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PubMed1991

The role of immunoglobulin and complement in enhancing the respiratory burst of neutrophils against Trichomonas vaginalis.

Human neutrophils, alone, did not kill Trichomonas vaginalis. More than 90% of T. vaginalis (10(5)/ml) survived in the presence of 10% normal human serum (NHS) while 90% of these organisms were killed in the presence of a combination of neutrophils (10(6)/ml) and 10% NHS. Mechanisms responsible for this serum-mediated neutrophil killing of T. vaginalis were demonstrated through a process of lucigenin-amplified neutrophil chemiluminescence. As evidenced by indirect immunofluorescence, NHS showed specific immunoglobulin G (IgG) titre of 1:8 for T. vaginalis. Purified IgG, at 1.6 mg/ml, showed no direct opsonizing or lytic effect on this organism. Formalin-fixed trichomonads opsonized by C2 deficient human serum promote 4 times more neutrophil chemiluminescence than those opsonized by Factor B deficient human serum. With the addition of purified IgG (5 mg/ml) neutrophil chemiluminescence was increased by 4 times and further improved trichomonal killing by neutrophils (from 5 +/- 4% to 78 +/- 16%) via activation of the classical complement pathway, but did not alter that due to activation of the alternative complement pathway. These studies indicate that both an IgG-enhanced classical complement pathway activation and an antibody-independent alternative complement pathway activation provide opsonin (C3) for T. vaginalis to facilitate the neutrophil killing mechanism.

Adult↗
PubMed1992

Binding of antibody and resistance to lysis of trypomastigotes of Trypanosoma cruzi.

Epimastigote forms of Trypanosoma cruzi are readily lysed by complement via the alternative pathway. Neither fibroblast-derived trypomastigotes nor blood-form trypomastigotes are lysed by complement alone and few (less than 30% of the Brazil strain) are lysed in the presence of parasite-specific antibody and complement. The mechanism by which trypomastigotes resist antibody-dependent, complement-mediated lysis is not clearly understood. In the present study, we have utilized flow cytometric analysis to examine the binding of parasite-specific antibody to epimastigotes, fibroblast-derived trypomastigotes and blood-form trypomastigotes of a Brazil strain of T. cruzi. We also determined the extent of lysis of these parasites in the presence of complement utilizing propidium iodide to determine cell death. It was found that all epimastigotes bind approximately the same amount of antibody but that there are subpopulations of trypomastigotes which bind antibody to varying degrees. When these subpopulations were sorted, and treated with complement, lysis was only minimally increased in the population of parasites which bound significantly greater amounts of antibody.

Animals↗
PubMed1989

Hereditary, complete deficiency of complement factor H associated with recurrent meningococcal disease.

Complement factor H (beta-1H globulin) is an important regulatory protein which inhibits the spontaneous complement activation via the alternative pathway. We describe a 15-year-old girl without any detectable factor H in plasma. She has had two episodes of meningococcal disease, but is otherwise completely healthy. Secondary to the factor-H deficiency, the levels of factor B, properdin, C3, and C5-C9 were strongly reduced due to spontaneous in vivo activation of the alternative complement pathway. Plasma C3dg was strongly elevated in spite of the factor-H deficiency; apparently erythrocyte CR1 substitutes for factor H in C3 degradation. Neither C3 nor complement lesions were demonstrable on her erythrocytes which did, however, show increased, spontaneous haemolysis in vitro in citrate plasma, but not in serum. The patient is a single child and her parents, who are unrelated and healthy, had half-normal levels of factor H. This reduction of factor H is sufficient to cause increased, spontaneous activation of the alternative pathway.

Antibodies, Bacterial↗
PubMed1994

Non-coordinated biosynthesis of early complement components in a deficiency of complement proteins C1r and C1s.

