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Surface phagocytosis and host defence in the peritoneal cavity during continuous ambulatory peritoneal dialysis.

Since patients on continuous ambulatory peritoneal dialysis are at high risk for peritonitis, the opsonins in peritoneal dialysis effluent responsible for phagocytosis and a neutrophil chemiluminescence response to surface-adherent Staphylococcus epidermidis were examined. In surface phagocytosis assays uninfected dialysate was as opsonic as 1% serum. The opsonic activity was heat stable and equal to that of purified IgG at the same concentration (0.1 mg/ml). In contrast, optimal chemiluminescence to surface-adherent Staphylococcus epidermidis was dependent on complement. C3 deposition on Staphylococcus epidermidis opsonized in dialysate was quantitated by an enzyme immunoassay (EIA) and represented 13% of control C3 deposited with opsonization in 10% serum. Unused dialysate was found to be inhibitory to neutrophil phagocytosis and complement deposition. A combination of the low pH and high dextrose concentration of dialysate was responsible, but restoration of the pH to 7.4 largely restored both indices. During peritonitis there was a parallel increase in IgG levels and C3 deposition (r = 0.8), and surface phagocytosis was also enhanced. On further analysis, subjects with a single episode of peritonitis had a significantly more opsonic peritoneal effluent than those who had two infections during the study. This latter group had a poor IgG response to infection. This study demonstrates the relative deficiencies of host defence in the peritoneal cavity and indicates that measures to improve opsonin delivery and reduce the inhibitory effects of dialysate would be beneficial.

Aged↗

The birth of immunology. III. The fate of the phagocytosis theory.

During the period of 1895-1910, immunology was preoccupied with defining the cellular (Elie Metchnikoff's phagocytosis theory) as opposed to the humoral basis of bactericidal defense. Although initial discovery of immunopathologic phenomena had been made (e.g., relating to transplantation, autoimmunity, allergy), focus on microbicidal therapy and diagnosis of infectious diseases remained the major stimuli of inquiry. The debate concerning the relative roles of phagocytes, complement, amboceptors (sensibilizing factors, antibody, antitoxins), various lysins (e.g., bacteriolysins, spermatolysins, hemolysins), agglutinins, stimulines, and then Almoth Wright's opsonins reflects the ambiguity of a scientific language being created in an era still struggling with a poorly defined experimental system, for the language, both its vocabulary (newly studied phenomena) and grammar (operational mechanisms) was yet to be codified. The joint award of the Nobel Prize to Metchnikoff and Paul Ehrlich in 1908 for their respective contributions to the "theory of immunity" appeared to proclaim a consensus, but the secret Nobel Committee reports that evaluated Metchnikoff's contributions reveal only a grudging acceptance of his position, and the award was clearly made on the basis of an apparent complementarity between the theoretical views of the humoralists and those elements of the phagocytosis theory that fit the then current discussion of immunity. In this regard, opsonins played an especially important role as both an experimental and conceptual bridge between the competing schools. What was no longer under consideration (and in fact never was explicitly debated) concerned the intellectual foundation of Metchnikoff's original concept of immunity as those activities that defined organismal identity, (developed from Metchnikoff's research in developmental biology) and which regarded host defense mechanisms as only subordinate to this primary function. Immunology in the first half of the 20th century pursued issues pertinent to chemically characterizing immune specificity and only later returned to the Metchnikovian question of how the immune identity was established. This latter venture has achieved molecular sophistication, but even such a formulation may be an inadequate answer to the Metchnikovian postulate. The theoretical discussion between cellularists and humoralists continues in new guises, for the essential debate remains unresolved.

Allergy and Immunology↗

Non-respiratory haemolymph proteins in the vineyard snail Helix pomatia. Changes after phagocytosis in vivo.

