Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Complement C5b”

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.

At least 1,045 records · Page 58Linked to original sources
PubMed1985

Monoclonal antibodies directed against gonococcal protein I vary in bactericidal activity.

Monoclonal antibodies (Mab) with specificity for protein I (PI) from Neisseria gonorrhoeae (GC) were examined for bactericidal activity. Mab 4G5 (gamma 3), ID3 (gamma 2a), and 1G6 (gamma 2a) bound to surface-exposed epitopes on PI of GC strain R11 (IA serotype) as assessed by co-agglutination and 125I protein A uptake. Mab 2H1 (gamma 3) that were directed against IB serotype strains and Mab 2E9 (gamma 2a) were negative in co-agglutination and protein A uptake assays and served as controls for some experiments. Only 4G5 and 1D3 were bactericidal for R11 when presensitized organisms were incubated in 10% absorbed, pooled normal human serum (PNHS) or 10% hypogammaglobulinemic serum (H gamma S) despite binding of nearly equivalent numbers of 4G5, 1D3, and 1G6 to R11 during presensitization, as assessed by 125I-protein A uptake. These Mab activated complement to a similar extent on GC R11, leading to deposition of 56.4 X 10(3), 61.9 X 1093), and 47.1 X 10(3) molecules of C3/organism during incubation in 10% C8-deficient serum. Deposition occurred almost exclusively via the classical complement pathway. Measurement of complement component C9 binding to R11 during incubation in H gamma S showed 35,700 molecules of C9/organism with 4G5, 32,600 C9/organism with 1D3, and surprisingly, 29,600 C9/organism with 1G6. Eight thousand four hundred molecules of C9/organism bound to 2E9-coated organisms, 6000 C9/organism to 2H1-coated bacteria, and 3600 C9/organism to nonpresensitized organisms. The C5b-9 complex deposited by 4G5 had a different sedimentation profile by sucrose density gradient analysis from the C5b-9 complex deposited by 1G6, consistent with a different molecular configuration of the bound complex. Mab 1G6 and 1D3, but not 2E9 or 2H1, were able to compete with 125I-4G5 for binding to GC R11. A Mab (2E6) directed against protein III of GC competed weakly with 125I-4G5 for binding to GC R11. Mab 1G6, but not 1D3, blocked 4G5-dependent killing in a dose-related fashion. Both 4G5 and IG6 reacted weakly with native PI of GC R11 by immunoblotting, but neither Mab recognized the 34,800 m.w. fragment of PI generated by trypsin and chymotrypsin treatment of outer membranes. In contrast, 2E9 reacted strongly by immunoblot with both native and cleaved PI of GC R11, suggesting binding to buried determinants of PI. These experiments show that Mab directed against identical or closely associated, surface-exposed epitopes on gonococcal PI differ markedly in bactericidal activity, despite leading to deposition of nearly equivalent numbers of C3 and C9 molecules per organism.

Anti-Bacterial Agents↗
PubMed1993

Interactions of soluble CD59 with the terminal complement complexes. CD59 and C9 compete for a nascent epitope on C8.

Human CD59-Ag is a glycophosphoinositol lipid-anchored inhibitor of the membrane attack complex of complement (MAC). We have examined the interactions of CD59 with the terminal complement complexes using soluble CD59 purified from human urine (CD59U). CD59U bound to SC5b-8, SC5b-9, and MAC complexes when present during their formation. When SC5b-8, SC5b-9, and MAC were allowed to perform, progressively less 125I-CD59U bound to the complexes. Terminal SC5b-9 complexes isolated from activated sera no longer bound CD59U, indicating that the binding sites had become inaccessible in the fully assembled SC5b-9 complex. Unlike acylated myocardial CD59 (CD59H) neither CD59U nor PIPLC-treated CD59H became incorporated into poly-C9 complexes, suggesting that the interaction of CD59 with C9 requires the lipid anchor. Human C9 and heterologous C9 from guinea pig serum, as well as the YTH53.1 anti-CD59 mAb, inhibited the binding of CD59U to SC5b-8. On the other hand, soluble CD59U did not inhibit binding of C9 to SC5b-8, although CD59E has been shown to limit the number of C9 molecules entering into MAC. This suggests that two interaction sites between C5b-8 and C9 exist: one conferring the initial binding of C9 into the C5b-8 complex, and a second directing the insertion of C9 into the lipid bilayer. The latter interaction is the prerequisite for C9 polymerization and the target for interference by CD59.