We report on a 60-year-old woman with systemic lupus erythematosus and a total (95%) C1r and a partial (36%) C1s deficiency. The patient complained about cutaneous lesions on forearms and legs without other systemic involvement. Elevated anti-nuclear, anti-native DNA and anti-SSA antibodies were present. The finding of persistently depressed levels of haemolytic complement activity (CH50) on both serum and plasma, associated with normal levels of C3, C4 and C2 components, and normal alternative pathway haemolytic activity showed a deficiency of an early component of the classical pathway. Indeed C1r component was below the limits of detection whereas C1s component was lowered (36%). The depressed CH50 was only corrected by purified C1r. Biosynthesis of C1r and C1s by patient's monocytes was spontaneously normal but not up-regulated by interferon-gamma for C1r alone, whereas the biosynthesis of C1s, but also of interleukin-6, was increased, indicating a specific disregulation of C1r. The deficiency was associated with a lupus syndrome and a fatal assumed septic shock. This is in agreement with other reported cases.

Cells, Cultured↗
PubMed2005

Endothelial cell protection and complement inhibition in xenotransplantation: a novel in vitro model using whole blood.

BACKGROUND: Studying the interactions between xenoreactive antibodies, complement and coagulation factors with the endothelium in hyperacute and acute vascular rejection usually necessitates the use of in vivo models. Conventional in vitro or ex vivo systems require either serum, plasma or anti-coagulated whole blood, making analysis of coagulation-mediated effects difficult. Here a novel in vitro microcarrier-based system for the study of endothelial cell (EC) activation and damage, using non-anticoagulated whole blood is described. Once established, the model was used to study the effect of the characterized complement- and coagulation inhibitor dextran sulfate (DXS, MW 5000) for its EC protective properties in a xenotransplantation setting. METHODS: Porcine aortic endothelial cells (PAEC), grown to confluence on microcarrier beads, were incubated with non-anticoagulated whole human blood until coagulation occurred or for a maximum of 90 min. PAEC-beads were either pre- or co-incubated with DXS. Phosphate buffered saline (PBS) experiments served as controls. Fluid phase and surface activation markers for complement and coagulation were analyzed as well as binding of DXS to PAEC-beads. RESULTS: Co- as well as pre-incubation of DXS, followed by washing of the beads, significantly prolonged time to coagulation from 39 +/- 12 min (PBS control) to 74 +/- 23 and 77 +/- 20 min, respectively (P < 0.005 vs. PBS). DXS treatment attenuated surface deposition of C1q, C4b/c, C3b/c and C5b-9 without affecting IgG or IgM deposition. Endothelial integrity, expressed by positivity for von Willebrand Factor, was maintained longer with DXS treatment. Compared with PBS controls, both pre- and co-incubation with DXS significantly prolonged activated partial thromboplastin time (>300 s, P < 0.05) and reduced production of thrombin-antithrombin complexes and fibrinopeptide A. Whilst DXS co-incubation completely blocked classical pathway complement activity (CH50 test) DXS pre-incubation or PBS control experiments showed no inhibition. DXS bound to PAEC-beads as visualized using fluorescein-labeled DXS. CONCLUSIONS: This novel in vitro microcarrier model can be used to study EC damage and the complex interactions with whole blood as well as screen ''endothelial protective'' substances in a xenotransplantation setting. DXS provides EC protection in this in vitro setting, attenuating damage of ECs as seen in hyperacute xenograft rejection.

Animals↗
PubMed2005

Apolipoprotein J (clusterin) activates rodent microglia in vivo and in vitro.

