-galactoside binding S-type animal lectins, which includes a group of highly conserved 14-kDa lectins isolated from human lung, heart, placenta, bovine heart, chicken skin, mouse fibroblasts, and the electric organ of the electric eel; CLC protein also showed sequence similarities to these 14-kDa animal lectins, including conservation of 7 of 16 invariant amino acid residues thought to comprise the carbohydrate-binding domain of these proteins, with conservative amino acid changes at others; thus, CLC protein could potentially possess carbohydrate or IgE-binding activities. Northern analyses revealed an approximately 900-bp mRNA species that was present in peripheral blood eosinophils from patients with eosinophilia, basophils from patients with chronic myelogenous leukemia, and in HL-60 cells induced towards eosinophilic differentiation with B cell growth factor-II (IL-5) or granulocytic differentiation with DMSO, but was absent in neutrophils, monocytes, T cells, B cells, or HL-60 cells induced towards monocytic differentiation with vitamin D3. Southern analyses revealed a gene of approximately 5 to 6 kb in length. The cDNA clone and complete amino acid sequence data for CLC protein will facilitate structure-function analyses of its unusual hydrophobic properties, unique propensity for crystallization, lysophospholipase, and potential lectin-like activities.

Amino Acid Sequence↗
PubMed1993

Transient changes in erythrocyte membrane permeability are induced by sublytic amounts of the complement membrane attack complex (C5b-9).

We have previously shown that sublytic heterologous complement induces large but transient increases in erythrocyte membrane permeability. We now report that when erythrocytes are bystanders in zymosan-activated autologous serum, they increase their Na+ permeability 10-fold, indicating that autologous complement can also induce transient membrane lesions. When we isolated the effect of the C5b-9 membrane attack complex of complement by using human C5b-9 assembled from purified components, we found there was minimal lysis but efficient Na+ uptake. Suspension of the sublytically damaged erythrocytes in K+ medium caused the cells to lyse, which is consistent with the cells recruiting a compensatory K+ efflux similar to that observed when human erythrocytes were exposed to heterologous complement. Sublytic C5b-9 exposure also became lytic when extracellular Ca2+ was limited and when the cells were exposed to charybdotoxin, an inhibitor of the Ca(2+)-activated K+ channel. This indicates that Ca2+ is required for the functional termination of the C5b-9 lesion. We also show that the membrane hyperpolarization resulting from activation of the Ca(2+)-dependent K+ efflux does not influence the termination of the C5b-9 lesion. Thus, the influx of Ca2+ through the complement lesion initiates at least two apparently independent adaptive responses: (1) a process that terminates the leak; and (2) a K+ efflux that has a volume regulatory function. Our data support the potential of the sublytic C5b-9 lesion to act as a physiologic mediator for autologous erythrocytes.

Calcium↗
PubMed1993

Membrane signaling by complement C5b-9, the membrane attack complex.

The terminal complement complexes C5b-7, C5b-8 and C5b-9 are able to generate nonlethal cell signals. One universal consequence of a cell being targeted by C5b-8 or C5b-9 is an influx of Ca2+. In addition, other second messengers, including cAMP, inositol phosphate intermediates and arachidonate metabolites, are generated by the terminal complement complexes in specific cell types. In vivo, terminal complement complexes have been found in a wide variety of inflammatory processes in humans and in experimental animal models. Some of these models of inflammation putatively induced by terminal complement complexes have been tested in complement-deficient animals, and indeed no inflammation results, which supports the critical role of the terminal complement complexes in the pathogenesis of the lesion.

Animals↗
PubMed1993

Terminal complement complex C5b-9 stimulates mitogenesis in 3T3 cells.

The membrane attack complex of complement (MAC) can induce reversible changes in cell membrane permeability resulting in significant but transient intracellular ionic changes in the absence of cell lysis. Because ion fluxes and cytosolic ionic changes are integral steps in the signaling cascade initiated when growth factors bind to their receptors, we hypothesized that the MAC-induced reversible changes in membrane permeability could stimulate cell proliferation. Using purified terminal complement components we have documented a mitogenic effect of the MAC for quiescent murine 3T3 cells. The MAC enhances the mitogenic effects of serum and PDGF, and also stimulates cell proliferation in the absence of other exogenous growth factors. MAC-induced mitogenesis represents a novel effect of the terminal complement complex that could contribute to focal tissue repair or pathological cell proliferation locally at sites of complement activation.

