Degree Of Complement Activation Research Articles

Hyper-inflammation and complement in COVID-19.

COVID-19 is a complex disease manifesting in a broad severity spectrum and involving distinct organs and systems. Hyperinflammation, including complement over-activation, has a pivotal role in severe COVID-19 pathobiology, stimulating the inflammatory response, causing microangiopathy, platelet-neutrophil activation, and hypercoagulability. SARS-CoV-2 can directly activate the complement system by the classic, alternative, and lectin pathways, and infected cells can produce intracellular complement (the complesome). COVID-19 severity appears to be associated with the degree of complement activation, and it has been hypothesized that patients with COVID-19 may benefit from therapeutic complement inhibition. Different complement cascade molecules may be targeted with potential advantages and disadvantages. Which target(s) is the most effective and when is the best timing for intervention remain open questions. Early phase I and phase II clinical trials have shown promising but conflicting results, warranting phase III controlled randomized trials. Upstream complement inhibition appears to better and more effectively block hyperinflammation with potential clinical significance. Understanding how SARS-CoV-2 exploits the complement system can add precious information about the pathogenesis of other infections, inflammatory, and autoimmune diseases beyond COVID-19.

Anti-PEG antibodies compromise the integrity of PEGylated lipid-based nanoparticles via complement

PEGylation of lipid-based nanoparticles and other nanocarriers is widely used to increase their stability and plasma half-life. However, either pre-existing or de novo formed anti-PEG antibodies can induce hypersensitivity reactions and accelerated blood clearance through binding to the nanoparticle surfaces, leading to activation of the complement system.In this study, we investigated the consequences and mechanisms of complement activation by anti-PEG antibodies interacting with different types of PEGylated lipid-based nanoparticles. By using both liposomes loaded with different (model) drugs and LNPs loaded with mRNA, we demonstrate that complement activation triggered by anti-PEG antibodies can compromise the bilayer/surface integrity, leading to premature drug release or exposure of their mRNA contents to serum proteins. Anti-PEG antibodies also can induce deposition of complement fragments onto the surface of PEGylated lipid-based nanoparticles and induce the release of fluid phase complement activation products.The role of the different complement pathways activated by lipid-based nanoparticles was studied using deficient sera and/or inhibitory antibodies. We identified a major role for the classical complement pathway in the early activation events leading to the activation of C3. Our data also confirm the essential role of amplification of C3 activation by alternative pathway components in the lysis of liposomes.Finally, the levels of pre-existing anti-PEG IgM antibodies in plasma of healthy donors correlated with the degree of complement activation (fixation and lysis) induced upon exposure to PEGylated liposomes and mRNA-LNPs.Taken together, anti-PEG antibodies trigger complement activation by PEGylated lipid-based nanoparticles, which can potentially compromise their integrity, leading to premature drug release or cargo exposure to serum proteins.

Complement Activation in Human Sepsis is Related to Sepsis-Induced Disseminated Intravascular Coagulation.

In human sepsis, little is known about the relationships between complement activation and the clinical characteristics of sepsis, including disseminated intravascular coagulation (DIC), interventions, and prognosis. Adult patients with sepsis admitted from November 2016 to December 2018 were included. We used the plasma levels of soluble C5b-9 (SC5b-9) as a marker of complement activation. We compared the clinical characteristics and complement components between patients with and without DIC. We also compared the clinical characteristics and each DIC parameter across quartile groups for the SC5b-9 value. Forty-nine sepsis patients were eligible. Thirty-four patients developed DIC, and eight patients died. The median (interquartile range) SC5b-9 value was 342 (261-501) ng/mL. Compared with patients without DIC, patients with DIC showed lower C3 levels (mean, 95.7 vs. 70.4 mg/dL, P < 0.01) and higher SC5b-9 levels (median, 287 vs. 400 ng/mL, P = 0.01). Patients were stratified by SC5b-9 quartile (ng/mL: low: < 260, moderate: 260-342, high: 343-501, highest: > 501). The mean Sequential Organ Failure Assessment score varied across these groups (P = 0.02). In the high and highest groups, many more patients received vasopressors and developed DIC. In the highest group, the coagulation parameters were severe, and thrombocytopenia was prolonged. In-hospital mortality tended to be high (33%) in the highest group. The degree of complement activation is related to DIC, severity, intensive interventions, and mortality. Further studies are needed to confirm the usefulness of SC5b-9 for stratifying sepsis patients.

