Formation Of Circulating Immune Complexes Research Articles

THE STUDY OF HUMORAL FACTORS OF ADAPTIVE IMMUNITY IN PATIENTS WITH NON-HODGKIN’S LYMPHOMAS

It is known that humoral factors of immunity play an important function of mediators in the cascade development of the immune response and can determine the effectiveness of the final, effector reactions of cellular immunity in the inactivation and elimination of antigens. Serum concentrations of major immunoglobulins IgG, IgA, IgM in NHL were analyzed. The highest serum content of IgG and IgA was revealed in the groups of patients with virus carrier. It has been established that one of the most important humoral markers of immunity is the circulating immune complexes (CIC). It has been established that one of the most important biological functions of immunoglobulins is antigen binding and the formation of CIC. A high level of CIC in NHL may be due not only to the activation of the immune response, but also to the suppression of the mechanisms of their elimination. Therefore, based on the results obtained, with NHL there is a pronounced imbalance of humoral immunity. CIC of large and small sizes are also increased, however, the highest increase in CIC was observed in groups of patients before chemotherapy with virus-bearing. An increase in the main immunoglobulins indicates the presence of humoral imbalance, and an increase in the CIC indicates the presence of intoxication of the body either due to the decay of the tumor cells themselves and infected cells, which almost always indicates the progression of the pathological process.

Peculiarities of the formation and subsequent removal of the circulating immune complexes from the bloodstream during the process of digestion

Background: Large protein aggregates, known as circulating immune complexes (CICs), are formed in biological fluids as a result of the development of the body's immune response to various provoking factors. The kinetic characteristics of the formation and removal of immune complexes (ICs), their physical parameters, the isotypic composition of immunoglobulins (Igs) and the antigenic component of the CICs may reflect certain aspects of certain pathological and metabolic processes taking place in humans and animals. The aim of this study is to assess the kinetic characteristics of the formation and removal of the CICs that form in blood after eating. We also analyze the changes in the isotypic composition of Igs of ICs that accompany this biological process in rodents and humans. Methods: We identified the CICs, which differed in size and class of Igs, using dynamic light scattering. To remove ICs from the plasma, we used immune-affinity sedimentation. Monoclonal antibodies for the Igs of different isotypes were added to the plasma samples to determine the isotypic composition of the ICs. Results: A large number of ICs were formed in the blood of rats and humans after eating (food CICs). In rats, food ICs are almost immediately filtered in the liver, without circulating in the bloodstream through the body. In humans, the level of food ICs in the blood increases for 3.5 h after ingestion, then within 7–8 h their gradual removal takes place. It was found that in the process of digestion in humans, the isotypic composition of Igs in the CICs changes and becomes more diverse. Conclusions: The molecular–cellular mechanisms of the formation and utilization of food CICs in humans and rodents do not match completely.

Peculiarities of the formation and subsequent removal of the circulating immune complexes from the bloodstream during the process of digestion.

Background: Large protein aggregates, known as circulating immune complexes (CICs), are formed in biological fluids as a result of the development of the body's immune response to various provoking factors. The kinetic characteristics of the formation and removal of immune complexes (ICs), their physical parameters, the isotypic composition of immunoglobulins (Igs) and the antigenic component of the CICs may reflect certain aspects of certain pathological and metabolic processes taking place in humans and animals. The aim of this study is to assess the kinetic characteristics of the formation and removal of the CICs that form in blood after eating. We also analyze the changes in the isotypic composition of Igs of ICs that accompany this biological process in rodents and humans. Methods: We identified the CICs, which differed in size and class of Igs, using dynamic light scattering. To remove ICs from the plasma, we used immune-affinity sedimentation. Monoclonal antibodies for the Igs of different isotypes were added to the plasma samples to determine the isotypic composition of the ICs. Results: A large number of ICs were formed in the blood of rats and humans after eating (food CICs). In rats, food ICs are almost immediately filtered in the liver, without circulating in the bloodstream through the body. In humans, the level of food ICs in the blood increases for 3.5 h after ingestion, then within 7-8 h their gradual removal takes place. It was found that in the process of digestion in humans, the isotypic composition of Igs in the CICs changes and becomes more diverse. Conclusions: The molecular-cellular mechanisms of the formation and utilization of food CICs in humans and rodents do not match completely.

