Concentration Of Immune Complexes Research Articles

#2314 Circulating immune complexes concentration and complement activation in sensitized kidney transplant recipients

Abstract Background and Aims Chronic antibody mediated rejection of the kidney transplant is the most common cause of graft loss in the late posttransplant period. Crucial in this process seems to be the role of classical pathway complement activation due to the formation of immune complexes. The formation of immune complexes is a benign process of a normally functioning immune system. In kidney transplant recipients (KTRs), the production of donor specific IgG/IgM antibodies (DSAs) against donor antigens (MHC molecules, ABO blood group antigens) leads to the attachment of immune complexes to cell membranes and subsequent tissue damage through complement activation. The attachment of C1q complement fragment to the immune complexes holds an important part in this process. This study aims to determine the levels of immune complexes as well as classical pathway complement activation in sensitized kidney transplant recipients. Method In this cross-sectional study 3 groups of KTRs were compared as to the levels of immunocomplexes concentration and classical pathway complement activation. Fourteen KTRs with preformed or de novo donor specific antibodies were compared to 28 kidney transplant recipients with no DSAs and to 22 subjects with no history of chronic kidney disease (controls). We used MicroVue Complement CIC-C1q EIA that is designed to detect immunocomplexes that bind to C1q in human serum or plasma. In order to estimate total classical pathway complement activation in serum, we used MicroVue Complement CH50 Eq EIA. Results Kidney function (eGFR) of KTRs with and without DSAs was similar (53.1 ± vs. 60.8 ± ml/min/1.73 m2, p = 0.54) but lower in comparison to controls (106.4 ml/min/1.73 m2, p < 0.001). We found a significantly higher concentration of C1q-immunocomplexes in the serum of KTRs with donor specific antibodies, in comparison to controls, as well as to the group of kidney transplant recipients with no DSAs (6.78 ± 2.7 vs. 4.83 ± 1.9 vs. 5.04 ± 1.2 μgEq/ml respectively, p = 0.0078). Furthermore, there was no significant difference of C1q-immunocomplexes between kidney transplant recipients with no DSAs and controls. Finally, a significant reduction of CH50 levels was observed in KTRs with DSAs as compared to controls and KTRs with no DSAs (39.2 ± 14.9 vs. 70.6 ± 34.3 vs. 67.9 ± 39.9 U/ml respectively, p = 0.003), which suggests higher levels of classical pathway complement activation. CH50 levels in KTRs with DSAs, who were further subcategorized in those with preformed and those with de novo DSAs were similar (39.68 ± 15.6 vs 39.08 ± 16.04 U/ml). Conclusion Kidney transplant recipients with donor specific antibodies have increased serum levels of immune complexes and increased classical pathway complement activation.

Evaluation of autoantibody profile in healthy subjects after mRNA vaccination against COVID-19.

SARS-CoV-2 severe acute respiratory syndrome has rapidly spread worldwide since 2019. All scientific and technological forces have concentrated towards the formulation of vaccines to contain the disease. In less than one year (December 2020) a first messenger RNA vaccine (Comirnaty, BioNTech/Pfizer) was authorized. However, the research community has wondered about possible side effects on the immune system, given the vaccines administration in phase 4. This study aims to evaluate the mRNA vaccine impact on the development of possible positive autoantibody profile in healthcare workers without any previous underlying pathology, after first, second and booster dose of Pfizer vaccine, by determining: circulating immune complexes concentrations (CIC); anti-myeloperoxidase (MPO) and anti-proteinase 3 (PR3) autoantibodies, the presence of antinuclear antibodies (ANA) and subsequent second level tests (extractable nuclear antigen (ENA) screen, double-strand DNA, extractable nuclear antigen (ANA) profile). The subjects were divided according to anti-SARS-CoV-2 IgG RBD antibodies increasing concentrations in: Group I<10 BAU/ml (N=114); Group II>1000 BAU/ml (N=112); Group III>2500 BAU/ml (N=78). Our data show no autoreactive response changes over time in healthy subjects after vaccination. In fact, evaluation of ANA, CIC, anti-MPO, anti-PR3 and the detection of specific autoantigens, did not display significant variations. The results suggest the exclusion of a correlation between the administration of the vaccine and the possible onset of autoimmune disorders. Nevertheless, further investigations will be needed to test for any long-term side effects on an ever-growing population.

