Complement activation and expression of membrane regulators in the middle ear mucosa in otitis media with effusion.
The aetiopathogenesis of chronic otitis media with effusion (OME) in children is not yet fully understood. OME is characterized by metaplasia of the epithelium and accumulation of sticky, glue-like effusion in the middle ear containing different mediators of inflammation, including activation fragments of the complement system. Here we examined whether the fluid phase complement activation is reflected in the middle ear mucosa and how the mucosa is protected against the cytolytic activity of complement. Mucosal biopsies from 18 middle ears of children with a history of chronic OME were taken. The biopsies were analysed by immunofluorescence microscopy after staining for complement fragments iC3b/C3c, C3d and C9, and regulators membrane cofactor protein (MCP; CD46), decay-accelerating factor (DAF; CD55) and protectin (CD59). There was a strong staining for iC3b/C3c, and a weaker one for C3d and C9 on the surface of the middle ear epithelial cells of OME patients but not in controls without OME. MCP was expressed on the hyperplastic three to four outer cell layers of the epithelium, while CD59 was expressed throughout the middle ear mucosa. The results suggest a strong ongoing complement activation and consequent inflammation in the middle ear cavity. Unrestricted complement damage of the epithelial lining is prevented by the strong expression of MCP and CD59. (+info)
Linkage analysis of systemic lupus erythematosus induced in diabetes-prone nonobese diabetic mice by Mycobacterium bovis.
Systemic lupus erythematosus induced by Mycobacterium bovis in diabetes-prone nonobese diabetic mice was mapped in a backcross to the BALB/c strain. The subphenotypes-hemolytic anemia, antinuclear autoantibodies, and glomerular immune complex deposition-did not cosegregate, and linkage analysis for each trait was performed independently. Hemolytic anemia mapped to two loci: Bah1 at the MHC on chromosome 17 and Bah2 on distal chromosome 16. Antinuclear autoantibodies mapped to three loci: Bana1 at the MHC on chromosome 17, Bana2 on chromosome 10, and Bana3 on distal chromosome 1. Glomerular immune complex deposition did not show significant linkage to any genomic region. Mapping of autoantibodies (Coombs' or antinuclear autoantibodies) identified two loci: Babs1 at the MHC and Babs2 on distal chromosome 1. It has previously been reported that genes conferring susceptibility to different autoimmune diseases map nonrandomly to defined regions of the genome. One possible explanation for this clustering is that some alleles at loci within these regions confer susceptibility to multiple autoimmune diseases-the "common gene" hypothesis. With the exception of the H2, this study failed to provide direct support for the common gene hypothesis, because the loci identified as conferring susceptibility to systemic lupus erythematosus did not colocalize with those previously implicated in diabetes. However, three of the four regions identified had been previously implicated in other autoimmune diseases. (+info)
Immunological and clinical follow up of hepatitis C virus associated cryoglobulinaemic vasculitis.
OBJECTIVE: To study immunological markers and compare these markers with standard measures for the clinical and immunological follow up of vasculitis activity in hepatitis C virus (HCV) associated cryoglobulinaemic vasculitis (CV). METHODS: Serial serum samples from eight patients with newly diagnosed HCV associated CV were followed during interferon alpha treatment induced remission of the CV. Vasculitis activity and disease extent were evaluated with the Birmingham vasculitis activity score (BVAS) and disease extent index (DEI). Cryoglobulinaemia, complement levels (C3c, C4, and CH50), rheumatoid factor (RF), autoantibodies such as antinuclear antibodies, soluble interleukin 2 receptor (sIL2r), soluble intercellular adhesion molecule-1 (sICAM-1), and soluble CD30 (sCD30) were determined. RESULTS: All patients achieved either complete or partial remission of their CV during interferon alpha treatment. There was a significant reduction in vasculitis activity and disease extent (BVAS, DEI), cryoglobulinaemia, RF, sIL2r, sICAM-1, and sCD30. Complement C3c levels increased significantly during this period. Erythrocyte sedimentation rate and levels of complement C4 and CH50 did not change significantly. Both clinical measures (BVAS and DEI) correlated significantly only with C3c and sCD30. CONCLUSIONS: Although this study was of only a small group of patients, it shows that BVAS and DEI as clinical measures and C3c and sCD30 as immunological markers may be useful in the follow up of disease activity of HCV associated CV. The data indicate that activity of the humoral (cryoglobulinaemia, RF, autoantibodies) and cellular (sIL2r, sICAM-1, sCD30) immune response and endothelial damage (sICAM-1) are found in HCV associated CV. (+info)
Complement activation in patients with rheumatoid arthritis mediated in part by C-reactive protein.