Circulating immune complexes and markers of systemic inflammation in RAO-affected horses.

The aim of this study was to investigate the levels of circulating immune complexes (CICs) and concentration of haptoglobin, fibrinogen and C-reactive protein in the serum of horses with recurrent airway obstruction and healthy controls. The study was conducted on a group of 14 adult Polish Konik horses, kept in uniform environmental and living conditions. Horses were divided into two groups: 7 horses were not affected by any respiratory problem (control group) and 7 horses had a history of recurrent airway obstruction (RAO) (study group). A clinical and laboratory evaluation, endoscopic examination and bronchoalveolar lavage (BAL) were performed in all horses. Levels of circulating immune complexes were significantly (p = 0.0057) increased in heaves-affected horses compared to healthy controls (median [25th-75th percentiles]) (3.96 [3.96-4.43] vs. 7.46 [5.13-11.9]). No significant difference was observed in the levels of the examined acute phase proteins between the groups. Moreover, all results were within the reference range established for horses. The results of this study indicate that heaves in horses is associated with the formation and high level of CICs. Haptoglobin, fibrinogen and C-reactive protein failed as markers of early stage systemic inflammation in the course of RAO.

Carbohydrate-rich high-molecular-mass antigens are strongly recognized during experimental Histoplasma capsulatum infection

During histoplasmosis, Histoplasma capsulatum soluble antigens (CFAg) can be naturally released by yeast cells. Because CFAg can be specifically targeted during infection, in the present study we investigated CFAg release in experimental murine histoplasmosis, and evaluated the host humoral immune response against high-molecular-mass antigens (hMMAg. >150 kDa), the more immunogenic CFAg fraction. Mice were infected with 2.2 x 10⁴ H. capsulatum IMT/HC128 yeast cells. The soluble CFAg, IgG anti-CFAg, IgG anti-hMMAg, and IgG-hMMAg circulating immune complexes (CIC) levels were determined by enzymelinked immunosorbent assay, at days 0, 7, 14, and 28 post-infection. We observed a progressive increase in circulating levels of CFAg, IgG anti-CFAg, IgG anti-hMMAg, and IgG-hMMAg CIC after H. capsulatum infection. The hMMAg showed a high percentage of carbohydrates and at least two main immunogenic components. We verified for the first time that hMMAg from H. capsulatum IMT/HC128 strain induce humoral immune response and lead to CIC formation during experimental histoplasmosis.

Autoantibodies associated with RNA are more enriched than anti-dsDNA antibodies in circulating immune complexes in SLE

Background and objectiveWe have earlier shown that polyethylene glycol (PEG)-precipitated circulating immune complexes (CIC) from systemic lupus erythematosus (SLE) patients induce cytokines in vitro, and that levels of both CIC...

Systemic adverse events following rituximab therapy in patients with Graves’ disease

Rituximab (RTX) therapy has shown promising results in Graves' disease (GD), with or without ophthalmopathy. We examined the occurrence of adverse events in GD patients treated with RTX. Ten patients received RTX and methimazole, while 10 patients received methimazole only. Adverse events were recorded, and the presence of circulating immune complexes (CIC) was measured as IgG, IgM and complement component 3 (C3) depositing on normal monocytes following incubation with patient plasma. Five patients had benign infusion-related adverse events at first infusion. Two patients developed a serum sickness-like reaction 11 days after the first RTX-infusion. One of these patients developed diarrhea, raised orosomucoid levels, lowgrade inflammation in colonoscopic biopsies, and iridocyclitis 1 yr later. At day 14, the most pronounced immunoglobulin/ C3-adherent to the test monocytes, indicative of CIC, was observed in the presence of plasma from these 2 patients (p=0.003 to p=0.01 vs asymptomatic patients). A 3rd patient had recurrent fever and symmetric polyarthritis from day 38, and colonoscopy-verified ulcerative colitis at day 68. This patient had the 3rd highest increase in Ig deposition on monocytes by day 14. The arthralgias persisted in 2 of the patients, despite glucocorticoid rescue therapy. We report articular adverse events in 3 and gastrointestinal symptoms in 2 out of 10 GD patients who received RTX without concurrent immunosupression. The joint symptoms were related to CIC formation.