Влияние иммобилизованной формы пробиотического препарата «Экофлор» на показатели неспецифического гуморального иммунитета осетровых рыб

Currently, due to the intensive development of aquaculture, interest in the study of the immune system and im-mune response in fish has increased. When growing fish in aquaculture, the conditions of their maintenance should contribute to maintaining the optimal activity of innate immunity. It has been established that the immunity of fish is more dependent on external influences, and environmental conditions are active regulators of fish immunoreactivity. Currently, various probiotic preparations with the ability to correct microbiological and immune parameters due to the action of symbiotic microorgasms have been widely used in aquaculture to increase and regulate the immune homeostasis of fish. Information is presented on the conduct of experimental studies to assess the effect of the probiotic drug “Ecoflor”, immobilized on enterosorbent, on the level of bactericidal activity of blood serum, the number of immunodeficient individuals, the content of nonspecific immune complexes in blood serum and immunocompetent organs, as well as on the level of C-reactive protein of sterlet yearlings (Acipenser ruthenus Linnaeus, 1758). It is noted that the use of a probiotic drug in an experi-mental dosage of 6 g/kg of compound feed has an effect on the indicators of nonspecific humoral immunity. Thus, sterlet grown on compound feeds with the specified rate of drug administration is characterized by an increased level of bactericidal activity of blood serum – 98% and the absence of immunodeficiency – 0%. In addition, this dosage helps to reduce the concentration of nonspecific immune complexes in the blood serum to 2.25 ± 1.65 and immunocompetent organs of sterlet yearlings: in the liver and spleen to 0.06 ± 0.02 and 0.30 ± 0.30, respectively. The results of the analysis of the level of C-reactive protein indicate that this drug does not cause inflammatory reactions in the body of fish.

Mycobacterium tuberculosis Cell Wall Antigens Induce the Formation of Immune Complexes and the Development of Vasculitis in an Experimental Murine Model.

Tuberculosis (TB) of the central nervous system (CNS) presents high mortality due to brain damage and inflammation events. The formation and deposition of immune complexes (ICs) in the brain microvasculature during Mycobacterium tuberculosis (Mtb) infection are crucial for its pathobiology. The relevance of ICs to Mtb antigens in the pathogenesis of CNS-TB has been poorly explored. Here, we aimed to establish a murine experimental model of ICs-mediated brain vasculitis induced by cell wall antigens of Mtb. We administered a cell wall extract of the prototype pathogenic Mtb strain H37Rv to male BALB/c mice by subcutaneous and intravenous routes. Serum concentration and deposition of ICs onto blood vessels were determined by polyethylene glycol precipitation, ELISA, and immunofluorescence. Histopathological changes in the brain, lung, spleen, liver, and kidney were evaluated by hematoxylin and eosin staining. Our results evidenced that vasculitis developed in the studied tissues. High serum levels of ICs and vascular deposition were evident in the brain, lung, and kidneys early after the last cell wall antigen administration. Cell wall Mtb antigens induce strong type III hypersensitivity reactions and the development of systemic vasculitis with brain vascular changes and meningitis, supporting a role for ICs in the pathogenesis of TB.

Immune complexome analysis of a rich variety of serum immune complexes identifies disease-characteristic immune complex antigens in systemic sclerosis

Systemic sclerosis (SSc) is an autoimmune disease characterized by vascular endothelial dysfunction and skin fibrosis. Recently, the presence and pathogenic role of immune complexes (ICs) of SSc patients were reported. However, the identities of antigens in these ICs are unknown. Therefore, we examined ICs in the serum of SSc patients to elucidate SSc pathogenesis. In this study, IC concentrations in serum samples from SSc and systemic lupus erythematosus (SLE) patients were measured by C1q enzyme-linked immunosorbent assays; immune complex analysis was used for comprehensive identification and comparison of antigens incorporated into ICs (IC-antigens). The expression patterns of SSc-specific IC-antigens in skin sections were investigated by immunohistochemistry. Compared with SLE patients who developed disease because of IC deposition, SSc patients had a greater number of IC-antigens and a smaller difference in IC concentrations, suggesting that SSc pathogenesis is affected by the proteins present in ICs. In contrast, the IC concentration and number of IC-antigens did not significantly differ according to the clinical phenotype of SSc. We identified 478 IC-antigens in SSc patients, including multiple RNAP II-associated proteins that were targeted by antibodies previously associated with SSc pathogenesis. The most frequently detected RNAP II-associated protein, RNA polymerase II transcription subunit 30 (MED30), was strongly expressed at lesion sites and reportedly regulates endothelial differentiation. Therefore, increased expression of MED30 in lesions may have an antigenic effect, and MED30 function may be impaired or inhibited by IC formation. RNAP II-associated proteins may SSc pathogenesis through mechanisms such as the MED30 pathway.

The role of the palatine tonsils in the regulation of the state of humoral local immunity in patients with chronic inflammatory diseases of the nose and perinasal sinuses when infected with viruses of the respiratory group