OBJECTIVE: Complement activation in patients with rheumatoid arthritis (RA) is considered to be triggered by immune complexes. Recently, it was shown that C-reactive protein (CRP) can activate the complement system in vivo. We therefore hypothesized that part of the complement activation in RA is due to CRP. The aim of this study was to investigate CRP-mediated complement activation in RA, and to assess its correlation with disease activity. METHODS: Complexes between CRP and the activated complement components C3d (C3d-CRP) and C4d (C4d-CRP), which reflect CRP-mediated complement activation, as well as the overall levels of activated C3 and C4 were measured in the plasma of 107 patients with active RA and 177 patients with inactive RA. Inactive RA was defined according to the American College of Rheumatology criteria for clinical remission. Disease activity was assessed by the modified Disease Activity Score (DAS28). RESULTS: Plasma levels of C3d-CRP and C4d-CRP were increased in the majority of the patients, and were significantly higher in patients with active disease versus those with inactive RA (P < 0.001). In patients with active RA, the plasma concentrations of C3d-CRP and C4d-CRP correlated significantly with the DAS28 (Spearman's rho 0.61 and 0.55, respectively; P < 0.001), whereas these correlations were less pronounced in patients with inactive RA (Spearman's rho 0.28 [P < 0.001] and 0.25 [P = 0.001], respectively). Levels of activated C3 and C4 were also increased in the majority of the patients, particularly in patients with active RA. CONCLUSION: Part of the activation of complement in RA is mediated by CRP and is correlated with disease activity. We suggest that this activation is involved in the pathogenesis of RA. (+info)
Kinetic analysis of the interactions of complement receptor 2 (CR2, CD21) with its ligands C3d, iC3b, and the EBV glycoprotein gp350/220.
The molecular mechanisms involved in the interaction of complement receptor 2 (CR2) with its natural ligands iC3b and C3d are still not well understood. In addition, studies regarding the binding site(s) of the receptor on C3 as well as the affinities of the C3 fragments for CR2 have produced contradictory results. In the present study, we have used surface plasmon resonance technology to study the interaction of CR2 with its ligands C3d, iC3b, and the EBV surface glycoprotein gp350/220. We measured the kinetics of binding of the receptor to its ligands, examined the influence of ionic contacts on these interactions, and assessed whether immobilized and soluble iC3b bound with similar kinetics to CR2. Our results indicate that 1) gp350 binding to CR2 follows a simple 1:1 interaction, whereas that of the C3 fragments is more complex and involves more than one intramolecular component; 2) kinetic differences exist between the binding of C3d and iC3b to CR2, which may be due to an additional binding site found on the C3c region of iC3b; and 3) iC3b binds to CR2 with different kinetics, depending on whether the iC3b is in solution or immobilized on the surface. These findings suggest that binding of CR2 to iC3b and C3d is more complex than previously thought. (+info)
Detection of immune deposits in skin lesions of patients with Wegener's granulomatosis.