Gliadin IgG antibodies and circulating immune complexes

Objective. Circulating immune complexes (CICs) in blood are associated with autoimmune-diseases such as systemic lupus erythematosus, immune complex glomerulonephritis, rheumatoid arthritis and vasculitis. However, slightly increased serum concentrations of such CICs are sometimes also found in healthy individuals. The objective of the current study was to assess whether food antigens could play a role in the formation of CICs. Material and methods. A total of 352 (265 F, 87 M), so far, healthy individuals were tested for CICs containing C1q and immunoglobulin G (IgG) as well as for gliadin IgG antibodies using the ELISA technique. Additionally, fructose and lactose malabsorption was assessed using hydrogen breath tests. Results. In our study, 15.3% (54/352) of the patients presented with elevated CIC concentrations (above 50 µg/ml) and 6.5% (23/352) of the study population were positive for gliadin IgG antibodies (above 20 U/ml). CIC concentration levels were significantly higher in the group with elevated gliadin IgG antibodies (CIC median: 49.0 µg/ml) compared with the group with normal levels of gliadin IgG antibodies (CIC median: 30.0 µg/ml; Mann-Whitney U-test, U=1992; p <0.001). As expected, there was no difference in CIC concentrations (Mann-Whitney U-test, U=6106; p=0.783) and gliadin IgG (Mann-Whitney U-test, U=3761; p=0.411) between patients in the fructose or lactose malabsorber groups and the subjects without malabsorption. Conclusions. The results of this study indicate that certain food antigens (e.g. gluten) could play a role in the formation of CICs. An association between CICs and fructose or lactose malabsorption seems to be improbable.

Radioprotection by Cu(II) Chelates of Nicotinyl-L-Aminoacidates

Essential metalloelement chelates presents a promising class of compounds for search and development of novel anti-radiation agents, study of which offers a new approach to overcome the pathological effects of ionizing radiation. In this study Cu(II) chelates of Nicotinyl-L-Tyrosinate and Nicotinyl-L-Tryptophanate Schiff Bases were synthesized and investigated as radioprotectors in animal experiments against injuries caused by ionizing irradiation. Based on the assessment of average life-span indices and 30-day animal survival after radiation exposure at LD50/30 (4.8 Gy) or LD100/30 (8.7 Gy) the radioprotective effects of 0 mg/kg (Vehicle), 10 mg/kg, 20 mg/kg, or 40 mg/kg Cu(II)(Nicotinyl-L-Tyrosinate)2 or Cu(II)(Nicotinyl-L-Tryptophanate)2 as well as their parents Schiff Bases were studied in case of single subcutaneous and oral administration to rats 1, 3, 6 and 24 hours prior to X-ray irradiation. The mixture of 4% propylene glycol and 1.4% polyvinyl alcohol in saline was used as a Vehicle for the administration of compounds to animal organism. According to the results obtained, administration of the metallochelates to the rat organism prior to X-ray irradiation provided strong radioprotective effects expressed upon application of all considered dose, mode and time treatment schedules: in groups of animals treated with Cu(II) chelates there was an remarkable increase in the indices of survival and average life-span in a 30-day period post exposure compared to the control Vehicle treated - irradiated rats. The appropriate parents Schiff Bases Nicotinyl-L-Tyrosinate and Nicotinyl-L-Tryptophanate also exerted the radioprotective action. However, these compounds were active as effective radioprotectors only in case X-ray irradiation at a dose level equal to or less than LD50/30. Data of analyses indicated that Cu(II) chelates as well as their initial amino acid Schiff Bases did not avert the development of disturbances of hematological indices of animals exposed at LD50/30. However, the findings of immunological studies have demonstrated the inhibiting effects of these compounds on circulating immune complexes (CICs) in blood plasma, which are the major mediators of immune response and are considered as indicators of the autoimmune and inflammatory components of radiation-induced up-regulated immune response. Unlike to Nicotinyl-L-Aminoacidate Schiff Bases their corresponding Cu(II) chelates diminished the harmful effects of radiation-induced CICs formation not only in case of irradiation at LD50/30, but also at much higher radiation dose levels. Thus, the single administration of Cu(II)(Nicotinyl-L-Tyrosinate)2 and Cu(II)(Nicotinyl-L-Tryptophanate)2 at non-toxic doses provides effective radiation protection and high level of survival of exposed animals. Research is performed in the frames of ISTC A-1321 Project.