Animals↗
PubMed1989

Multimeric C9 within C5b-9 deposits in unique locations in the cell wall of Salmonella typhimurium.

We have shown previously that multimeric C9 within C5b-9 (C9:C5b-8 greater than 3:1) is needed for killing of a rough strain of Escherichia coli. We now extend these studies using serum sensitive, rough (R) and serum resistant, wild type (WT) strains of Salmonella typhimurium as well as a mutant S. typhimurium strain (TS) with a temperature sensitive mutation in synthesis of keto-deoxy-octulosonate, a constituent within the deep core structure of Salmonella LPS. Both R and TS required multimeric C9 within C5b-9 to be killed. Addition at 37 degrees C of increasing inputs of C9 to TS or R bearing C5b-9 led to a dose-related increase in C9 binding and killing. In contrast, addition of high inputs of C9 to the same strains at 4 degrees C, a procedure that limits the C9:C5b-8 ratio to 1:1, resulted in low C9 binding and minimal killing. Bactericidal C5b-9 formed at 37 degrees C on R and TS with high inputs of C9 co-sedimented with the bacterial outer membrane on sucrose density gradient analysis. Non-bactericidal C5b-9 on R, WT, and TS co-sedimented near the inner membrane, despite the presumed lack of association between these constituents. Whereas 125I C9 within the non-bactericidal pools immunoprecipitate with anti-C5, 125I C9 within bactericidal pools did not immunoprecipitate with anti-C5, anti-C7, or anti-C9. These findings suggest that bactericidal C5b-9 may be deposited in a unique location or configuration within the bacterial cell wall.

Cell Wall↗
PubMed1984

Molecular composition of the tubular structure of the membrane attack complex of complement.

The composition of the tubular structure of the membrane attack complex of complement (MAC) which migrates as a high molecular weight band (Mr approximately 1.2- 1.3 X 10(6) upon sodium dodecyl sulfate, polyacrylamide gel electrophoresis under reducing conditions was analyzed and compared to the high molecular weight band (Mr approximately 1.1 X 10(6] of tubular poly(C9). The sodium dodecyl sulfate-resistant band of the MAC, designated MAC-poly(C9), is composed of C6, C7, C8 alpha-gamma, and poly(C9), in approximate molar ratios of the protomers of 1:1:1:10-18. This conclusion is based 1) on the results of the incorporation of labeled proteins into MAC-poly(C9); 2) on the immunostaining of MAC-poly(C9) with anti-C6, anti-C7, anti-C8 alpha-gamma, and anti-C9 and its lack of immunostaining with anti-C5 and anti-C8 beta; and 3) on the dissociation of MAC-poly(C9) to 1 mol of C6, C7, C8 alpha-gamma and 10 to 18 mol of C9 upon treatment with 8 M guanidine isothiocyanate. Ultrastructurally the sodium dodecyl sulfate-resistant poly(C9) tubule and MAC-poly(C9) tubule are indistinguishable, suggesting a similar ultrastructure of the C6, C7, C8 alpha-gamma, and C9 subunits in the MAC-poly(C9) tubule. Further analogies among these four proteins are their tendency to form disulfide-linked dimers. It is concluded that the transmembrane channel of the MAC is formed by a tubule in which C6, C7, C8 alpha-gamma are copolymerized with poly(C9), whereas the C5b and C8 beta subunits are not part of the tubule structure and may form the 170-A long appendage of the MAC. This appendage is dissociated upon boiling in sodium dodecyl sulfate whereas the tubule remains stable.

Complement Membrane Attack Complex↗
PubMed2005

Role of the endoplasmic reticulum unfolded protein response in glomerular epithelial cell injury.

