Ion channels: structure and function.
Explore the source record for details and available documents.
Adenosine Triphosphatases↗
SEARCH · Search PubMed
Results for “Complement Membrane Attack Complex”
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Expression of a hybrid complement regulatory protein, membrane cofactor protein decay accelerating factor on Chinese hamster ovary. Comparison of its regulatory effect with those of decay accelerating factor and membrane cofactor protein.
C activation on the cell surface is supposedly regulated by membrane cofactor protein (MCP) and decay accelerating factor (DAF). These are complementary in function: MCP acts as a cofactor in factor I-mediated C3b and C4b inactivation, thus preventing the assembly of C3 convertases, whereas DAF accelerates spontaneous decay of the assembled C3 convertase. In this report, a hybrid MCP-DAF was expressed on Chinese hamster ovary cells by transfecting cDNA, and its regulatory activity was compared with those of MCP and DAF transfectants and with a transfectant expressing both MCP and DAF (MCP + DAF). The C3 deposition on sensitized CHO cells through activation of the classical pathway was blocked to a different degree with these transfectants, the order being MCP + DAF > DAF > hybrid MCP-DAF > MCP. Likewise, the C3 deposition via the alternative pathway was blocked efficiently in the order hybrid > MCP + DAF > MCP. The C-mediated cytolysis of CHO cells virtually reflected the degree of C3 fragment deposition. The MCP-DAF transfectant acquired additive protective activity against alternative pathway-mediated C3 deposition and cytolysis but was less potent in circumventing classical pathway attack than cells that expressed DAF alone or DAF + MCP. Hybrid MCP-DAF may be useful for alleviating C-mediated cell damage, especially via the alternative pathway.
Animals↗
The sheep analogue of human CD59: purification and characterization of its complement inhibitory activity.
An inhibitor of the membrane attack complex of complement was isolated from the membranes of sheep erythrocytes. Fast protein liquid chromatography (FPLC) and affinity purification procedures for this sheep complement-inhibiting protein (SCIP) both yielded a pure protein with an apparent M(r) of 19,000 under reducing and non-reducing conditions. Incubation of the denatured protein with neuraminidase and Endo-F reduced the apparent M(r) to 18,000 and 15,000 respectively, while treatment with O-deglycosidase or phosphatidylinositol-specific phospholipase C (PIPLC) did not affect the apparent M(r). SCIP was detectable on erythrocytes and lymphocytes but not on platelets and could partially be removed by PIPLC treatment. Deglycosylation of the pure protein markedly reduced and PIPLC treatment abolished its activity. A monoclonal antibody (mAb) raised against sheep complement-inhibiting protein (SCIP) enhanced the susceptibility of sheep erythrocytes to lysis by homologous complement. SCIP inhibited complement after the stage of C5b-7 formation. Amino-terminal protein sequence was obtained and was shown to be similar to that of human CD59. All these features suggest that SCIP is the sheep equivalent of human CD59. Human CD59 has been reported to be species selective in that it inhibits complement from relatively few species. However, SCIP efficiently inhibited lysis of guinea-pig erythrocytes by complement from a wide range of species tested indicating that it is a potent and non-selective inhibitor of the membrane attack complex of complement (MAC).
Amino Acid Sequence↗
Elimination of terminal complement complexes in the plasma membrane of nucleated cells: influence of extracellular Ca2+ and association with cellular Ca2+.
Nucleated cells, unlike erythrocytes, are able to survive limited complement attack by eliminating potentially cytolytic complement channels from the plasma membrane (PM) by processes that involve, plasma membrane (PM) by processes that involve, but may not be limited to, endocytosis. The observation that C5b-9 channels, as well as C5b-8 and C5b-7 intermediates, are rapidly eliminated from the cell surface of nucleated cells has prompted us to examine whether terminal complement complexes stimulate membrane events that lead to accelerated elimination of these complexes. We have suggested previously that ion flux through terminal complement complexes might influence the rate of elimination on the basis of our finding that terminal complement complexes with larger functional channel sizes are more rapidly eliminated. In this study, we examined the role of Ca2+ on the elimination rate of terminal complement complexes in the PM of Ehrlich cells, because changes in Ca2+ flux across the PM are known to influence many metabolic activities including endocytosis. To determine the elimination rate for terminal complement complexes by functional analysis, cells bearing C5b-7 or C5b-8 complexes with or without a sublytic dose of C9 were incubated at 37 degrees C for various time intervals before converting the remaining complexes to lytic C5b-9 channels. The initial elimination rates for the terminal complement complexes were compared in the presence of 0.015, 0.15, and 1.5 mM CaCl2 in the medium. Sufficient lowering of the extracellular Ca2+ concentration, (Ca2+)o, resulted in prolonging the elimination of each of the terminal complement complexes to a different extent. The effect of (Ca2+)o on the elimination rate was most pronounced for C5b-8 in the presence of a sublytic number of C5b-9, with less of an effect on C5b-8 alone, and the least effect with C5b-7. The elimination rates for terminal complement complexes were also determined by measuring the persistence of C5b antigen on the cell surface at 37 degrees C in the presence of various (Ca2+)o by using fluorescence-activated cell sorter analysis and were comparable with that obtained by functional analysis. Examination of the effect of terminal complement complexes on the cellular Ca2+ concentration, (Ca2+)i, revealed that these complexes increased the (Ca2+)i in proportion with the known functional pore size of the terminal complement complex in the PM. In addition, Quin 2, which can buffer internal Ca2+ transients, was found to increase the susceptibility of Ehrlich cells to lysis by C5b-9, further suggesting a relationship between the (Ca2+)i and the elimination process.(ABSTRACT TRUNCATED AT 400 WORDS)
Aminoquinolines↗
Channel fluctuations induced by membrane attack complex C5B-9.
