Complement: a host defense mechanism ready for pharmacological manipulation?
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Murine cell-derived MLV vector particles usually are highly sensitive to human complement-mediated lysis. Expression of the human complement inhibitor CD59 on murine packaging cells resulted in partial protection of these cells from lysis caused by human complement proteins. Furthermore, CD59 was incorporated into MLV vector particles released by these packaging cells, leading to an improved resistance of the virions against human complement-mediated inactivation.
Anticomplementary activity (ACA) of aggregates in intravenous immunoglobulin preparations (IVIG) was investigated using the modified Kabat and Meyer classical complement consumption method recommended by the European Pharmacopoeia and a C1q-coated microtitre enzyme-linked immunosorbent assay (ELISA). The physical characteristics of aggregates were found to affect complement binding. Aggregates formed by heating IVIG preparations at acid pH bound complement poorly, while aggregates formed by heating IVIG at neutral pH showed high ACA. This suggests that analysis of complement binding capacity provides a level of aggregate characterization of aggregates which is additional to quantitation by High-performance liquid chromatography ((HPLC). The correlation (r = 0.98) between the two tests was good when aggregates formed at neutral pH were compared, but decreased (r = 0.57) when aggregates formed at acid pH were included. A comparison of the results showed that there were no significant differences in the classification of aggregates with acceptable/unacceptable (i.e. pass/fail outcome) values of ACA. Between assay variation (CV = 7.6%) was lower in the ELISA test compared with the complement consumption assay (where percentage binding varied from 78.9% to 100%). Both assays are justified for the evaluation of ACA in therapeutic IVIG. The ELISA had the advantage in being more precise, less dependent on reagent source and requiring less technical expertise.
Previous studies have documented that liposome-encapsulated hemoglobin (LEH) can cause a rapid and transient thrombocytopenia following intravenous injection into small animals. The present study evaluated the role of complement during the LEH-induced thrombocytopenia in rats. We have compared changes in platelet levels in the blood, platelet organ distribution, and total hemolytic complement levels following intravenous administration of LEH in control and complement-depleted rats. Changes in platelet organ distribution at various times after LEH administration were monitored by labeling autologous platelets with indium-111 (111In)-oxine and imaging the 111In-platelets with a gamma camera after reinjection. Platelet counts were determined by light-scattering methods and by following 111In radioactivity at various times after LEH administration. Platelet levels did not significantly change for the complement-depleted rats during the 60 min following an injection of LEH, whereas thrombocytopenia (40% decrease) was noted within 4 min post-LEH-injection for control rats with a gradual return to baseline circulating platelet levels within 60 min. This drop in circulating platelets was correlated with a rapid redistribution of 111In-platelets from the circulation to the lungs and liver, whereas complement-depleted rats showed no transient movement of the 111In-platelets from the circulation. Baseline complement levels of 21.6 +/- 2.2 CH50/ml for control rats and 0.2 +/- 0.1 CH50/ml for complement-depleted rats did not significantly change during the 60 min following LEH administration. This study suggests that complement must be present during LEH-induced transient thrombocytopenia, as complement-depleted rats underwent no thrombocytopenia, and that the transient LEH-induced thrombocytopenia may be associated with complement activation.
The substantial virus lysis was induced by HIV-1-infected patient serum and normal human complement serum in the presence of purified patient IgG. Non-infected CD4+ T cells coated with the whole virus or with a recombinant HIV-1 envelope gp120 and sensitised with patient IgG were also shown to be susceptible to complement-dependent lysis. The serum level of complement regulatory protein in a fluid phase, the C1-esterase inhibitor, was significantly correlated with serum concentration of C1q-circulating immune complexes (P=0.0062), but inversely with CD4+ T cell count (P < 0.0001). Accordingly, the disease progression in HIV-1-infected patients was significantly correlated with the level of complement activation as determined by serum level of C1-esterase inhibitor (P=0.0001), and inversely correlated with CD4+ cell count (P < 0. 0001) and gp120-specific antibody titre (P=0.0086). These results strongly suggest that the complement activation by gp120-specific antibodies play a very important role in virus clearance, but also in depletion of infected as well as gp120-coated non-infected CD4+ bystander T cells during the course of HIV-1 infection.
Vaccinia virus complement control protein (VCP) is encoded by vaccinia virus, with its homolog encoded by other pathogenic poxviruses including variola virus. Since rodents are the primary reservoir hosts of cowpox virus (CPV) and since CPV encodes a highly conserved functional homolog of VCP, termed here the inflammation modulatory protein (IMP), the effects of injection of CPV into the footpads of mice was determined in order to study the precise in vivo effects of IMP. Macroscopic examination of the site of injection with a recombinant virus lacking IMP (CPV-IMP) showed greater tissue damage, with more hemorrhage and induration, than sites injected with the wild-type cowpox virus. In addition, the measurement of the specific swelling response carried out for several weeks revealed significantly greater swelling in mice injected with CPV-IMP. Thus, IMP modulates the complement-activated inflammatory response in vivo. Furthermore, the diminished destruction of host tissue observed in the presence of IMP indicates symbiosis in which the virus ensures the preservation of surrounding host tissue, possibly to support the growth of its progeny.