Detection of microfilarial antigen in circulating immune complex from sera of Wuchereria bancrofti-infected individuals.

Polyethylene glycol-precipitated circulating immune complexes (CICs) from the sera of patients with Bancroftian filariasis were examined for parasite antigen content by an ELISA. Of the 227 patients, 214 were asymptomatic microfilariae carriers with microfilaremias ranging between six and 14,000/ml blood, and 13 were symptomatic patients with chronic filariasis without microfilaremia. In addition, the sera of 10 of the patients treated with diethyl carbamazine were also examined. These assays were devised using mouse monoclonal antibody raised against microfilariae of Wuchereria bancrofti. Using an acid-phase covalent-bound plate for the ELISA makes it possible to assay the antigen contents simply. Microfilarial antigens were detected from 92 (43.0%) of microfilaria (mf)-positive individuals. Furthermore, the level of antigen in CIC was correlated with mf counts from night blood Nuclepore filtration results. Antigen was not detected in CIC from patients with a variety of nonfilarial helminth infections. These findings indicate that microfilariae play an important role of CIC formation in lymphatic filariasis.

Identification of Myelin Basic Proteins in Circulating Immune Complexes Associated with Lepromatous Leprosy

Circulating immune complexes (CIC) were first measured in lepromatous patients (LL) by the 125I-C 1q binding assay and the polyethylene glycol (PEG) precipitation test. High levels were found by both methods (95 and 90% of positives, respectively). LL-CIC were investigated for the presence of neural antigens. CIC were precipitated in 3.5% PEG, filtered through protein A-Sepharose affinity chromatography, eluted with glycine-HCI, pH 2.8, and washed with PBS; fractions after CIC dissociation were studied by SDS-PAGE and Western blotting. The LL-CIC PEG precipitates and the glycine-HCI eluates were positive in 76 and 71% respectively against anti-myelin basic proteins (MBP) monoclonal antibody, showing a single band at 15-25 kDa similar to the one obtained incubating MBP with anti-MBP. No reaction was detected with CIC-PBS fractions; strips were incubated with other anti-neural antibodies such as anti-glial fibrillary acidic proteins, anti-S-100, and anti-neuarofilaments, without any reactivity. Our results demonstrate that LL-CIC contain MBP as an antigen; its significance could be related to the pathogenesis of leprosy since the liberation of MBP after Mycobacterium leprae nerve damage may elicit anti-MBP autoantibodies to myelin breakdown, which reacts with peripheral nerve MBP inducing CIC formation. This mechanism may be important in demyelination and destruction of nerve in leprosy.

Autoantibodies and circulating immune complexes in sera from patients with hepatitis B virus-related chronic liver disease

Sera from 56 patients with hepatitis B virus-related chronic liver disease (CLD) and 30 normal individuals as controls were examined by enzyme immunoassay (EIA) for the presence of autoantibodies directed against actin, tubulin, myosin, double-stranded (ds) DNA, polymerized human albumin, thyroglobulin, and trinitrophenyl coupled to bovine serum albumin (TNP-BSA). Patients with CLD had consistently elevated levels of IgG, IgA, and IgM antibodies directed against all the panel antigens. The percentage of patients with autoantibodies of the IgA class was particularly high: respectively, 88 and 78% of the patients had strikingly high levels of anti-actin and anti-TNP-BSA IgA autoantibodies. High amounts of IgA and IgG antibodies to polymerized albumin as well as IgG to thyroglobulin were also detected. Circulating immune complexes (CIC) were isolated from patients' sera and their autoantibody activities were tested on the same antigen panel. The autoantibodies thus detected were of the same class and possessed the same activities, although at higher values than those present in the homologous sera. These results indicate that, regardless of their origin, autoantibodies are present in high amounts in the sera of these patients. Moreover, autoantibodies participating in the formation of CIC might play a pathological role.

Participation of antigens related to the psoriasis associated antigen, pso p27, in immune complex formation in patients with ankylosing spondylitis.

