Molecular Weight Immune Complexes Research Articles

CHANGES IN THE CONTENT OF IMMUNOGLOBULINS AND CIRCULATING IMMUNE COMPLEXES IN THE BLOOD SERUM OF RABBIT RECIPIENTS DURING ALLOGENEOUS WHOLE BLOOD TRANSFUSION

The paper presents the results of research on changes in the content of immunoglobulins and circulating immune complexes in the blood serum of recipient rabbits after allogeneic whole blood transfusion. The relevance of the work is determined by the importance of monitoring the state of immunoglobulins and CIC, since these indicators inform about the success of the blood transfusion procedure and the removal of the material by the recipient's immune system. Thus, the aim of the work was to study changes in the content of immunoglobulins and CIC in the blood serum of recipient rabbits. The material for research was blood serum samples obtained from 5 rabbits on 3, 8 and 23 days after transfusion. In the experiments, clinically healthy animals were used, which were transfused with whole blood. The feeding diet of the experimental animals met the need for nutrients and biologically active substances, the animals had free access to water. Blood from donor animals was collected from the jugular vein using a semi-closed method. Donor blood samples of rabbits were prepared in polymer containers with the anticoagulant Tsfda. Keeping animals, transfusions, manipulations and conducting research were carried out on the basis and in the conditions of Vedmedservis NSC, Animal Blood Bank NNL and the Department of Surgery and Pathophysiology named after Acad. I.O. Povazhenka, NULES of Ukraine. The study of the content of immunoglobulins in the blood serum of rabbits was carried out according to the Mancini method, the CIC was determined by the method based on the selective precipitation of high molecular weight immune complexes contained in the blood serum with polyethylene glycol with a molecular weight of 6000Da, followed by the determination of the optical density by spectrophotometry at λ = 450 nm. Based on the results of the research, it was established that with allogeneic transfusion of whole blood to recipient rabbits, there is a significant increase in the content of immunoglobulins of class M in blood serum on the 3rd day of the experiment and a decrease in their content on the 23rd day of the experiment. It was shown that with allogeneic transfusion of whole blood to recipient rabbits, the content of immunoglobulins of class G and A in blood serum decreases during the 23 days of the experiment. Allogeneic transfusion of whole blood in the body of recipient rabbits activates the formation of circulating immune complexes in the blood serum of animals, which can be deposited in the perivascular space and cortical layer of the kidneys, causing complement activation and inflammatory processes.

Program extracorporeal photophoresis in complex treatment of psoriatic arthritis

The results of complex treatment with the programmed procedures of extracorporeal photopheresis and methotrexate in patients with psoriatic arthritis for 12 months are presented. It has been established that program extracorporeal photopheresis in combination with basic therapy with methotrexate in patients with psoriatic arthritis have a significant effect on the relief of articular syndrome (reduction of morning stiffness, pain intensity, joint swelling). It was revealed that the inclusion of programmed extracorporeal photopheresis and methotrexate in the complex treatment of psoriatic arthritis reduces the level of circulating low molecular weight, medium molecular weight and high molecular weight immune complexes and in most cases leads to clinical and immunological remission. This is confirmed by a decrease in complaints of the severity of articular syndrome and a decrease in the clinical and laboratory activity of the disease (a significant decrease in the level of C-reactive protein, erythrocyte sedimentation rate). A scheme has been developed for the programmed use of extracorporeal photopheresis procedures in the complex treatment of psoriatic arthritis, which consists of 2 procedures every other day with an interval of 2,5-3 months for 12 months. Combined therapy with the programmed application of extracorporeal photopheresis procedures and methotrexate prolongs the period of disease remission and can be recommended for patients with psoriatic arthritis with high activity, insufficient effect of basic therapy and intolerance to higher doses of immunosuppressive drugs.