Celiac disease and complement activation in response to Streptococcus pneumoniae

Individuals with celiac disease (CD) are at increased risk of invasive pneumococcal disease (IPD). The aim of this study was to explore whether the complement response to Streptococcus pneumoniae differed according to CD status, and could serve as an explanation for the excess risk of IPD in CD. Twenty-two children with CD and 18 controls, born 1999–2008, were included at Kalmar County Hospital, Sweden. The degree of complement activation was evaluated by comparing levels of activation products C3a and sC5b-9 in plasma incubated for 30 min with Streptococcus pneumoniae and in non-incubated plasma. Complement analyses were performed with enzyme-linked immunosorbent assay (ELISA). Pneumococcal stimulation caused a statistically significant increase in C3a as well as sC5b-9 in both children with CD and controls but there was no difference in response between the groups. After incubation, C3a increased on average 4.6 times and sC5b-9 22 times in both the CD and the control group (p = 0.497 and p = 0.724 respectively).Conclusion: Complement response to Streptococcus pneumoniae seems to be similar in children with and without CD and is thus unlikely to contribute to the increased susceptibility to invasive pneumococcal disease in CD.What is Known:• An excess risk of pneumococcal infections has been demonstrated in individuals with celiac disease.• Infectious complications can depend on hyposplenism but alternative mechanisms are sparsely examined.What is New:• Complement activation in response to Streptococcus pneumoniae was examined in children with and without celiac disease but no differences could be demonstrated.

Modeling Hemolytic-Uremic Syndrome: In-Depth Characterization of Distinct Murine Models Reflecting Different Features of Human Disease.

Diarrhea-positive hemolytic-uremic syndrome (HUS) is a renal disorder that results from infections with Shiga-toxin (Stx)-producing Escherichia coli. The aim of this study was to establish well-defined refined murine models of HUS that can serve as preclinical tools to elucidate molecular mechanisms of disease development. C57BL/6J mice were subjected to different doses of Stx2 purified from an E. coli O157:H7 patient isolate. Animals received 300 ng/kg Stx2 and were sacrificed on day 3 to establish an acute model with fast disease progression. Alternatively, mice received 25 ng/kg Stx2 on days 0, 3, and 6, and were sacrificed on day 7 to establish a subacute model with moderate disease progression. Indicated by a rise in hematocrit, we observed dehydration despite volume substitution in both models, which was less pronounced in mice that underwent the 7-day regime. Compared with sham-treated animals, mice subjected to Stx2 developed profound weight loss, kidney dysfunction (elevation of plasma urea, creatinine, and neutrophil gelatinase-associated lipocalin), kidney injury (tubular injury and loss of endothelial cells), thrombotic microangiopathy (arteriolar microthrombi), and hemolysis (elevation of plasma bilirubin, lactate dehydrogenase, and free hemoglobin). The degree of complement activation (C3c deposition), immune cell invasion (macrophages and T lymphocytes), apoptosis, and proliferation were significantly increased in kidneys of mice subjected to the 7-day but not in kidneys of mice subjected to the 3-day regime. However, glomerular and kidney volume remained mainly unchanged, as assessed by 3D analysis of whole mount kidneys using CD31 staining with light sheet fluorescence microscopy. Gene expression analysis of kidneys revealed a total of only 91 overlapping genes altered in both Stx2 models. In conclusion, we have developed two refined mouse models with different disease progression, both leading to hemolysis, thrombotic microangiopathy, and acute kidney dysfunction and damage as key clinical features of human HUS. While intrarenal changes (apoptosis, proliferation, complement deposition, and immune cell invasion) mainly contribute to the pathophysiology of the subacute model, prerenal pathomechanisms (hypovolemia) play a predominant role in the acute model. Both models allow the further study of the pathomechanisms of most aspects of human HUS and the testing of distinct novel treatment strategies.

Complement Regulatory Genetic Mutations in the Setting of Autoimmune Thrombotic Thrombocytopenic Purpura: A Case Series.

