Polyclonal IgA Research Articles

Polyclonal and oligoclonal IgA synthesis in the cerebrospinal fluid of neurological patients: An immunoaffinity-mediated capillary blot study

An intrathecal synthesis of IgA has been reported in various neurological disorders. However, the frequency of its occurence and the electrophoretic characteristics of the locally produced IgA remained a matter of controversy. We developed a sensitive immunoaffinity-mediated capillary blot technique for the detection of polyclonal and oligoclonal IgA in the CSF of 115 patients with various neurological disorders. Paired CSF and serum samples containing 50 ng IgA after appropriate dilutions were submitted to isoelectric focusing in agarose gels; IgA was then blotted onto a polyvinylidene difluoride sheet coated by an anti-IgA antiserum or by infectious antigens. The immunoblots were revealed by an alkaline phosphatase-conjugated anti-IgA antiserum. Only five samples displayed CSF-restricted oligoclonal IgA bands, including two out of 33 from MS patients. In herpetic encephalitis ( n = 5) and varicella-zoster meningitis ( n = 2), a strong intrathecal production of virus-specific IgA antibodies was detectable. In such cases, faint oligoclonal IgA antibodies were superimposed on a polyclonal background. A weak local production of anti- Borrelia burgdorferi IgA antibodies was present in two out of four cases of neuroborreliosis.

Charge distribution of serum IgA in IgA nephropathy with episodes of macrohematuria

In order to examine further the role of the electrical change of IgA in the pathogenesis of IgAN, we studied the charge distribution of serum IgA in IgAN with episodes of macrohematuria. Blood samples were taken from 20 IgAN patients during bouts of macrohematuria and again more than 3 months later. Twenty healthy adults were used as controls. All samples were focused by Multiphor II flatbet electrofocusing unit apparatus (LKB) on an agarose gel (0.8% isogel agarose-EF, 2.5% pharmalyte 4.0-6.5). The focused gels were incubated with polyclonal goat anti-human IgA and were washed in several changes of saline for 48 hours. Immunofixed materials remaining in the gels after washing were fixed and stained with coomassie brilliant blue R250. The stained gels were analysed by densitometry. The relative areas of the individual peaks and their proportional contributions to the total IgA spectrotype were determined by computer. The results were as follows: 1. The percentage of proportional contribution of IgA peaks (pI 4.7-5.15) in IgAN were higher in patients than in controls (p less than 0.01). 2. The percentage of proportional contribution of IgA peaks (pI 4.9-5.05) in IgAN at bouts of macrohematuria was higher than in the same patients when not undergoing episodes of macrohematuria (p less than 0.01). These results suggest that the excess amount of anionic IgA may contribute to the pathogenesis of IgAN.

Intravascular coagulation necrosis of the skin associated with cryofibrinogenemia, diabetes mellitus, and cardiolipin autoantibodies

Intravascular coagulation necrosis of the skin is rare and appears as hemorrhagic infiltrates that may develop ulcerating necrosis, mainly on the acral areas. The face, arms, and legs were severely involved in our patient. In this patient intravascular coagulation necrosis was associated with cryofibrinogenemia, diabetes mellitus, and IgM cardiolipin autoantibodies. In addition, rheumatoid factor, elevated polyclonal IgA, and haptoglobin were present as risk factors for the vasculopathy. Skin biopsy specimens showed plugging of dermal venules by thrombi formed of fibrin and erythrocytes. Immunohistologic staining revealed a strong positive reaction for fibrinogen, with some positivity for C3, C4, IgG, IgA, and IgM. Erythrocyte extravasation occurred in late lesions without being accompanied by perivascular leukocytic infiltrates. Detailed clinical examination failed to identify an underlying malignancy. Treatment with heparin and prednisolone produced only a brief remission. However, the combination of chlorambucil (7 mg/day orally) with low-dose oral prednisolone (10 mg/day) for several weeks controlled the disease and greatly reduced the cryofibrinogen. No relapse occurred after discontinuation of treatment.

Human IgA and IgG F(ab')2 that bind to staphylococcal protein A belong to the VHIII subgroup.