ETHIOPATHOGENETIC BIOMARKERS FOR THE FORMATION OF HEPATOSPLENOMEGAL SYNDROME AND THE CHOICE OF TREATMENT OF PATIENTS WITH ESOPHAGEAL PHLEBECTASIS BLEEDING

Summary. A range of specific biomarkers to evaluate the severity of the condition, the choice of tactics, effectiveness and prognosis of treatment are used.
 Goal. Evaluation of biomarkers characterizing the etiopathogenetic disorders in hepatosplenomegaly syndrome complicated by recurrent bleeding from esophageal phlebectasis to select individual treatment tactics.
 Materials and methods. The digestive function of phagocytes is investigated, the concentration of opsonizing, membranotropic cytotoxic factors and the degree of endogenous intoxication in 3 groups of patients with different etiological factors of hepatosplenomegaly, complicated by bleeding are evaluated.
 Results. The presence of specific trigger factors (viral hepatitis, herpes viruses, hereditary-related disorders of lysosomal enzymes) made it possible to classify patients into three groups. In the first group of patients with viral hepatitis B and C the induction of phagocytic activity of neutrophils, a significant increase in lymphocytotoxicity on the background of disorders of protein and lipid metabolism showed. In the second group of patients with the presence of herpes group viruses and autoimmune component an imbalance of opsonizing factors revealed, in particular proteins of the complement system, which caused impaired clearance of low molecular weight immune complexes and products of their degradation. In the third group of patients with hepatitis a significant decrease in the activity of lysosomal enzymes of phagocytic cells are found. The identified pathogenetic biomarkers indicate the presence in this group of patients with congenital enzymopathy characteristic of accumulation diseases. For patients with a pronounced manifestation of intercellular interactions disorders, as an effective alternative to liver transplantation the use of transfusions of mesenchymal stem cells, which are capable of trans differentiation under the influence of mediator microenvironment factors may be.

Non-enzymatic glucosylation induced neo-epitopes on human serum albumin: A concentration based study.

Hyperglycaemia induced non enzymatic glycation is accelerated in diabetic patients and aggressively involved in diabetes progression. Human serum albumin (HSA) is the most abundant protein in blood circulation. In hyperglycaemia, it undergoes fast glycation and results in the impairment of structure. Our previous study has demonstrated structural alterations in Amadori-albumin modified with different glucose concentrations from physiological to pathophysiological range. Here, we focused on immunological characterization of Amadori-albumin. Immunogenicity of Amadori-albumin was analysed by direct binding and competitive ELISA. Amadori-albumin was found to be highly immunogenic (expect albumin modified with 5mM) and induced high titre antibodies depending upon the extent of modification. Very high titre antibodies were obtained with albumin modified with 75mM glucose as compared to native albumin. Anti-Amadori-albumin-IgG from rabbit sera exhibited increased recognition of Amadori-albumin than native albumin in competitive immunoassay. Alteration induced in albumin after glucosylation has made it highly immunogenic. Induced antibodies were quite specific for respective immunogens but showed cross-reaction with other Amadori/native proteins. It suggests that glucosylation has generated highly immunogenic epitopes on albumin. Formation of high molecular weight immune complex with retarded mobility further supports specificity of anti-Amadori-albumin-IgG towards Amadori-albumin. It may be concluded that due to early glycation, an array of modification occurred in HSA structure. Such gross structural changes might favour polymerization of most of the native epitopes into potent immunogenic neo-epitopes, but some original epitopes were still active and has contributed in the immunogenicity. It could be concluded that induction of anti-Amadori-albumin antibodies may be due to protection of glucose modified albumin from protiolytic breakdown. We assumed that this type of protein modifications might occur in diabetic patients in hyperglycaemic conditions that may be recognised as foreign molecules and can induce autoantibodies. Increased level of anti-Amadori-albumin autoantibodies may be used as a biomarker in disease diagnosis and its progression.

Changes in the Plasma Proteome of Manduca sexta Larvae in Relation to the Transcriptome Variations after an Immune Challenge: Evidence for High Molecular Weight Immune Complex Formation

Manduca sextais a lepidopteran model widely used to study insect physiological processes, including innate immunity. In this study, we explored the proteomes of cell-free hemolymph from larvae injected with a sterile buffer (C for control) or a mixture of bacteria (I for induced). Of the 654 proteins identified, 70 showed 1.67 to >200-fold abundance increases after the immune challenge; 51 decreased to 0-60% of the control levels. While there was no strong parallel between plasma protein levels and their transcript levels in hemocytes or fat body, the mRNA level changes (i.e.I/C ratios of normalized read numbers) in the tissues concurred with their protein level changes (i.e.I/C ratios of normalized spectral counts) with correlation coefficients of 0.44 and 0.57, respectively. Better correlations support that fat body contributes a more significant portion of the plasma proteins involved in various aspects of innate immunity. Consistently, ratios of mRNA and protein levels were better correlated for immunity-related proteins than unrelated ones. There is a set of proteins whose apparent molecular masses differ considerably from the calculatedMr's, suggestive of posttranslational modifications. In addition, some lowMrproteins were detected in the range of 80 to >300 kDa on a reducing SDS-polyacrylamide gel, indicating the existence of highMrcovalent complexes. We identified 30 serine proteases and their homologs, 11 of which are known members of an extracellular immune signaling network. Along with our quantitative transcriptome data, the protein identification, inducibility, and association provide leads toward a focused exploration of humoral immunity inM. sexta.

