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Serum amyloid P component and C-reactive protein mediate phagocytosis through murine Fc gamma Rs.

The pentraxins, serum amyloid P component (SAP) and C-reactive protein (CRP) are acute-phase serum proteins in mice and humans, respectively. Although SAP binds to DNA and chromatin and affects clearance of these autoantigens, no specific receptor for SAP has been identified. CRP is an opsonin, and we have shown that it binds to FcgammaR. Mice deficient in FcgammaR were used to assess the role of these receptors in phagocytosis by pentraxins using zymosan as a ligand. Phagocytosis of zymosan by bone marrow macrophages (BMM) was enhanced by opsonization with SAP or CRP. BMM from mice deficient in all three FcgammaR or in gamma-chain ingested unopsonized zymosan, but phagocytosis of SAP- or CRP-opsonized zymosan was not enhanced. SAP binding to BMM from gamma-chain-deficient mice was also greatly reduced, indicating little or no binding of SAP to FcgammaRII. SAP and CRP opsonized zymosan for phagocytosis by BMM from mice deficient in FcgammaRII or FcgammaRIII. SAP, but not CRP, opsonized zymosan for uptake by neutrophils that express only low levels of FcgammaRI. Together these results indicate that FcgammaRI and FcgammaRIII are receptors for SAP in the mouse. Opsonization of zymosan by CRP is mediated through FcgammaRI. Pentraxins are major proteins of the innate immune system and arose earlier in evolution than Igs. The use of FcgammaR by the pentraxins links innate and adaptive immunity and may have important consequences for processing, presentation, and clearance of the self-Ags to which these proteins bind.

Adjuvants, Immunologic↗

Role of complement component C1q in the IgG-independent opsonophagocytosis of group B streptococcus.

We investigated the role of complement component C1q in the IgG-independent opsonophagocytosis of type III group B Streptococcus (GBS) by peripheral blood leukocytes. We report that C1q binds to type III GBS both in normal human serum deficient in IgG specific for type III capsular polysaccharide and in a low-ionic strength buffer. The dissociation constant Kd ranged from 2.0 to 5.5 nM, and the number of binding sites Bmax ranged from 630 to 1360 molecules of C1q per bacterium (CFU). An acapsular mutant strain of GBS bound C1q even better than the wild type, indicating that the polysaccharide capsule is not the receptor for C1q. In serum, binding of C1q to GBS was associated with activation of the classical complement pathway. However, normal human serum retained significant opsonic activity after complete depletion of C1q, suggesting that the serum contains a molecule that is able to replace C1q in opsonization and/or complement activation. Mannan-binding lectin, known to share some functions with C1q, appeared not to be involved, since its depletion from serum had little effect on opsonic activity. Excess soluble C1q or its collagen-like fragment inhibited phagocytosis mediated by normal human serum, suggesting that C1q may compete with other opsonins for binding to receptor(s) on phagocytes. We conclude that, although C1q binds directly to GBS, C1q binding is neither necessary nor sufficient for IgG-independent opsonophagocytosis. The results raise the possibility that additional unknown serum factor(s) may contribute to opsonization of GBS directly or via a novel mechanism of complement activation.

Carrier Proteins↗

Bovine venereal vibriosis: activity of inflammatory cells in protective immunity.

The capability of bovine polymorphonuclear (PMN) and mononuclear phagocytes to kill Campylobacter (Vibrio) fetus venerealis was tested under various conditions in order to judge their roles in protective immunity. Bovine PMN killed C fetus in the presence of immunoglobulin G (IgG) but not immunoglobulin A (IgA) antibodies. Some phagocytosis occurred in the absence of antibody in glass-adherent cultures, but not in suspension cultures--indicating that surface phagocytosis by the neutrophils normally present during estrus could account for natural protection at this time. Mononuclear phagocytes also killed C fetus in the presence of opsonins, making this a likely factor in protection during the later stages of inflammation when mononuclear cells predominate. The presence of numerous lymphocytes in this mononuclear infiltration and the capability of C fetus antigens to induce delayed hypersensitivity (DH) are consistent with the possibility that cell-mediated immunity (CMI) also may be involved in protective immunity.

