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PubMedDate not supplied

Plasma levels of endotoxin and anaphylatoxin in patients with multiple organ failure.

Plasma concentrations of endotoxin and anaphylatoxins were studied inpatients with multiple organ failure (MOF). Thirty one blood samples from nine patients with MOF and twenty samples from preoperative patients as control were determined. The average plasma concentration of endotoxin was 188.7 +/- 37.0 pg-ml-1 (Mean + S.E.) in MOF patients and 12.4 +/- 3.3 pg.ml-1 in control patients (p less than 0.05). Plasma anaphylatoxin (C3a, C5a) levels increased significantly and C4 level decreased in MOF patients as compared with those in control patients. However, no significant correlation was found between plasma endotoxin and anaphylatoxin levels. Endotoxin does not seem to be a major factor in activating of the complement system in patients with MOF.

Adult↗
PubMed1996

Reduction of granulocyte activation during hemodialysis with regional citrate anticoagulation: dissociation of complement activation and neutropenia from neutrophil degranulation.

Neutropenia and degranulation of neutrophils during hemodialysis with cellulosic membranes have been linked to complement activation, whereas in the synthetic polymethyl methacrylate (PMMA) membrane, degranulation occurs without notable complement activation. The mechanisms of neutrophil degranulation under these conditions have not yet been elucidated. Ionized calcium is an important prerequisite of granulocyte activation during in vitro blood contact with both types of artificial surfaces. This study compared the effect of normal ionized calcium during heparin anticoagulation with the effect of extracorporeal calcium depletion during regional citrate anticoagulation on activation of blood components. Because ionized calcium is reduced only in the extra-corporeal circuit, citrate anticoagulation in addition helps to differentiate between extracorporeal and systemic activation phenomena. Twelve chronic hemodialysis patients were dialyzed with polymethyl methacrylate (PMMA, 16 treatments) or cuprophane (CUP, 16 treatments) membranes either during regional citrate anticoagulation or while anticoagulated with heparin. During hemodialysis with CUP, anticoagulation with citrate significantly reduced neutropenia, C3a levels, and lactoferrin release. Elastase concentrations, however, were not reduced by citrate, probably because elastase release occurred not locally in the cuprophane dialyzer, but mostly in the systemic circulation of the patient. PMMA did not elevate C3a levels, and neutropenia was only mild. Both parameters were not influenced by citrate anti-coagulation. However, PMMA profoundly induced elastase and lactoferrin release during heparin anti-coagulation. Depletion of ionized calcium markedly reduced PMMA-mediated neutrophil degranulation in the extracorporeal circuit. The results indicate that ionized calcium is a requirement for neutrophil degranulation during hemodialysis. In PMMA membranes, neutrophil degranulation occurs independent of high complement levels, occurs at least partially inside the dialyzer, and requires the presence of ionized calcium in the extracorporeal circuit. In cuprophane membranes, degranulation was uncoupled from neutropenia and did not correlate with the degree of complement activation. Even in cuprophane dialysis, degranulation of secondary granules was markedly dependent on ionized calcium levels in the extracorporeal circuit.

Anticoagulants↗
PubMed2006

Identification of a complement receptor 1 peptide for inhibition of immune hemolysis.

Complement sensitization of red blood cells (RBCs) can cause life-threatening hemolytic anemias. We have previously shown that complement receptor 1 (CR1) derivatives specifically the N-terminal region with decay accelerating activity (DAA) for inactivation of a key enzyme in the complement cascade can reduce complement-mediated RBC destruction in vitro and in an in vivo mouse model of hemolytic transfusion reaction. In the present study, we have modeled the N-terminal CR1 molecule based on the X-ray crystal structure of decay accelerating factor and the NMR structure of a homologous CR1 domain. Based on the homology model, we identified a 34-mer peptide encompassing the putative DAA which in vitro reduced hemolysis, C3a release and surface C3 deposition. More importantly, this peptide at 0.6 mM was effective in prolonging survival of transfused incompatible RBCs in vivo. Our results indicate that CR1-based structure-function studies may provide insights for developing structure-derived transfusion therapeutics in the future.