Immunogenicity, Inflammation, and Lipid Accumulation in Cynomolgus Monkeys Infused with a Lipidated Tetranectin-ApoA-I Fusion Protein

High density lipoprotein (HDL)-targeted therapies, which promote cholesterol efflux from cells, are currently in development for reducing cardiovascular events in acute coronary syndrome. Human apolipoprotein A-I (apoA-I), the major HDL protein, was fused to the trimerization domain of tetranectin (TN) and complexed with phospholipids to generate a HDL mimetic (lipidated TN-ApoA-I) with reduced renal clearance and enhanced efficacy. Cynomolgus monkeys received 24-h intravenous infusions of control, 100 mg/kg or 400 mg/kg lipidated TN-ApoA-I every 4 days for 3 weeks, followed by a 6-week recovery period. After multiple infusions of lipidated TN-ApoA-I, clinical condition deteriorated and was accompanied by changes indicative of a progressive inflammatory response; increased levels of cytokines, C-reactive protein and vascular/perivascular infiltrates in multiple tissues. Rapid formation of antidrug antibodies occurred in all animals receiving lipidated TN-ApoA-I. Enhanced drug clearance corresponding to a relative lack of high molecular weight immune complexes in blood, suggestive of preferred removal/clearance, was observed in some animals. Expected dose-dependent increases in serum lipids were accompanied by vacuolated monocytes/macrophages in multiple organs, which in the glomeruli were shown to be CD68-positive, contain lipid and co-localized with granular IgG deposits. Lipid accumulation may have been a direct result of a high drug load, possibly enhanced by immune complex formation, inflammation, and altered lipid metabolism. Noteworthy was the inter- individual inconsistency in the severity of clinical and histopathologic findings, drug clearance and inflammatory markers. In conclusion, multiple infusions of lipidated TN-ApoA-I resulted in high immunogenicity, lipid accumulation and were not well tolerated in nonhuman primates.

Studies on peroxynitrite-modified H1 histone: Implications in systemic lupus erythematosus

Peroxynitrite is a powerful nitrating and oxidizing molecule and capable of modifying proteins' structure. Hyper-nitration of tyrosine residues has been seen in various pathological states, including autoimmune disorders like systemic lupus erythematosus (SLE) and rheumatoid arthritis. SLE, a chronic autoimmune disease, is primarily characterized by increased levels of autoantibodies, predominantly against ds-DNA. However, the initial antigenic stimulus for the disease etiopathogenesis has remained elusive. Carbonyl and nitrotyrosine have been extensively used as a biomarker of oxidative and nitrosative stress. In this study, commercially available H1 histone was exposed to increasing concentrations of peroxynitrite for 30 min. The peroxynitrite-mediated structural changes in histone were studied by ultraviolet & fluorescence spectroscopy, CD, HPLC, 1-anilinonaphthalene-8-sulfonic acid binding and polyacrylamide gel electrophoresis. Analysis of results revealed that carbonyl and nitrotyrosine contents were significantly increased in peroxynitrite-modified H1 compared to native H1. In experimental animal, peroxynitrite-modified H1 induced high titre antibodies as compared to native H1, and the immunogenicity was found to be directly proportional to nitrotyrosine content. Further, the induced antibodies showed specificity for the immunogen and appreciable cross-reactions with tyrosine rich nitrated proteins. Formation of high molecular weight immune complex with retarded mobility further supports the specificity of induced anti-peroxynitrite-modified H1 antibodies for the immunogen. Binding of SLE anti-DNA autoantibodies with peroxynitrite-modified H1 was analyzed by direct binding and competition ELISA. The data show preferential binding of SLE autoantibodies to peroxynitrite-modified H1 as compared to native H1 histone and native DNA. The results point towards the possible role of peroxynitrite-modified H1 histone in SLE etiopathogenesis.