Apolipoprotein J (apoJ; also known as clusterin and sulfated glycoprotein (SGP)-2) is associated with senile plaques in degenerating regions of Alzheimer's disease brains, where activated microglia are also prominent. We show a functional link between apoJ and activated microglia by demonstrating that exogenous apoJ activates rodent microglia in vivo and in vitro. Intracerebroventricular infusion of purified human plasma apoJ ( approximately 4 microg over 28 days) activated parenchymal microglia to a phenotype characterized by enlarged cell bodies and processes (phosphotyrosine immunostaining). In vitro, primary rat microglia were also activated by apoJ, with changes in morphology and induction of major histocompatibility complex class II (MHCII) antigen. ApoJ increased the secretion of reactive nitrogen intermediates in a dose-dependent manner (EC(50) 112 nm), which was completely blocked by aminoguanidine (AG), a nitric oxide synthase inhibitor. However, AG did not block the increased secretion of tumor necrosis factor-alpha by apoJ (EC(50) 55 nm). Microglial activation by apoJ was also blocked by an anti-apoJ monoclonal antibody (G7), and by chemical cleavage of apoJ with 2-nitro-5-thiocyanobenzoate. The mitogen-activated protein kinase kinase and protein kinase C inhibitors PD98059 and H7 inhibited apoJ-mediated induction of reactive nitrogen intermediate secretion from cultured microglia. As a functional measure, apoJ-activated microglia secreted neurotoxic agents in a microglia-neuron co-culture model. We hypothesize that ApoJ contributes to chronic inflammation and neurotoxicity through direct effects on microglia.

Animals↗
PubMed1990

Serum haemolytic classical and alternative pathways of complement in infancy: age-related changes.

The haemolytic activity of complement was evaluated in the serum of healthy children from birth to 2 years of age using the kinetic method for the determination of the time needed to lyse 50% of target red cells (t 1/2). No sex-linked differences were observed in any of the age groups studied and the lowest lytic activity levels for both complement pathways were detected in neonates. The two pathways, however, showed different maturation patterns, i.e., lytic activity levels similar to those of adults were reached between the 1st and 3rd month of life (classical pathway) and around the 13th month (alternative pathway). In the age group of 7 to 24 months, the lytic activity of the classical pathway was higher than in adults. The present data permitted us to establish normal ranges of t 1/2 values for the classical and alternative pathways in serum of healthy neonates and children aged 1 to 24 months.

Age Factors↗
PubMed1993

Hereditary properdin deficiency in three families of Tunisian Jews.

Hereditary properdin deficiency is a rare genetic disorder of the complement system. Three propositi and six additional family members with properdin deficiency have been found following analysis of the hemolytic activity of the classical (CH50) and the alternative (AP50) complement pathways in the sera of 101 survivors of meningococcal infections and 59 survivors of severe pneumococcal and Haemophilus influenza infections. All the properdin-deficient individuals had undetectable levels of properdin by radial immunodiffusion and by Western blotting. They belonged to three non-related families of Tunisian Jews who came from different parts of Tunisia. Two patients had a meningococcal infection at 15 and 16 years of age, respectively, and one had Haemophilus influenza meningitis at 1.5 years of age. In contrast to the fulminant and fatal course of meningococcal infection which was previously described in some properdin-deficient patients, our patients had a relatively mild disease. Properdin deficiency may not be as rare as previously thought. Analysis of AP50, in addition to CH50, in sera of patients who had meningococcal infection, will probably disclose many more cases of hereditary properdin deficiency. In addition, our findings indicate that, as in other complement abnormalities, hereditary properdin deficiency may also be associated with the ethnic origin of the patient.

Adolescent↗
PubMed1996

Receptor (CD46) modulation and complement-mediated lysis of uninfected cells after contact with measles virus-infected cells.

Recently, it has been observed that the infection of human target cells with certain measles virus (MV) strains leads to the downregulation of the major MV receptor CD46. Here we report that CD46 downregulation can be rapidly induced in uninfected cells after surface contact with MV particles or MV-infected cells. Receptor modulation is detectable after 30 min of cocultivation of uninfected cells with MV-infected cells and is complete after 2 to 4 h, a time after which newly synthesized MV hemagglutinin (MV-H) cannot be detected in freshly infected target cells. This contact-mediated receptor modulation is also induced by recombinant MV-H expressed by vaccinia virus and is inhibitable with antibodies against CD46 and MV-H. By titrating the effect with MV Edmonston strain-infected cells, a significant contact-mediated CD46 modulation was detectable up to a ratio of 1 infected to 64 uninfected cells. As a result of CD46 downregulation, an increased susceptibility of uninfected cells for complement-mediated lysis was observed. This phenomenon, however, is MV strain dependent, as observed for the downregulation of CD46 after MV infection. These data suggest that in acute measles or following measles vaccination, uninfected cells might also be destroyed by complement after contacting an MV-infected cell.