Amino Acid Sequence↗
PubMed1983

Stimulation of complement production in mice by N alpha-(N-acetylmuramyl-L-alanyl-D-isoglutamine)-N epsilon-stearoyl-L-lysine.

N alpha-(N-acetylmuramyl-L-alanyl-D-isoglutamine)-N epsilon-stearoyl-L-lysine, a synthetic muramyl dipeptide analog, stimulated the production of the third component of complement (C3) in mice. The serum concentration of C3 was elevated significantly by subcutaneous treatment with a single dose (10 to 100 micrograms per mouse) of the adjuvant 24 h before assay of the serum. Thereafter, the concentration decreased gradually with time and returned to the normal level on day 4 to 5. Immunoelectrophoretic analysis of the serum revealed that the decrease in serum C3 could not be accounted for by the cleavage to C3a and C3b. By intermittent treatment with the adjuvant on every fifth day, a significant increase in serum C3 was repeated. However, no continuous retention of the serum level of C3 was established even during continuous treatment with the adjuvant once a day for 10 consecutive days. Instead, in this case, the level of C3 increased repeatedly at almost 5-day intervals.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed2001

Coagulation and complement activation.

UNLABELLED: The purpose of this investigation was to assess the effect of heparin coating of a new stent construction (Stent Graft, Jomed Implantate GmbH, Germany) on platelet and coagulation activity. METHODS: Stent grafts with an ePTFE membrane interfoliated between two stents were deployed in tubings to form Chandler loops. Fresh human blood with a low concentration of heparin was rotated for 1 h, then collected and used for measurements of platelet number, thrombin-antithrombin complex (TAT), CD11b, C3a and C5b-9. There were five study groups: Group 1, conventional unmodified stents (n = 8); Group 2, untreated stent grafts (n = 8); Group 3, heparin-coated stents and untreated membrane (n = 7); Group 4, heparin-coated stents and membrane (n = 8); Group 5, heparin-coated PVC tubings with no stents (n = 8). RESULTS: There was a significant drop in platelet count, increase in TAT-values and CD11b expression in Groups 1-3 but not in Group 4 compared to Group 5. Examination by scanning electron microscopy revealed extensive activation on non-modified stents but almost no deposition of thrombotic material on heparin-modified stent grafts. CONCLUSIONS: With unmodified stents and membrane there were signs of significant activation of platelets and coagulation. In contrast, the heparin-coated stent graft induced much less alterations, indicating improved blood compatibility.

Blood Coagulation↗
PubMed1986

Neutrophil activation in thermal injury as assessed by increased expression of complement receptors.

We studied neutrophil activation in patients with burns by serial immunofluorescent measurement of neutrophil expression of the complement opsonin receptors CR1 and CR3. CR1-dependent fluorescence was initially (days 0 through 5 after the burn) elevated (mean +/- SEM, 294 +/- 42 vs. 63 +/- 6 in the controls; P less than 0.001) and gradually returned to normal (days 6 through 8, 270 +/- 62, P less than 0.001; days 9 through 13, 185 +/- 38, P less than 0.001; days 14 through 19, 143 +/- 27, P less than 0.001; and days 20 through 50, 93 +/- 5, P less than 0.04). CR3-dependent fluorescence paralleled that of CR1. Neutrophil chemotaxis in response to zymosan-activated serum, a source of C5a, was depressed (days 0 through 5, 77 +/- 4 percent of control, P less than 0.001; days 6 through 8, 70 +/- 4 percent, P less than 0.001; days 9 through 13, 74 +/- 3 percent, P less than 0.001; days 14 through 19, 90 +/- 4 percent, P less than 0.01; and days 20 through 50, 97 +/- 3 percent, P not significant) and inversely correlated with CR1- and CR3-dependent fluorescence (r = -0.559, P less than 0.001; and r = -0.709, P less than 0.001, respectively). Plasma C3a desArg levels were above normal (100 +/- 5 ng per milliliter) throughout (days 0 through 5, 305 +/- 42; days 6 through 8, 546 +/- 69; days 9 through 13, 490 +/- 72; days 14 through 19, 409 +/- 54; and days 20 through 50, 260 +/- 36; all P less than 0.005). Thus, neutrophils in burned patients were activated as indicated by increased expression of complement receptors. The correlation between this increase and the depression of chemotaxis in response to zymosan-activated serum suggests that C5a is responsible for systemic neutrophil activation, which may contribute to the increased susceptibility to infection of patients with burns.