3T3 Cells↗
PubMed1992

Decay-accelerating factor expression on either effector or target cells inhibits cytotoxicity by human natural killer cells.

Previous studies have shown that freshly isolated CD16+ NK cells are deficient in the expression of decay-accelerating factor (DAF), or CD55, a membrane regulator of C3 activation. In this study we investigated the significance, for NK cell-mediated lysis, of DAF expression on the target and effector cells. The effect of DAF expression on the susceptibility of NK cell targets was investigated by several means: first, DAF- cell lines were cloned from K562; second, the cloned DAF- cells were reconstituted with exogenous purified DAF; and third, anti-DAF F(ab')2 was used to block DAF function on the DAF+ K562 cells. Consistently, the presence of DAF in the target cell membrane, either naturally occurring or experimentally incorporated, afforded the target cell protection against lysis, and this protection could be blocked with anti-DAF. Similarly, targets for antibody-dependent cell-mediated cytotoxicity with exogenous DAF incorporated in their plasma membrane became less sensitive to antibody-dependent cell-mediated cytotoxicity by NK cells compared with the same target cells without incorporated DAF in their membranes. DAF incorporated in the plasma membranes of the effector NK cells made the NK cells less effective at killing K562 targets. The known function of DAF is to regulate C3 activation, and we were able to demonstrate that the isolated NK cell is capable of releasing C3. It is also possible that the participation of DAF in NK cell function represents a new, noncomplement-dependent function for DAF.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1992

Defective glycosylphosphatidylinositol anchor synthesis in paroxysmal nocturnal hemoglobinuria granulocytes.

To investigate the biosynthesis of the glycosylphosphatidylinositol (GPI) anchor in the granulocytes of paroxysmal nocturnal hemoglobinuria (PNH), the glycolipids of granulocytes from PNH patients and normal volunteers were biosynthetically labeled with [3H]mannose in the presence of tunicamycin. Extracted glycolipids were examined by thin-layer chromatography and compared with known biosynthetic intermediates. Normal granulocytes consistently showed [3H]mannose incorporation into the complete GPI core, several GPI biosynthetic intermediates, and dolichol phosphate mannose (DPM). The granulocytes of 10 patients with PNH that had no expression of CD55 and CD59 on greater than 95% of the cells were able to incorporate [3H]mannose into DPM, but were not able to incorporate detectable amounts into the complete GPI core. THus, PNH granulocytes do not synthesize detectable amounts of the complete GPI core and this defect likely accounts for the absence of GPI-linked membrane proteins on hematopoietic cells in this syndrome.

Antigens, CD↗
PubMed1992

Paramyosin inhibits complement C1.

We report here the results of studies showing that inhibition of C is a property of several invertebrate paramyosins. Paramyosins from Taenia solium, Schistosoma mansoni, and the mussel Mytilus edulis bind polymeric collagen and can be isolated from crude extracts of tissues by collagen affinity. These paramyosins inhibit C1 function whether the C1 is isolated or present in C2-deficient serum. Because T. solium paramyosin was the best inhibitor, we concentrated further studies on this molecule. T. solium paramyosin binds purified C1q in solution with a dose/response similar to C1r2S2. Further studies of the C1-paramyosin interaction indicate that: 1) C4 is not activated, 2) C4b2a decay is not affected, and 3) there is no effect on the efficiency of C3-9, as provided in EDTA-chelated guinea pig serum, in lysing SRBC. Thus, paramyosin inhibition is directed at the initiation of the classical pathway. The results suggest that paramyosins of helminthic parasites may have a role as modulators of the host immune response through C inhibition at C1.

Animals↗
PubMed1990

Homologous species restriction of the complement-mediated killing of nucleated cells.