C5b-9-induced glomerular epithelial cell (GEC) injury in vivo (in passive Heymann nephritis) and in culture is associated with damage to the endoplasmic reticulum (ER) and increased expression of ER stress proteins. Induction of ER stress proteins is enhanced via cytosolic phospholipase A(2) (cPLA(2)) and limits complement-dependent cytotoxicity. The present study addresses another aspect of the ER unfolded protein response, i.e. activation of protein kinase R-like ER kinase (PERK or pancreatic ER kinase), which phosphorylates eukaryotic translation initiation factor 2-alpha (eIF2alpha), thereby generally suppressing translation and decreasing the protein load on a damaged ER. Phosphorylation of eIF2alpha was enhanced significantly in glomeruli of proteinuric rats with passive Heymann nephritis, compared with control. In cultured GECs, complement induced phosphorylation of eIF2alpha and reduced protein synthesis, and complement-stimulated phosphorylation of eIF2alpha was enhanced by overexpression of cPLA(2). Ischemia-reperfusion in vitro (deoxyglucose plus antimycin A followed by glucose re-exposure) also stimulated eIF2alpha phosphorylation and reduced protein synthesis. Complement and ischemia-reperfusion induced phosphorylation of PERK (which correlates with activation), and fibroblasts from PERK knock-out mice were more susceptible to complement- and ischemia-reperfusion-mediated cytotoxicity, as compared with wild type fibroblasts. The GEC protein, nephrin, plays a key role in maintaining glomerular permselectivity. In contrast to a general reduction in protein synthesis, translation regulated by the 5'-end of mouse nephrin mRNA during ER stress was paradoxically maintained, probably due to the presence of short open reading frames in this mRNA segment. Thus, phosphorylation of eIF2alpha and consequent general reduction in protein synthesis may be a novel mechanism for limiting complement- or ischemia-reperfusion-dependent GEC injury.

Animals↗
PubMed1989

Membrane permeability to macromolecules mediated by the membrane attack complex.

A simple and well-defined system of purified phospholipids and human complement proteins was used to study membrane permeability to macromolecules mediated by the membrane attack complex (MAC) of complement. Large unilamellar vesicles (LUVs) of phosphatidylcholine (PC) or phosphatidylserine (PS) containing trapped macromolecules [bovine pancreatic trypsin inhibitor (BPTI), thrombin, glucose-6-phosphate dehydrogenase (G6PD), and larger molecules] were used to monitor permeability. Membrane permeability to macromolecules was measured by thrombin inhibition by an external inhibitor or by separation of released molecules by gel filtration. Membrane-bound intermediates (C5b-8 or C5b-93) were stable for hours, and macromolecular permeability occurred without fragmentation, fusion, or aggregation of the vesicles. Quantitative membrane binding by C5b-7 as well as essentially quantitative release of thrombin was obtained for PS vesicles. MAC binding to PS-LUVs approximated the theoretical Poisson distribution curve for full release of vesicle contents by one complex per vesicle. Reactions with PC-LUVs occurred with some fluid-phase MAC assembly. Therefore, results from experiments with these vesicles were interpreted in a relative manner. However, the values obtained closely corroborated those obtained with PS-LUVs. At low C9/C5b-8 ratios, the size of the lesion was proportional to the C9 content of the MAC. Half-maximum release of BPTI, thrombin, and G6PD, by a single MAC per vesicle, required approximately 3,5, and 7 C9/C5b-8 (mol/mol), respectively. Larger molecules (greater than or equal to 118-A diameter) were not released from the vesicles. Release of G6PD (95.4-A diameter) required 45% of saturating C9. Therefore, it appeared that the last half of the bound C9 molecules did not increase pore size and the pore which released G6PD approached the diameter of the closed circular lesion measured (by others) in electron micrographs (approximately 100 A). The results were consistent with the formation of a stable membrane pore by a single complex per vesicle in which C9 molecules line only one side of the pore at low C9/C5b-8 ratios and maximum pore size is attained by incomplete, noncircular polymers of C9.

Cell Membrane Permeability↗
PubMed1992

Role of complement in endotoxin/platelet-activating factor-induced lung injury.

C receptor-1 is a protein involved in the regulation of C3 and C5-convertases. Recombinant human soluble C receptor-1 has recently been produced and shown to reduce infarct size in a rat model of myocardial ischemia/reperfusion injury. The present study aimed to investigate whether recombinant human soluble C receptor-1 exerts any protective effect on pulmonary injury produced in a rodent model of adult respiratory distress syndrome. In this model, Escherichia coli endotoxin (LPS, 0.1 microgram/kg) combined with platelet-activating factor (1 pmol/kg/min over 60 min, n = 10) caused microvascular lung injury characterized by elevation of myeloperoxidase activity, deposition of C3 and C5b-9 on the endothelium of pulmonary vessels, and pulmonary edema. Furthermore, bronchoalveolar lavage revealed increased neutrophil count and elevated protein concentration. These pulmonary responses were associated with elevated serum TNF-alpha. Pretreatment (10 min, i.v.) with recombinant human soluble C receptor-1 at 10 mg/kg (n = 13), but not at 1 mg/kg, prevented the LPS/platelet-activating factor-induced pulmonary edema (p less than 0.01) and changes in the bronchoalveolar lavage fluid cell count (p less than 0.01) and protein concentration (p less than 0.05), and attenuated the deposition of C3 and C5b-9 to lung vessels. There was no effect on lung myeloperoxidase activity and serum TNF-alpha. Also, C depletion by cobra venom factor (500 U/kg, i.v.) eliminated the pulmonary edema and elevated leukocyte count in bronchoalveolar lavage fluid, but had no effect on lung myeloperoxidase activity and serum TNF-alpha. These data suggest that C factors may play an important role in the pathophysiology of adult respiratory distress syndrome.