Impact of Peroxynitrite Modification on Structure and Immunogenicity of H2A Histone

Peroxynitrite is an extremely reactive entity and has in vivo existence. Its interaction with biomolecules may cause oxidation and nitration. In this study, commercially available H2A histone was exposed to peroxynitrite generated in vitro. The peroxynitrite-mediated structural changes in histone were studied by ultraviolet & fluorescence spectroscopy, high performance liquid chromatography, anilinonaphthalene-8-sulfonic acid binding and polyacrylamide gel electrophoresis. Analysis of results revealed that carbonyl, nitrotyrosine and dityrosine contents were significantly increased in peroxynitrite-modified H2A compared with native H2A. Rabbits challenged with peroxynitrite-modified H2A induced high titre antibodies. The immunogenicity of peroxynitrite-modified H2A was directly proportional to protein nitrotyrosine content and induced antibodies showed specificity for the immunogen and good cross-reaction with nitrated epitopes of other modified proteins. Formation of high molecular weight immune complex with retarded mobility further supports the specificity of induced anti-100 mum peroxynitrite-modified H2A antibodies for the immunogen. It may be concluded that induction of anti-H2A histone antibodies could be due to protection of peroxynitrite-modified histone from proteolytic breakdown and its subsequent recognition by immunoregulatory cells as foreign molecule.

Blood Group O and ABO Non-Identical Platelet Transfusions Are Associated with Increased Red Cell Utilization in Non-Trauma Surgical Patients.

The transfusion of ABO non-identical platelets is an accepted practice, involving infusion of antigen and/or antibody that is ABO incompatible with the recipient. This results in the formation of high molecular weight immune complexes, which could, in theory, adversely affect platelet and coagulation factor function. Studies have demonstrated putative deleterious effects of ABO non-identical transfusions. In cardiac surgery patients receiving ABO non-identical platelets required 50% more red blood cell (RBC) transfusions than those who received only ABO identical platelets (Transfusion 41:790, 2001). Furthermore, blood group O and A patients have genetically determined lower levels of von Willebrand factor (vWF) than B and AB patients (Blood 69:1691, 1987), and might be expected to have more bleeding and require more RBC transfusions. Our institution implemented a protocol of transfusing only ABO identical components in April, 2005. We report here the potential effects of transfusing ABO non-identical platelets, by comparing RBC usage in surgical patients before and after introduction of a protocol of “only ABO identical” transfusions. We also report the role of ABO blood group in red cell usage. Because non-group O trauma patients by necessity receive group O red cells we excluded them from this analysis. Transfusion records for all non-trauma surgical patients receiving any platelet transfusions for one year after April, 2005 were reviewed. Patients receiving only ABO identical platelet transfusions were analyzed by blood groups: B/AB versus O/A. Group B/AB patients (n=72) required a mean of 12.5 +/− 11.7 RBCs, whereas group A/O patients (n=313) required 17.8 +/− 20.5 (p=0.036). These findings are the reverse of the patient populations receiving a mixture of ABO identical and non-identical platelets prior to April, 2005, before the implementation of an “ only ABO identical” transfusion policy. Group B/AB patients, who received more ABO non-identical platelet transfusions than A/O patients (mean 7.5 +/− 11 versus 3.3 +/− 6.2, p=0.0001), required significantly more RBCs (mean 19 +/− 25 versus 13 +/− 13, p=0.0086). Amongst those receiving only ABO identical transfusions group O patients required the largest number of RBCs, (mean 19.6 +/− 24.1), followed by A (16.0 +/− 15.8, p=0.096), B (13.2 +/− 13.1, p=0.041), and AB (11.2 +/− 7.8, p=0.05). One plausible explanation is that blood group O patients have the lowest levels of vWF and other clotting factors, while AB patients have the highest levels (AB > B > A > O). Our data in recipients of ABO identical transfusions confirm findings in the literature that group O patients may be at greater risk of hemorrhage than groups B and AB patients. However, when ABO non-identical platelets are transfused, significantly more bleeding is seen in group AB and B patients than in group O patients. We hypothesize that ABO non-identical transfusions may impair, rather than improve, hemostasis in some patients.RBC Usage (Units) Pre and Post ABO Identical Only PolicyBlood Groups A/OBlood Groups B/ABP-ValuePre-ABO Identical Only Policy13 +/− 1319 +/− 250.0086Post-ABO Identical Only Policy17.8 +/− 20.512.5 +/− 11.70.036