Animals↗

Association of blood proteins with large unilamellar liposomes in vivo. Relation to circulation lifetimes.

The proteins associated with liposomes in the circulation of mice were analyzed in order to determine whether bound proteins significantly influence the fate of liposomes in vivo. Liposomes were administered intravenously via the dorsal tail vein of CD1 mice and were isolated from blood after 2 min in the absence of coagulation inhibitors using a rapid "spin column" procedure. Various negatively charged liposomes exhibiting markedly different clearance properties were studied; notably, these included liposomes containing 10 mol % ganglioside GM1 which has been previously shown to effectively limit liposomal uptake by the fixed macrophages of the reticuloendothelial system. The protein binding ability (PB; g of protein/mol of lipid) of the liposomes was quantitated and related to the circulation half-life (tau 1/2) of the liposomes. Liposomes having similar membrane surface charge imparted by different anionic phospholipids were found to exhibit markedly different protein binding potentials. Furthermore, PB values determined from the in vivo experiments were found to be inversely related to circulation half-lives. PB values in excess of 50 g of protein/mol of lipid were observed for rapidly cleared liposomes such as those containing cardiolipin or phosphatidic acid (tau 1/2 less than 2 min). PB values for ganglioside GM1-containing liposomes (tau 1/2 greater than 2 h) were significantly less (PB less than 15 g of total protein/mol of total lipid). PB values were also determined for liposomes recovered from in vitro incubations with isolated human serum; relative PB values obtained from these in vitro experiments were in agreement with relative PB values measured from in vivo experiments. PB values, therefore, could be a useful parameter for predicting the clearance behavior of liposomes in the circulation. Liposomes exhibiting increased PB values in vivo were shown by immunoblot analysis to bind more immune opsonins, leading to a higher probability of phagocytic uptake. Finally, based on results obtained using the in vitro system, it is suggested that the mechanism by which ganglioside GM1 prolongs the murine circulation half-life of liposomes is by reducing the total amount of blood protein bound to the liposomes in a relatively nonspecific manner.

Animals↗

[The response of human lymphocytes to Staphylococcus aureus peptidoglycan in a system of opsonic cooperation].

Interaction of lymphocytes with S. aureus peptidoglycan treated with normal human serum and its fractions was studied on the basis of luminol-dependent chemiluminescent reaction. Treatment with whole serum led to the considerable increase and acceleration of lymphocytic reactions. The opsonic effect mainly depended on antibodies and complement, the contribution of other opsonins (which could be partially attributed to fibronectin) did not exceed 30%. IgG and fibronectin in concentrations, similar to their concentration in normal human plasma, enhanced the reactions 3.4 +/- 0.3 (p less than 0.05) and 1.5 +/- 0.005 (p greater than 0.05) times, respectively. The problem of the intrapopulation profile of opsonic-lymphocytic reactions and their role in the system of cell-mediated and humoral interactions is discussed.

Adult↗

Proteolysis of defensive proteins in peritonitis exudate: pathobiochemical aspects and therapeutical approach.

Peritonitis exudate reveals strong proteolytic activity which is paralleled by deficient opsonic capacity and high concentrations of lysosomal proteinases (elastase and cathepsin B). Lysosomal serine and cysteine proteinases (elastase, cathepsins B, L) were shown to degrade immunoglobulin G(IgG) and seem to be at least partially responsible for the observed proteolytic inactivation of IgG in peritonitis exudates. Intraabdominal serum application seems to restore opsonic activity by substitution of opsonins and proteinase inhibitors.

Ascitic Fluid↗

[Immune defenses of newborn infants].

Host defenses to bacterial infection are deficient in the neonate. This deficiency contributes to severe systemic infections in newborns. Phagocytosis of bacteria is decreased due to deficient activity of phagocytic cells and opsonines. Immunoglobulin secretion depending on B and helper T cell activities is strongly depressed due to immaturity of both populations. Therefore, maternal immunoglobulins of IgG type which cross placenta since 32 weeks of gestation play an important role in newborn defenses to bacteria even if this protection is incomplete.