Adult↗
PubMed2001

The orphan receptor C5L2 has high affinity binding sites for complement fragments C5a and C5a des-Arg(74).

The substantial variations in the responses of cells to the anaphylatoxin C5a and its desarginated form, C5adR(74), suggest that more than one type of cell surface receptor for these ligands might exist. However, only a single receptor for C5a and C5adR(74), CD88, has been characterized to date. Here we report that the orphan receptor C5L2/gpr77, which shares 35% amino acid identity with CD88, binds C5a with high affinity but has a 10-fold higher affinity for C5adR(74) than CD88. C5L2 also has a moderate affinity for anaphylatoxin C3a, but cross-competition studies suggest that C3a binds to a distinct site from C5a. C4a was able to displace C3a, suggesting that C5L2, like the C3a receptor, may have a low binding affinity for this anaphylatoxin. Unlike CD88 and C3a receptor, C5L2 transfected into RBL-2H3 cells does not support degranulation or increases in intracellular [Ca(2+)] and is not rapidly internalized in response to ligand binding. However, ligation of C5L2 by anaphylatoxin did potentiate the degranulation response to cross-linkage of the high affinity IgE receptor by a pertussis toxin-sensitive mechanism. These results suggest that C5L2 is an anaphylatoxin-binding protein with unique ligand binding and signaling properties.

Amino Acid Sequence↗
PubMed1985

Sodium morrhuate stimulates granulocytes and damages erythrocytes and endothelial cells: probable mechanism of an adverse reaction during sclerotherapy.

Stimulated by a patient with dyspnea, thrombocytopenia, and leukopenia after sodium morrhuate sclerotherapy, we studied the effect of this agent on the plasma coagulation and complement systems, the formed elements of the blood, and cultured human endothelial cells. The addition of sodium morrhuate to citrated plasma did not cause clotting or shorten the prothrombin time or partial thromboplastin time. Incubation of a 1:100 dilution of the clinical sodium morrhuate preparation in heparinized plasma led to a modest rise in [C3a]. The addition of the drug (dilutions 1:50 to 1:300) to granulocytes caused prompt aggregation (and, at the higher concentrations, granulocyte cytotoxicity [trypan blue exclusion; lactate dehydrogenase release]), but the same dilutions failed to aggregate platelets. However, 0.05% morrhuate added to washed red blood cells caused a prompt 84.0% (+/- 0.8% SEM) hemolysis, rendering the supernatant buffer a potent platelet aggregant. Not only was this sclerosing agent toxic to granulocytes and red cells, but a 1:1000 dilution of the drug also caused the destruction of 35.5% (+/- 6.6%) of cultured endothelial cells as measured by chromium 51 release. Three other agents in current use (ethanolamine oleate, sodium tetradecyl sulfate, and polidocanol) were studied and found to cause effects qualitatively similar to those of sodium morrhuate. We conclude that these drugs cause phlebosclerosis not primarily through induction of plasma coagulation, but by directly damaging endothelium and red cells, triggering platelets, and aggregating granulocytes at the venous wall endothelium. These effects likely derive from the surfactant properties of sodium morrhuate as well as its high arachidonate content.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood Coagulation↗
PubMed1995

Attenuation of hyperoxic lung injury in rabbits with superoxide dismutase: effects on inflammatory mediators.