Staphylococcal protein A (SPA) is a bacterial membrane protein that possesses, in addition to its Fc gamma-binding activity, a distinct specificity for the Fab region of some IgM, IgA, IgG, and IgE. The Fab site that binds to SPA has been localized to the V region of the Ig H chain. In a previous study of human monoclonal and polyclonal IgM, we demonstrated that binding to SPA was highly restricted to molecules of the VHIII subgroup, and that nearly all VHIII IgM were able to bind SPA. The present study examines the VH composition of SPA-binding and SPA-nonbinding fractions of purified human polyclonal IgA, and IgG F(ab')2 fragments. We found that 22% of the IgA and 15% of the IgG F(ab')2 bound to SPA-agarose. Analysis with VH subgroup-specific antisera indicated that the SPA-binding fraction of IgA was dominated by the VHIII subgroup, and the SPA-binding fraction of IgG F(ab')2 contained only VHIII molecules. Furthermore, substantial portions of the total VHIII protein in IgA and in IgG F(ab')2 bound to SPA. We conclude that Fab binding to SPA is both restricted to and highly prevalent among human VHIII molecules, regardless of Ig class. These results suggest that protein A is an Ig superantigen.

Biochemical studies on the interaction of fibronectin with Ig.

We have previously biochemically characterized three separate sites on the fibronectin (Fn) molecule that interact with IgG. These studies have been extended to examine the interaction of Fn with other classes and subclasses of Ig. By ELISA, a preferential quantitative binding order of Fn to the major Ig classes and subclasses was obtained as follows: IgG greater than IgM greater than IgA, IgG1 greater than IgG3 = IgG4 greater than IgG2, and IgA1 = IgA2. Using fragments of Fn obtained by subtilisin digestion followed by IgM and IgA affinity chromatography, immunoblot analysis using monospecific antisera to separate regions of the Fn molecule, and amino acid sequence analysis, these studies indicate that polyclonal IgA and IgM interact with Fn in the same three regions and under the same ionic conditions as previously described for IgG. Site 1 is a 22-kDa fragment that commences at residue 1 of the Fn molecule. Sites 2 (16 kDa) and 3 (26-29 kDa) begin at residues 588 and 1597, respectively. Under physiological conditions a monoclonal antibody that recognizes site 1 completely inhibited the interaction of intact Fn with IgG, IgM, and IgA. Therefore, this is the only physiologically active site in the intact molecule. Aggregated but not monomeric IgG competitively inhibited the binding of Fn to IgG-coated microtiter ELISA plates; thus, this interaction can take place in a fluid-phase system. These results indicate that Fn can potentially interact with immune complexes and aggregates of all Ig in the circulation and thus may play a significant role in both their clearance and deposition in Fn-containing tissues, such as occurs in immune-complex-related disorders.

Cytokine regulation of localized inflammation. Induction of activated B cells and IL-6-mediated polyclonal IgG and IgA synthesis in inflamed human gingiva.

Abstract It is well established that increased numbers of plasma cells occur in the localized tissues of chronic inflammatory diseases such as adult periodontitis, and enzymatic isolation has shown that most B lineage cells produce IgG-subclass with some IgA-subclass responses. It would be of importance to determine if excess production of cytokines in the localized lesion account for these responses and in the present study we have assessed gingival mononuclear cell (GMC) supernatants for cytokines that activate B cells including IL-6R expression and for levels of IL-6 present. Inasmuch as limited numbers (approximately 1 to 3 x 10(6) cells) of GMC were obtained from surgically removed tissues (approximately 400 mg), we have focused on the analysis of IL-6 production by GMC in this study. Further, initial evidence of additional cytokines that are produced by GMC and induce expression of IL-6R on resting B cells has been obtained. The GMC and PBMC from individual patients were cultured in the presence (or absence) of Con A. Higher levels of IL-6 were produced spontaneously by GMC when compared with Con A-stimulated PBMC. When PBMC cultures were supplemented with GMC supernatants obtained from the same patient, high numbers of spot-forming cells (SFC), mainly of IgG followed by IgA isotype, were seen. The induction of SFC by GMC supernatants was inhibited by incubation with a goat anti-human IL-6 antibody. When the effect of GMC supernatants on subclasses of PBMC SFC was determined, the response was IgG1 greater than IgG2 greater than IgG3 = IgG4 and IgA1 greater than IgA2, a pattern remarkably similar to the distribution of plasma cells in the GMC itself. To assess for cytokines in GMC supernatants that mediated B cell activation, supernatants containing anti-IL-6 were cultured with PBMC or purified B cells for 72 h. This treatment induced small proliferative B cell responses and elevated expression of IL-6R on B cells, but did not induce SFC responses. Further, incubation of B cells with GMC supernatants induced resting B cells (G0/G1) to enter the cell cycle (S and G2/M). Addition of human rIL-6 to these cultures on day 3 restored IgG- and IgA-subclass SFC responses by day 7. Cytokine-induced IL-6R expression also occurred in vivo because freshly isolated GMC expressed high levels of this receptor.(ABSTRACT TRUNCATED AT 400 WORDS)