Antibody Formation↗

[Protective effect of glycyrrhizine in mice with systemic Candida albicans infection and its mechanism].

We proved that a Chinese herb extract--Glycyrrhizine, the sweet component of Licorice root--protected mice from Candida albicans infection in doses of 0.5 mg/(kg.d) for 15-20 d. After administration of the above doses, the mortality rate of infected mice was decreased from 100% to 65%; the survival time was extended from 6.76 +/- 1.95 to 10.99 +/- 7.24 days after infection with lethal spores; and the toxic symptoms were also alleviated. The mechanism of protection is probably related to several factors, such as plasma opsonin activity promotion, enhancement of NK cell activity, increasing levels of plasma SOD, etc. All of this indicates that GCZ might be used as an accessory drug in the treatment of severe fungal infection.

Animals↗

Interaction between Pseudomonas aeruginosa and host defenses in cystic fibrosis.

The major causes of morbidity and mortality in cystic fibrosis are chronic pulmonary obstruction and infection. Mucoid Pseudomonas aeruginosa is the primary pathogen in up to 90% of these patients. Once Pseudomonas organisms colonize the airways, they are virtually never eradicated. No defect in systemic host defense has been elucidated, however, several mechanisms contribute to the breakdown in host defenses that allow persistence of this organism in the endobronchial space. These mechanisms involve both bacterial adaptation to an unfavorable host environment and impaired host response. P aeruginosa adapts to the host by expressing excessive mucoid exopolysaccharide and a less virulent form of lipopolysaccharide. These features make it less likely to cause systemic infection, yet still enable it to resist local host defenses. Mucociliary clearance becomes impaired due to abnormal viscoelastic properties of sputum, squamous metaplasia of the respiratory epithelium, and bronchiectasis. Despite a brisk antibody response to a variety of Pseudomonas antigens, several defects in antibody-mediated opsonophagocytosis have been identified. These include (1) development of antibody isotypes that are suboptimal at promoting phagocytosis, (2) formation of immune complexes that inhibit phagocytosis, and (3) proteolytic fragmentation of immunoglobulins in the endobronchial space. Complement-mediated opsonophagocytosis is also compromised by proteolytic cleavage of complement receptors from the cell surface of neutrophils and complement opsonins from the surface of Pseudomonas. The resultant chronic inflammation and infection lead to eventual obliteration of the airways.

Antibodies, Bacterial↗

Phagocytosis of synthetic particles in earthworms. Effect of antigenic stimulation and opsonization.

We examined the phagocytic activity of coelomocytes in earthworms Lumbricus terrestris. The synthetic polymeric HEMA particles were used in this study. The phagocytic capacity was increased after the protein administration or preincubation of HEMA particles with coelomic fluid. Furthermore, we isolated and characterized the proteins adsorbed on the surface of the particles which have opsonin-like properties.

Animals↗

Role of humoral factors in host resistance to the Bacteroides fragilis group.

The alternative complement pathway and antibody play an important role in host resistance to members of the Bacteroides fragilis group. Clinical isolates of the B. fragilis group are typically resistant to the bactericidal action of serum. Naturally occurring antibodies in serum directed against these bacteria belong primarily to the IgM class. These antibodies are not required for activation of the alternative pathway by the bacteria but rather potentiate alternative pathway activation. Activation of the alternative pathway by the bacteria leads to the generation of chemotactic activity for neutrophils and to C3 deposition on the bacterial surfaces. Both iC3b, the predominant molecular form of the bacteria-bound C3, and IgM antibody are necessary for phagocytosis and killing of the bacteria by neutrophils. IgM acts synergistically with iC3b to facilitate adherence of the bacteria to the neutrophils. Certain Bacteroides strains require additional as yet uncharacterized auxiliary opsonins for maximal adherence to neutrophils.