BACKGROUND: Superoxide dismutase (SOD) has been shown to attenuate hyperoxic lung injury. This effect is thought to be exhibited by scavenging superoxide released from neutrophils and other aerobic cells. The aim of the current study was to investigate the effect of pre-treatment with SOD on the chemical mediators in hyperoxic lung injury. METHODS: Thirty male anesthetized rabbits were allocated to receive one of three treatments (n = 10 for each group): ventilated with 100% oxygen for 36 h with or without recombinant human SOD (rhSOD) treatment, and ventilated with air for 36 h without rhSOD. In the rhSOD-treated group, a single intravenous dose of rhSOD 10,000 U kg BW-1 was administered immediately after the start of exposure to 100% oxygen and thereafter infused at a rate of 340,000 U kg BW-1 day-1 for 36 h until the animals were sacrificed. The lungs of all rabbits were ventilated with 100% oxygen or air. Haemodynamics, PaO2, and lung mechanics were recorded during the ventilation period. After exposure to 100% oxygen, lung mechanics, cell fraction of bronchoalveolar lavage fluid (BALF), activated complements, cytokines, and arachidonic acid metabolite concentrations in BALF were measured and analyzed. The lung wet-to-dry (W/D) weight ratio and albumin concentrations in BALF were determined as indices of pulmonary oedema. RESULTS: Exposure to the high concentration of oxygen for 36 h caused no significant changes in haemodynamics but decreased compliance and increased A-aDo2. In the rhSOD-treated group, the decrease in compliance was not observed. At the end of the 36 hr-exposure period, however, hyperoxia significantly increased the lung W/D weight ratio, influx of neutrophils into the lung, BALF concentrations of C3a, C5a, tumor necrosis factor-alpha, interleukin (IL)-1 beta, IL-6, IL-8, thromboxane B2, and albumin. Pre-treatment with rhSOD attenuated these increases. Exposure to 100% oxygen caused extensive morphologic lung damage (alveolar haemorrhage and hyaline membrane formation), which was lessened by rhSOD. CONCLUSION: These results indicate that intravenous rhSOD prevented hyperoxic lung injury (decreases in PaO2 and compliance, increased pulmonary vascular permeability, histopathological damage) in rabbits. This prophylactic effect of rhSOD amy be due, in part, to decreased chemical mediators such as activated complements, cytokines, and arachidonic acid metabolites.

Animals↗
PubMed1980

Relation of putative thioester bond in C3 to activation of the alternative pathway and the binding of C3b to biological targets of complement.

The reaction of [14C]methylamine with native human C3 led to the stoichiometric incorporation of methylamine, loss of hemolytic activity, and the concomitant exposure of a sulfhydryl group that could be labeled with [14C]iodoacetamide. Both labeled sites were located in the C3d portion of the alpha-chain, which is known to contain the metastable binding of C3b. The methylamine-modified C3 [C3(CH3NH2)] was shown to exhibit many of the functional properties of C3b, although the C3a portion of the molecule remained covalently attached. C3(CH3NH2) bound Factor B and beta 1H, and could be cleaved by C3b inactivator in the presence of beta 1H. C3(CH3NH2) added to human serum caused activation of the alternative pathway and consumption of C3. In presence of Factors B and D and Mg++, C3(CH2NH2) formed a C3 convertase. The convertase-forming material could be removed from solution by anti-C3a Sepharose and the preformed convertase was completely inhibited by purified antibody to C3a. This antibody did not affect the function of the C3 convertase that contained C3b. Similar functional properties were exhibited by C3 exposed for short periods of time to relatively low concentrations of chaotropic reagents, such as KSCN or guanidine. These results suggest that the initial C3 convertase of the alternative pathway may be formed from native C3, without proteolysis, by the attack of a variety of nucleophiles including water. The C3 convertase formed from this altered C3 then generates by proteolytic cleavage the initial metastable C3b that is capable of attaching to receptive surfaces. Conversion of C3 to C3b exposes one sulfhydryl residue as does modification of C3 with methylamine. When the C3d portion of C3b bound to zymosan particles via the metastable binding site was treated with radiolabeled methylamine, the fragment was released from the particles in radiolabeled form. These findings are consistent with the concept that native C3 contains an active carbonyl group, probably in the form of a thioester, which can either react with water to form functionally C3b-l;ike C3 or, upon enzymatic conversion of C3 to C3b, allows C3b to form an ester bond with hydroxyl groups on the target surface.

Complement Activation↗
PubMed2004

Reversal of diabetes in non-immunosuppressed rhesus macaques by intraportal porcine islet xenografts precedes acute cellular rejection.