Topicality: Acute respiratory diseases of the upper respiratory tract, which in the vast majority of cases are caused by viruses, especially with frequent recurrences of diseases, reduce local and system immunity, which often leads to the accumulation of bacterial infection due to the activation of endogenous microflora or exogenous infection and causes the development of viral-bacterial diseases. Recurrent respiratory diseases contribute to the formation of chronic pathology of the oro- and nasopharynx, which in turn supports the violation of the body's protective mechanisms. Previous studies have shown that the tonsils of the lymphadenoid pharyngeal ring perform the function of immunological protection, and the absence of palatine tonsils (PT) negatively affects the state of both local and system immunity. Aim: to determine the role of the palatine tonsils in the realization of local humoral reactions in patients with chronic inflammatory diseases of nose and paranasal sinuses when infected with viruses of the respiratory group. Materials and methods: Studies were conducted in 134 patients with chronic inflammatory diseases (CID) of nose and paranasal sinuses (PNS) in remission, among whom 93 had a diagnosis of chronic rhinosinusitis, 27 – chronic catarrhal rhinitis, 14 – allergic rhinitis with chronic bronchitis. The control group consisted of 47 practically healthy persons without pathology from the ENT organs. The examined persons were divided into those who had PT and those from whom they were surgically removed no earlier than five years ago. All examinees were divided into those who had anamnestic antibodies in their blood to one or more of the respiratory viruses, such as: adenovirus, respiratory syncytial virus and influenza A virus in a clinically significant titer, and those in whom antibodies were not detected. The local humoral immunity state of patients was assessed by the content of immune complexes (IC) and concentrations of secretory immunoglobulin A (sIgA), interferons alpha- (α-IFN) and gamma- (γ-IFN) in the oropharyngeal secretion (OS), which were determined by the immunoenzymatic method using sets of reagents for enzyme-linked immunosorbent assays ("Cytokin", RF) and the Stat Fax-2100 analyzer (USA). All examined patients had a healthy state of the oral cavity. Results: The presence or absence of palatine tonsils changes the levels of number of humoral factors of local immunity in oropharyngeal secretion in practically healthy donors and in patients with chronic inflammatory diseases of the nose and PNS. The production of sIgA and α-IFN in OS depends on PM, decreasing in practically healthy persons of the control group who underwent tonsillectomy. Chronic inflammatory diseases of the nose and PNS caused a decrease in the concentration of IC and α-IFN in the OS of patients of both groups- with or without PT. In patients with normal PMs who got sick with viral infections, in contrast to those with removed PMs, α-IFN level in OS increased and practically did not differ from those in healthy donors. The greatest suppression of α-IFN production was determined in patients with CID of the nose and PNS with removed PM and with the absence of clinically significant titers of antiviral antibodies. The level of γ-IFN production did not reliably change in the OS of patients with CID of nose and PNS in the remission phase both with normal PT and after tonsillectomy, but depended to a greater extent on the transferred infectious disease caused by viruses of the respiratory group. Conclusion: PTs play a significant role in the formation of a number of reactions of local immunity in the naso- and oropharynx, both in practically healthy individuals and in patients with chronic inflammatory disease of nose and paranasal sinuses. Therefore, the determination of local immunity indices in patients with chronic inflammatory diseases of the upper respiratory tract with different status of the palatine tonsils, who are in contact or had contact with respiratory viruses, provides an opportunity to identify the most vulnerable links in the system of their anti-infective protection, which should first of all be paid attention to during examination and treatment of patients with chronic inflammatory diseases of the upper respiratory tract, especially in cases of frequent recurrence of acute respiratory diseases.

Recurrence of IgA nephropathy after kidney transplantation: experience from the Swiss transplant cohort study

BackgroundRecurrence of IgA nephropathy (IgAN) after kidney transplantation occurs in about 30% of patients. The relevance of recurrence for the long-term graft survival is expected to increase, since graft survival continues to improve.MethodsIn a nested study within the Swiss Transplant Cohort Study the incidence of IgAN recurrence, predictive factors, graft function and graft and patient survival were evaluated. Serum concentration of total IgA, total IgG, Gd-IgA1 and IgA-IgG immune complex were measured using ELISA-based immunologic assays.ResultsBetween May 2008 and December 2016, 28 women and 133 men received their kidney allograft for end-stage kidney disease due to IgAN in Switzerland. Over a median follow-up time of 7 years after transplantation, 43 out of 161 patients (26.7%) developed an IgAN recurrence, of which six (13.9%) had an allograft failure afterwards and further four patients (9.3%) died. During the same follow-up period, 6 out of 118 patients (5%) each experienced allograft failure or died without prior IgAN recurrence. After 11 years the risk for IgAN recurrence was 27.7% (95%-CI: 20.6–35.3%). Renal function was similar in patients with and without recurrence up to 7 years after transplantation, but worsened thereafter in patients with recurrence (eGFR median (interquartile range) at 8 years: 49 ml/min/1.73m2 (29–68) vs. 60 ml/min/1.73m2 (38–78)). Serum concentration of total IgA, total IgG, Gd-IgA1 and IgA-IgG immune complex within the first year posttransplant showed no significant effect on the recurrence of IgAN. Younger recipients and women had a higher risk of recurrence, but the latter only in the short term.ConclusionsOur study showed a recurrence risk of 28% at 11 years after transplantation, which is consistent with previous literature. However, the predictive value of known biomarkers, such as serum Gd-IgA1 and IgA-IgG IC, for IgAN recurrence could not be confirmed.