Subcorneal pustular dermatosis associated with rheumatoid arthritis and raised IgA: simultaneous remission of skin and joint involvements with dapsone treatment.

A 44 year old white woman with rheumatoid arthritis for 19 years developed subcorneal pustular dermatosis. She had increased polyclonal IgA and IgA rheumatoid factor. After 4 months' treatment with dapsone 100 mg daily the patient had neither skin lesions nor active joint disease.

Anti-inflammatory activity of human IgA antibodies and their Fab alpha fragments: inhibition of IgG-mediated complement activation.

The interaction of human IgA antibodies with the classical pathway of complement activation was investigated in a homologous human system, by means of two IgA1 and three IgG1 myeloma proteins having antibody activity against a defined antigen, staphylococcal alpha-toxin. In a solid-phase antigen-dependent C3b-binding ELISA system, the monoclonal IgG antibodies were previously shown to activate the classical complement pathway synergistically, resembling polyclonal IgG antibodies, whereas IgA antibodies were unable to activate complement by either pathway. In the present study, IgA antibodies were found to inhibit significantly the activation of complement initiated by antigen-bound polyclonal or mixed monoclonal IgG antibodies, in relation to the amount of IgA antibodies applied and bound to antigen. IgA1 myeloma proteins devoid of antigen-binding activity were without effect. Inhibition was independent of the ability of the IgA antibodies to compete against the IgG antibodies in binding to antigen, and was demonstrable with physiological concentrations of antibodies. Similar results were obtained with polyclonal serum IgA having antigen-binding activity. However, the binding of C1q to antigen-complexed IgG was inhibited only by a monoclonal IgA antibody that could compete against one of the three monoclonal IgG antibodies that bound C1q synergistically. This observation implied that at least two mechanisms were involved in the inhibition of C3b fixation. Fab alpha fragments of monoclonal IgA antibodies, obtained by cleavage with IgA1 protease from Haemophilus influenzae type b, were found to have a similar inhibitory effect on C3b fixation to the intact IgA1 antibodies. This observation supports the hypothesis that IgA1 proteases contribute to the invasive pathogenicity of certain mucosal bacteria, by cleaving secretory IgA1 antibodies to antigen-binding Fab alpha fragments, which are not only defective in mucosal defense properties, but which also protect the organisms from other immune effector systems, such as complement activation.

Transforming growth factor beta induces IgA production and acts additively with interleukin 5 for IgA production.

Effects of transforming growth factor beta (TGF-beta) on IgA production by LPS-stimulated B cells have been studied. TGF-beta itself could augment polyclonal IgA production in concomitant inhibition of polyclonal IgM and IgG1 production. Furthermore, TGF-beta and IL-5 additively augmented IgA production. TGF-beta exerted its activity early in the culture (by 2 d in a 5-d culture) and IL-5 was required late in the culture. Surface IgA- (sIgA-) B cells responded to TGF-beta for the development of IgA-secreting cells. By contrast, sIgA+ B cells, but not sIgA- B cells, responded to IL-5 for IgA production. These results suggest that TGF-beta has a differential role in the induction of IgA production from IL-5 on murine-activated B cells.