BACKGROUND: The functional response and immunobiology of primarily non-vascularized islet cell xenografts remain poorly defined in non-human primates. METHODS: We transplanted 20,000 adult porcine islet equivalents/kg (purified and cultured for 48-h) intraportally into six streptozotocin-diabetic and two non-diabetic rhesus macaques. Two recipients were killed at various intervals post-transplant for histologic examination of livers bearing xenografts. RESULTS: Plasma glucose levels in diabetic recipients averaged 94 mg/dl at 12 h, 92 mg/dl at 24 h, 147 mg/dl at 48 h, and 157 mg/dl at 72 h post-transplant. Serum porcine C-peptide was present in eight of eight recipients at 12 h, in five of six at 24 h, in four of four at 48 h, and in one of two at 72 h post-transplant. C3a and SC5b-9 plasma levels increased at 12 h post-transplant and returned to pre-transplant levels by 24 h. IgG, IgM anti-pig and anti-Gal IgG serum antibody levels did not increase post-transplant. Rejection was initiated by IgM and complement deposition on islets. Neutrophils dominated the cellular infiltrate at 12 h; CD4+ and CD8+ T cells were the main infiltrating cells at 24, 48, and 72 h; and macrophages increasingly infiltrated xenografts starting at 24 h post-transplant. Numerous xenoislets were present at all time points; their proportion without intraislet infiltrates decreased from 65% at 24 h to 17% at 72 h post-transplant. CONCLUSIONS: Pig-to-primate intraportal islet xenografts reverse diabetes and the majority of intraportally transplanted xenogeneic islets are not subject to hyperacute rejection. They undergo acute cellular rejection mediated by CD4+- and CD8+ T cells and macrophages.

Acute Disease↗
PubMed1979

Anaphylatoxin-induced histamine release with human leukocytes: studies of C3a leukocyte binding and histamine release.

Purified human C3a and synthetic COOH-terminal peptides of C3a, i.e., a pentapeptide, Leu-Gly-Leu-Ala-Arg (5R), and an octapeptide, Ala-Ala-Ala-Leu-Gly-Leu-Ala-Arg (8R) induced histamine release from human basophil granulocytes. On a molar basis, 5R was one-tenth and 8R was one-fifth as active as C3a in causing histamine release. It was found that 125I-C3a binds to whole leukocytes, interacting with both mononuclear cells and neutrophils and the binding was inhibited by preincubation of cells with unlabeled C3a, but not by C5a. 5R and 8R also inhibited the binding of 125I-C3a to the cells. However, on a molar basis, 2,000 times more 8R or 6,000 times more 5R is required for 50% inhibition of 125I-C3a binding as compared with native C3a. Autoradiography of cells using 125I-C3a and 125I-C5a showed preferential binding of 125I-C3a to eosinophils and basophils, whereas 125I-C5a binds primarily to neutrophils and eosinophils and to a lesser extent to basophils. The preferential binding of C3a and C5a to different cell types may herald significance related to their physiological functions.

Anaphylatoxins↗
PubMed2005

Complement activation-related pseudoallergy: a new class of drug-induced acute immune toxicity.

A major goal in modern pharmacotechnology is to increase the therapeutic index of drugs by using nanoparticulate vehicle systems in order to ensure slow release or targeted delivery of drugs. With all great benefits, however, these innovative therapies can carry a risk for acute immune toxicity manifested in hypersensitivity reactions (HSRs) that do not involve IgE but arises as a consequence of activation of the complement (C) system. These anaphylactoid reactions can be distinguished within the Type I category of HSRs as "C activation-related pseudoallergy" (CARPA). Drugs and agents causing CARPA include radiocontrast media (RCM), liposomal drugs (Doxil, Ambisome and DaunoXome) and micellar solvents containing amphiphilic lipids (e.g., Cremophor EL, the vehicle of Taxol). These agents activate C through both the classical and the alternative pathways, giving rise to C3a and C5a anaphylatoxins that trigger mast cells and basophils for secretory response that underlies HSRs. Pigs provide a useful model for liposome-induced CARPA as minute amounts of reactogenic lipomes cause C activation with consequent dramatic cardiovascular and laboratory abnormalities that mimic some of the human symptoms. Consistent with the causal role of C activation in liposome-induced HSRs, a recent clinical study demonstrated correlation between the formation of C terminal complex (SC5b-9) in blood and the presence of HSRs in patients treated with liposomal doxorubicin (Doxil). Overall, the CARPA concept may help in the prediction, prevention and treatment of the acute immune toxicity of numerous state-of-the-art drugs.