Animals↗
PubMed2005

C5b-9 does not mediate chronic tubulointerstitial disease in the absence of proteinuria.

BACKGROUND: In nephrotic glomerular diseases, the intratubular assembly of the membrane attack complex (C5b-9) is one of the principal mediators of chronic tubulointerstitial damage. Here, we examined whether C5b-9 has a pathogenic role in tubulointerstitial disease in the absence of proteinuria. METHODS: Three pathophysiologically distinct models of nonproteinuric chronic tubulointerstitial disease were induced in Piebald-Viral-Glaxo (PVG) rats, with or without C6 deficiency (C6+ and C6): (1) unilateral ureteric obstruction (UUO, days 1, 3, 6, 14, and 21; N= 5-6/group); (2) cyclosporine (CsA) nephropathy (15 mg/kg SC daily with 0.05% sodium diet; day 14, 35 N= 9/group); and (3) streptozotocin (STZ)-induced diabetes (day 90, N= 8/group). RESULTS: The peritubular deposition of C5b-9 increased in all three models. In UUO, the number of vimentin-positive tubules, interstitial volume expansion, and monocyte accumulation were similar in both the C6+ and C6- groups at all time points. There was a trend toward an earlier peak in myofibroblast accumulation in C6- rats with UUO (d3 vs. d6; P= 0.05), but this did not prevent fibrosis at later time points. In CsA nephropathy, cortical tubulointerstitial damage was also similar in both C6+ and C6- groups on day 14, despite equivalent CsA trough levels. Finally, in STZ-induced diabetes, rats did not develop proteinuria, and tubulointerstitial disease (distal tubule glycogen nephrosis, interstitial volume expansion, and tubular dilatation) was not altered by C6 deficiency. CONCLUSION: These data suggest that, in contrast to proteinuric states, C5b-9 does not have a significant impact on the progression of tubulointerstitial damage in nonproteinuric chronic renal disease.

Animals↗
PubMed1997

Sublytic concentrations of the membrane attack complex of complement induce endothelial interleukin-8 and monocyte chemoattractant protein-1 through nuclear factor-kappa B activation.

Activation of the complement cascade and subsequent assembly of the membrane attack complex (MAC) occur in a number of pathophysiological settings. When formed on the surface of endothelial cells in sublytic concentrations, the MAC can induce a number of proinflammatory activities, including the secretion of soluble mediators (eg, interleukin (IL)-8 and monocyte chemoattractant protein (MCP)-1) and the up-regulation of cell surface adhesion molecules. Available data indicate that MAC-induced cell activation may occur through several complex signal transduction pathways, but little is known about the intranuclear mechanisms by which complement-derived products promote the up-regulation of inflammatory mediators. Using purified distal complement proteins (C5-9) to assemble functional MAC on early-passage human umbilical vein endothelial cells (HUVECs), we examined mechanisms of MCP-1 and IL-8 induction. Formation of sublytic concentrations of MAC promoted an increase in nuclear factor (NF)-kappa B DNA binding activity within 60 minutes as determined by serial electrophoretic mobility shift assay. Cytosolic to nuclear translocation of NF-kappa B was confirmed by Western immunoblot and immunocytochemical analyses. Formation of the C5b-8 complex also promoted NF-kappa B translocation but to a lesser degree than observed in HUVECs containing complete MAC. No cytosolic to nuclear translocation of the p65 NF-kappa B subunit was observed in unstimulated HUVECs or in cells incubated with the MAC components devoid of C7. Preincubation of HUVECs with pyrrolidine dithiocarbamate prevented MAC-induced increases in IL-8 and MCP-1 mRNA concentrations and protein secretion. A direct cause and effect linkage between MAC assembly and NF-kappa B activation was established through examination of the pharmacological effect of the peptide SN50 on IL-8 and MCP-1 expression. SN50 is a recently engineered 26-amino-acid peptide that contains a lipophilic cell-membrane-permeable motif and a nuclear localization sequence that specifically competes with the nuclear localization sequence of the NF-kappa B p50 subunit. This study provides direct in vitro evidence that the distal complement system (MAC) can promote proinflammatory endothelial cell activation, specifically, increases in IL-8 and MCP-1 mRNA concentrations and protein secretion, and that cytosolic to nuclear translocation of NF-kappa B is necessary for this response.