The assembly of complement (C) components C5b-9 in membranes results in the formation of transmembrane lesions. The C9 component has been shown to be mainly responsible for formation of the ultrastructurally visible tubules associated with C5b-9 complexes. Several studies have disputed the role of C9 polymerization in C-mediated cytolysis on the grounds that C5b-9 lyses cells in the absence of tubular formation. Here, C5b-9 complexes were reconstituted into high-impedance planar lipid bilayers and shown to form channels which are heterogenous in size. The smallest channels had unitary conductances of 15 picoSiemens (pS) in 0.1 M NaCl. The closing of these channels showed voltage-dependence at membrane potentials exceeding 40 mV. These channels were more cation-selective, with K+ ions being favored over Na+. The 15-pS channels described here are much smaller than the channels attributed previously to either C5b-9 or polymerized C9 complexes but resemble channels formed by the C9b fragment, which does not polymerize into tubules. These results indicate that C5b-9 complexes are capable of damaging membranes by forming initially small ion channels which then aggregate in the membrane to form tubular lesions with much larger conductances. Like C5b-9, C5b-8 also increased membrane permeability. However, this increase in membrane conductance could not be resolved into single channels, suggesting that C5b-8 may induce membrane leakiness by perturbing the packing of membrane lipids, whereas addition of C9 results in authentic production of ion channels.
Cell Membrane Permeability↗
Complement proteins C5b-9 cause release of membrane vesicles from the platelet surface that are enriched in the membrane receptor for coagulation factor Va and express prothrombinase activity.
We have investigated the composition and function of membrane microparticles released from platelets exposed to the C5b-9 proteins of the complement system. Gel-filtered human platelets were incubated with sub-lytic amounts of the purified C5b-9 proteins and the distribution of surface antigens was analyzed using monoclonal antibodies and flow cytometry. C5b-9 assembly caused secretory fusion of the alpha-granule membrane with the plasma membrane and the release of membrane vesicles (approximately 0.1-micron diameter) that contained the plasma membrane glycoproteins (GP) GP Ib and GP IIb-IIIa as well as the alpha-granule membrane protein GMP-140. These microparticles were highly enriched in the C9 neoantigen of the C5b-9 complex. The apparent surface density of C5b-9 on the microparticles was approximately 10(3)-fold higher than on the platelet itself, suggesting that the vesicles were selectively shed from the plasma membrane at the site of C5b-9 insertion. C5b-9 induced the expression of an activation-dependent epitope (recognized by monoclonal antibody, PAC1) in GP IIb-IIIa on the platelet surface but not in GP IIb-IIIa on the microparticles. The surface of the microparticles was also highly enriched in alpha-granule-derived coagulation factor V (or Va), accounting for nearly half of all the membrane-bound factor V detected. The number of potential membrane binding sites for factor Va was probed by adding saturating concentrations of factor Va light chain. Under these conditions, the density of factor Va binding sites on the microparticle surface exceeded that on the C5b-9-treated platelet by three to four orders of magnitude. Moreover, the microparticles provided most of the membrane surface for conversion of prothrombin to thrombin by VaXa. These studies demonstrate that the microparticles shed by C5b-9-treated platelets (and not the platelets themselves) provide the principal binding sites for coagulation factor Va and the principal catalytic surface for the prothrombinase complex. Platelet-derived microparticles formed during complement activation in vivo could provide a membrane surface that facilitates the assembly and dissemination of procoagulant enzyme complexes.