Antibodies, Bacterial↗

[Anti-keratin filament autoantibodies].

IgG anti-keratin-filament autoantibodies occur in all human sera and reach high titres in the sera of patients with various cutaneous and non-cutaneous diseases. At indirect immunofluorescence, these autoantibodies may, depending on their titre, appear as 'upper-cytoplasmic' antibodies or 'stratum-corneum' antibodies. In vitro, IgG anti-keratin-filament autoantibodies significantly enhance (as opsonins) the phagocytosis of keratin-filament aggregates by human monocytes and polymorphonuclear leucocytes. They may therefore, also in vivo, play a role in the removal of insoluble extracellular keratin-filament aggregates generated by necrotic or apoptotic cell death. Keratin (Civatte) bodies and deposits of localized cutaneous amyloid are examples of such keratin-filament aggregates that are found in various skin diseases but are also regularly present in large amounts in normal human skin. IgM anti-keratin-filament autoantibodies, which may, at indirect immunofluorescence, appear as 'general cytoplasmic' antibodies, also occur in all human sera and reach high titres in the sera of patients with various diseases. The regular presence of IgM in keratin bodies or deposits of localized cutaneous amyloid therefore probably represents the binding of IgM anti-keratin-filament autoantibodies to their respective antigens. To what extent these in vivo-bound autoantibodies prevent any further damaging reaction by the immune system in response to the liberation of keratin-filament autoantigen is as yet unclear.

Amyloid↗

[Active endogenous detoxication in the complex treatment of peritonitis].

A method of selective endogenous detoxication is described. Its aim is to enhance the elimination of large- and midmolecular pathological substances from the patient's organism by means of stimulation of the cell link of the mononuclear phagocyting system and correction of the level of blood opsonins. An examination of 150 patients with peritonitis of different etiology, spread and phases of the process has shown a pronounced improvement of clinico-laboratory indices in patients whose complex treatment included additional selective endogenous detoxication. Rapid arrest of the endogenous intoxication is associated with diminution of large- and midmolecular pathological substances in blood.

Adolescent↗

[The fibronectin deficiency in normal newborn infants corrects itself very quickly].

Fibronectin (FN) is a protein which acts as an opsonin and participates in anti-infective mechanisms. We measured FN in cord blood from 39 normal newborns by Laurell's electroimmunoassay. The mean value of FN was 13.5 +/- 6.8 mg/dl, lesser then mean value of 26 normal children 1 mo. to 7 y. old, 27.3 +/- 8.7 mg/dl (p less than 0.001). FN continued being low, 14.1 +/- 8.1 mg/dl al 48 h. of age. Nevertheless it was normal at 15 days of age (24.6 +/- 9.7 mg/dl). FN levels were lower at 15 days of age in 19 newborns who received vitamin K (12.2 +/- 6.1 mg/dl) than in the group which was not treated (18.1 +/- 9.9 mg/dl) but the difference was not significant (p greater than 0.05). There was a close correlation between cord blood levels and levels at 48 h. (p = 0.001) and levels at 15 d. (p = 0.003). There was no relationship either with the gestational age and birth weight, no prothrombin and protein C levels. The FN decrease in the newborn can be one of the causes of the opsonization deficiency, nevertheless normalization occurred very soon, before the second week.

Age Factors↗

Relative importance of Salmonella-specific antibody isotypes in phagocytosis of Salmonella typhimurium by ovine mammary neutrophils.

The comparative opsonic efficiency of ovine salmonella-specific antibody isotypes was determined by measurement of specific phagocytic uptake of opsonised virulent Salmonella typhimurium by ovine mammary neutrophils. An in vitro phagocytosis assay revealed that IgM was superior to IgG2 in promoting the phagocytosis of opsonised virulent organisms. IgG1, on the other hand, was non-opsonic. Superiority of the IgM isotype over IgG2 as an opsonin was also evident in studies on the viability of opsonised S typhimurium upon phagocytosis. It was revealed that the percentage of organisms killed was appreciably greater when opsonisation was carried out with the IgM than with the IgG2 isotype, although after ingestion by neutrophils there was essentially no difference in the efficiency with which the ingested organisms were killed.