ObjectiveTo explore the benefits of adding eculizumab for the treatment of refractory autoimmune thrombotic thrombocytopenic purpura (iTTP) with complement dysregulation.Patients and MethodsFrom January 1, 2014, through July 1, 2017, we identified patients with iTTP defined by ADAMTS13 (disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) levels less than 5% and the presence of ADAMTS13 inhibitor. Patients who progressed after receiving standard of care management for iTTP were subjected to a comprehensive evaluation to look for evidence of complement activation. Herein, we share our single-institute experience regarding the clinical course and treatment algorithm for 3 patients with refractory iTTP.ResultsAll the patients had clinical deterioration despite treatment with plasma exchange, corticosteroids, rituximab, and vincristine, which prompted us to look for evidence of complement activation and associated genetic mutations. Complement-related genetic aberrations were present in all 3 patients, who had had different degrees of complement activation. The first 2 patients did not benefit from eculizumab when treatment was started before complete clearance of inhibitors to ADAMTS13. However, they had durable remissions when eculizumab was introduced after clearance of ADAMTS13 inhibitors. The third patient started eculizumab therapy after inhibitor levels were undetectable.ConclusionWe found eculizumab therapy to be effective in all 3 patients. However, its efficacy was prominent only after clearance of antibodies against ADAMTS13 via therapeutic plasma exchange.

Co-Complexes of MASP-1 and MASP-2 Associated with the Soluble Pattern-Recognition Molecules Drive Lectin Pathway Activation in a Manner Inhibitable by MAp44

The lectin pathway of complement is an integral component of innate immunity. It is activated upon binding of mannan-binding lectin (MBL) or ficolins (H-, L-, and M-ficolin) to suitable ligand patterns on microorganisms. MBL and ficolins are polydisperse homo-oligomeric molecules, found in complexes with MBL-associated serine proteases (MASP-1, -2, and -3) and MBL-associated proteins (MAp19 and MAp44). This scenario is far more complex than the well-defined activation complex of the classical pathway, C1qC1r(2)C1s(2), and the composition of the activating complexes of the lectin pathway is ill defined. We and other investigators recently demonstrated that both MASP-1 and MASP-2 are crucial to lectin pathway activation. MASP-1 transactivates MASP-2 and, although MASP-1 also cleaves C2, MASP-2 cleaves both C4 and C2, allowing formation of the C3 convertase, C4bC2a. Juxtaposition of MASP-1 and MASP-2 during activation must be required for transactivation. We previously presented a possible scenario, which parallels that of the classical pathway, in which MASP-1 and MASP-2 are found together in the same MBL or ficolin complex. In this study, we demonstrate that, although MASPs do not directly form heterodimers, the addition of MBL or ficolins allows the formation of MASP-1-MASP-2 co-complexes. We find that such co-complexes have a functional role in activating complement and are present in serum at varying levels, impacting on the degree of complement activation. This raises the novel possibility that MAp44 may inhibit complement, not simply by brute force displacement of MASP-2 from MBL or ficolins, but by disruption of co-complexes, hence impairing transactivation. We present support for this contention.

Natural IgM antibodies are involved in the activation of complement by hypoxic human tubular cells

Ischemia-reperfusion injury (IRI) has a major impact on graft survival after transplantation. Renal proximal tubular epithelial cells (PTEC) located at the corticomedullary zone are relatively susceptible to IRI and have been identified as one of the main targets of complement activation. Studies in mice have shown an important role for the alternative pathway of complement activation in renal IRI. However, it is unclear whether experimental data obtained in mice can be extrapolated to humans. Therefore, we developed an in vitro model to induce hypoxia-reoxygenation in human and mouse PTEC and studied the role of the different pathways of complement activation. Exposure of human PTEC to hypoxia followed by reoxygenation in human serum resulted in extensive complement activation. Inhibition studies using different complement inhibitors revealed no involvement of the alternative or lectin pathway of complement activation by hypoxic human PTEC. In contrast, complement activation by hypoxic murine PTEC was shown to be exclusively dependent on the alternative pathway. Hypoxic human PTEC induced classic pathway activation, supported by studies in C1q-depleted serum and the use of blocking antibodies to C1q. The activation of the classic pathway was mediated by IgM through interaction with modified phosphomonoesters exposed on hypoxic PTEC. Studies with different human sera showed a strong correlation between IgM binding to hypoxic human PTEC and the degree of complement activation. These results demonstrate important species-specific differences in complement activation by hypoxic PTEC and provide clues for directed complement inhibition strategies in the treatment and prevention of IRI in the human kidney.