The homologous restriction of complement (C) lysis is attributed to membrane proteins: decay-accelerating factor (DAF), C8 binding protein (C8bp) and P18/CD59. Since these proteins are also expressed on peripheral blood cells, species restriction was tested for in the complement-mediated killing of antibody-coated human leucocytes by human or rabbit complement. Killing was more efficient when rabbit complement was used. Preincubation of cells with an antibody to DAF abolished the difference. When C1-7 sites were first attached to the cells and either rabbit or human C8, C9 were added, the killing of monocytes and lymphocytes was equally efficient; only in polymorphonuclear neutrophils was a higher efficiency of rabbit C8, C9 seen. Thus, in contrast to haemolysis, restriction occurred predominantly at the C3 level and the action of the terminal complement components was not inhibited. Since C8bp isolated from peripheral blood cells showed essentially similar characteristics as the erythrocyte-derived C8bp, the failure of C8bp to inhibit the action of the terminal components on nucleated cells might reflect differences of the complement membrane interactions between erythrocytes or nucleated cells, respectively.

Animals↗
PubMed1989

Factor J: isolation and characterization of a new polypeptide inhibitor of complement C1.

An Mr 20,000 protein inhibitor of C1, the first component of complement, has been purified from human urine and characterized. This inhibitor, tentatively designated factor J, is apparently distinct from known complement inhibitors. During purification on QAE-Sephadex, Mono Q, and heparin-Sepharose, factor J was detected by its ability to inhibit the complement-mediated lysis of sheep erythrocytes bearing antibody, C1, and activated C4 (EAC14). The purity of factor J was documented by the concordant elution from a hydroxylapatite column of functional activity and the UV absorbance as measured at three different wavelengths (220, 254, and 280 nm). The relative Mr of 20,000 was determined by sodium dodecyl sulfate-slab gel electrophoresis of radioiodinated protein. Amino acid analysis indicated a high cysteine content and allowed calculations of a specific activity of 7 functional units/pmol. The target of factor J inhibitory activity on the lysis of EAC14 was localized to C1 by the following criteria: factor J inhibited C1 in a C1 transfer assay, but had no effect on C42 activity or decay, and had no effect on the efficiency of isolated C2 or C3-C9 as provided in serum-EDTA. Factor J inhibition was rapid and not significantly influenced by temperature. In a second functional assay, factor J inhibited the association of the tetrameric complex C1r2s2 with 125I-C1q, and the results, when analyzed graphically by a reciprocal plot, were consistent with noncompetitive inhibition (Ki = 529-760 pM range). Functional and/or antigenic data indicated that factor J is distinct from the other known inhibitors of C1, namely the C1 inhibitor and the C1q inhibitor. Antihuman serum precipitated radioiodinated factor J, indicating that an antigen identical or cross-reacting with factor J exists in serum. In summary, factor J is a newly described potent inhibitor of C1 function.

Animals↗
PubMed1989

Decay accelerating factor regulates complement activation on glomerular epithelial cells.

Epithelial cells of the glomerular capillary are the site of C5b-9 mediated injury in rat membranous nephropathy. We investigated the regulation of C activation by cultured glomerular epithelial cells (GEC). Rat and human GEC were more resistant to C injury by homologous C than heterologous C. In human GEC homologous C cytotoxicity was enhanced by antiserum to decay accelerating factor (DAF) indicating that homologous C activation was, at least in part, restricted by membrane DAF. Anti-DAF immunoprecipitated a 67-kDa protein from human glomeruli. In rat GEC, pronase and phosphatidylinositol-specific phospholipase C (which are known to inactivate human DAF) enhanced cytotoxicity by homologous C. Thus, DAF is present on human GEC in culture and in human kidney glomeruli, and a DAF-like protein is present on cultured rat GEC. These proteins regulate C activation in vitro and may play a role in controlling C activation on GEC in vivo.

Animals↗
PubMed1989

Human eosinophils express CD4 protein and bind human immunodeficiency virus 1 gp120.