Anaphylaxis↗
PubMed1987

Neutrophil activation after burn injury: contributions of the classic complement pathway and of endotoxin.

We attempt to elucidate the mechanisms of neutrophil (PMN) activation after burn injury. We previously reported prolonged elevations of PMN cell surface complement (C) opsonin receptor levels after burn trauma with a corresponding period of depressed PMN chemotaxis to C5a, which suggests that the C product, C5a, was responsible for PMN activation. However, a lack of direct correlation of C activation with C receptor levels soon after injury raised the possibility of a second PMN-activating substance. We therefore investigated the effect of endotoxin (LPS) on the expression of the C receptors (CR1 and CR3) by normal human PMNs. Concentrations from 0 to 50 ng/ml of LPS 026:B6 caused a dose response increase in the PMN surface expression of CR1 and CR3 as assessed by monoclonal antibody binding and indirect immunofluorescence. The relative CR1-dependent fluorescence rose from a mean of 50 to 385 and CR3 from 50 to 300. Chelation by ethylenediaminetetra acetic acid (EDTA) did not influence this dose response, thus ruling out the possibility of C activation by LPS--an inference supported by the lack of complement activation observed with these concentrations of LPS in normal serum. A similar dose response was obtained in the absence of other cell types or serum, which implies a direct effect that mimicked that of C5a. To determine the mechanism of the later, prolonged C activation after burn injury, we next examined C activation products in 22 patients with burn injuries. Elevations of plasma C3a desArg were present and persisted for 50 days. Elevations were at maximum levels on days 9 through 13 postburn (mean +/- standard error of mean [SEM], 496 +/- 47 ng/ml versus normal 113 +/- 32; p less than 0.01). These were accompanied by elevations of C4a desArg (917 +/- 154 ng/ml versus normal 424 +/- 50; p less than 0.01), which are indicative of classic pathway activation. Finally, we examined PMN function, phagocytosis and percentage killing of Staphylococcus aureus, and found PMN function to be unaltered in the 22 patients. Thus PMN activation after burn injury appears to be caused by LPS soon after injury and by C5a later after injury and affects only selected PMN functions.

Adolescent↗
PubMed1999

Consumption of C4b-binding protein (C4BP) during in vivo activation of the classical complement pathway.

C4BP has a central role in regulating the classical complement (C') pathway, but it is still uncertain whether or not it is consumed during in vivo complement activation. Attempts to demonstrate changes in C4BP plasma levels in systemic lupus erythematosus and essential mixed cryoglobulinaemia have failed, probably due to up-regulation of this protein during the inflammatory reaction. We have studied one patient with severe post-transfusion complement-mediated anaphylaxis (CMA), and 67 patients with hereditary C1 inhibitor deficiency (hereditary angioedema (HAE)). The first of these two conditions is characterized by the absence of systemic inflammatory reaction and the second by acute and chronic activation of the C' classical pathway. C4BP, C4BP-C4b complex, and soluble terminal C' complex (sC5b-9) were measured in the patients' plasmas by ELISA techniques and C3a and C4a by radioimmunoassays. In CMA, 15 min after the transfusion, there was a massive C' activation, with increases in C4a, C3a, sC5b-9, C4BP-C4b complexes and decreases in C4, C3 and C4BP. All parameters reverted to preinfusion values within 24 h. Depletion of C4 was correlated with that of C4BP. In patients with HAE, the median value of C4BP (83% range 54-165) was significantly lower (P < 0.0001) than in normal controls (99% range 70-159), with no difference between patients in remission or during acute attacks. C4BP-C4b complexes could not be detected in HAE patients. The results of this study indicate that C4BP is consumed in vivo during acute, and possibly during chronic activation of the C' classical pathway, and that this protein, after interaction with C4b, not longer circulates in plasma.