Anti‐ganglioside Antibodies and the Presynaptic Motor Nerve Terminal

The Guillain Barré syndromes (GBS) are the world's leading cause of acute autoimmune neuromuscular paralysis. Understanding the pathophysiological events of GBS, and improving immunotherapies are fundamental to improving the clinical outcome. Recent research into GBS and the Miller Fisher syndrome (MFS) variant has focused on the forms mediated by anti-ganglioside antibodies in which correlations have been established between anti-ganglioside antibodies and specific clinical phenotypes, notably between anti-GM1/GD1a antibodies and the acute motor axonal variant and anti-GQ1b/GT1a antibodies and MFS. Anti-ganglioside antibodies can arise through molecular mimicry with GBS-associated Campylobacter jejuni oligosaccharides. Our work has focused on axonal and glial components of the motor nerve terminal as a model site of injury, and through combined active and passive immunization paradigms in glycosyltransferase knockout mice we have developed murine neuropathy phenotypes mediated by anti-ganglioside antibodies. Several determinants influence disease expression including the level of immunological tolerance to microbial glycans that mimic self gangliosides, the degree of complement activation, and the ganglioside density in target tissue. Such studies provide us with clear information on an antibody-mediated pathogenesis model for GBS and should lead to rational therapeutic testing of agents that are potentially suitable for use in man.

Use of Serum or Buffer-Changed EDTA-Plasma in a Rapid, Inexpensive, and Easy-To-Perform Hemolytic Complement Assay for Differential Diagnosis of Systemic Lupus Erythematosus and Monitoring of Patients with the Disease

We previously described a simplified quantitative hemolytic assay for classical pathway (CP) hemolytic function in serum that has been shown to correlate with the 50% hemolytic complement (CH50) assay. In the present study, we used this assay to compare CP functions; plasma levels of C3, C4, and C3dg; and ratios of C3dg to C3 in healthy individuals and patients with systemic lupus erythematosus (SLE) or rheumatoid arthritis (RA) with different degrees of complement activation. A significant depression in CP function and levels of C4 and C3 and increased C3dg levels and C3dg/C3 ratios were observed in the SLE patients. In patients with RA, CP function was normal, whereas C3, C4, and C3dg levels and the C3dg/C3 ratio were elevated. The SLE results are compatible with systemic complement consumption, whereas the RA data suggest an acute-phase reaction with a normal C3 catabolic rate. To facilitate the handling of patient samples, we also developed a method to restore the hemolytic function of EDTA-plasma by transferring it to Veronal-buffered saline containing the thrombin inhibitor lepirudin. This process inhibits coagulation and enables complement activation, allowing a longer time lag between sample harvesting and testing. These results, combined with previous correlation studies, suggest that the CP hemolytic assay can effectively replace the CH50 assay for routine SLE differential diagnosis and monitoring of disease activity.

Cytokine induction by circulating immune complexes and signs ofin-vivocomplement activation in systemic lupus erythematosus are associated with the occurrence of anti-Sjögren's syndrome A antibodies

Circulating immune complexes (IC) and levels of IC-induced cytokines have been correlated with complement activation and autoantibody profiles in systemic lupus erythematosus (SLE). SLE sera were analysed concerning levels of immune complexes (IC), classical complement function and different antinuclear and anti-C-reactive protein (CRP) autoantibodies. Blood mononuclear cells from healthy donors were stimulated with isolated IC and production of interleukin (IL)-10, IL-6 and IL-12p40 was measured. Functional experiments revealed that increased levels of IC-induced cytokines were associated with both increased classical complement activation and the occurrence of anti-Sjögren's syndrome A (SSA) and anti-SSB but not other autoantibodies. Biochemical measurement of circulating IC showed that the degree of complement activation and the occurrence of anti-SSA were synergistically associated with levels of circulating IC in SLE sera, as complement activation was a prerequisite for the enhancing effect of anti-SSA. Anti-CRP was associated with complement activation, but not with other autoantibodies. Our results indicate that anti-SSA and possibly anti-SSB antibodies influence IC formation and subsequent IC-induced cytokine induction, and that they thereby participate in the inflammatory process in active SLE.