Animals↗
PubMed1994

Effect of some phytogenic agents and synthetic compounds on complement cascade-mediated hemolysis.

A variety of anti-inflammatory compounds obtained from various medicinal plants (phytogenic) as well as some synthetic compounds were tested on the complement cascade in vitro. Bovine erythrocytes were treated with rabbit antibovine red blood cell antibody. Rabbit plasma was diluted with veronal buffer and mixed with erythrocytes in the presence or absence of drugs and incubated. Erythrocytes were pelleted and the absorbance of the supernatant at 412 nm determined. Drugs could be grouped into three categories; (a) those with no effect whatsoever; (b) those which produced definite enhancement of the release of hemoglobin by the complement cascade, and (c) prednisolone, which inhibited the release of hemoglobin by the complement cascade. We suggest that prednisolone and the drugs which had no effect on the complement cascade are safer to use as anti-inflammatory agents, while drugs enhancing the complement cascade may have potential adverse properties.

Animals↗
PubMed1991

Complement function and the synthesis of lung surfactant may be a regulation which preterm infants have in common.

The levels of CH50, the complement components, C1q, C4, and C3, C3 degradation fragments, and factor B were determined in the cord blood of 128 newborn infants. The levels of C3, C4, and C3d3 (an index of chronic in vivo complement activation) were clearly lower in the 28 infants with respiratory distress syndrome (RDS) than in the infants with other lung diseases or with normal lungs. CH50 and factor B levels were also low in RDS. Levels of C1q and other serum components in RDS infants were similar to the average levels in other infants without RDS at a corresponding gestational age. Lung surfactant is synthesized in alveolar type 2 cells, in which the complement components C4, C3, and factor B, but not C1q, have been reported to be synthesized. It seems possible that common factors regulate the synthesis of some complement components and surfactant.

Complement C1q↗
PubMed1991

Activation of complement by Treponema denticola.

Oral spirochetes have been shown to be associated with periodontal diseases and are present in increased numbers in lesions of greater severity. In this study, the interaction of Treponema denticola with human complement, a major antibacterial defense system, was examined. For each of two strains of T. denticola, it was found that both the classical and alternative pathways of human complement were activated in human serum upon incubation at 37 degrees C. C3 fragments were deposited on the surface of this organism following complement activation; the fragments bound included both of the major C3-derived opsonic fragments C3b and iC3b. Under incubation conditions identical to those carried out for complement activation in serum, T. denticola failed to degrade purified, hemolytically-active C3, although it readily degraded inactivated C3. Thus, despite the documented proteolytic activity of this organism, complement activation and deposition of complement-derived opsonins may be important defense mechanisms in the control of infections with T. denticola.

Complement C3↗
PubMed1996

Autoantibodies to the collagen-like region of C1q are strongly associated with classical pathway-mediated hypocomplementemia in systemic lupus erythematosus.

We have investigated the presence of autoantibodies to the collagen-like region (CLR) of C1q and its relationship with classical pathway-dependent hypocomplementemia in patients with SLE. Antibodies to CLR/C1q were quantitated in the plasma of 113 unselected patients with SLE by ELISA, using purified CLR as antigen. Plasma concentration of C3, CH50 activity and C2 hemolytic activity were determined according to standard procedures. The prevalence of IgG antibodies to CLR/C1q in the study population was 33.6%. Plasma titers of anti-CLR/C1q autoantibodies showed a strong negative correlation with CH50 activity (p < 0.0001) and with plasma levels of C3 (p < 0.0001). Eighty-five percent of patients with severe complement consumption exhibited high titers of anti-CLR/C1q antibodies in plasma, independently of clinical disease activity. Anti-CLR/C1q antibodies were present in the plasma of 38% of patients with moderate classical pathway consumption and 14% of patients with no evidence of complement consumption. Analysis of sequential samples from six patients over a period of 18 to 24 months demonstrated that changes in CH50 activity mirrored those of the plasma titers of anti-CLR/C1q antibodies. Acquired hypocomplementemia through the classical pathway is strongly associated with the presence of anti-CLR/C1q autoantibodies in SLE. The results suggest that anti-CLR/C1q antibodies may perpetuate classical pathway activation, independently of clinical disease activity.