Haemolymph proteins from the vineyard snail Helix pomatia were studied by chromatographic and immune-electrophoretic techniques with the dual purpose of characterizing the normal composition of haemolymph and to look for possible opsonins. The oxygen-carrying proteins, alpha- and beta-haemocyanin, were by far the most abundant proteins, but at least three non-respiratory proteins could be demonstrated. We found consistent changes in the appearance of the immunoprecipitation pattern of these non-respiratory proteins after the injection of particulate foreign matter into the snail's circulation, and we suggest that they may be opsonins. We also found a haemagglutinin in haemolymph. It was present in very low concentration and was similar to, but not identical with the haemagglutinin which is present in the albumin gland.

Animals↗

Immunorecognition in the freshwater bivalve, Corbicula fluminea. I. Electrophoretic and immunologic analyses of opsonic plasma components.

Plasma (cell-free hemolymph) from the freshwater clam, Corbicula fluminea, has been found to possess opsonizing activity which could be completely eliminated by preabsorption of plasma with aldehyde-fixed rabbit red blood cells (RRBC). The detergents, SDS and Chapso, consistently dissociated several major proteins (Mr 20, 40, 66 and over 200 kd) from opsonized RRBC. Moreover, a partial recovery of opsonizing activity was attributed to this pool of desorbed proteins after removal of detergent by dialysis. Some of these proteins failed to bind to RRBC if the plasma was pretreated with heat, EGTA or preabsorbed with RRBC. These treatments were also found to eliminate plasma-opsonizing activity. A monoclonal antibody raised against opsonized RRBC bound to several of the same proteins seen in SDS-PAGE and could block opsonin-mediated RRBC phagocytosis by clam hemocytes. The blocking antibody also was crossreactive with epitopes expressed at the surface of washed hemocytes. Taken together these results suggest that the candidate opsonin(s) is a divalent cation-dependent, heat sensitive protein(s) with an Mr ranging from 20 to 66 kd (under reducing conditions). Furthermore, antigenically related forms of the molecules are found both in the plasma and associated with the surface membrane of circulating hemocytes.

Animals↗

Serum opsonic activity and neutrophil phagocytic capacity of newborn lambs before and 24-36 h after colostrum uptake.

Neutrophil (PMN) counts, immune complex (IC) uptake by PMN, and serum opsonising activity for promoting yeast uptake were used to evaluate infection clearing capacity in 16 lambs prior to colostrum feeding (two lambs fed bovine colostrum, 14 suckled lambs) and at 2 days of age. At 2 days of age lambs had more circulating PMN than they had prior to colostrum uptake (P less than 0.01). Colostrum feeding caused a significant increase in the percent of lamb PMN phagocytosing IC, although at Day 2 the percent phagocytosis was significantly lower (32.2%) than for adult controls (90%). Yeast opsonophagocytosis was greater when 24-36 h post-feeding serum was the source of opsonin than when pre-feeding serum was used (P less than 0.001). When adult serum was the opsonin, yeast opsonophagocytosis was approximately twice the phagocytosis mediated by 24-36 h post-feeding serum. The peripheral neutrocytosis and the enhancement of opsonophagocytosis generated by absorption of either ovine or bovine colostrum did not differ. The results of this study suggest that the parameters evaluated may be used for indicating the presence (or absence) of passively acquired protective immunity.

Animals↗

The opsonic activity of bovine milk whey for the phagocytosis and killing by neutrophils of encapsulated and non-encapsulated Escherichia coli.

Encapsulated strains of Escherichia coli were found to be more resistant to phagocytosis and killing by bovine neutrophils; requiring in the order of 100 times more serum opsonins than non-encapsulated strains. Mid-lactation pooled whey from cows with no history of mastitis was opsonic for non-encapsulated strains, but had no effect on encapsulated organisms. In contrast, early lactation pooled whey (5-10 days post-partum) was opsonic for all strains of E. coli. It is concluded that since early lactation milk contains sufficient opsonins, severe E. coli mastitis at this stage of lactation is not due to opsonic deficiency.

Animals↗

Opsonization, biodistribution, and pharmacokinetics of polymeric nanoparticles.