Analysis of five serum samples and three synovial fluids from patients with ankylosing spondylitis (AS) and five serum samples from healthy blood donors for the presence of antibodies cross reacting with the Fc part of rabbit IgG (rheumatoid factors (RFs] using an isotype specific, enzyme linked immunosorbent assay (ELISA) showed only insignificant amounts of free RFs, while IgG RFs were observed in alkaline dissociated circulating immune complexes (CICs). Only insignificant amounts of free antibodies reacting with the psoriasis associated antigen pso p27 could be detected in the samples, while extensive amounts of IgG antibodies and moderate amounts of IgM antibodies reacting with pso p27 were detected in alkaline dissociated CICs from the patients. Pso p27 has been reported to share a common determinant with the Fc part of human IgG. Removal of the RF activity from the CICs of patients with AS by absorption with IgG resulted in a decrease of the anti-pso p27 activity. Monoclonal anti-pso p27 antibodies in a sandwich ELISA were used to detect antigens cross reacting with pso p27. A positive reaction was observed in all serum CICs and in one of the synovial fluid CICs. The data indicate that antigens related to pso p27 participate in CIC formation in AS and may also be responsible for the elicitation of rheumatoid factors in patients with AS.

Detection of bovine serum albumin in the circulating IgA immune complexes of patients with IgA nephropathy

Alimentary antigenic challenge has been postulated to have a role in the genesis of IgA circulating immune complexes (CIC), resulting in mesangial IgA disease. In this study, we examined the relationship between bovine serum albumin (BSA) and IgA CIC in patients with IgA nephropathy. Of the 47 patients studied, elevated IgA CIC levels were found in 32% by the F(ab′) 2 anti-C 3 and Raji cell enzyme immunoassays (EIA). Elevated IgA anti-BSA antibody levels were found in 9 patients, and there was a positive correlation between these levels and IgA CIC as measured in the Raji cell EIA ( R = 0.60, P < 0.001). In 4 patients with elevation of both IgA CIC and IgA anti-BSA antibody levels, solubilization experiments were done to demonstrate the presence of BSA antigen in the IgA CIC. Using the Raji cell EIA, the IgA CIC levels decreased significantly after preincubating the sera with serial concentrations of excess BSA. No corresponding effect was seen with human serum albumin used as control. Hence, BSA may be the antigenic stimulus in the formation of IgA CIC in selected patients with IgA nephropathy. The pathogenic capacity of these IgA-BSA CIC remains to be determined.

Antigen-antibody complexes in hemophilia.

Antigen‐antibody complexes have been found by several groups of investigators in patients with hemophilia. The amount of circulating immune complexes (CIC) has varied with the assay method, the largest amount has been found with the Staph binding assay and the smallest with the Raji cell method. We have correlated the level of immune complexes detected by both the Raji cell and the Staph binding assay with clinical disease status: joint disease, positive inhibitor status, and hematuria but not hepatitis. The complexes are formed with different antigens, Factor VIII, the Factor VIII inhibitor, and antibody to the hepatitis B surface antigen for example have been detected in isolated CIC. CIC have not been detected in synovium or vasculature and rarely in the synovial fluid. The effect of the infusion of concentrate on the levels of the immune complexes is variable among patients but consistent within individuals over time. Studies by our group show a rise in CIC in 3 patients and a fall in six following concentrate infusion with return to baseline in 24 hours. In vitro studies of the monomuclear phagocyte system Fc receptor function showed enhanced function in those six patients whose levels decreased post infusion while the three with rise in CIC showed a prolonged clearance and decreased mononuclear phagocyte system function. This effect was not correlated with the amount of infusion or the yearly use of the concentrate. Changes in CIC were not due to infusion of preformed complexes in the Factor VIII concentrate nor to de novo formation of CIC since in vitro incubation of pre‐infusion serum with concentrate revealed no change in CIC levels. However, change in reticulo endothelial Fc receptor function correlated with the changes in CIC suggesting a modulation of monocyte function by Factor VIII concentrate infusion. In summary, circulating immune complexes are present in transfused hemophiliacs which may be associated with secondary disease, and regulation of the immune function in these patients.

Detection and partial characterization of circulating immune complexes in hydatid disease.

Thirty sera from eight patients with disseminated or localized hydatid disease have been examined for the presence of circulating immune complexes (CICs) by the conglutinin-binding assay and for immunoglobulin levels. The highest levels of CICs were of the immunoglobulin A (IgA) class, with lower values of IgG-CIC and IgM-CIC; these results did not correlate, except for IgG, with the free immunoglobulin levels. Efforts to identify parasitic antigen(s) involved in the CIC formation with different methods have been unsuccessful. In the follow-up of each patient, CIC appeared to be better correlated to clinical conditions than to hemagglutination titers. We have concluded that the presence of CIC in hydatid disease is probably an expression of B-cell polyclonal activation and that these complexes are valuable in the clinical monitoring of the disease.