Blotting, Northern↗
PubMed1980

Inherited deficiency of the ninth component of complement in man.

A 76-year-old man was found to have low (33% normal) serum complement (C) hemolytic activity, although C3 and C4 protein levels were normal. Further evaluation of his serum and plasma indicated that all C components were present in normal or elevated amounts except for C9, which was undetectable by both antigenic and functional assays. Addition of purified human C9 led to full restoration of the hemolytic activity. Family studies demonstrated that the deficiency was inherited as an autosomal codominant trait and was not linked with alleles at the HLA-A or HLA-B loci. The patient had no history of recurrent or unusual infections and no evidence of autoimmune disease. The availability of serum totally lacking in C9 permitted an investigation of the lytic capacity of the C5b-8 segment of the C attack mechanism, which was pursued in kinetic studies on the hemolysis of erythrocyte intermediates. These studies indicated that hemolysis occurred approximately 100 times slower in patient than in normal serum, using either EA or EAC1-7 intermediates as target cells. Serum bactericidal activity also was slower in patient serum, occurring at a rate about 1/35 that observed in normal serum. These studies provide direct independent evidence that cytolysis of erythrocytes and bacteria can be mediated by C5b-8, and allow a quantitative estimation of the increment in the rates of these reactions provided by normal serum levels of C9. The presence of readily detectable though slow hemolytic activity of C9-deficient serum may account for the difficulty in identifying individuals with this defect.

Aged↗
PubMed2003

ONO-6818, a novel, potent neutrophil elastase inhibitor, reduces inflammatory mediators during simulated extracorporeal circulation.

BACKGROUND: Among the serine proteases, neutrophil elastase is a powerful cytotoxic enzyme and plays a pivotal role in the inflammatory response associated with cardiopulmonary bypass. This study assesses the effects of the specific inhibition of neutrophil elastase by a novel, potent, low-molecular-weight neutrophil elastase inhibitor, ONO-6818. We hypothesized that ONO-6818 reduces inflammatory mediators and modulates adhesion molecules and the deformability of neutrophils during simulated extracorporeal circulation. METHODS: Simulated extracorporeal circulation was established by recirculating fresh heparinized (3.75 U/mL) human blood for 120 minutes in a membrane oxygenator and a roller pump with and without 1.0 micromol/L of ONO-6818 (n = 9 for control group, n = 7 for ONO-6818 group). The neutrophil adhesion molecules, CD11b and L-selectin, and the cytoplasmic F-actin of neutrophils were measured by flow cytometry. Neutrophil deformability was evaluated using simulated silicon microcapillaries. Neutrophil elastase, interleukin 8, and C5b-9 were measured using enzyme immunoassay. RESULTS: Neutrophil elastase levels were significantly lower in the ONO-6818 group. ONO-6818 significantly reduced interleukin 8 and C5b-9 production. ONO-6818 did not modulate changes of CD11b and L-selectin during recirculation. Cytoplasmic F-actin content and changes of neutrophil deformability did not significantly differ between the groups. CONCLUSIONS: Inhibition of neutrophil elastase activity with ONO-6818 reduces further interleukin 8 production and the formation of the complement membrane attack complex, and this results in a reduction of neutrophil elastase levels during simulated extracorporeal circulation. This study suggests that specific neutrophil elastase inhibition with ONO-6818 is a feasible therapeutic option to attenuate the exaggerated inflammatory response associated with cardiopulmonary bypass.

Actins↗
PubMed2003

Adverse outcome of human islet-allogeneic blood interaction.