Blood Platelets↗
Cerebrospinal fluid concentrations of the complement MAC inhibitor CD59 in multiple sclerosis and patients with other neurological disorders.
Rodent oligodendrocytes have a unique susceptibility among glia to the lytic effects of complement, due in part to a deficiency in CD59 (protectin), a key surface inhibitor of the complement membrane attack complex (MAC). The possibility that shedding of CD59 by human oligodendrocytes contributes to complement-mediated oligodendrocyte injury in inflammatory demyelinating disease has been investigated by estimating levels of CD59 in cerebrospinal fluid samples from 12 patients with demyelinating disease of the central nervous system and 13 with other neurological diseases. No significant differences were found between patients and controls, or between patients with active and those with clinically inactive demyelinating disease, providing no direct support for oligodendrocyte shedding of CD59 in multiple sclerosis.
Antigens, CD↗
Activation of complement in the central nervous system: roles in neurodegeneration and neuroprotection.
The complement system is an essential effector of the humoral and cellular immunity involved in cytolysis and immune/inflammatory responses. Complement participates in host defense against pathogens by triggering the formation of the membrane attack complex. Complement opsonins (C1q, C3b, and iC3b) interact with surface complement receptors to promote phagocytosis, whereas complement anaphylatoxins C3a and C5a initiate local inflammatory responses that ultimately contribute to the protection and healing of the host. However, activation of complement to an inappropriate extent has been proposed to promote tissue injury. There is now compelling evidence that complement activation in the brain is a double-edged sword in that it can exert beneficial or detrimental effects depending on the pathophysiological context. This review focuses on the roles of the complement system in the pathogenesis of acute brain injury (cerebral ischemia and trauma) and chronic neurodegeneration (Alzheimer's disease). Because many effects of the complement appear to promote neuronal survival and tissue remodeling, directing activation of the complement system in the brain may provide a better therapeutic rationale than inhibiting it.
Alzheimer Disease↗
Immunocytochemical localisation of complement components C8 and C9 in human diseased muscle. The role of complement in muscle fibre damage.
The localisation of the complement components C8 and C9 was studied immunocytochemically in human diseased muscle to determine the role of complement in muscle fibre damage. Monoclonal antibodies to 2 epitopes of C9 and a monoclonal antibody to the alpha subunit of C8 were applied to frozen sections of muscle biopsies from 9 cases of dermatomyositis, 5 cases of polymyositis, 7 cases of Duchenne muscular dystrophy and 4 cases of Becker muscular dystrophy. These were compared with 6 control biopsies which were morphologically normal. In all cases of inflammatory myopathies several non-necrotic fibres showed discrete peripheral patches of C9 and to a lesser extent C8. In the muscular dystrophies peripheral C9 was observed on a few non-necrotic fibres and basophilic fibres showed C9 between the fibres as well as at the periphery. In all cases necrotic fibres labelled intensely with C9 and C8 but intensities varied with the different monoclonal antibodies. This was thought to result from differences in the polymerisation of the C9 molecule in the membrane attack complex. Complement C8 and C9 were also localised to blood vessels in 3 cases of muscular dystrophy, 2 cases of polymyositis and all cases of juvenile dermatomyositis. No complement was observed in the control samples. Our results provide evidence for the sublytic formation of the membrane attack complex (MAC) on non-necrotic fibres in inflammatory myopathies and muscular dystrophy. This sublytic formation of the MAC may induce sublethal metabolic damage, mediated by calcium, and suggests a primary role of complement in muscle damage not only in inflammatory disorders but also muscular dystrophy.
Adolescent↗
Localization of clusterin in the epimembranous deposits of passive Heymann nephritis.
The membrane attack complex of complement (MAC) plays an important role in the mediation of proteinuria in experimental membranous nephropathy induced by Heymann antiserum. SP-40,40 is a recently described serum protein which appears to inhibit the formation of cytolytic MAC in a manner analogous to S protein/vitronectin. SP-40,40 is homologous to proteins originally isolated from rat and ram seminal fluid (sulfated glycoprotein 2 and clusterin, respectively). By current convention, these proteins are considered clusterin homologues. The objective of this study was to examine the participation of rat clusterin in passive Heymann nephritis. Using an antibody to rat clusterin as an immunofluorescent probe, clusterin deposits were demonstrated along the glomerular capillary wall in an identical pattern to rat C3 and C5b-9. Decomplementation using cobra venom factor prevented proteinuria and intraglomerular MAC formation. The epimembranous clusterin were not detected in the complement-depleted animals. The role of clusterin in the mediation of glomerular injury remains unknown, but it is probably related to in situ formation of the terminal complement cascade where it may play a regulatory role.