The influence of parenteral influenza vaccination on local immunity indices and microbiota of oropharyngeal secretion in patients with chronic inflammatory diseases of the upper respiratory tract

When studying the effect of vaccines against influenza, attention is mainly paid to obtaining high titers of protective antibodies in the blood and reducing the incidence of respiratory infections among vaccinated people. At the same time, the changes occurring in patient,s body from the factors of local specific and innate immunity remain insufficiently studied. The aim of the study was to determine the effect of parenteral influenza vaccination on the state of local immunity, cytology and microbiota of oropharyngeal secretion (OS) in patients with chronic inflammatory diseases of the upper respiratory tract. The study of immunological and microbiological parameters was performed in 32 patients with chronic inflammatory diseases of the upper respiratory tract, including 11 diagnosed with chronic rhinosinusitis, 9 – with chronic tonsillitis, 12 – with chronic pharyngitis, 3 and 12 weeks after vaccination with trivalent inactivated influenza-vaccine (PASTEUR, SA, France), which was administered intramuscularly. Single vaccination against influenza A and B has been shown to normalize reduced local humoral immunity indices, in particular sIgA and immune complexes concentrations, increase lymphocyte output to oropharyngeal secretions and cause a significant decrease in the representation of OS transient microflora without affecting the overall level of bacterial contamination. In both periods after the vaccination the reduced content of interferon-α in the OS of patients with chronic inflammatory diseases of the upper respiratory tract did not change. The obtained data allow to recommend vaccination against influenza virus in the period up to 3 months before the the beginning of mass infections as an effective means of stimulating the protective reactions of local immunity of oropha­rynx and nasopharynx mucous membranes in patients with chronic inflammatory diseases of the upper respiratory tract.

Nonspecific protective factors in patients with early latent syphilis (report 1)

The objective of this work is to study nonspecific protective factors in patients with latent early syphilis. The results of the study are based on the data of a comprehensive examination before treatment of 142 patients with early latent syphilis and 20 patients of the comparison group. Methods of investigation: serological methods for the diagnosis of syphilis complex of serological reactions, enzyme-linked immunosorbent assay (IgM, IgG), Indirect immunofluorescence reaction (RIF) RIF-200, RIF-abs; determination of cytokines IL-2, IL-6, IL-10, TNFα and INFγ in blood serum by enzyme-linked immunosorbent assay; determination of the phenotype of lymphocytes (CD-receptors), the concentration of circulating immune complexes, phagocytic number, phagocytic index, Nitroblue tetrazolium test. In the blood serum of patients the concentration of IL-2, IL-6, IL-10 and TNFα, INFγ was significantly increased. The concentration of IL-10 and IL-6 was also significantly increased in patients with more than 1 year of infection. The most significant disorders of the parameters of the metabolic activity of neutrophils were found in patients with an infection of more than 1 year; an imbalance in the circulating immune complexes concentration was also established. Under the increasing duration of the infection, the concentration of CD16+ in the blood serum progressively decreases. The content of pro-inflammatory and anti-inflammatory cytokines in the blood serum of patients with early latent syphilis was significantly increased and depended on the timing of infection. This relationship was most pronounced for IL-6 and IL-10. In patients with early latent syphilis there is also an imbalance between CD25+ and late activation factor HLA-DR against the background of a progressive decrease in the number of CD16+ lymphocytes. A direct relationship was established between the time of infection and the number of NK-cells. The revealed disorders may be the consequence of significant disorders on the part of neutrophils and may be one of the factors of the latent course of syphilitic infection.

Immunological status in pregnant goats

In order to form the animal immunity and develop individual immunological features certain environmental conditions and diet are required. These immunological defense specificities are also defined by animal species, gender, age and other personal traits. Such physiological process as pregnancy has a great impact upon the immune system so the immunity assessment and the patterns identification of changes during this period are an important issue of the goat husbandry. The aim of this research was to study immunological features in Saanen goats depending on month of pregnancy. The research was conducted in North-Western region of Russian Federation, in Saanen goats. The experimental group included 30 pregnant Saanen goats, 1-4 years of age, selected using matched pairs method; control group included 30 non-pregnant Saanen goats, same age. The blood samples were taken 5 times during pregnancy, each month once. Results presented as mean±standard error of the mean. Student's t-test was used after proving normal distribution. Level of significance is labeled as p<0.05. The results are displayed in table 1. After analyzing the received data the significant lysozyme activity decreasing during the whole period of pregnancy was detected – the maximum values of this indicator are observed in the 1st month of pregnancy, the minimum in the 5th month of pregnancy. There was a tendency to serum bactericidal activity decrease for the entire period of pregnancy, the maximum values were observed in the 2nd month of pregnancy, the minimum – in the 5th month. The tendencies to phagocytosis indicators reduction were observed as well. It is noticeable that the maximum values for the phagocytosis indicators were noted during the 2nd month of pregnancy, the minimum – during the 5th month of pregnancy. And for the entire period of pregnancy these values were decreased. Concerning the different immunoglobulin classes blood levels the nonlinear dynamics were observed. So, immunoglobulin A during the 2nd month of pregnancy reached minimal values, and during the following months immunoglobulin A concentration increases, reaching a maximum by the 5th month. The differences during this period are significant when compared with the control group. The immunoglobulin M concentration didn't have significant differences during the entire pregnancy between the both groups. The immunoglobulin G concentration was minimal during the 1st month of pregnancy, and in the 2nd and 3rd months, an increase in this indicator in pregnant animals when compare with control group animals was observed. By the 4th month of pregnancy, the immunoglobulin G concentration of was at the maximum level, and in the 5th month of pregnancy a tendency to decrease the concentration was observed. A linear increase in the blood circulating immune complexes concentration was observed during the entire period of pregnancy. The differences were significant. Thus, during the entire pregnancy, the dynamics of humoral immunity markers increasing were observed. Consequently this data is important to be taken into consideration while keeping pregnant goats.