The process of opsonization is one of the most important biological barriers to controlled drug delivery. Injectable polymeric nanoparticle carriers have the ability to revolutionize disease treatment via spatially and temporally controlled drug delivery. However, opsonin proteins present in the blood serum quickly bind to conventional non-stealth nanoparticles, allowing macrophages of the mononuclear phagocytic system (MPS) to easily recognize and remove these drug delivery devices before they can perform their designed therapeutic function. To address these limitations, several methods have been developed to mask or camouflage nanoparticles from the MPS. Of these methods, the most preferred is the adsorption or grafting of poly(ethylene glycol) (PEG) to the surface of nanoparticles. Addition of PEG and PEG-containing copolymers to the surface of nanoparticles results in an increase in the blood circulation half-life of the particles by several orders of magnitude. This method creates a hydrophilic protective layer around the nanoparticles that is able to repel the absorption of opsonin proteins via steric repulsion forces, thereby blocking and delaying the first step in the opsonization process.

Animals↗

Cardiopulmonary bypass and host defense functions in human beings: I. Serum levels and role of immunoglobulins and complement in phagocytosis.

In patients undergoing open-heart surgical procedures, the serum levels of immunoglobulins and complement were determined as well as the functional capacity of these defense proteins as opsonins to facilitate phagocytosis by polymorphonuclear leukocytes. A considerable decrease in serum levels of the proteins studied was found after cardiopulmonary bypass (CPB). As a result, the opsonic capacity of post-CPB plasma was diminished. After correction for hemodilution, however, no difference between pre- and post-CPB plasma, as measured by the activity of thermolabile (e.g., complement C3) and thermostabile (e.g., immunoglobulin IgG) opsonins, could be demonstrated. It is concluded that CPB causes a quantitative but no functional decrease in levels of IgG and C3.

Adult↗

Altered tissue-specific opsonic activities and opsono-recognition of liposomes in tumour-bearing rats.

Reticuloendothelial phagocytic and serum opsonic activity was evaluated at terminal stages of tumour growth in rats transplanted subcutaneously with chondrosarcoma in an attempt to evaluate the role of opsonic protein(s) in governing liposome recognition and clearance by the macrophage system. The liver of the tumour-bearing animals manifested a decline in the uptake of multilamellar vesicles composed of egg phosphatidylcholine: cholesterol: dicetyl phosphate (mole ratio 7:2:1) from the blood when compared to healthy animals. In contrast, an increase in splenic clearance of liposomes was encountered in tumour-bearing rats. Studies with isolated liver non-parenchymal cells suggested that liposome recognition in both health and at terminal stages of cancer growth is influenced by a serum opsonin, which can be precipitated by 35-50% ammonium sulphate, as well as the concentration of calcium levels in serum. Serum of healthy animals equally enhanced liposome recognition by the hepatic macrophages of both normal and tumour-bearing rats. In contrast, both cell populations manifested poor liposome recognition in the presence of serum pooled from tumour-bearing animals and the results were comparable to the corresponding liposome-cell interaction in the absence of serum. The opsonic activity of serum derived from tumour-bearing rats could be demonstrated either by prior dialysis of serum against de-ionized water or by addition of EGTA. Liver phagocytes of healthy animals recognized more liposome in the presence of dialysed or EGTA-chelated tumour-serum than that of liver cells derived from tumour transplanted rats. A significant increase in serum calcium concentration was found in all tumour-bearing rats. When the concentration of calcium in the serum of normal animals was increased to the level that is encountered in tumour-bearing rats, a sharp drop in liposome recognition by liver phagocytes was observed. This drop in opsonic activity was not related to changes in the ionic strength of serum. The ammonium sulphate precipitated opsonin was also calcium-sensitive and its opsonic activity was abolished in the presence of calcium. Studies with isolated splenic phagocytes suggested that an increase in the opsonic activity of serum, but not the elevated calcium level, was responsible for hyperphagocytosis of liposomes by the splenic phagocytes of tumour-transplanted animals. The opsonic molecule which enhanced liposome recognition by liver non-parenchymal cells failed to enhance liposome clearance by the splenic phagocytes. These findings suggest that the alteration in macrophage clearance of liposomes during the terminal growth of cancer may be mediated in part by changes in the opsonic capacity of serum.