Autoantibodies↗
PubMedDate not supplied

Transplantation of xenogeneic cells secreting beta-endorphin for pain treatment: analysis of the ability of components of complement to penetrate through polymer capsules.

The permeation of component of complement and secreted peptides through polymer capsules (PM30, K6305, and K5708) were examined. To analyze permeability by complement, the degree of hemolysis of sensitized sheep erythrocytes (EA) (1 x 10(9)/ml) enclosed in each type of capsule was examined after 24-h incubation in culture medium containing 10% human serum. PM30 and K6305 prevented the permeation of complement well, while K5708 did not. EA suspended in alginate prevented hemolysis even in K5708. Peptide permeation through the capsules was assessed by measuring the concentration of ACTH secreted by proopiomelanocortin (POMC)-gene-transfected-Neuro2A in the culture medium on days 4, 7, 14, 21, and 28 after encapsulation. The ACTH levels in the culture medium remained high until day 28. Alginate appeared to prevent the secretion, because ACTH levels decreased in alginate-suspended cells after day 14. The PM30-K6306 double capsules containing cell lines, Neuro2A, BHK21 (hamster fibroblasts), L929 (mouse fibroblasts), and HF-SKFII (human fibroblasts) were transplanted into the cerebrospinal fluid (CSF) space of the monkeys in the lumber region. The morphological examination showed the partial survival of Neuro2A, and BHK21 and HF-SKFII, which were cells concordant with the monkeys. On the other hand, L929 cells, which were discordant with the monkeys, could not survive at all. Because these results suggest that the complement components penetrate the polymer capsules, concordant cells are preferable for xenografting with polymer capsules into the CSF space.

Adrenocorticotropic Hormone↗
PubMed2004

Hypocomplementemia in systemic lupus erythematosus and primary antiphospholipid syndrome: prevalence and clinical significance in 667 patients.

The objective of the study was to analyse the prevalence and clinical significance of hypocomplementemia in a large series of patients diagnosed either with systemic lupus erythematosus (SLE) or with primary antiphospholipid syndrome (APS) and its association with the main clinical, hematological and immunological features of these diseases. Between 1992 and 2003, complement determinations (C3 and C4 levels, CH50 activity) were performed in 597 consecutive patients diagnosed with SLE (530 women and 67 men, mean age 32.6 years) and 70 with primary APS (57 women and 13 men, mean age 38.7) visited in our department. Complement determinations are routinely made at the first visit of patients and yearly during the follow-up. SLE and primary APS were diagnosed according to current classification criteria. Hypocomplementemia was detected in 371 (62%) of SLE patients. Compared with patients with normal complement values, those with hypocomplementemia showed a higher prevalence of female gender (P < 0.001), fever (P = 0.021), nephropathy (P < 0.001), cutaneous vasculitis (P = 0.023), positive anti-dsDNA antibodies (P = 0.012) and cryoglobulinemia (P < 0.001). In addition, patients with hypocomplementemia showed a higher prevalence of APS-related features such as hemolytic anemia (P = 0.001) and antiphospholipid antibodies (P < 0.001). Hypocomplementemia was prospectively related to accumulated hospitalization in SLE patients but not with the accumulated number of lupus flares or with the survival after follow-up of five years. In contrast, 33 (47%) patients with primary APS presented low complement values, which were associated with a higher prevalence of livedo reticularis (P = 0.022), thrombocytopenia (P = 0.004), lupus anticoagulant (P = 0.013), positive IgM-aCL (P = 0.039), positive ANA (P = 0.002) and anti-dsDNA (P = 0.046). The diagnostic value of hypocomplementemia in patients with SLE is based on the association with disease activity, immune-complex mediated manifestations (glomerulonephritis, cryoglobulinemia) and APS-related features (livedo reticularis, hemolytic anemia and aPL). Hypocomplementemia was found in nearly half of patients with primary APS, and was associated with some APS features (livedo reticularis, thrombocytopenia, aPL) but also with SLE-related immunological markers (ANA and anti-dsDNA), identifying a subset of patients with primary APS with a higher risk of evolving to SLE. These results clearly support the routine determination of complement factors in the clinical follow-up of patients with SLE and primary APS.