Importance of intestinal mucosal immunity and luminal bacterial cell wall polymers in the aetiology of inflammatory joint diseases

The distal intestine contains bacterial cell wall polymers capable of inducing acute and chronic polyarthritis if systemically distributed. Parenteral injection of peptidoglycan-polysaccharide (PG-PS) polymers from certain bacterial species produces spontaneously relapsing erosive synovitis in susceptible rat strains, and normally subarthropathic amounts of PG-PS and lipopolysaccharide (endotoxin) can reactivate arthritis initially induced by PG-PS. These experimental results illustrate the inflammatory potential of luminal bacterial products and the importance of genetically determined host susceptibility factors in the pathogenesis of arthritis. Normally, luminal complexing by secretory IgA and an intact epithelial barrier limits uptake of luminal antigen; however, intestinal inflammation enhances mucosal uptake and systemic distribution of potentially injurious macromolecules, including PG-PS and lipopolysaccharide. Occult intestinal inflammation, which may be related to non-steroidal anti-inflammatory drugs or may be disease-associated, occurs in approximately two thirds of patients with rheumatoid arthritis, idiopathic reactive arthritis and ankylosing spondylitis. Enhanced mucosal permeability to macromolecules occurs in rheumatoid arthritis, enteric infections and idiopathic inflammatory bowel disease. Intestinal inflammation is associated with increased mucosal IgG production and circulating immune complexes. Hyperactive IgA synthesis occurs in many types of inflammatory joint disease. Polyclonal IgA is increased in rheumatoid arthritis, Sjögren's syndrome, ankylosing spondylitis, Reiter's syndrome, and reactive arthritis following Yersinia infection. Anti-Klebsiella IgA cross-reacts with HLA-B27 antigen, and antibodies to enteric bacteria are able to lyse lymphocytes from HLA-B27 patients with ankylosing spondylitis. Anti-Yersinia IgA is produced at the mucosa in increased quantities in patients who develop arthritis following Yersinia enteritis, possibly as a consequence of defective cellular immunity. Serum concentrations of IgA correlate with activity of rheumatoid arthritis and ankylosing spondylitis, and serum IgA immune complexes are associated with rheumatoid vasculitis, suggesting that IgA contributes to the pathogenesis of arthritis. We speculate that intestinal injury may also induce or perpetuate arthritis by systemic distribution of inflammatory mediators produced by intestinal immune effector cells.

Anti-histone autoantibodies in ddY mice, an animal model for spontaneous IgA nephritis

The ddY mice are known to develop spontaneous glomerulonephritis resembling human IgA nephritis after 40 weeks of age. A sharp rise of circulating polyclonal IgG and IgA is also observed at this stage. Since these overproduced immunoglobulins seem to be related to the development of murine glomerulopathy, antigen-antibody interactions between renal tissue proteins and serum immunoglobulins were analyzed by Western blotting in ddY mice before and after 40 weeks of age. Serum IgG at 50 weeks reacted with an 18-kDa renal tissue protein which was identified as histone H3, as well as with histone H1. Renal histones were extracted along with IgG from the murine kidney at 50 weeks in a high salt soluble fraction. Serial studies of anti-histone antibodies by enzymelinked immunosorbent assay showed that IgG class antibodies markedly increased after 40 weeks of age. IgA class antibodies mildly increased after 56 weeks of age. Anti-DNA antibodies were not detected. These results demonstrate that ddY mice also develop mainly IgG class and partly IgA class anti-histone autoantibody after 40 weeks of age, and that histone-anti-histone complexes may contribute to the development of murine glomerulopathy. Although anti-histone antibodies have been reported in lupus mice, ddY mice differ from these mice in that no anti-DNA antibodies develop.

Complement-fixing properties of human IgA antibodies. Alternative pathway complement activation by plastic-bound, but not specific antigen-bound, IgA.

The complement-fixing properties of human IgA antibodies bound to specific antigen, or coated directly on plastic surfaces, were examined in comparison with those of IgG antibodies. Use was made of antigen-binding (anti-staphylococcal alpha-toxin) IgA and IgG monoclonal antibodies and normal polyclonal IgA and IgG, purified greater than 99.9% by avoidance of denaturing processes. Complement-fixation ELISA was used, with a high density of biotin-conjugated staphylococcal alpha-toxin bound to avidin-coated plates for the efficient capture of antibodies, and conditions were adjusted for the assessment of classical and alternative pathways of complement activation. Although IgA coated directly on plastic surfaces activated the alternative complement pathway in a dose-dependent manner, IgA antibodies bound to antigen failed to fix complement by either classical or alternative pathways. In contrast, IgG antibodies, either bound to antigen or coated directly on plastic, activated complement mainly by the classical pathway. It was concluded that the complexation of IgA antibodies with antigen is insufficient to elicit complement activation: rather a degree of denaturation seems to play a part in the expression of alternative complement pathway-activating properties by IgA.