Animals↗
The double-edged flower: roles of complement protein C1q in neurodegenerative diseases.
A role for the complement cascade in AD neuropathology was hypothesized over a decade ago, and the results of a significant number of in vitro studies are consistent with the involvement of this pathway in AD pathogenesis (reviewed in). Since C1q is colocalized with thioflavine-positive plaques and the C5b-9 complement membrane attack complex is detected in AD brain at autopsy, it is reasonable to hypothesize that complement activation has a role in the manifestation of AD either by its lytic capacity or as a trigger of glial infiltration and initiation of potentially damaging inflammation. The observed diminished glial activation and reduced loss of neuronal integrity in a murine model overexpressing mutant human APP but lacking the ability to activate the classical complement cascade provide the first direct evidence for a detrimental role of C1q, and presumably activation of the classical complement pathway in an animal model of AD. Research is now focused on generating mouse models that more closely mimic the human disease, so that the role of complement activation and inflammation on the behavioral/learning and memory dysfunction that occurs in this disease can be assessed. In addition, candidate therapies such as targeted inhibition of complement activation will need to be tested in these animal models as a step toward treatment of humans with the disease. However, it is important that the potential for a protective effect of C1q early on in disease progression should not be overlooked. Rather, strategies that enhance or mimic the protective effects of C1q as well as strategies that inhibit the detrimental processes should be fully investigated.
Amyloid beta-Peptides↗
Immune evasion of tumor cells using membrane-bound complement regulatory proteins.
Membrane-bound complement regulatory proteins (mCRPs) play an important role in the protection of cells from complement-mediated injury. It is now apparent that malignant tumor cells also express these proteins to escape complement attack. Here, Arko Gorter and Seppo Meri discuss the implications of complement resistance for the immunotherapeutic treatment of solid tumors with monoclonal antibodies.
Animals↗
Complementary therapies for inflammation.
Explore the source record for details and available documents.
Animals↗
[Activation of complement by hemodialysis membrane].
Incubation of normal human serum with hemodialysis membranes in vitro resulted in conversion of C3 which was detected by crossed immunoelectrophoresis. In addition, formation of C3d was also observed, as was detected by double-decker rocket immunoelectrophoresis. Furthermore, breakdown products of complement (iC3b, C4d & Bb) in plasma samples were measured by ELISA. The microassay plates were coated with monoclonal antibodies which bind specifically to human iC3b, C4d and Bb, respectively, while the plasma samples were drawn from five polycystic kidney patients during initial hemodialysis. As a result, the iC3b and Bb levels in plasma were seen to increase during hemodialysis but the C4d levels revealed no significant changes. It was also observed that the Bb levels in patients undergoing hemodialysis were significantly higher than those in various renal and collagen diseases. Terminal complement complex (TCC) was not present in detectable amounts in normal human serum when measured by ELISA. However, incubation of normal human serum with hemodialysis membranes in vitro resulted in detection of TCC. TCC was present in the same plasma samples drawned from the five polycystic kidney patients during initial hemodialysis. It is suggestive that activation of complement by hemodialysis membrane is predominantly mediated through the alternative pathway and products such as TCC and anaphylatoxin are formed by this activation.
Complement Activation↗
Characterization of homologous restriction factor (HRF20) in human skin and leucocytes.
Homologous restriction factor with a molecular weight of 20 kD (HRF20) is a membrane protein that inhibits assembly of the membrane attack complex of homologous complement. Distribution of HRF20 in normal human skin was studied. The plasma membrane of keratinocytes was stained, and the intensity of the staining pattern was higher in the basal cell layer than in the granular layer. Endothelial cells of blood vessels in the dermis were also stained. The molecular weight of HRF20 on erythrocytes and epidermis is 16 kD, determined by Western blot analysis. Those of polymorphonuclear cells and lymphocytes appeared as two bands, a major band of 20 kD and a minor band of 16 kD. Susceptibility of HRF20 to phosphatidylinositol-specific phospholipase C (PIPLC) was examined. After PIPLC treatment of the sections, HRF20 was not detected on the epidermis and was very slightly expressed on the blood vessels. These results indicate that HRF20 attaches to keratinocytes and blood vessels via phosphatidylinositol, regulating the formation of membrane attack complexes of homologous complement on the cell membrane.
Blood Proteins↗
Effect of factor Xa inhibitors on thrombin formation and complement and neutrophil activation during in vitro extracorporeal circulation.
