Triggering of the dsRNA Sensors TLR3, MDA5, and RIG-I Induces CD55 Expression in Synovial Fibroblasts

Background: CD55 (decay-accelerating factor) is a complement-regulatory protein highly expressed on fibroblast-like synoviocytes (FLS). CD55 is also a ligand for CD97, an adhesion-type G protein-coupled receptor abundantly present on leukocytes. Little is known regarding the regulation of CD55 expression in FLS. Methods: FLS isolated from arthritis patients were stimulated with pro-inflammatory cytokines and Toll-like receptor (TLR) ligands. Transfection with polyinosinic-polycytidylic acid (poly(I:C)) and 5'-triphosphate RNA were used to activate the cytoplasmic double-stranded (ds)RNA sensors melanoma differentiation-associated gene 5 (MDA5) and retinoic acid-inducible gene-I (RIG-I). CD55 expression, cell viability, and binding of CD97-loaded beads were quantified by flow cytometry. Results: CD55 was expressed at equal levels on FLS isolated from patients with rheumatoid arthritis (RA), osteoarthritis, psoriatic arthritis and spondyloarthritis. CD55 expression in RA FLS was significantly induced by IL-1 beta and especially by the TLR3 ligand poly(I:C). Activation of MDA5 and RIG-I also enhanced CD55 expression. Notably, activation of MDA5 dose-dependently induced cell death, while triggering of TLR3 or RIG-I had a minor effect on viability. Upregulation of CD55 enhanced the binding capacity of FLS to CD97-loaded beads, which could be blocked by antibodies against CD55. Conclusions: Activation of dsRNA sensors enhances the expression of CD55 in cultured FLS, which increases the binding to CD97. Our findings suggest that dsRNA promotes the interaction between FLS and CD97-expressing leukocyte

A functional SNP in the regulatory region of the decay-accelerating factor gene associates with extraocular muscle pareses in myasthenia gravis

Complement activation in myasthenia gravis (MG) may damage muscle endplate and complement regulatory proteins such as decay-accelerating factor (DAF) or CD55 may be protective. We hypothesize that the increased prevalence of severe extraocular muscle (EOM) dysfunction among African MG subjects reported earlier may result from altered DAF expression. To test this hypothesis, we screened the DAF gene sequences relevant to the classical complement pathway and found an association between myasthenics with EOM paresis and the DAF regulatory region c.-198C>G SNP (odds ratio=8.6; P=0.0003). This single nucleotide polymorphism (SNP) results in a twofold activation of a DAF 5′-flanking region luciferase reporter transfected into three different cell lines. Direct matching of the surrounding SNP sequence within the DAF regulatory region with the known transcription factor-binding sites suggests a loss of an Sp1-binding site. This was supported by the observation that the c.-198C>G SNP did not show the normal lipopolysaccharide-induced DAF transcriptional upregulation in lymphoblasts from four patients. Our findings suggest that at critical periods during autoimmune MG, this SNP may result in inadequate DAF upregulation with consequent complement-mediated EOM damage. Susceptible individuals may benefit from anti-complement therapy in addition to immunosuppression

The role of the complement system in traumatic brain injury: a review

Traumatic brain injury (TBI) is an important cause of disability and mortality in the western world. While the initial injury sustained results in damage, it is the subsequent secondary cascade that is thought to be the significant determinant of subsequent outcomes. The changes associated with the secondary injury do not become irreversible until some time after the start of the cascade. This may present a window of opportunity for therapeutic interventions aiming to improve outcomes subsequent to TBI. A prominent contributor to the secondary injury is a multifaceted inflammatory reaction. The complement system plays a notable role in this inflammatory reaction; however, it has often been overlooked in the context of TBI secondary injury. The complement system has homeostatic functions in the uninjured central nervous system (CNS), playing a part in neurodevelopment as well as having protective functions in the fully developed CNS, including protection from infection and inflammation. In the context of CNS injury, it can have a number of deleterious effects, evidence for which primarily comes not only from animal models but also, to a lesser extent, from human post-mortem studies. In stark contrast to this, complement may also promote neurogenesis and plasticity subsequent to CNS injury. This review aims to explore the role of the complement system in TBI secondary injury, by examining evidence from both clinical and animal studies. We examine whether specific complement activation pathways play more prominent roles in TBI than others. We also explore the potential role of complement in post-TBI neuroprotection and CNS repair/regeneration. Finally, we highlight the therapeutic potential of targeting the complement system in the context of TBI and point out certain areas on which future research is needed

Genetics and complement in atypical HUS

Central to the pathogenesis of atypical hemolytic uremic syndrome (aHUS) is over-activation of the alternative pathway of complement. Following the initial discovery of mutations in the complement regulatory protein, factor H, mutations have been described in factor I, membrane cofactor protein and thrombomodulin, which also result in decreased complement regulation. Autoantibodies to factor H have also been reported to impair complement regulation in aHUS. More recently, gain of function mutations in the complement components C3 and Factor B have been seen. This review focuses on the genetic causes of aHUS, their functional consequences, and clinical effect