BACKGROUND: Wegener's granulomatosis (WG) is considered a pauci-immune systemic vasculitis based on the absence of immune deposits in renal biopsies of patients with active disease. In animal models of antineutrophil cytoplasmic antibody (ANCA) associated glomerulonephritis, immune deposits along the glomerular capillary wall are present at early stages of lesion development. These deposits are degraded rapidly, resulting in "pauci-immune" lesions. OBJECTIVE: To test the hypothesis that immune deposits can also be detected in early lesions of patients with WG, thereby initiating an inflammatory reaction that, in time, is augmented in the presence of ANCA, resulting in pauci-immune lesions later on. METHODS: The presence of immune deposits in skin biopsies taken within 48 hours of lesion development was investigated. Direct immunofluorescence was used to examine 32 skin biopsies for the presence of immune deposits (IgG, IgA, IgM, C3c). When possible, a comparison was made between the immunofluorescence findings in renal and skin biopsies taken at the same time. RESULTS: Four of 11 biopsies taken at initial presentation and four of 21 biopsies taken at the onset of a relapse of WG showed IgG and/or IgA containing immune deposits in the subepidermal blood vessels. All nine renal biopsies showed pauci-immune glomerulonephritis, irrespective of the presence (n=5) or absence (n=4) of immune deposits in the skin biopsy. CONCLUSION: A substantial number of skin biopsies showed immune deposits during active disease. These results could support the hypothesis that immune complexes may trigger vasculitic lesions in WG. (+info)
Human alpha galactosidase and alpha 1,2 fucosyltransferase concordantly inhibit xenoreactivity of NIH 3T3 cells with human serum.
AIM: To study the influence of the expression of human alpha galactosidase and alpha1,2 fucosyltransferase on Gal alpha 1,3 Gal and consequent xenoreactivity in NIH3T3 cells. METHODS: The expression levels of G antigen and H antigen and binding of human natural antibodies (IgG and IgM) and complement (C3c) to NIH3T3 cells were analyzed by flow cytometry. Western blot was employed to further determine the expression of glycoproteins of G antigen. Cytolysis assay with normal human serum was performed by MTT assay. RESULTS: Western blot showed that glycoproteins with molecular weight of 107 kDa, 98 kDa, 88 kDa, 56 kDa, 40 kDa, and 37 kDa were inhibited and even abrogated totally in alpha galactosidase transfectants and alpha 1,2 fucosyltransferase transfectants. The combined transfection of the two enzymes led to a much stronger inhibition of the glycoproteins. The binding of GS-IB4 was decreased by 57.4 % in alpha galactosidase transfectants, 28.8 % in alpha 1,2 fucosyltransferase transfectants, and 72.1 % in combined transfectants, respectively. In contrast, UEA-1 binding was increased about 6.7-fold, 6.0-fold, and 8.0-fold respectively. The xenoreactivity with human IgG was also reduced by 61.4 %, 67.0 %, and 73.4 %, respectively in the three kinds of transfectants. The resistance to cytolysis mediated by human serum was enhanced by 42.4 % in alpha galactosidase transfectants, 51.9 % in alpha 1,2 fucosyltranferase, and even 65.5 % in the combined transfectants. CONCLUSION: Although alpha galactosidase and alpha 1,2 fucosyltransferase had different biochemical properties, they could inhibit the expression of Gal alpha 1,3 Gal synergistically, leading to stronger resistance of xenograft against cytolysis. (+info)
Progesterone and RU486 regulation of uterine complement C3 after prior induction with estradiol.
Previous results demonstrated that progesterone (P4) given simultaneously with estradiol (E2) prevented stimulation by E2 of complement C3 expression in the immature rat uterus. Northern blot analysis revealed that simultaneous administration of P4 was able to prevent the E2-stimulated increase in C3 mRNA concentration in the luminal epithelial cells. The purpose of the present investigation was to determine whether progesterone modulates C3 expression after the gene has been induced by prior administration of E2 and also to determine the reversibility of this effect by the concomitant administration of RU38486, 17 beta-hydroxy-11 beta-[4-dimethylaminophenyl]estra-4,9,-dien-3-one (RU486). This regulation was studied by examination of protein synthesis as well as mRNA concentrations. Immature 21-day-old female rats were treated with E2 for 2 days (1 microgram/day), followed 24 h later by P4 (500 micrograms) or vehicle. Uteri were removed 6, 9, and 18 h after progesterone treatment and the radiolabeled secreted proteins were analyzed by SDS-PAGE and immunoprecipitation using a goat anti-rat C3 antibody. In animals treated with vehicle, E2-stimulated C3 synthesis remained elevated at 6 and 9 h and returned to control values by 18 h. In contrast, the administration of P4 resulted in a decrease in C3 synthesis at 6 and 9 h with the greatest decrease observed at 9 h. Similar results were obtained when C3 mRNA concentrations were examined. E2-stimulated C3 mRNA concentrations were decreased in rats treated with progesterone compared to those treated with vehicle alone.2 (+info)