Animals↗

Functional assays for evaluation of serogroup B meningococcal structures as mediators of human opsonophagocytosis.

Functional flow cytometry and chemiluminescence (CL) assays have been modified to identify serogroup B meningococcal structures that mediate anti-meningococcal opsonophagocytosis. Serogroup B meningococcal outer membrane vesicles (OMV) were adsorbed to fluorescent latex beads (OMV-beads) and opsonized with acute phase and convalescence sera from patients with serogroup B meningococcal disease. Phagocytosis of these beads by human monocytes and polymorphonuclear leukocytes (non-lymphocytes) was dependent on both antigen exposure on the bead surface and on serum opsonization. OMV-beads opsonized with serum from a patient recovering from meningococcal disease, caused 97% of the non-lymphocytes to phagocytose an average of 15.8 beads per cell with a CL response of 46,550 mVs, whereas opsonized control beads were phagocytosed by 19% of the non-lymphocytes with 1.1 beads per cell and a CL response of 53 mVs. Increased amounts of functional, anti-OMV opsonins were detected during infection, and opsonized OMV-beads elicited phagocyte responses of similar magnitude to those of opsonized whole meningococci. Phagocyte internalization of OMV-beads was confirmed by confocal laser scanning microscopy. We conclude that epitopes on the meningococcal outer membrane are recognized by anti-meningococcal opsonins in these functional phagocytosis assays, which provide a basis for subsequent evaluation of various purified bacterial components as mediators of human opsonophagocytic responses and hence future vaccine constituents.

Antigens, Bacterial↗

Blood-transfusion in group-B streptococcal sepsis.

Transfusion of fresh whole blood was evaluated as a means of supplying opsonins and lessening the high mortality of group-B streptococcal sepsis in neonates. Pre-transfusion and post-transfusion sera from 22 infants were examined for the presence of opsonins against group-B organisms. Opsonic activity rose only when donor blood containing heat-stable antibody was administered in high volume (greater than 40% of blood-volume). 9 of 9 infants transfused with blood containing antibody to their infecting strain survived septic episodes. 3 of 6 who received blood lacking antibody to their infecting strain died.

Antibodies, Bacterial↗

Assessment of certain neutrophil receptors, opsonophagocytosis and soluble intercellular adhesion molecule-1 (ICAM-1) following thermal injury.

Polymorphonuclear leukocytes (PMLs) play a key role in host defense, and phagocyte dysfunction has been associated with increased susceptibility to infections in patients with thermal injury. Intercellular adhesion molecule-1 (ICAM-1) plays a role in leukocyte accumulation and extravasation. Therefore, the aim of the present study was to assess the PMLs expression of opsonin receptors: Fc gamma RIII, CR1 and CR3; opsonophagocytosis of PMLs and plasma soluble ICAM-1. Flow cytometric analysis (FCM) was used to study PMLs expression of IgG Fc-receptor III (Fc gamma RIII) as well as the complement receptors CR1 (receptor for C3b) and CR3 (receptor for C3bi) in 23 patients with large burns. Analysis of PML complement- and immunoglobulin-mediated phagocytosis of Candida albicans were performed in parallel using the phagocytic index. Plasma sICAM-1 was determined using ELISA. This study revealed a significant increase, with variable degrees, in CR1 and CR3-dependent fluorescence, complement-mediated phagocytosis of C. albicans and plasma sIGAM-1 that started at day 2 and remained for about 20 days before normalization. In contrast, Fc gamma RIII-dependent fluorescence and Ig-mediated phagocytosis were significantly decreased versus the control values. These results demonstrate significant changes of PMLs opsonin receptors expression and opsonophagocytosis documenting systemic activator of PMLs after large burns. In addition, elevation of plasma sICAM-1 may enhance the harmful effect of neutrophil activation through leukocyte accumulation and extravasation through endothelial damage in skin and in lung.