Distribution of radiolabelled anti-CA125 monoclonal antibody OC125-F(ab′)2-fragment following resection guided by antibodies (REGAJ) in ovarian cancer patients

Ovarian cancer is a highly malignant tumor of mainly postmenopausal women. The long-term prognosis of this malignancy is largely determined by micrometastasis present at the time of second-look surgery. In general, patients face a poor outcome. New radio-immunoscintigraphic methods to target tumor tissue specifically via antigen-antibody binding were developed. However, few studies so far investigated the pattern of in vivo distribution of radiolabelled mAbs and/ or the specificity of antigen-antibody interaction. In this study we examined the immunological interaction and distribution of 131l-OC125-F(ab')2-fragment, an anti-CA-125 mAb, in patients with CA-125 positive ovarian malignancies. Sixteen patients with primarily CA-125 positive gynecological tumors underwent REGAJ surgery. Biopsies of tumor tissue and not tumor infiltrated tissue, serum, and ascites were sampled during or prior to REGAJ surgery, respectively. After preparation of tissue cytosols, samples were assessed for CA-125 and radioactive uptake. By radiochromatography immunological analysis for presence of the target antigen CA-125, the mAb 131l-OC125-F(ab')2-fragment, and immune complexes was performed on different specimen. CA-125 concentrations were higher in serum samples, ascites, and malignant tissue biopsies of malignoma patients compared to those without signs of malignant disease. CA-125 was higher in the tissue cytosol than in the cell membrane fraction. Gel filtration revealed CA-125 with moieties of 75,000 to > 600,000 d. Accumulation of radioactivity was more frequently associated with the presence of unbound 131l-OC125-F(ab')2-fragment or high molecular weight immune complexes. Radioactive uptake, however, was not confined to tissue of high CA-125 expression. Moreover, both immune complex as well as 131l-OC125-F(ab')2-fragment could be isolated from cytosols of tissue not infiltrated by tumor cells as well. Our study demonstrates that the majority of CA-125 is located intracellularly and thus inaccessible to 131l-OC125-F(ab')2-fragment per se. The uptake of 131l-OC125-F(ab')2-fragment into the cytosol of tumor-free and malignant tissue samples prompts us to speculate that certain mechanisms for antigen-specific and nonspecific cellular trafficking of mAbs do exist. We present a model to explain our observations.

Rapid assay for immune complex bound antigens from urine of cancer patients.

Concentrated urine from patients with various types of cancer was fractionated by S-500 gel filtration chromatography to yield high molecular weight (mw) immune complexes (IC) and lower mw immunoglobulin (Ig) fractions. Column fractions were assayed for the presence of IgM and IgG by immunoblot probing using anti-human IgM and anti-human IgG alkaline phosphatase conjugates. These results were used to define IC fractions (those from high mw S-500 fractions which were positive for immunoglobulin) and Ig fractions (those from medium mw S-500 fractions which were positive for immunoglobulin). Antigen components of high mw S-500 IC fractions were then determined by immunoblot probe using medium mw S-500 Ig fractions as the antibody probe (i.e. autoantibody). This method for identification of immune complex antigens has the potential to probe for tumor-associated antigens, autoantigens, or foreign antigens from starting material which contains both immune complex and free immunoglobulin.

Fcγ Receptor Recognition of IgG Ligand by Human Monocytes and Macrophages

We examined the binding characteristics of human monocytes and macrophages with the IgG ligands, human monomeric IgG and a small human IgG aggregate, trimeric IgG. Our purpose was to utilize fresh monocytes, in vitro cultured monocytes, and alveolar macrophages in direct and indirect binding experiments. Freshly isolated monocytes expressed only a single binding site for IgG monomer and IgG trimer. In contrast, in vitro cultured monocytes, gamma-interferon-treated monocytes, and freshly isolated alveolar macrophages expressed a single binding site for IgG monomer and, in addition, a high and low affinity binding site for IgG trimer. The high affinity binding site for IgG trimer (Kd approximately equal to 1 nM) appeared identical to the binding site for IgG monomer. The low affinity binding site for IgG trimer (Kd = 50 to 250 nM) appeared to be due to Fc gamma RII, because antibody to Fc gamma RII inhibited its expression. Since Fc gamma RII, in contrast to Fc gamma RI, does not bind monomeric IgG, the data suggest that this low affinity receptor for trimeric IgG, Fc gamma RII, can bind low molecular weight circulating immune complexes at concentrations 10- to 100-fold lower than Fc gamma RI. Thus, these studies suggest that at 37 degrees C, macrophage Fc gamma RII may play a functional role in the recognition of small molecular weight immune complexes.