No Evidence that Knops Blood Group Polymorphisms Affect Complement Receptor 1 Clustering on Erythrocytes

Clustering of Complement Receptor 1 (CR1) in the erythrocyte membrane is important for immune-complex transfer and clearance. CR1 contains the Knops blood group antigens, including the antithetical pairs Swain-Langley 1 and 2 (Sl1 and Sl2) and McCoy a and b (McCa and McCb), whose functional effects are unknown. We tested the hypothesis that the Sl and McC polymorphisms might influence CR1 clustering on erythrocyte membranes. Blood samples from 125 healthy Kenyan children were analysed by immunofluorescence and confocal microscopy to determine CR1 cluster number and volume. In agreement with previous reports, CR1 cluster number and volume were positively associated with CR1 copy number (mean number of CR1 molecules per erythrocyte). Individuals with the McCb/McCb genotype had more clusters per cell than McCa/McCa individuals. However, this association was lost when the strong effect of CR1 copy number was included in the model. No association was observed between Sl genotype, sickle cell genotype, α+thalassaemia genotype, gender or age and CR1 cluster number or volume. Therefore, after correction for CR1 copy number, the Sl and McCoy polymorphisms did not influence erythrocyte CR1 clustering, and the effects of the Knops polymorphisms on CR1 function remains unknown

Study of circulating immune complexes in thyroid disease. Comparison of Raji cell RIA and specific thyroglobulin-anti-thyroglobulin radioassay

Circulating soluble immune complexes (ICs) were studied by Raji cell RIA in a series of patients with and without thyroid disorders. Clearly elevated IC levels (>41.2 εgeq/ml; i.e. >3 SD above the mean value of the normal controls) were found in 3 of 18 patients with Hashimoto's thyroiditis, in 4 of 39 with differentiated thyroid carcinoma, and in none of 21 patients with Graves' disease as well as in none of 21 normal healthy controls. Thyroglobulin (Tg) was not found in the ICs bound to Raji cells when sought by a sensitive radiolabeled antibody technique. Furthermore, experiments carried out with radiolabeled Tg-anti- Tg ICs (Tg-ICs) obtained by mixing Tg with homologous (Hashimoto's sera) antibody revealed that homologous Tg-ICs did not bind to Raji cells, while the ICs prepared from heterologous (rabbit) antibody did bind. Tg-ICs were also assayed in the same sera and in additional samples from 29 normal controls and 12 patients with Graves' disease by a recently developed specific immunoradiometric assay (Takeda, Y., and J. P. Kriss, J Clin Endocrinol Metab 44: 46, 1977). Lower levels of Tg-ICs (22–262 ng⁄ml) than those detectable by Raji cell RIA were shown in 5.5‰ of the patients with Hashimoto's thyroiditis, 38.2‰ of those with Graves' disease, and 33.2‰ of those with differentiated thyroid carcinoma. No correlation was found between these results and the IC levels assessed by Raji cell RIA. Our data suggest that both Tg-ICs and complexes unrelated to Tg are present in sera of patients with thyroid autoimmune disorders and thyroid carcinoma. The ICs detected by the Raji cell method must not contain Tg, since 1) they occur in patients who are negative by the specific Tg-IC assay, 2) they are present in concentrations nearly 1000-fold higher than Tg-ICs, 3) Tg cannot be detected in the ICs bound to the Raji cells, and 4) Tg- ICs prepared from human antibody did not bind to Raji cells. The possible pathogenic role of these substances remains to be established. The failure of Tg-ICs to fix complement (and thus bind to Raji cells) may explain why IC disease is not a characteristic feature of autoimmune thyroid diseas

Comparison of complement fixation and radioassay techniques to detect solubilized human thyroid microsomal antigenic activity

Although recently the human thyroid microsomal antigen (M-Ag) has been possibly identified as the thyroid peroxidase, its nature remained unknown over almost three decades. One of the difficulties encountered in the identification of M-Ag derived from the conflicting data obtained in the attempts to solubilize active antigenic material from thyroid subcellular fractions. In particular, following detergent treatment, M-Ag could not be detected by complement fixation, while a full recovery of the antigen has been observed using a radioassay technique. In the present investigation, the antigenic activity of Triton X-100 solubilized thyroid microsomes was assessed in parallel by complement fixation and radioassay methods employing the same anti-microsomal antibody (anti-M Ab) preparation for antigen detection. In untreated microsomes antigenic activity was detected by both methods. In contrast, detergent solubilized M-Ag was detected by radioassay, but could not be detected by complement fixation. These data indicate that detergent solubilization diminishes the complement fixing capacity of M-Ag, while the solubilized antigen can still be fully detected by its binding reaction with the autoantibody, and explain the discrepant results obtained in previous studie