Various surgical techniques influence on the local immunity state in the postoperative period in children's tonsils hypertrophy

Topicality: There are many conflicting opinions in the literature regarding the best surgical technique for the treatment of OSAS related to palatine hypertrophy – partial (tonsillotomy) or complete (tonsillectomy) removal of lymphoid tissue of the tonsils. Also, there are many opinions about the use of a variety of traditional and modern surgical instruments to reduce the volume of the tonsils, including using a laser, a radiofrequency knife, an ultrasonic knife, a shaver (microdebrider) and a coblator. Aim of the study: The aim of the study was to compare the surgical methods effectiveness of tonsils hypertrophy treatment which the based on the restoration of the mucous membrane's local immunity function. Materials and methods: Children who undergoing surgical treatment for palatine tonsils hypertrophy 2-3 degrees according to the Brodsky classification and the presence of OSAS took part in the study. The patients were randomized and divided into four comparison groups using different surgical techniques. The first group, which used the surgical technique of classical tonsillotomy (TT) included 28 patients, the second group, which used the surgical technique of intracapsular shavertonsillotomy(STT) included 18 patients, the third group, which used surgical technique of intracapsular coblationtonsillotomy (CTT), and groups which using surgical technique of tonsilloplasty included 12 patients (TP). Determined the state of local immunity before surgery and 1 month after surgery by concentration levels of sIgA, α-IFN, IL-1β, TGF-1β and general-purpose immune complexes. Results: Recovery of the concentration of the major marker of local immunity sIgA occurs significantly (P&lt;0.05) faster when using the surgical technique of CTT and STT. Recovery of the concentration of the major marker of antiviral immunity α-IFN occurs significantly (P &lt;0.05) faster when using the surgical technique of CTT, STT and TP. Recovery of the concentration of the anti-inflammatory cytokine IL-1β occurs significantly (P&lt;0.05) faster with the use of the surgical technique of STT and TP. Indicators of the regeneration factor TGF-1β are significantly the highest when using the surgical techniques of STT, TT and TP. The concentration of immune complexes in the oropharyngeal secretions of patients with tonsil hypertrophy at 1 month after surgery significantly decreased with the use of surgical techniques TT and TP. Significant (P &lt;0.05) increase in IgG level was de- termined in all comparison groups except the group where tonsilloplasty was used. Conclusions: The safety surgical technique of palatine tonsils volume reduction – TP and STT promotes significantly (P&lt;0,05) faster restoration of concentration of the main mucous membrane's local immunity markers.

LOCAL IMMUNITY STATUS OF OROPHARYNX IN PATIENTS WITH SCLEROMA

Scleroma is a rare chronic granulomatous disease of the upper respiratory tract caused by Klebsiella pneumoniae subsp. rhinoscleromatis. To date its pathogenesis is as yet little understood. At the same time, scleroma is associated with a number of immune system disturbances. The aim: To study local immunity status of oropharynx in patients with scleroma, and to compare its parameters in various clinical forms of the disease. 20 apparently healthy subjects and 92 patients with scleroma (33 males, 59 females) underwent clinical immunologic evaluation. There were 31 patients with dominating infiltrative form of scleroma, 30 – with dominating atrophic form, 31 – with dominating scarring form. Concentration of secretory and monomeric immunoglobulin A, immunoglobulin G, α-interferon, interleukin 1β in oropharyngeal secretion was determined by enzyme immunoassay. Patients with scleroma were found to have altered local immunity of oropharyngeal secretion. There was a strong tendency for decreased concentration of secretory immunoglobulin A - 1.3-2.0 times, and decreased immunoglobulin G level – 1.5-2.3 times (р < 0.05) as compared to the values in healthy subjects. Specific features of local immunity in oropharyngeal secretion in various forms of scleromatous inflammatory process in upper respiratory tract were found: the most significant decrease of α-interferon concentration in atrophic and scarring forms of the disease, and the largest increase of anti-inflammatory interleukin 1β and immune complex concentration in infiltrative form of scleroma. The study revealed deficiency of local immunity factors in oropharynx, being indicative of immunopathogenetic role of diagnosed disturbances in development and persistence of chronic inflammation in scleroma, and emphasizing the necessity of immunocorrection in complex therapy of the disease.