Adolescent↗

Influence of age and plasma treatment on neutrophil phagocytosis and CD18 expression in foals.

The aim of this study was to evaluate the influence of age and plasma treatment on neutrophil phagocytosis, CD18 expression and serum opsonic capacity in foals in field settings. Microbial infections constitute a large threat in young foals and neutrophil functions are crucial for the defense. Blood samples were obtained from 13 foals at seven time points between the ages of 2 and 56 days and once from 16 adult horses. Six of the foals were treated with adult plasma at the age of 1 week. Neutrophil phagocytosis of yeast after various opsonizations and the expression of complement adhesion receptor CD18 were analysed by flow cytometry. Autologous serum opsonization resulted in 52+/-6.1% phagocytic neutrophils in 2-day-old foals (n = 12), a significantly lower rate than in adult horses (mean 84+/-3.1%; n = 16). In foals, yeast ingestion per neutrophil was also lower than in adults. Opsonic capacity increased with age (p < 0.05), reaching adult levels at 3-4 weeks. An increase in serum opsonic capacity followed plasma treatment (p < 0.05). The phagocytic capacity of foal neutrophils at the time-points studied was equal to or higher than that in the adults, when pooled adult horse serum or anti-yeast IgG was used as opsonin. In foals, serum IgG concentration was negatively correlated to serum opsonic capacity. CD18 receptor expression was higher in neutrophils from foals (<21 days old) than in those from adult horses (p < 0.05). The results indicate that foals are transiently deficient in serum opsonic capacity, which negatively affects their capacity for neutrophil phagocytosis. These changes in serum opsonins, unrelated to IgG, may be important factors in susceptibility to infections in foals.

Actinomycetales Infections↗

Species difference in the disposition of liposomes among mice, rats, and rabbits: allometric relationship and species dependent hepatic uptake mechanism.

PURPOSE: The species difference in the pharmacokinetics of liposomes was investigated in mice, rats and rabbits. METHODS: Liposomes were intravenously injected at doses of 1, 10 and 100 (nmol/g body weight), and the time courses of liposomes in blood, liver and spleen were measured. Pharmacokinetic parameters were regressed as a function of body weight (BW) and dose of liposomes (D). The uptake mechanism of liposomes was also examined with the isolated perfused liver between rats and mice. RESULTS: Mean residence time increased with the increase of BW and D of liposomes. This increase of mean residence time resulted from the decreased total body clearance, which was principally explained by the species difference in the hepatic uptake clearance (CLh) of liposomes. The parameter CLh was regressed well by a multiple regression as a function of BW and D. In this analysis, an exponent for BW was around 0.5, which clearly indicates that smaller animals have higher uptake clearance per unit BW. Immunohistochemical analysis revealed that there was no significant difference in the density of Kupffer cells among these species. This suggest that the species difference in CLh resulted not from the density of Kupffer cells but from the uptake ability of Kupffer cells among species. In the isolated perfused liver, the hepatic uptake of liposomes was mainly explained by opsonin dependent uptake in rats, while opsonin independent uptake in mice. CONCLUSIONS: These quantitative and qualitative information on the species difference of liposome disposition will provide an useful information for constructing a drug delivery system using liposomes.

Animals↗

Enhanced hepatic uptake of liposomes through complement activation depending on the size of liposomes.