Complement activation in acute coronary syndromes

The complement system is part of the host defence response. However, considerable evidence suggests that complement plays an important role in the pathophysiology of ischemic heart disease. The aim of this study was to evaluate complement activation in patients with all forms of acute coronary syndromes (ACS) and to examine the relationship between the degree of complement activation and myocardial injury. The study population included 152 subjects (26 females): 82 with ACS (35 acute myocardial infarction (AMI), 22 non-Q wave MI (NQMI), 25 unstable angina (UAP)) (Group A), 35 stable angina (SA) (Group B), and 35 healty control subjects (Group C). Complement 3 (C3), Complement 4 (C4), C-reactive protein (CRP), troponin I (TnI) as well as creatine kinase MB (CK-MB) were evaluated. Patients' blood samples were taken on admission (day 1) and after 2, 3 and 7 days in group A. However, only one measurement was performed in the groups B and C. Plasma C3 and C4 peak levels were significantly higher in patients with AMI (141+/-29 and 35+/-11 mg/dl) and NQMI (136+/-13 and 35+/-7 mg/dl) than in patients with SA (128+/-14 and 27+/-10 mg/dl) and the control subjects (114+/-22 and 22+/-7 mg/dl) (p<0.03). Also, C3 and C4 serum levels in patients with SA and UAP (126+/-16 and 31+/-7 mg/dl) were significantly higher than those in control subjects (p<0.01, p<0.03, respectively). At 1-week follow-up, there were no significant differences between the plasma levels of C3 and C4 in patients with UAP (p>0.05). However, plasma levels of C3 and C4 were significantly different between days in patients with AMI and NQMI (p<0.0001). Plasma C3 and C4 levels in ACS showed a relationship with peak CK-MB and Tn I levels (p<0.01). Plasma CRP level in ACS showed positive correlation with C3 (p<0.01) and C4 (p<0.001). In this study, we determined that plasma C3 and C4 levels were elevated in ACS and SA. Although C3 and C4 were higher in ACS and SA, the systemic levels of inflammatory markers in patients with SA and UAP were lower than those found in the AMI and NQMI groups. The relationship between C3, C4 levels and ACS further suggests that the complement activation is related to necrosis within the myocardium.

Activation of Complement Pathways after Contusion-Induced Spinal Cord Injury

Previous studies have shown that a cellular inflammatory response is initiated, and inflammatory cytokines are synthesized, following experimental spinal cord injury (SCI). In the present study, we tested the hypothesis that the complement cascade, a major component of both the innate and adaptive immune response, is also activated following experimental SCI. We investigated the pathways, cellular localization, timecourse, and degree of complement activation in rat spinal cord following acute contusion-induced SCI using the New York University (NYU) weight drop impactor. Mild and severe injuries (12.5 and 50 mm drop heights) at 1, 7, and 42 days post injury time points were evaluated. Classical (C1q and C4), alternative (Factor B) and terminal (C5b-9) complement pathways were strongly activated within 1 day of SCI. Complement protein immunoreactivity was predominantly found in cell types vulnerable to degeneration, neurons and oligodendrocytes, and was not generally observed in inflammatory or astroglial cells. Surprisingly, immunoreactivity for complement proteins was also evident 6 weeks after injury, and complement activation was observed as far as 20 mm rostral to the site of injury. Axonal staining by C1q and Factor B was also observed, suggesting a potential role for the complement cascade in demyelination or axonal degeneration. These data support the hypothesis that complement activation plays a role in SCI.

Activation of Complement Pathways after Contusion-Induced Spinal Cord Injury

Previous studies have shown that a cellular inflammatory response is initiated, and inflammatory cytokines are synthesized, following experimental spinal cord injury (SCI). In the present study, we tested the hypothesis that the complement cascade, a major component of both the innate and adaptive immune response, is also activated following experimental SCI. We investigated the pathways, cellular localization, timecourse, and degree of complement activation in rat spinal cord following acute contusion-induced SCI using the New York University (NYU) weight drop impactor. Mild and severe injuries (12.5 and 50 mm drop heights) at 1, 7, and 42 days post injury time points were evaluated. Classical (C1q and C4), alternative (Factor B) and terminal (C5b-9) complement pathways were strongly activated within 1 day of SCI. Complement protein immunoreactivity was predominantly found in cell types vulnerable to degeneration, neurons and oligodendrocytes, and was not generally observed in inflammatory or astroglial cells. Surprisingly, immunoreactivity for complement proteins was also evident 6 weeks after injury, and complement activation was observed as far as 20 mm rostral to the site of injury. Axonal staining by C1q and Factor B was also observed, suggesting a potential role for the complement cascade in demyelination or axonal degeneration. These data support the hypothesis that complement activation plays a role in SCI.