Role of p300 in the pathogenesis of Henoch-Schonlein purpura nephritis and as a new target of glucocorticoid therapy in mice

Background:Henoch-Schonlein purpura nephritis (HSPN) is a very common secondary kidney disease of childhood. Its pathogenesis and the treatment mechanism of glucocorticoid have not been fully elucidated. The aim of this study was to determine the relationship between p300 and the pathogenesis, glucocorticoid therapy in mice with HSPN, respectively.Methods:Forty-eight C57BL/6N male mice, weighing 18 to 20 g, were selected (3–4 weeks old, n = 8 per group). The mice in the normal control group (Group I) were given normal solvent and the HSPN model group (Group II) were given sensitizing drugs. The mice in Group III were injected intraperitoneally with dexamethasone after being given sensitizing drugs. Meanwhile, mice in Groups IV, V and VI with conditional knockout of p300 were also given normal solvent, sensitizing drugs and dexamethasone.The levels of serum IgA, creatinine, and circulating immune complex (CIC) concentrations, 24 h urinary protein and urinary erythrocyte in C57 wild mice, and p300 conditional knockout mice in each group were measured. The expression of p300 in renal tissues and the expression of glucocorticoid receptor (GR) α and β, transforming growth factor (TGF)-β1, and activator protein (AP)-1 after dexamethasone treatment were determined by real-time polymerase chain reaction and Western blotting.Results:Compared with the normal solvent control group (Group I), the expression of p300 mRNA in the model group (Group II) was significantly up-regulated. Western blotting further confirmed the result. Urinary erythrocyte count, 24 h urinary protein quantification, serum IgA, CIC, and renal pathologic score in Group V were distinctly decreased compared with non-knockout mice in Group II (9.7 ± 3.8 per high-power field [/HP] vs. 18.7 ± 6.2/HP, t = 1.828, P = 0.043; 0.18 ± 0.06 g/24 h vs. 0.36 ± 0.08 g/24 h, t = 1.837, P = 0.042; 18.78 ± 0.85 mg/mL vs. 38.46 ± 0.46 mg/mL, t = 1.925, P = 0.038; 0.80 ± 0.27 μg/mL vs. 1.64 ± 0.47 μg/mL, t = 1.892, P = 0.041; 7.0 ± 0.5 vs. 18.0 ± 0.5, t = 1.908, P = 0.039). Compared with non-knockout mice (Group III), the level of urinary erythrocyte count and serum IgA in knockout mice (Group VI) increased significantly after treatment with dexamethasone (3.7 ± 0.6/HP vs. 9.2 ± 3.5/HP, t = 2.186, P = 0.024; 12.38 ± 0.26 mg/mL vs. 27.85 ± 0.65 mg/mL, t = 1.852, P = 0.041). The expression level of GRα was considerably increased in the knockout group after dexamethasone treatment compared with non-knockout mice in mRNA and protein level (t = 2.085, P = 0.026; t = 1.928, P = 0.035), but there was no statistically significant difference in the expression level of GRβ between condition knockout and non-knockout mice (t = 0.059, P = 0.087; t = 0.038, P = 1.12). Furthermore, the expression levels of glucocorticoid resistance genes (AP-1 and TGF-β1) were notably increased after p300 knockout compared with non-knockout mice in mRNA and protein level (TGF-β1: t = 1.945, P = 0.034; t = 1.902, P = 0.039; AP-1: t = 1.914, P = 0.038; t = 1.802, P = 0.041).Conclusions:p300 plays a crucial role in the pathogenesis of HSPN. p300 can down-regulate the expression of resistance genes (AP-1 and TGF-β1) by binding with GRα to prevent further renal injury and glucocorticoid resistance. Therefore, p300 is a promising new target in glucocorticoid therapy in HSPN.

Asparaginase Immune Complexes Induce FcγRIII‐Dependent Basophil Activation and Concentration‐Dependent Hypersensitivity Reactions

BackgroundAsparaginase (ASNase) is an important drug for treatment of pediatric acute lymphoblastic leukemia. However, antibody responses to ASNase increase the risk of leukemia relapse by mediating ASNase‐induced hypersensitivity and blocking ASNase's therapeutic effects. Previously, we have demonstrated two mechanism of ASNase hypersensitivity: one involving an anti‐ASNase IgG response, and another involving cell‐associated anti‐ASNase IgE antibodies. These studies suggested that formation of ASNase immune complexes (ICs) is sufficient to induce ASNase hypersensitivity.ObjectiveTo detect ASNase ICs in mice after the onset of ASNase hypersensitivity and determine whether FcγRIII engagement by ASNase ICs can lead to immune cell activation.MethodsMice were sensitized to native E. coli ASNase using aluminum hydroxide adjuvant. Six μm protein G‐coated polystyrene beads were used to detect plasma ASNase ICs by flow cytometry after the onset of ASNase‐induced hypersensitivity. Fluorochrome‐labeled ASNase was used to detect ASNase IC binding to immune cells. The basophil activation test (BAT) was used to evaluate ASNase IC effects on basophils.ResultsASNase ICs were detected after the onset of murine ASNase hypersensitivity reactions and ASNase IC concentration strongly correlated with reaction severity (r2= 0.796, P=0.0005). ASNase ICs formed ex vivo at high anti‐ASNase IgG1 antibody concentrations. ASNase IC binding to naïve and sensitized peripheral blood neutrophils, macrophages/monocytes, and basophils was FcγRIII‐dependent. Consistent with these results, the BAT demonstrated that ASNase IC activation of naïve and sensitized basophils was FcγRIII‐dependent.ConclusionASNase‐sensitized mice form ICs after ASNase administration, which activate murine basophils, and likely mast cells, via FcγRIII. This suggests that ASNase ICs can contribute to the onset and severity of ASNase hypersensitivity.Support or Funding InformationUniversity of Pittsburgh School of Pharmacy and NIH Grants RO1 CA216815 to CAFThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Antibody Dependent Enhancement of Acinetobacter baumannii Infection in a Mouse Pneumonia Model.