Adult↗
PubMed1991

Leishmanial protein kinases phosphorylate components of the complement system.

Externally oriented protein kinases are present on the plasma membrane of the human parasite, Leishmania. Since activation of complement plays an important role in the survival of these parasites, we examined the ability of protein kinases from Leishmania major to phosphorylate components of the human complement system. The leishmanial protein kinase-1 (LPK-1) isolated from promastigotes of L. major was able to phosphorylate purified human C3, C5 and C9. Only the alpha-chain of C3 and C5 was phosphorylated. The beta-chain appeared not to be a substrate for this enzyme. C3b which is formed by proteolytic cleavage of C3 was not phosphorylated by LPK-1. Trypsin treatment of phosphorylated C3 (P-C3) resulted in the disappearance of 32P from the alpha-chain. This was correlated with the conversion of the C3 alpha-chain to the alpha'-chain of C3b, and the appearance of a 9 kDa 32P fragment comigrating with the C3a fragment of C3. P-C3 was more resistant to cleavage by trypsin than nonphosphorylated C3. LPK-1 phosphorylated purified C3a and two synthetic peptides, C3a21R and YA-C3a10R, derived from its COOH-terminal end, which contain the C3a binding site to leukocytes and platelets. LPK-1 did not phosphorylate C3a8R. Phosphoamino acid analysis of the synthetic peptides indicated that serine 71 of C3a was phosphorylated by LPK-1. Treatment of C3 with either methylamine or freeze-thaw C3 (H2O) prevented phosphorylation by the LPK-1 suggesting that substrate conformation may be involved in recognition by the leishmanial enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1984

Role of C3b receptors in the enhancement of interleukin-2-dependent T-cell proliferation.

The mechanism by which the complement system influences immune responses to T-cell-dependent antigens has not yet been clarified. That is why we studied the effect of the third complement component (C3) on different T-cell-dependent processes using well-defined mouse T-cell lines. While C3 did not influence the interleukin-2 (IL-2) production of the ST2/K-9 helper T-cells, the IL-2-dependent proliferation of the ST1 line was shown to be dose-dependently enhanced by C3. It is proved that neither the haemolytic activity of C3 nor the C3a fragment had any role in the process. The effect of C3 on the IL-2-dependent T-cell growth is even more enhanced (up to five-fold) when using polymerised C3. When the ST1 cell line is cultured in the presence of the cross-linked ligand, T-cells formed 80% less rosettes with red blood cells coated with antibody and mouse or human C3b. It is strongly suggested that C3--particularly when aggregated--exerts its enhancing effect on the growth of IL-2-dependent cell lines by binding to C3b receptors present on such T-cells.

Animals↗
PubMed1985

Common evolutionary origin of alpha 2-macroglobulin and complement components C3 and C4.

A comparison of the sequence of the subunit of human alpha 2-macroglobulin (alpha 2M; 1451 amino acid residues) with that of murine complement component pro-C3 (1639 amino acid residues) reveals eight extended regions of sequence similarity. These regions contain between 19% and 31% identically placed residues and account for 75% and 67%, respectively, of the polypeptide chains of alpha 2M and pro-C3. Published sequence data for complement component C4 show that segments of this protein match well with corresponding stretches in alpha 2M and pro-C3. It is proposed that alpha 2M, C3 and C4, which all contain a unique activatable beta-cysteinyl-gamma-glutamyl thiol ester, have a common evolutionary origin and are homologous proteins. Several larger regions of low sequence similarity indicate the presence of structural domains in each of these proteins that specifically modify an underlying common gross structure. The quartets of basic residues in pro-C3 and pro-C4, at which cleavage takes place to produce the mature subunits of these proteins, and most of the residues forming the anaphylatoxin peptides of C3 and C4 (C3a and C4a) are absent in alpha 2M. In addition, C3 and C4 contain large portions, which extend beyond the COOH terminus of alpha 2M.

Amino Acid Sequence↗