Modulation of Fcγ receptors on the human macrophage cell line U-937

Macrophage receptors for the Fc portion of IgG play an important role in host defense, inflammation, and the pathophysiology of autoimmune disorders. We studied one important function of Fc gamma receptors--the ability to bind IgG ligand. Direct binding experiments analyzed by nonlinear regression were consistent with monomeric and trimeric IgG binding to a single class of receptors. Indirect binding experiments were also consistent with this interpretation and revealed that both IgG ligands completely inhibited the binding of the other. In addition, we used an anti-Fc gamma RII monoclonal antibody known to compete for the Fc gamma RII ligand binding site and known to inhibit IgG trimer binding to other cells. At concentrations of antibody which saturated all Fc gamma RII sites, no inhibition of IgG trimer binding to U-937 was observed. This was evident despite the observation that the numbers of Fc gamma RI and Fc gamma RII, determined by equilibrium binding of monomeric IgG and anti-Fc gamma RII antibody, respectively, were similar on U-937. Monoclonal antibodies were used to compare the expression and modulation of Fc gamma receptor proteins with their ability to bind monomeric and trimeric IgG ligands. Dexamethasone and gamma-interferon regulated U-937 Fc gamma RI protein expression and IgG ligand binding to a similar degree. In contrast, the expression of Fc gamma RII was not altered by dexamethasone. Interferon-gamma primarily stimulated Fc gamma RI, as determined both by reactivity with monoclonal antibody (227 +/- 26%) and by monomeric IgG ligand binding (350 +/- 151%). In addition, dexamethasone inhibited by 33% the gamma-interferon effect on Fc gamma RI protein and by 56% the effect on Fc gamma RI binding of monomeric IgG. Preincubation of U-937 with anti-Fc gamma RII antibody did not alter the effect of dexamethasone or gamma-interferon on IgG trimer binding. These data indicate that on U-937 cells Fc gamma RII does not function in the recognition of small molecular weight immune complexes and that Fc gamma RI is the Fc gamma receptor responsible for the binding of both monomeric and trimeric human IgG. Furthermore, Fc gamma RI is the major Fc gamma receptor on U-937 that is modulated by both gamma-interferon and glucocorticoids.

Biological property of immune complex and its pathogenicity to persist otitis media with effusion

This study was designed to evaluate the biological property of light modecular weight immune complex, in comparison with that of heavy immune complex. Soluble immune complexes were obtained by mixing horseradish peroxidase (HRP) with guinea pig's antiserum to HRP and were devided into several fractions according to their molecular weights by a gel filtration. After the chemotactic activity of each fraction was examined in vitro, the lightest and the heaviest molecular weight immune complexes were injected into a guinea pig's left and right superior bulla, respectively. Although a middle ear effusion developed in both sides of the bulla, the histopathological feature of an experimentally induced otitis media with effusion varied with the molecular weight of the injected immune complex. The light molecular weight immune complex was proved to be a stronger inducer for macrophages than the heavy molecular weight immune complex both in vitro and in vivo, and to have a tendency to persist in the tympanic cavity for a longer duration than the heavy molecular weight immune complex. These results suggested that an otitis media was susceptible to persist when a light molecular weight immune complex was formed in the middle ear. The light molecular weight immune complex was suspected to play a role in the pathogenesis of otitis media with effusion.

Biological property of immune complex to prolong the duration of otitis media with effusion.

This study was designed to evaluate the biological properties of a light molecular weight immune complex, in comparison with that of a heavy molecular weight immune complex. Soluble immune complexes were obtained by mixing horseradish peroxidase (HRP) with guinea pig antiserum to HRP and were divided by gel filtration into several fractions according to their molecular weight (MW). After the chemotactic activity of each fraction was examined in vitro, the lightest and the heaviest MW immune complexes were injected into a guinea pig's left and right superior bulla, respectively. Although a middle ear effusion developed in both the bulla, the histopathological feature of an experimentally induced otitis media with effusion varied with the MW of the injected immune complex. The light MW immune complex proved to be a stronger inducer for a macrophage than the heavy MW immune complex, both in vitro and in vivo, and to have a tendency to persist longer in the tympanic cavity than the heavy MW immune complex. These results suggested that otitis media had a tendency to persist when a light MW immune complex was formed in a middle ear. The light MW immune complex was suspected to be involved in the pathogenesis of otitis media with effusion.