Acinetobacter baumannii has become a pathogen of increasing medical importance because of the emergence of multidrug-resistant strains and the high rate of mortality of infected patients. Promising animal study results have been reported recently with active and passive immunization against A. baumannii virulence factors. In the present study, a monoclonal IgG3 antibody, 8E3, was developed with specificity for the K2 capsular polysaccharide of A. baumannii, and its therapeutic potential was assessed. 8E3 enhanced macrophage-mediated bactericidal activity against the A. baumannii clinical strain AB899. However, 8E3 treatment (passive immunization) of AB899-infected mice led to a substantial increase in mortality and to substantial increases in bacterial load in blood, lung, and in splenic samples. In vitro investigations showed a large binding capacity in the supernatant of bacterial cultures, suggesting that shed capsule components act as a binding sink for 8E3. Investigations of 8E3 pharmacokinetics in mice demonstrated that unbound concentrations of the antibody dropped below detection limits within 24 hours after a 200 mg/kg dose. However, total concentrations of antibody declined slowly, with an apparent terminal half-life (t 1/2) of 6.7-8.0 days, suggesting that the vast majority of 8E3 in blood is bound (e.g., with soluble capsule components in blood). We hypothesize that high concentrations of 8E3-capsule immune complexes act to inhibit bacterial clearance in vivo. To the best of our knowledge, this is the first demonstration of antibody-dependent enhancement of A. baumannii infection, and these observations highlight the complexity of antibody-based therapy for A. baumannii infections.

Abstract 579: Anti-apolipoprotein A-I Antibody Profile Correlates With Cardiovascular Disease Outcomes

Apolipoprotein A-I (ApoA-I) is a target of IgG autoantibody induction in patients, but the role of these antibodies has not been fully elucidated. Previous research has characterized anti-ApoA-I IgG antibodies targeting delipidated ApoA-I as a biomarker of cardiovascular progression, but only a moderate association was observed. We hypothesize that free anti-ApoA-I IgG is a single component of the anti-ApoA-I response and characterization of anti-ApoA-I antibody profiles will be more predictive of adverse cardiovascular outcomes. Given the relative concentrations of ApoA-I and anti-ApoA-I antibodies, we examined sera samples from 375 patients with coronary artery disease (CAD) to quantify soluble ApoA-I/IgG immune complexes (ICs). We found a range of ApoA-I/IgG IC concentrations in patients, irrespective of free anti-ApoA-I antibodies. While free antibodies failed to predict outcomes in this CAD cohort, a median Cox regression analysis over 6 years of follow-up determined a hazard ratio of 1.5 (95% CI: 1.03-2.18, p = 0.03) for patients with below median ApoA-I/IgG ICs levels after adjusting for 11 traditional cardiovascular risk factors. In comparison, a cohort of healthy subjects exhibited significantly higher ApoA-I/IgG ICs. Pearson correlation analysis between ApoA-I/IgG ICs in the 375 patients with CAD and 25 patient characteristics found that only hypertension showed a significant association with ApoA-I/IgG ICs (r = -0.154, p = 0.003). In addition, no significant relationship between ApoA-I/IgG ICs and total ApoA-I concentration (r = -0.0601, p = 0.51) or total IgG concentration (r = 0.134, p = 0.137) was observed. Continued characterization of the molecular characteristics and immunologic sequelae associated with the immune complexes will help determine the functional implications of ApoA-I/IgG ICs in vivo . Current data using affinity column purification shows co-elution of ApoA-I and IgG in patients with elevated levels of ApoA-I/IgG ICs. Furthermore, ApoA-I/IgG ICs elute through size exclusion chromatography in the HDL fraction. The identification and characterization of ApoA-I/IgG ICs has the potential to guide clinical diagnosis and intervention strategies in patients with atherosclerotic cardiovascular disease.

A generalized quantitative antibody homeostasis model: maintenance of global antibody equilibrium by effector functions.