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In Alzheimer's disease (AD) patients, the complement components Clq, C4 and C3 can be detected in different types of beta/A4 plaques, one of the hallmarks of AD. Contradictory findings on the presence of late complement components in AD brains have been reported. Nevertheless, it was suggested in recent studies that in AD brain complement activation results in complement membrane attack complex (MAC) formation and that complement activation may act as an intermediate between beta/A4 deposits and the neurotoxicity observed in AD. In the present study the presence of a number of complement components and regulatory proteins in AD temporal cortex and, for comparison, in glomerulonephritis (GN) was analysed. In GN kidneys, besides Clq, Clr, Cls and C3, the late components and the C5b-9 complex are also associated with capillary basement membrane and mesangial immune complex deposits. In AD temporal cortex Clq, C4 and C3 are co-localized with beta/A4 deposits. However, in contrast to the GN kidney, the late complement components C5, C7 and C9, as well as the C5b-9 membrane attack complex cannot be detected in beta/A4 positive plaques. The absence of the cytolytic C5b-9 complex in AD brain suggests that in AD, the complement MAC does not function as the proposed inflammatory mediator between beta/A4 deposits and the neurofibrillary changes.
The potential suppressive effect of oil-pollution in the aquatic environment on fish immune responses was investigated by injecting rainbow trout (Oncorhychus mykiss) with an extract obtained from diesel oil-based drilling mud. To investigate the effect of the extract dose, 4 groups of 6 fish were exposed to 0, 0.6, 1.2, and 2.4 mL extract/Kg body weight (B.W.). To keep the injection volume constant, each extract dose was made up 2.4 mL/Kg B.W. with olive oil. Six weeks later the fish were sacrificed and a number of immmune parameters monitored. In a second experiment, the effect of exposure times was investigated. Fish wee exposed to 2.4 mL extract/Kg B.W., 2.4 mL olive oil/Kg B.W., or 2.4 mL saline/Kg B.W. and immune parameters were monitored 2, 4, 6 and 8 weeks post-injection. In the dose response experiment, there was no significant effect of the extract on serum immunoglobulin (Ig) levels and haemolytic complement (CH50) activity, but a suppressive effect on serum lysozyme level was found using the 0.6 mL/Kg dose. This dose also elevated proliferative activity of head kidney lymphocytes in response to the mitogen PHA. In the second experiment, investigating exposure time, again no significant overall effect on serum Ig or lysozyme level and CH50 activity was observed, although Ig levels were significantly lower at week two relative to the other groups. However, both extract- and olive oil-injected fish showed significantly lower lysozyme levels compared with the saline-injected (control) fish throughout the experiment. In contrast, at week six post-exposure, head kidney lymphocyte proliferation was significantly elevated in both the extract- and olive oil-exposed fish relative to the control fish.
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The levels of IgM, IgG and IgA were measured in the serum of 337 psychiatric inpatients (92 patients with bipolar depression, 150 patients with unipolar depression and 95 schizophrenics) and compared to 150 healthy subjects. A significantly higher level of IgM was found in all psychiatric groups compared with the controls, and IgM levels were more elevated in female patients than in male patients for the bipolar and unipolar groups. There were no differences for the other immunoglobulins (IgG, IgA) among the groups studied. In 51 of the patients (17 bipolar, 34 unipolar), other measurements were performed (C-reactive protein, antinuclear antibodies, lymphocyte antibodies, thyroid antibodies, complement and the third and fourth factors of complement). The unipolar group showed a significant rise of C-reactive protein values and the presence of antinuclear antibodies. Interestingly all patients with antinuclear antibodies were females. No difference was found between psychiatric patients and controls in lymphocyte antibodies, thyroid antibodies and values of complement.
Serum concentrations of acute-phase-proteins C-reactive protein (CRP), alpha 1-antitrypsin (AAT), alpha 1-acid glycoprotein (AGP) as well as levels of immunoglobulins G, A and M and of complement components C3 and C4 were evaluated in 15 patients with advanced (stages III and IV) Hodgkin's disease. Of these patients 9 suffered from B symptoms including pruritus, night sweats and fever. While all patients had highly increased concentrations of CRP and AAT and 11 patients also had elevated levels of AGP in their sera, these concentrations were significantly (P less than 0.001) reducible by the administration of chemotherapy. Patients with B symptoms also had significantly higher concentrations of CRP (P less than 0.02), AAT (P less than 0.05) and AGP (P less than 0.05) in their sera than patients without. Plasmapheresis which was performed in 3 patients did not achieve a long-lasting reduction of serum concentrations of any acute-phase-protein tested. Complement components C3 and C4 exhibited a similar behaviour as acute-phase-proteins in that they were elevated in patients with B symptoms and reducible by the administration of chemotherapy (P less than 0.001 and P less than 0.02, respectively). We conclude that serum concentrations of CRP, AAT and AGP can serve as useful markers for the assessment of tumour activity in patients with advanced Hodgkin's disease. Whereas the concentrations of immunoglobulins G and A in patients were comparable to normal controls, IgM was significantly (P less than 0.05) reduced in patients who had received chemotherapy, but not in those who were newly diagnosed and had not received any treatment. Thus, chemotherapy lowered serum concentrations of IgM without influencing levels of IgG and IgA.
The intravenous injection of the anticomplementary protein from cobra venom, cobra factor (CoF),2 induces decreases in mean arterial blood pressure and circulating platelets in rabbits. The changes are rapidly reversed. Both changes require the presence of C3 and occur in rabbits genetically deficient in the sixth component of complement. The hypotensive effects of CoF were blocked by the histamine H2-receptor antagonist burimamide. An acute C3-dependent change in blood pressure and circulating platelets also was demonstrated following the intravenous injection of S. marcescens endotoxin. However, abrogation of these acute changes by C3 depletion did not alter the extent of a second, prolonged fall in blood pressure and platelets induced by S. marcescens endotoxin occurring after 60--90 min. C3 depletion also did not alter the lethal effects of the S. marcescens endotoxin.
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