Adult↗
PubMed1996

Leakage of protein into lungs of preterm ventilated rabbits is correlated with activation of clotting, complement, and polymorphonuclear leukocytes in plasma.

We investigated whether leakage of protein in lungs of preterm ventilated rabbits of 28- and 29-d gestational age is correlated with activation of clotting, complement, and polymorphonuclear leukocytes (PMN) in plasma. We found signs of systemic activation of clotting, complement and PMN in ventilated 28-d gestational age rabbits, as indicated, respectively, by increased median plasma fibrin monomer concentrations (83 versus 40% of normal adult rabbit plasma in nonventilated 28-d gestational age rabbits, p < 0.01), decreased median plasma CH50 activity (112 versus 122 U/L in nonventilated 28-d gestational age rabbits, p < 0.05), and increased median plasma beta-glucuronidase concentrations (159 versus 97% of maximal activated adult rabbit plasma in nonventilated 28-d gestational age rabbits p < 0.05). We did not find signs of systemic activation in the ventilated 29-d gestational age group. Higher median total protein concentrations in alveolar wash of the ventilated 28-d gestational age rabbits (2.7 versus 1.3 mg/mL in the nonventilated rabbits. p < 0.01) indicated protein leakage into the lungs, and this protein leakage was more pronounced in the lungs of ventilated 28-d gestational age rabbits than in those of ventilated 29-d gestational age rabbits (2.1 mg/mL, p < 0.01). The total protein concentration in the alveolar wash of all 28-d gestational age rabbits was correlated with the concentration of fibrin monomers (p = 0.51, p = 0.035) and beta-glucuronidase (p = 0.61, p = 0.011), and the CH50 activity (p = -0.73, p = 0.002) in plasma. We conclude that leakage of protein in lungs of preterm ventilated rabbits of 28-d gestational age is correlated with activation of clotting, complement, and PMN in plasma. This activation process may contribute to lung injury by intravascular and intraalveolar deposition of fibrin and formation of proteinaceous edema.

Animals↗
PubMed1994

A glucan having reticuloendothelial system-potentiating and anti-complementary activities from the tuber of Pinellia ternata.

A glucan, called pinellian G, was isolated from the tuber of Pinellia ternata BREIT. It was homogenous on electrophoresis and gel chromatography, and its molecular mass was estimated to be 1.5 x 10(4). It is composed solely of D-glucose, in addition to a few O-acetyl groups. Methylation analysis, nuclear magnetic resonance and enzymic degradation studies indicated that it is a branched glucan mainly composed of alpha-1,4-linked D-glucopyranose residues with partially alpha-1,3-linked units and 4,6-branching points. The glucan showed significant reticuloendothelial system-potentiating activity in a carbon clearance test, as well as pronounced anti-complementary activity.

Acetylation↗
PubMed1994

Structural features and anti-complementary activity of rehmannan SA, a polysaccharide from the root of Rehmannia glutinosa.

The structural features of rehmannan SA, a polysaccharide with remarkable reticuloendothelial system-potentiating activity obtained from the root of Rehmannia glutinosa, were investigated by methylation analysis and periodate oxidation. Rehmannan SA is mainly made up of arabino-3,6-galactan type structural units. Both rehmannan SA and rehmannan SB showed pronounced anti-complementary activity.

Animals↗