Antibodies to polyclonal IgA, IgA1, and IgA2 and isotype-specific immune complexes in IgA nephropathy.

The concentrations of serum IgG and IgM antibodies to polyclonal IgA (IgAp), IgA1, and IgA2 were determined by enzyme immunoassay in 31 patients with IgA nephropathy and 30 healthy controls. Patients with IgA nephropathy had significantly raised concentrations of serum IgA compared to controls (Mann-Whitney U test, P = 0.001) and increased concentrations of conglutinin-binding IgA immune complexes (P = 0.024). No differences in the median concentrations of IgG and IgM anti-IgA antibodies were found between the patients and the controls. In serum samples from healthy controls there was a significant positive correlation between IgM anti-IgAp and IgA immune complex concentrations (P = 0.05), which contrasted with the finding of an inverse correlation between IgM anti-IgAp and IgA immune complex concentrations in patients with IgA nephropathy (P less than 0.05). In addition, the concentrations of conglutinin binding IgM immune complexes in serum were found to correlate with the concentration of IgM anti-IgAp (0.010 less than P less than 0.025), IgM anti-IgA1, and IgM anti-IgA2 (P much less than 0.005 for both) in patients with IgA nephropathy but not in controls. IgM anti-IgA antibodies may be important in augmenting the clearance of IgA immune complexes from the serum of patients with IgA nephropathy.

Receptor for IgA in group A streptococci: cloning of the gene and characterization of the protein expressed in Escherichia coli.

The gene for an IgA-binding protein from a group A streptococcal strain was cloned and expressed in Escherichia coli. The IgA-binding protein, called protein Arp, was purified on IgA-Sepharose, allowing complete purification in a single step. Analysis of protein Arp by Western immunoblotting demonstrated a major IgA-binding band, with an apparent molecular weight of 42 kD. The purified protein was shown to bind serum IgA and secretory IgA, as well as monoclonal IgA of both subclasses. There was no binding to IgM, IgD or IgE, but a weak binding to IgG. Inhibition experiments with whole bacteria indicated that IgA and IgG bind at separate sites. Experiments with immunoglobulin fragments showed that protein Arp binds to the Fc region of both IgA and IgG. The equilibrium constant of the reaction between protein Arp and polyclonal human IgA was determined to be 5.6 x 10(8) M-1. Amino acid sequencing of protein Arp demonstrated a direct repeat of 7 amino acids in the NH2-terminal region, a feature previously found in several streptococcal M proteins. This suggests that protein Arp, like M proteins, may be a streptococcal virulence factor.

In vitro and in vivo activity of polyclonal and monoclonal human immunoglobulins G, M, and A against Pseudomonas aeruginosa lipopolysaccharide

We evaluated the in vitro opsonophagocytic killing activity of monoclonal human immunoglobulin G (IgG), IgM, and IgA specific for Pseudomonas aeruginosa lipopolysaccharide and the in vivo protective capacity in neutropenic mice of both monoclonal and purified polyclonal IgG, IgM, and IgA. Monoclonal IgM was efficacious in mediating opsonophagocytic killing only in conjunction with complement, whereas monoclonal IgG opsonic killing was potentiated by complement, and monoclonal IgA opsonic killing was independent of complement. These findings are similar to those previously reported for purified polyclonal IgM, IgG, and IgA. The monoclonal and polyclonal immunoglobulins had comparable 50% protective doses in neutropenic mice (range, 0.28 to 0.46 microgram per mouse). The protective activity of IgM in neutropenic mice was abolished by cobra venom factor treatment, whereas IgG and IgA maintained efficacy in cobra venom factor-treated mice. These data indicate that all three major human serum immunoglobulin isotypes have opsonophagocytic and protective activities against P. aeruginosa, with a critical role for complement in the function of IgM.

Anti-IgA antibodies in IgA-deficient children.