Animals↗

Phagocytosis of Cryptococcus neoformans by rat alveolar macrophages.

A radiolabeling method was developed to investigate the interaction in vitro between Cryptococcus neoformans and Lewis rat alveolar macrophages (AM phi). AM phi were harvested by lung lavage, monolayers of adherent cells were established in wells of microtiter plates and [51Cr]-labeled yeast cells were added to the monolayers. After removal of extracellular yeasts, the adherent radioactivity associated with the AM phi was directly proportional both to the number of yeasts added and to the number of yeasts per AM phi as determined by microscopic examination of Giemsa-stained monolayers. Phagocytosis (attachment and/or ingestion) of radiolabeled C. neoformans by AM phi was a sensitive, quantitative and reproducible assay for the evaluation of the AM phi-C. neoformans interaction. AM phi were able to phagocytose encapsulated strains of C. neoformans. The extent of phagocytosis was inversely related to the capsule size. Normal rat serum (NRS) was an excellent source of opsonins for the ingestion. Inactivation of serum complement or depletion of C3 by affinity chromatography removed most of the opsonic activity of NRS. Specific antibodies against C. neoformans did not increase phagocytosis.

Agglutination Tests↗

Blood bactericidal assay (Pasteurella haemolytica) comparison of morbidity in marketed feeder calves.

An in vitro bactericidal assay that used bovine heparinized blood was investigated for its usefulness in detecting differences in the bactericidal immunity of calves against Pasteurella haemolytica serotype 1 (Ph1). Greater than 90% of killing occurred within 30 minutes. The substitution of fetal calf serum for autologous calf plasma caused loss of bactericidal activity of the blood. Decomplemented calf serum also was low in bactericidal activity. The blood bactericidal assay appears to be opsonin antibody-dependent and complement-dependent. The coefficient of variation (CV) that can be expected with this assay was established by use of a group of 8 calves; within-day CV maximum was 0.9, and between-day CV maximum was 2.1. The blood bactericidal assay was used to evaluate 30 calves under typical market stress from 4 farms in eastern Tennessee. All calves had decreased bactericidal activity, as they moved into a feedyard in Texas. The bactericidal activity was reduced among sick calves, based on the severity of clinical signs. Morbidity was highest during the first 14 days in the feedlot. During this period, healthy calves had a decreased bactericidal index (BI) of 4 points, and calves with clinical signs of bovine respiratory tract disease for 3 days had a decreased BI of 8 points. The average reduction in the BI of calves with clinical signs of bovine respiratory tract disease for 6 or more days was 14 points.

Animals↗

Antibody to the outer membrane proteins is the dominant opsonic antibody in normal human serum against H. influenzae type b.

We hypothesized that the predominant opsonic antibody of normal serum is directed against outer membrane proteins (OMP). Sera from 10 normal adults were tested for their opsonic capacity against Haemophilus influenzae b (Eagan strain) by the luminol-enhanced neutrophil chemiluminescence elicited on incubation with serum-opsonized bacteria, and the ability to deposit C3 on the bacterial surface. Peak chemiluminescence correlated with the amount of C3 on the bacterial surface (r = 0.71, P less than 0.025) and this, in turn, correlated with the concentration of IgG directed against outer membrane proteins, (r = 0.75, P less than 0.01), but not with the concentration of anticapsular polysaccharide antibody. Two groups of sera were easily distinguished based on the chemiluminescence experiments: a high opsonic group (greater than 50,000 peak counts per second; c.p.s.) and a low opsonic group (less than 10,000 c.p.s.). The IgG fraction from the high opsonic sera could augment C3 deposition when added to a low opsonic serum, but could not after absorption of the anti-OMP antibody by affinity chromatography. We conclude that the predominant opsonin of normal serum is antibody to outer membrane proteins, a finding which could be significant for the development of future vaccines against H. influenzae b.

Antibodies, Bacterial↗