Statin drugs do not affect serum complement activation in vitro.

Statin drugs prevent coronary heart disease through anti-inflammatory mechanisms in addition to the well-known reduction of low-density lipoproteins. The complement system plays an essential role in the inflammatory response and has been postulated to be modified by statins. A direct role for statins in complement activation, however, has not been previously investigated. We therefore studied the effect of statins on in vitro complement activation. Pravastatin, atorvastatin and the active metabolite of the latter, ortho-hydroxy atorvastatin, were added to normal human serum and incubated for 1 h in the absence or presence of aggregated immunoglobulin (classical pathway activation) or cobra venom factor (alternative pathway activation). The degree of complement activation, as detected by specific complement-activation products for the classical pathway (C1rs-C1-inhibitor complexes), the combined classical and lectin pathway (C4bc), the alternative pathway (C3bBbP) and the final common pathway (C3bc and TCC), was not affected by pre-incubation of the serum with any of the statins. Statins do not affect complement activation directly, but indirect effects in vivo may well be operative.

Complement activation reaches maximum during equilibrium between antigen and antibody in an in vitro model for thrombolysis with streptokinase.

Treatment with streptokinase (SK) of patients with acute myocardial infarction (AMI) is known to activate the complement system. In a previous investigation we found a great variation in the degree of complement activation taking place during infusion of SK in patients with AMI (range: 0-293% increase in pretreatment complement split product 3d (C3d) levels). In this in vitro study we added SK to serum with different concentrations of SK-antibodies (SK-Ab) and found that both the relative concentrations and the total concentrations of SK and Ab determined the degree of complement activation elicited by SK. This finding was in accordance with the Kendall-Heidelberger equation of antigen-antibody equivalence. Furthermore, we found parallel rises in C3d and C4d, indicating that complement was activated via the classical pathway. A condition for maximal complement activation is antigen-antibody equivalence.

Modulation of fluid-phase complement activation inhibits hyperacute rejection in a porcine-to-human xenograft model

YPERACUTE rejection (HAR) of porcine organs is due to classical complement activation by naturally occurring human antibodies to the endothelium. Pigs that are transgenic for human regulators of complement (RCA) seem promising with respect to avoiding HAR, and their use in various studies has brought xenotransplantation closer to clinical reality. However, RCAs are usually expressed on the endothelial cells as clusters and the activation of complement may take place at some distance from RCA localisation. Thus, it should be emphasised that, although RCA pig xenografts may be protected against HAR, a certain degree of complement activation may still contribute to an inflammatory reaction with subsequent impaired graft function. Therefore, efforts should be taken to control the fluid-phase activation of complement, which was the main topic of our studies. We have established an ex vivo perfusion model 1 to investigate the effect of complement manipulation by intravenous immunoglobulin (IVIG), a C3-binding peptide (Compstatin) and C1-inhibitor (C1-INH) on HAR.

Inflammatory cell recruitment, distribution, and chemiluminescence response at IgG precoated- and thiol functionalized gold surfaces.

The role of complement activation by artificial surfaces relative to inflammatory response is not well understood. This study was performed to evaluate the inflammatory cell recruitment, distribution, and ex vivo metabolic activation of surfaces with different plasma protein adsorption and complement activation properties in vitro. The implants were (1) pure gold (reference), (2) albumin-precoated (3) IgG-precoated gold, and (4) 3-mercapto-1, 2-propanediol [mercaptoglycerol (MG)] and (5) glutathione (GSH) immobilized to gold. The implant disks were inserted subcutaneously in rats for 24 h, and the number of inflammatory cells that were recruited to the implant adjacent to the surrounding fluid phase (exudate) and the surfaces were quantified by DNA measurements. The oxidative burst was analyzed ex vivo using spontaneous and phorbol myristate acetate (PMA)-stimulated, luminol-enhanced chemiluminescence (CL). The in vitro surface-induced anti-rat C3 binding was evaluated by ellipsometry and antibody techniques after plasma incubations for 1 and 30 min. The ellipsometric results showed that immobilized mercaptoglycerol and IgG-coated, but not the immobilized glutathione or the reference Au, bound anti-C3. The in vivo results revealed that the largest amount of cells was associated with the IgG-coated surfaces, followed by immobilized GSH and MG, albumin-coated, and gold surfaces, respectively. No spontaneous ex vivo luminol-enhanced CL was recorded from the cells irrespective of surface functionality or localization. A down-regulation of surface-associated and exudate leukocyte CL was observed ex vivo, irrespective of surface functionality. The results do not indicate a clear relationship between the degree of complement activation in vitro and leukocyte recruitment and adhesion in vivo for differently functionalized surfaces.