Antibody response of mice to lactate dehydrogenase-elevating virus during infection and immunization with inactivated virus

BALB/c and Swiss mice were infected with lactate dehydrogenase-elevating virus (LDV) or immunized with glutaraldehyde-inactivated or ether-extracted virus and their plasma was monitored for anti-LDV IgG and IgM levels by ELISA and indirect fluorescent antibody staining, for neutralizing antibodies, for sensitized antibody-virus complexes, for immune complexes, and for total plasma IgG and IgM. In infected mice, anti-LDV IgM was transiently formed during the first 2 weeks post infection (p.i.) but only at a low level. Anti-LDV IgG was produced in a biphasic manner with an initial peak at about 10 days p.i. and a secondary rise reaching a maximum level 30–80 days p.i. which was retained throughout the persistent phase of infection. The concomitant appearance of comparable levels of low molecular weight immune complexes suggests that most anti-LDV IgG was complexed with LDV proteins. Also, as early as 10 days p.i., infectious antibody-LDV complexes developed, which were neutralizable by rabbit anti-mouse IgG, whereas antibodies that neutralize the infectivity of exogenously added LDV appeared only 1–2 months p.i. Throughout infection, most of the anti-LDV IgG was directed to VP-3, the envelope glycoprotein of LDV, which was found to exist in at least 10 distinct forms ranging in molecular weight from 24 to 42 kDa. Anti-LDV IgG levels as high as those observed in infected mice developed in mice immunized with inactivated LDV. Antibodies to glutaraldehyde-inactivated LDV were also mainly directed to VP-3, but exhibited no neutralizing activity. The polyclonal B cell activation associated with a persistent LDV infection and the formation of immune complexes were not observed in mice immunized with inactivated virus.

Thrombocytopenic purpura in narcotics addicts.

Since November 1982 we have seen an association of thrombocytopenic purpura with chronic narcotic addiction in 70 patients with a mean platelet count of 53000 +/- 4000 (SE); 33 had stopped taking intravenous drugs for an average of 21.2 +/- 4.7 months; 13 of 15 had elevated antibody titers for a virus related to the acquired immunodeficiency syndrome. Platelet-bound IgG, IgM, and complement levels were 16.7-, 5.6-, and 3.1-fold greater than control values, respectively, and 2.6-, 1.9-, and 2.4-fold greater than values in 25 patients with classic autoimmune thrombocytopenic purpura studied at the same time. Thirty-three of thirty-six addicts had elevated circulating immune complexes, whereas 8 patients with autoimmune thrombocytopenia had no elevation. Eleven of eighteen addicts had positive serum platelet-reactive IgG titers, compared to 5 of 19 patients with classic autoimmune thrombocytopenia. The platelet-reactive IgG in sera of addicts was composed of 7S IgG antibody as well as high molecular weight (immune complex) IgG. Thus, chronic addicts appear to have a new immunologic platelet disorder associated with the presence of 7S IgG antiplatelet antibody, like patients with classic autoimmune thrombocytopenic purpura, and immune complex associated nonspecific platelet IgG, like male homosexual patients with thrombocytopenia.

Immune complexes and abnormal liver function in haemophilia.

Abnormal 125I-Clq-binding activity was found in the sera of 94% of 55 haemophiliacs. Sera from 66% of these patients inhibited macrophage uptake of labelled aggregated human IgG in a competitive radiobioassay. These results suggest that large molecular weight immune complexes are present in these sera. Analysis of the precipitates obtained directly from the sera by addition of 4% polyethylene glycol showed either a mixture of IgG and IgM or IgM alone. There was poor correlation between the radiobioassay results and the Clq-binding activities in the whole group of 55 patients, suggesting a heterogeneous population of complexes. Nevertheless, a significant correlation was found between Clq-binding activities and the radiobioassay results in seven patients with antibodies to factor VIII, suggesting a homogeneous population of complexes. There was poor correlation between the level of immune complexes and the amount of replacement therapy the patient had received in the previous six months. Abnormal liver function tests were found in 55% of the patients studied but there was poor correlation between these abnormal levels and the Clq binding activities and radiobioassay results. Only two patients had clinical evidence of liver disease.