The objective of this study was to differentiate the roles of opsonins and phagocytic cells in the size-dependent hepatic uptake of liposomes in the submicron region. The extent of opsonization decreased with the decrease in size of liposomes (from 800 to 200 nm in diameter) and no enhancement of uptake was observed at 200 nm. There was no effect of liposome size on the uptake of unopsonized liposomes. Serum was pretreated with empty liposomes of each size and its opsonic activity was measured in the perfused liver. The small liposomes could not consume the opsonic activity, while the larger ones did so substantially. These results suggest that opsonins bind to liposomes depending on the size of liposomes and phagocytic cells take up liposomes in proportion to the extent of opsonization. Size-dependent liposome degradation in serum was also found, which was consistent with the size-dependent complement activation, because liposomes with this composition have been shown to be degraded by complement. The mechanism of opsonization was examined by treating serum at 56 degrees C for 30 min or with anti-C3 antiserum. Since both treatments inhibited the opsonic activity, the hepatic uptake of liposomes is considered to occur via complement receptor. In conclusion, the size of liposomes affected complement recognition, and the liposomes were taken up by the liver depending on the extent of opsonization.

Animals↗

Difference in binding of killed and live Streptococcus pneumoniae serotypes by C-reactive protein.

The binding and opsonic properties of C-reactive protein (CRP) for various species of bacteria were investigated. CRP bound more avidly to killed than to live Streptococcus pneumoniae, the binding varying among various serotypes; CRP hardly bound to a number of other bacterial species studied. CRP enhanced complement-dependent phagocytosis of live S. pneumoniae by granulocytes but did not enhance the phagocytosis of live Staphylococcus aureus or group B streptococci. We suppose that CRP may serve as an opsonin for killed bacteria and bacterial debris but is probably not an important opsonin for live bacteria other than S. pneumoniae.

Adult↗

Phagocytosis and destruction of Staphylococcus aureus.

A review is presented for the phagocytosis and intracellular killing of Staphylococcus aureus by polymorphonuclear and mononuclear leukocytes. Recruitment of adequate numbers of leukocytes to the site of infection occurs through the process of chemotaxis. Recognition of invading staphylococci by the phagocytic cells is mediated through bacterial opsonization. Both processes depend upon the activation of the heat-labile complement system which generates the majority of chemotactic (C5a) and opsonic (C3b) molecules for S. aureus phagocytosis. The key role of peptidoglycan in the cell wall of staphylococci in these events is stressed. Attachment and ingestion of opsonized staphylococci occurs via poorly-defined receptors for opsonins in the membrane of the leukocyte. The greater phagocytic capacity of neutrophils as compared to monocytes is not reflected in differences in their membrane receptors for staphylococcal opsonins. Once ingested, staphylococci are rapidly destroyed by oxygen-dependent and oxygen-independent bactericidal mechanisms of the phagocytes. Small numbers of S. aureus may survive within the leukocyte. Special attention is focused on the numerous ways S. aureus is able to hinder, evade, and directly damage the phagocytic defense mechanisms of the host.

Cell Membrane↗

Phagocytosis of Giardia lamblia trophozoites by human colostral leukocytes.

AIM: To determine the phagocytic activity of the polymorphonuclear and mononuclear cells present in human colostrum, and to verify the influence of opsonins in the adherence, ingestion and killing of Giardia lamblia trophozoites. METHODS: Polymorphonuclear and mononuclear phagocytes were incubated with G. lamblia trophozoites, in the presence as well as the absence of supernatant of human colostrum (the source of opsonins) for 30, 60 and 120 min. The trophozoites/phagocytes ratio was 1:1, and the percentage of phagocytosed trophozoites was determined by microscopic examination of acridine orange-stained cells. RESULTS: The mononuclear phagocytes presented more functional activity than the polymorphonuclear. The highest indexes of adherence (77.6 +/- 5.1), ingestion (68.9 +/- 5.5) and killing (48.5 +/- 4.9) were obtained through the incubation of mononuclear cells in the presence of colostrum supernatant for 120 min. CONCLUSION: The phagocytes of human colostrum were able to ingest G.lamblia trophozoites and presented microbicidal activity in vitro, suggesting that these phagocytes may act as an additional mechanism of protection against infant giardiasis through breastfeeding.

Adolescent↗