Anti-TNF treatment of baboons with sepsis reduces TNF-alpha, IL-6 and IL-8, but not the degree of complement activation.

The activation of complement and the release of TNF-alpha, IL-6 and IL-8 are important pathogenic factors behind organ dysfunction in sepsis. The aim of this study was to determine whether infusion of anti-TNF antibodies alters complement activation and plasma concentrations of pro-inflammatory cytokines at high doses of Escherichia coli. Six baboons received intravenously 2 x 10(9) live E. coli bacteria per kg body weight (group 1), in addition five received pretreatment with 1 mg per kg body weight anti-TNF antibodies (group 2), and seven received 5 x 10(8) live E. coli bacteria per kg body weight (group 3). Two hours after the start of infusion of the bacteria, plasma concentrations of C3 activation products, C5a and the terminal SC5b-9 complement complex were increased in groups 1 and 2 (P < 0.05), but there was no significant difference between the groups. At 2 h the levels of TNF-alpha, IL-6 and IL-8 were lower in group 2 compared with group 1 (P<0.05). In group 2 compared with group 1 the TNF-alpha concentrations were, however, higher at 4, 8 and 24 h. The explanation for this phenomenon is probably that TNF-alpha binds to the anti-TNF antibody complex and is released slowly after it has been bound. The study showed that infusion of anti-TNF antibodies reduced the concentrations of TNF-alpha, IL-6 and IL-8, without any detectable influence on complement activation.

Complement activation in patients undergoing mechanical circulatory support.

Contact of blood with artificial surfaces activates pro-inflammatory responses and the complement cascade. This may have broad implications on the post implantation fate of patients needing mechanical circulatory support. Therefore, we investigated the course and prognostic value of complement factors C3a and C5a in 66 patients supported with pulsatile ventricular assist devices. All patients were in severe cardiogenic shock, i.e., catecholamine dependent and in the intensive care unit, before implementation of mechanical circulatory support. Isolated left ventricular support (Novacor [Oakland, CA] or Thermo Cardiosystems, Inc. [TCI; Woburn, MA]) was used in 28 patients, and biventricular support (Berlin Heart [Mediport, Berlin, Germany]) in 38 patients. Before initiation of mechanical circulatory support, no statistically significant differences in C3a or C5a between surviving and nonsurviving patients with left ventricular assist devices (LVADs) were found. Patients with biventricular assist devices (BVADs) had significantly higher C3a (804 +/- 364 ng/L) levels than patients with LVADs (536 +/- 204 ng/L, p = 0.02) before mechanical circulatory support. Only C5a, only in the BVAD group, was able to predict patients' post implantation course before implantation of a ventricular assist device (p = 0.02). Three weeks after initiation of mechanical circulatory support, complement factors remained increased in all groups. There was no difference, however, in complement activation between patients with LVADs and those with BVADs. Patients not reaching transplantation had significantly higher C3a levels at this point than those successfully supported (p = 0.007). The degree of complement activation mainly depends on the severity of cardiogenic shock before initiation of mechanical circulatory support, and not on the device used. Patients with extremely high levels of complement activation before implantation of the device could be saved with BVAD rather than LVAD support. Patients who continued to have highly elevated complement levels 3 weeks after initiation of mechanical circulatory support had unfavorable prognoses. Complement activation indicates the severity of cardiogenic shock before implementation of mechanical circulatory support and the degree of recovery from secondary organ dysfunction while on the device. It is fairly independent of the system used for mechanical circulatory support, and therefore can be applied to predict patients' post implantation course and outcome.