BACKGROUND: A report of inflammatory damage when islets come into contact with allogeneic blood prompted us to confirm the finding. METHODS: Fresh handpicked human islets were incubated in blood group matched, nonsensitized allogeneic blood. Destruction was quantified by assaying the supernatants for proinsulin release and by blood clot histology. The effect on global hemostasis was assessed by thromboelastography (TEG), and heparin-bonded tubing was used to assess the effect on blood cellular counts. In separate experiments, islets were incubated in allogeneic blood with heparin or Reopro (monoclonal anti-GpIIbIIIa). Islets were also incubated in serum, and cryosections were stained for C1q, C4, C3, C5b-9, immunoglobulin (Ig)M, and IgG binding using immunohistochemistry. RESULTS: Histologic assessment showed severe destruction in 37% of islets in contact with allogeneic blood versus none in controls and a sevenfold increase in proinsulin release from controls (n = 6)(P < 0.005). TEG (n = 11) showed accelerated coagulation in the presence of islets (P < 0.001). Analysis of blood cellular counts (n = 3) showed consumption of platelets, neutrophils, and monocytes in the presence of islets (P < 0.001). Inhibition of coagulation with heparin (n = 3) or inhibition of platelet aggregation with Reopro (n = 3), separately or together (n = 3), did not make a substantial improvement in the destruction in terms of histology or proinsulin release. Immunohistochemical staining (n = 4) revealed C1q, C4, C3, and C5b-9 deposition along with IgG binding. IgM binding was weak if anything. CONCLUSION: This study confirms and extends the finding that human islet-allogeneic blood interaction results in significant destruction of islet tissue with activation of the coagulation cascade and platelet, neutrophil, and monocyte consumption. There was evidence for activation of complement by the classical pathway along with IgG binding.

Abciximab↗
PubMed1997

Antiproteinuric effect of angiotensin-converting enzyme inhibition and C5b-9 urinary excretion in membranous glomerulonephritis.

BACKGROUND: Angiotensin-converting enzyme (ACE) inhibitors have an antiproteinuric effect in membranous glomerulonephritis (MGN). However, no studies have investigated whether this antiproteinuric effect is influenced by urinary C5b-9 excretion, a marker of immunological activity in this disease. METHODS: Eleven patients with biopsy-proven MGN were treated with captopril for 8 weeks. The evolution of several clinical and biochemical parameters, including 24-h urinary protein excretion was evaluated every 4 weeks. Urinary C5b-9 excretion was measured at the onset and at the end of captopril treatment. RESULTS: Patients with MGN had significantly higher C5b-9 excretions than a group of 14 healthy controls (89 +/- 23 vs 3.7 +/- 1.4 ng/mg UCr; P < 0.001). A significant correlation was found between urinary C5b-9 and the magnitude of proteinuria, both at the onset and at the end of treatment. After 8 weeks of captopril treatment, proteinuria had decreased from 8 +/- 1.8 to 5.2 +/- 1.3 g/day (P < 0.05). Four weeks after captopril discontinuation, proteinuria rose to 7.3 +/- 1.7 g/day (P < 0.05). A marked variability in the antiproteinuric response was observed, ranging from 0 to 85% with respect to baseline values. No correlation between decrease in proteinuria and baseline urinary C5b-9 levels was observed. Several patients with elevated urinary C5b-9 levels had captopril-induced decrease in proteinuria. CONCLUSIONS: ACE inhibition induces an antiproteinuric effect in patients with MGN. The urinary C5b-9 excretion does not predict the magnitude of this response.

Adult↗
PubMed2000

Complement components of the innate immune system in health and disease in the CNS.

The innate immune system and notably the complement (C) system play important roles in host defense to recognise and kill deleterious invaders or toxic entities, but activation at inappropriate sites or to an excessive degree can cause severe tissue damage. C has been implicated as a factor in the exacerbation and propagation of tissue injury in numerous diseases including neurodegenerative disorders. In this article, we review the evidence indicating that brain cells can synthesise a full lytic C system and also express specific C inhibitors (to protect from C activation and C lysis) and C receptors (involved in cell activation, chemotaxis and phagocytosis). We also summarise the mechanisms involved in the antibody-independent activation of the classical pathway of C in Alzheimer's disease, Huntington's disease and Pick's disease. Although the primary role of C activation on a target cell is to induce cell lysis (particularly of neurons), we present evidence indicating that C (C3a, C5a, sublytic level of C5b-9) may also be involved in pro- as well as anti-inflammatory activities. Moreover, we discuss evidence suggesting that local C activation may contribute to tissue remodelling activities during repair in the CNS.

Central Nervous System↗
PubMed1976

Isolation of the terminal complement complex from target sheep erythrocyte membranes.