The CD4 glycoprotein, expressed on leukocytes belonging to subsets of T lymphocytes and to cells of monocyte/macrophage lineage, participates in the functioning of T cells and serves as a receptor for HIV-1 and HIV-2. Human eosinophils, a class of granulocytic leukocytes, have been found to express CD4. With anti-CD4 mAbs CD4 was demonstrable on eosinophils from both normal and eosinophilic donors. Eosinophils synthesized a 55-kD CD4 polypeptide immunoprecipitable with two anti-CD4 mAbs. Eosinophil CD4 bound HIV-1 gp120 as assessed by competition for anti-OKT4A, but not anti-OKT4, mAb binding. Eosinophils, normally rich in gastrointestinal and genitourinary tract tissues, increase in numbers in patients with metazoan parasitic infections. In these sites and diseases, CD4 expression by eosinophils may be pertinent to their immunologic functions and could make these cells susceptible to HIV infection.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1989

Mature human eosinophils have the capacity to express HLA-DR.

Human eosinophils are known to lose Ia antigen expression as they mature, and, accordingly, eosinophils obtained from the blood of five eosinophilic donors and three of four normal donors failed to display the major histocompatibility complex class II antigen HLA-DR, as determined by flow cytometry. However, when eosinophils from these nine donors were maintained in culture with recombinant human granulocyte-macrophage colony-stimulating factor and murine 3T3 fibroblasts, HLA-DR consistently developed on the eosinophils. By days 4-6 of culture, 24-97% of eosinophils were HLA-DR+, and the eosinophils remained morphologically mature. In contrast, another class II antigen, HLA-DQ, was not detectable by flow cytometry on eosinophils from eight of nine donors. Cultured eosinophils were able to synthesize HLA-DR, as documented by the incorporation of [35S]methionine into immunoprecipitable HLA-DR heavy and light chains. These findings show that mature eosinophils can synthesize and express HLA-DR and provide a means whereby eosinophils may interact with CD4+ lymphocytes.

Cells, Cultured↗
PubMed1989

Phosphatidylinositol-specific phospholipase C, a possible virulence factor of Staphylococcus aureus.

Phosphatidylinositol-specific phospholipase C (PIPLC), an enzyme that can specifically release phosphatidylinositol-linked proteins from host cells, is one of the extracellular enzymes produced by Staphylococcus aureus. To investigate whether PIPLC might be a virulence factor, we assessed PIPLC production by S. aureus strains that had been isolated from healthy carriers and from infected patients with or without toxic shock syndrome. Although none of five vaginal isolates from healthy women was a PIPLC producer, only 10 of 32 selected pathogenic strains that caused significant infections or toxic shock syndrome elaborated PIPLC enzyme activity. Seven of 24 toxic-shock-associated strains, compared with 3 of 8 non-toxic-shock-associated strains, were positive for PIPLC. The majority of strains that produced PIPLC were negative for toxic shock syndrome toxin 1 (P less than 0.05); this association between PIPLC production and strains negative for toxic shock syndrome toxin 1 was even stronger among strains isolated only from patients with toxic shock syndrome (P less than 0.01). Among all 32 pathogenic isolates, PIPLC-producing S. aureus strains were isolated from four of four patients developing adult respiratory distress syndrome and four of five patients with disseminated intravascular coagulation, suggesting a significant association between PIPLC production and adult respiratory distress syndrome and/or disseminated intravascular coagulation (P less than 0.002). On the basis of these results, we propose that PIPLC is a virulence factor of S. aureus and is implicated in the development of adult respiratory distress syndrome and disseminated intravascular coagulation.

Carrier State↗
PubMed1989

Paroxysmal nocturnal hemoglobinuria. A complement-mediated disease.

Paroxysmal nocturnal hemoglobinuria (PNH) is a hemolytic disease characterized by an increased sensitivity of erythrocytes to the hemolytic action of complement. Two membrane proteins, the decay-accelerating factor and the C8-binding protein, which protect normal erythrocytes from the hemolytic action of complement, are deficient on the abnormal blood cells from patients with PNH. Other membrane proteins unrelated to complement regulation, but which share with the decay-accelerating factor and the C8-binding protein a common post-translational modification, namely a glycan-phosphatidylinositol linkage to the cell membrane, are also missing from PNH cells. In the present review, clinical, biological, and molecular aspects of PNH are discussed. In addition, diagnostic tests in clinical use are discussed, and new tests using indirect immunofluorescent assays are proposed.

Complement System Proteins↗