Dietary Gluten Challenge Does Not Influence the Levels of Circulating Immune Complexes in Patients with Dermatitis Herpetiformis

To examine the relationship between the gluten-sensitive enteropathy (GSE) and IgA circulating immune complexes (CIC) in dermatitis herpetiformis (DH) a series of dietary gluten-challenge studies were performed in patients with DH and patients with ordinary GSE. Serial serum samples were monitored for IgA-, IgG-, and IgM-containing CIC levels. In the first study, 9 DH patients and 5 controls were fed 20 g of gluten flour as a breakfast meal on 1 of 2 consecutive study days. DH patients did not develop or increase their levels of CIC after gluten-challenge or gluten-free meals. There was no significant difference between the DH patients and the control group in regard to development of CIC. To evaluate the effect of dietary gluten in another form, 8 DH patients were given meals containing 100 g of boiled Canadian cracked wheat. Two patients with ordinary GSE were also challenged with cracked wheat. Again there was no elevation or induction of CIC above baseline determinations by gluten-challenge meals. These studies suggest that dietary gluten does not induce the formation of CIC in patients with DH.

ANTI-RETINAL AUTOIMMUNITY AND CIRCULATING IMMUNE COMPLEXES IN PATIENTS WITH RETINAL VASCULITIS

Sera from 44 patients with isolated retinal vasculitis (RV), 38 patients with retinal vasculitis accompanying systemic inflammatory diseases (RV+SID), and 33 patients with a similar range of systemic inflammatory diseases without eye involvement (SID alone) were assayed for circulating immune complexes (CIC) and for anti-retinal autoantibodies. CIC were present in 41% of patients with isolated RV and 55% of patients with RV + SID, whilst anti-retinal antibodies were present in about 70% of all patients with RV. 42% of those with SID alone had CIC and 30% of those with SID alone had retinal autoantibodies. Titres of anti-retinal antibodies were higher in patients with RV than in those with SID alone. In isolated RV there was an inverse relation between pronounced retinal autoimmunity and the occurrence of CIC—i.e., the more severe autoimmune retinal disease occurred in CIC-negative patients. Most patients with RV + SID tended to have mild or moderate retinal disease accompanied by both retinal autoantibodies and CIC, but severe retinal disease occurred in CIC-positive patients who did not have circulating anti-retinal antibodies. Patients with SID alone had high titres of retinal antibodies only when they were CIC-positive. It is suggested that the formation of CIC, possibly of an idiotype/anti-idiotype nature, may be a compensatory mechanism accompanying anti-retinal autoimmunity and that an imbalance between autoimmunity and immune complex formation may be an important predisposing factor in the development of retinal inflammatory disease.

Bullous pemphigoid: a correlative study of autoantibodies, circulating immune complexes and dermo-epidermal deposits.

Twenty bullous pemphigoid (BP) patients were studied to establish any correlation between free anti-basement membrane zone (BMZ) antibodies, circulating immune complexes (CIC) and dermo-epidermal junction deposits. CIC levels were evaluated by 2% polyethylene glycol (PEG) precipitation. The twenty patients were found to have IgG and/or C3 deposited in the BMZ. Eight of the twelve patients who had no free anti-BMZ antibodies displayed a positive in vivo C4 and/or CIq staining and high levels of CIC. Moreover, CIC were detected in only one patient with positive circulating free anti-BMZ antibodies. The presence of free anti-BMZ antibodies was generally found to correlate with the absence of cutaneous deposits of CIq and /or C4 and with negative CIC; on the other hand, the absence of free anti-BMZ antibodies was generally found to correlate with high levels of CIC and with deposits of C3 and CIq and/or C4. The absence of circulating free anti-BMZ antibodies in BP patients, could be explained by the formation of CIC. It is possible that BMZ antigens released from damaged tissue could combine with free antibodies and form complexes in the blood. The release could involve locally formed immune complexes. Elevated CIC levels were generally found to correlate with the presence of active disease.