The homeostasis of antibodies can be characterized as a balanced production, target-binding and receptor-mediated elimination regulated by an interaction network, which controls B-cell development and selection. Recently, we proposed a quantitative model to describe how the concentration and affinity of interacting partners generates a network. Here we argue that this physical, quantitative approach can be extended for the interpretation of effector functions of antibodies. We define global antibody equilibrium as the zone of molar equivalence of free antibody, free antigen and immune complex concentrations and of dissociation constant of apparent affinity: [Ab]=[Ag]=[AbAg]=KD. This zone corresponds to the biologically relevant KD range of reversible interactions. We show that thermodynamic and kinetic properties of antibody–antigen interactions correlate with immunological functions. The formation of stable, long-lived immune complexes correspond to a decrease of entropy and is a prerequisite for the generation of higher-order complexes. As the energy of formation of complexes increases, we observe a gradual shift from silent clearance to inflammatory reactions. These rules can also be applied to complement activation-related immune effector processes, linking the physicochemical principles of innate and adaptive humoral responses. Affinity of the receptors mediating effector functions shows a wide range of affinities, allowing the continuous sampling of antibody-bound antigen over the complete range of concentrations. The generation of multivalent, multicomponent complexes triggers effector functions by crosslinking these receptors on effector cells with increasing enzymatic degradation potential. Thus, antibody homeostasis is a thermodynamic system with complex network properties, nested into the host organism by proper immunoregulatory and effector pathways. Maintenance of global antibody equilibrium is achieved by innate qualitative signals modulating a quantitative adaptive immune system, which regulates molecular integrity of the host by tuning the degradation and recycling of molecules from silent removal to inflammatory elimination.

Hydrolysis and Dissolution of Amyloids by Catabodies.

Catalytic antibodies (catabodies) hold potential for superior immunotherapy because of their turnover capability and no or minimal induction of inflammatory responses. Catabodies neutralize and remove target antigens more potently than conventional antibodies. Depending on the catalytic rate constant, a single catabody molecule degrades thousands to millions of target molecules over its useful lifespan, whereas conventional antibodies only form reversibly associated, stoichiometric complexes with the target. Thus, removal of the antibody-bound target requires accessory phagocytic cells that ingest the immune complexes, which is usually accompanied by release of inflammatory mediators. In comparison, catabodies bind the target only transiently, and the rapid and direct target destruction reduces the concentration of immune complexes that can activate inflammatory processes. These features are especially pertinent when large target amounts at anatomically vulnerable sites must be removed, e.g., amyloids. We reported specific catabodies to misfolded transthyretin (misTTR) amyloid and amyloid β peptide (Aβ). Accumulation of the oligomeric and fibrillized amyloid TTR forms causes diverse systemic pathologies, including cardiomyopathy, polyneuropathy, and skeletal diseases. Brain Aβ aggregates are thought to cause central nervous system degenerative disease, chiefly Alzheimer's disease. We describe methods for testing catabody-mediated degradation and dissolution of Aβ and TTR.

NEUROSENSOR DEAFNESS AFTER MILD CRANIO CEREBRAL TRAUMAS

The clinical features and pathogenesis are studied after mild cranio-cerebral trauma (MCCT). As a result of immunological and biochemical examinations, during the 1st day after MCCT the concentration of TBA-active products are statistically reliable raised compared to the control group. The increased concentration of circulant immune complexes (CIC) after mild cranio-cerebral trauma in adults can be linked to the involvement of immunopathological processes. The results of study showed that biochemical and immunological disorders remaining after recovery of disease. It is showed that pathogenic treatment of the mild brain contusion to be longer than the generally accepted time period.

Influence of Duration of Heroin Dependence on Humoral Immunologic Indicators.

The incidence of autoantibodies may be associated with the duration of drug use. In this study, we assessed the association between the duration of heroin dependence and various humoral immunologic indicators, including IgA, IgG, IgM, complement component 3, complement component 4, rheumatoid factor, anti-β2-glycoprotein 1 (IgA, IgG, IgM), antinuclear antibody, circulating immune complexes, and cryoglobulins. A total of 363 patients with heroin dependence were enrolled in this cross-sectional and prospective study over a 3.5-year period. Depending on the duration of heroin use, participants were divided into 3 groups: up to 3 years, 4 to 7 years, and more than 7 years of heroin dependence. All patients were analyzed for the indicators. There was a significant difference between the duration of heroin dependence and increased concentration of IgA (P = 0.0000), IgG (P = 0.0000), IgM (P = 0.0001), complement component 3 (P = 0.042), rheumatoid factor (P = 0.0001), anti-β2-glycoprotein 1 (IgA, P = 0.0098; IgG, P = 0.0000; IgM, P = 0.0000), the presence of antinuclear antibody (P = 0.01) and cryoglobulins (P = 0.0000), and decreased concentration of complement component 4 (P = 0.002). There was no significant difference in circulating immune complex concentration (P = 0.097). A longer duration of heroin dependence was associated with increased concentrations of IgA, IgG, IgM, complement component 3, rheumatoid factor, anti-β2-glycoprotein 1 (IgA, IgG, IgM), presence of antinuclear antibodies and cryoglobulins, and decreased concentrations of complement component 4, but there was no influence on circulating immune complex values.