BACKGROUND: Even when large doses of heparin are administered during cardiopulmonary bypass, thrombin is produced. Thrombin is a powerful protease that is associated with the thrombotic and bleeding complications of open heart surgery and is produced by cleavage of prothrombin by factor Xa. This study assessed the ability of a specific inhibitor of factor Xa, recombinant tick anticoagulant peptide (rTAP), alone or in combination with standard heparin and a low-molecular-weight heparin, enoxaparin, to suppress thrombin formation and activity during in vitro extracorporeal circulation. METHODS AND RESULTS: Fresh, anticoagulated human blood was recirculated for 2 hours in an extracorporeal membrane oxygenator perfusion circuit at 37 degrees C. Four anticoagulant protocols were evaluated; porcine heparin (3.75 U/mL); enoxaparin (17.5 U/mL); rTAP (4 mumol/L); and porcine heparin plus rTAP (2 mumol/L). Blood samples were obtained for analysis from the donor, after anticoagulation, and after 5, 30, 60, and 120 minutes of recirculation. There were no significant differences between groups in platelet count, response to adenosine diphosphate, or prothrombin fragment (F1.2) production. rTAP plus heparin reduced beta-thromboglobulin release; fibrinopeptide A concentrations were significantly higher with rTAP alone. Enoxaparin strongly and significantly inhibited complement C5b9 production and neutrophil elastase release and was associated with significantly increased concentrations of C1-C1 inhibitor and kallikrein-C1 inhibitor complexes. CONCLUSIONS: rTAP does not reduce thrombin formation or activity during in vitro extracorporeal circulation. Enoxaparin markedly inhibits formation of the complement membrane attack complex and neutrophil elastase release, possibly by accelerating C1 inhibitor activity.
Adenosine Diphosphate↗
Molecular aspects of complement-mediated bacterial killing. Periplasmic conversion of C9 from a protoxin to a toxin.
As part of the membrane attack complex complement protein C9 is responsible for direct killing of bacteria. Here we show that in the periplasmic space of an Escherichia coli cell C9 is converted from a protoxin to a toxin by periplasmic conditions missing in spheroplasts. This conversion is independent of the pathway by which C9 enters the periplasm. Both, C9 shocked into the periplasm and plasmid-expressed C9 targeted to the periplasm via a signal sequence are toxic. Toxicity requires disulfide-linked C9 because export into the periplasm of cells defective in disulfide bond synthesis (dsbA and dsbB mutants) is not toxic unless N-acetylcysteine is added externally to promote cystines. A N-terminal fragment, C9[1-144], is not toxic nor is cytoplasmically expressed C9, even in trxB mutants that are able to form disulfide bonds in the cytoplasm. Importantly, expression of full-length C9 in complement-resistant cells has no effect on their viability. Expression and translocation into the periplasm may provide a novel model to identify molecular mechanisms of other bactericidal disulfide-linked proteins and to investigate the nature of bacterial complement resistance.
Amino Acid Sequence↗
Cytosolic calcium and protein kinase C reduce complement-mediated glomerular epithelial injury.
In rat membranous nephropathy, protein-uria is due to formation of the C5b-9 membrane attack complex of complement (C), and is associated with morphological evidence of glomerular epithelial cell (GEC) injury. Analogous morphological changes are induced by C5b-9 in cultured GEC. In addition, in cultured GEC C5b-9 induces Ca2+ influx, as well as Ca2+ mobilization and increased 1,2-diacylglycerol due to the activation of phospholipase C. In this study we investigated how this GEC activation pattern might influence C-mediated GEC injury. We demonstrate that the C5b-9-induced increase in cytosolic Ca2+ concentration ([Ca2+]i) did not impair ATP generation by mitochondria, suggesting that it does not contribute to cytotoxicity. Moreover, this increase in [Ca2+]i protected GEC from C-mediated cytolysis. However, a large increase in [Ca2+]i (produced by the Ca2+ ionophore A23187) impaired ATP generation and aggravated C-mediated cytotoxicity, suggesting that intact mitochondrial activity is necessary for GEC to withstand C attack. Activation of protein kinase C (PKC) by phorbol myristate acetate (PMA) also decreased C-mediated cytolysis. Conversely, C lysis was enhanced in GEC that had been pretreated for 18 hours with a high dose of PMA to deplete PKC, and following PKC inhibition with H-7. Therefore, PKC activation, possibly resulting from C5b-9-induced increase in 1,2-diacylglycerol, triggered mechanisms that protected GEC from C-mediated injury. Thus, as a consequence of C5b-9-induced phospholipase activation, the amount of C-induced GEC injury is diminished.
Adenosine Triphosphate↗