(1) Membranes from sheep erythrocytes lysed with antibody and human complement were solubilized in Triton X-100 and subjected to isoelectric focusing in polyacrylamide gels containing 1% Triton X-100. Membrane-bound serum proteins were located in the gels by subsequent immunoelectrophoresis against antisera to human serum proteins. Monospecific antisera against C9 and C5 were used to locate the terminal complement complex, which is not dissociated by Triton X-100. The complex focused between pH 5.8 and pH 6.5 and was separated from the bulk of other membrane-bound serum proteins, which focused at pH ranges below than 6.0. (2) In a second step, proteins electrophoretically eluted from the gel sections containing the terminal complement complex were chromatographed on Sepharose 6B equilibrated with 0.05% Triton X-100. Fused rocket immunoelectrophoresis was used to monitor separations. This step separated the terminal complement complex from the remaining contaminating proteins. The complex eluted in a broad peak corresponding to a molecular weight range of 800000-4000000. (3) The terminal complement complex thus obtained migrated with alpha-mobility and yielded a single precipitation arc in crossed immunoelectrophoresis using polyvalent antisera to human serum proteins. A distinct precipitate was obtained with monospecific anti-C9. The presence of C5 and C6, in complex with one another and with C9 was demonstrable by immuno-double-diffusion. No immunoprecipitate was obtained with antisera to sheep erythrocyte membrane proteins. (4) Dodecyl sulfate gel electrophoresis of the complex revealed seven protein bands of 190000, 160000, 115000, 93000, 85000, 68000 and 60000 daltons. Planimetric quantitation of densitometric scans gave a molar ratio of approx. 0.7:0.3:1:1:1:2:1 for these bands, respectively. All bands stained faintly with periodate-Schiff. Two-dimensional dodecyl sulfate gel electrophoresis showed that the first two bands (190000 and 160000 daltons, probably C5b and C5c) represented proteins possessing more than one peptide chain linked by disulfide bonds. The main subunit for both bands was a protein of approximately 68000 daltons. Band 5 (83000 daltons, probably C8alpha) was split into two peptide chains of approximately 68000 and 15000 daltons. The other components were not affected by dithiothreitol treatment. (5) The dodecyl sulfate gel electrophoretograms obtained were very similar to that described by Kolb and Müller-Eberhard (Kolb, W.P. and Müller-Eberhard, H.J. (1975) J. Exp. Med. 141, 724-735) for the terminal complement complex isolated from inulin-activated serum. However, certain minor but consistent deviations were observed. A preliminary correction of the electrophoretograms is presented.

Animals↗
PubMed1991

Evaluation of thromboxane production and complement activation during myocardial ischemia in patients with angina pectoris.

BACKGROUND: The complement system and arachidonic acid metabolites are involved in severe myocardial ischemia such as myocardial infarction. Furthermore, there is experimental evidence for C5a participation in thromboxane production. METHODS AND RESULTS: We examined whether C5a and thromboxane are produced during brief and reversible episodes of myocardial ischemia induced in patients with stable angina. Twenty-five patients underwent either atrial pacing or percutaneous transluminal coronary angioplasty associated with arterial and coronary sinus blood sampling. Rapid atrial stimulation of patients with effort angina caused significant ST segment depression (delta ST = -1.7 +/- 0.2 mm), decreased fractional lactate extraction (from +12.8 +/- 2.5% baseline to -13.7 +/- 4.6% at peak ischemia, n = 13, p less than 0.001), and increased coronary sinus plasma thromboxane B2 levels (from 345 +/- 85 pg/ml baseline to 1,684 +/- 64 pg/ml at peak ischemia, p less than 0.01). Changes of fractional lactate extraction correlated significantly with changes of coronary sinus plasma levels of thromboxane B2. There was no change of coronary sinus 6-keto-PGF1 alpha levels. Similar pacing of control subjects (n = 6) did not cause release of lactate or thromboxane. Seventeen other patients underwent exercise testing with noninvasive measurements of thromboxane and prostacyclin metabolites in urinary samples collected before and after the test. No detectable increase of urinary 11-dehydrothromboxane B2 was measured in patients with stable angina after exercise-induced myocardial ischemia. However, basal 11-dehydrothromboxane B2 levels were significantly higher in patients with angina (105 +/- 25 pg/mmol creatinine, n = 9) than in control patients (45 +/- 8 pg/mmol creatinine, n = 8, p less than 0.05 between groups). Coronary sinus plasma levels of the anaphylatoxin C5a always remained below 4 ng/ml in patients undergoing pacing. More severe myocardial ischemia after coronary angioplasty (percent lactate extraction decreased from +24.8 +/- 2.7% baseline to -41.6 +/- 22.4% at peak ischemia, p less than 0.05) was not associated with C3a or C5b-9 generation. In all patients, there was neither platelet sequestration nor platelet alpha-granule release (no changes of beta-thromboglobulin/platelet factor 4 levels) into the coronary sinus plasma. CONCLUSIONS: Patients with stable angina have chronically increased thromboxane synthesis as assessed by excretion of urinary metabolites. Thromboxane is acutely released into the coronary sinus during pacing-induced ischemia without significant intracoronary platelet aggregation. Complement does not appear to be activated in stable angina during brief and reversible episodes of myocardial ischemia and does not contribute to thromboxane production.

Angina Pectoris↗
PubMed2005

Immune complement activation on polystyrene and silicon dioxide surfaces. Impact of reversible IgG adsorption.

We have studied aspects of the molecular background to immune complement activation on solid surfaces. Quartz crystal microbalance with dissipation monitoring (QCM-D) sensor surfaces were modified by means of spin coating with polystyrene (PS) or sputtering of silicon dioxide (SiO2). The IC activation on modified QCM-D surfaces was investigated by incubation in serum, followed by determinations of the amounts of bound C3 fragments (C3c) at the surface. Determinations of soluble C3a and soluble C5b-9 complex (sC5b-9) were made with enzyme immunoassay (EIA) method. We found that IC activation was high on PS surfaces, independent of the method used for measurements. On the SiO2 surfaces, IC activation was generally lower, but still detectable with anti-C3c as well as sC5b-9 and C3a determinations. Pre-coating the surfaces with a layer of IgG resulted in that IC activation became very high on PS surface, while the IC response remained low on SiO2 surfaces. The lower level of IC activation on the SiO2 surfaces was explained by a low surface concentration of IgG as measured with QCM-D. This was a result of the high reversibility of the IgG protein adsorption as well as absence of sufficient conformational changes of adsorbed IgG molecules. The QCM-D method was as sensitive as the C3a and sC5b-9 determinations to reveal surface associated IC-activation on these model surfaces. Additional advantages of the QCM-D method are the broad dynamic measurement window, i.e. the high precision and the ability to perform time resolved measurements and the ease of making different surface modifications.

Adsorption↗
PubMed2001

Sublytic C5b-9-stimulated Schwann cell survival through PI 3-kinase-mediated phosphorylation of BAD.

Sublytic C5b-9 induces cell cycle activation, proliferation, and rescue from apoptosis in Schwann cells. The signaling pathways for C5b-9-mediated rescue were investigated. Following serum withdrawal, DNA fragmentation, detected by TUNEL and FACS analysis, was 56.7% +/- 7.3 and 91.9% +/- 2.4 in cultured sciatic nerve Schwann cells from 6-day-old rats after 18 h and 24 h, respectively. Apoptosis was confirmed by inhibition of DNA fragmentation in a dose-dependent manner by DMQD-CHO, a caspase-3 inhibitor. Treatment with sublytic C5b-9 generated with purified components (C5*9) or Ab+C7-depleted serum (C7dHS)+C7 rescued 89% and 86% of Schwann cells, respectively, as compared with cells treated with C5*6, C8, C9, or Ab+C7dHS. Sublytic C5b-9 increased Schwann cell PI-3 kinase and Akt activity maximally at 5 min 3.14 +/- 0.5-fold and 3.56 +/- 0.4-fold, respectively, over controls. ERK-1 activity was maximally stimulated 2.98-fold at 15 min. Inhibition of PI-3 kinase by LY294002 abrogated the C5b-9-mediated Schwann cell rescue from apoptosis, while inhibition of ERK-1 with PD098,059 did not. PI-3 kinase-Akt pathway activation by C5b-9 induced, within 15 min, a 6.34 +/- 1.2-fold increase in BAD phosphorylation at Ser 136, but not at Ser 112. Downstream Bcl-x(L) protein was increased 2.61-fold +/- 0.34-fold by 18 h and 3.9-fold +/- 0.84-fold by 24 h over controls. LY294002 prevented both BAD phosphorylation at Ser 136 and Bcl-x(L) protein induction, while PD098,059 did not. Our data indicated that sublytic C5b-9 rescued Schwann cell from apoptosis via activation of PI-3 kinase-Akt, BAD phosphorylation on Ser 136 and increased expression of Bcl-x(L). Sublytic C5b-9 detected on Schwann cell in vivo during inflammatory neuropathy may facilitate survival of Schwann cell capable of remyelination.

Animals↗