IgG and IgM isotype antibodies to polyclonal human IgA, myeloma IgA1, and myeloma IgA2 were estimated in 38 IgA-deficient children aged between 0.9 and 15 years. All children had IgM anti-IgA antibodies. IgG antibodies against either polyclonal IgA, IgA1, or IgA2 were present in 63% of the IgA-deficient children. IgG anti-IgA antibodies were detected against all three antigens in 8 of 11 severely IgA-deficient children and in 7 of 27 partially IgA-deficient children, but in only 1 of 23 healthy adult controls. The proportion of children with IgG anti-IgA antibodies was significantly greater in the severely IgA-deficient group in comparison with the partially IgA-deficient group and the adult controls (chi-square test, P less than 0.01 and P less than 0.005, respectively). There was a strong correlation within each IgG subclass between antibody responses toward each of the three IgA antigens. Twenty-four children were followed over a period ranging from 0.9 to 11 years (mean, 2.3 years). Three children who were initially IgG anti-IgA antibody negative became antibody positive and three who were antibody positive became antibody negative. Five children with severe IgA deficiency remained severely IgA deficient and IgG antibodies to IgA persisted in all five at follow-up. The presence of IgG anti-IgA antibodies did not influence the normalization of serum IgA at follow-up in 14 of 19 children who were initially partially IgA deficient.

Nonimmune binding of Ig to Clostridium perfringens. Preferential binding of IgM and aggregated IgG.

In an attempt to find a bacterial IgM receptor, a large number of bacterial strains of different species were screened for the ability to bind human IgM. Certain strains of the anaerobic bacterium Clostridium perfringens were found to bind a major fraction of polyclonal IgM. One bacterial strain showed a particularly high binding capacity and was studied in more detail. This strain is also able to bind a minor fraction of polyclonal IgA and IgG. Inhibition experiments indicate that the different Ig classes bind to one and the same R structure. The ability of the strain to bind polyclonal Ig is correlated to the number of subunits in the Ig. This correlation can most simply be explained by increasing avidity with increasing number of subunits. In agreement with this hypotheses, experiments with aggregated IgG show that binding ability increases with aggregate size. Experiments with Ig fragments indicate that the binding structure in Ig is located in the F(ab')2 region. The ability of this bacterial strain to bind a majority of IgM molecules as well as aggregated IgG is potentially useful in immunologic work and represents a new type of Ig binding to bacteria.

The identification of murine Peyer's patch T cell-derived factors which enhance IgA responses.

The humoral immune response to ingested antigens is dominated by antibody of the IgA isotype. In the mouse as in other higher mammals the major inductive site of secretory IgA responses is the Peyer’s patch (1). Work in this and other laboratories has indicated that regulatory T cells (2) and accessory cells (3) within the PP microenvironment play an important role in controlling the isotype specificity of mucosal immune responses. We have previously shown that treatment of whole PP cell populations, derived by enzymatic disaggregation, with the oxidative mitogen sodium periodate results in the formation of cell clusters enriched in L3T4+ T cells and dendritic cells (DC) (4). Co-culture of these cluster cells with either PP B cells or bone marrow mononuclear cells enhances polyclonal IgA production(5). Furthermore, the IgA inductive effect of the cluster cells is also found in 7 day culture supernatants of isolated clusters, indicating that the effect is mediated by soluble factors. T cell hybridomas have been produced from cluster T cells and the culture supernatants screened for IgA enhancing activity on whole PP cultures. Culture supernatants which showed IgA enhancing activity were tested for well characterized lymphokines including IL-2, IL-4 and IL-5. Attempts were also made to generate DC hybridomas by fusion of DC enriched cluster populations with a HAT sensitive subclone of the macrophage line P388D1. These initial fusions however, produced further T cell hybridomas as demonstrated by expression of surface Thy 1.2 antigen and the production of multiple T cell lymphokines. Culture supernatants from several of these hybridomas also showed potent IgA enhancing activity when added to whole PP cell cultures or LPS stimulated spleen B cell cultures. In all cases, hybridoma supernatants exhibiting IgA enhancing activity were positive for IL-4 and IL-5. To further investigate the role of these lymphokines in IgA synthesis, LPS stimulated spleen B cells and nonstimulated PP B cells were cultured with recombinant IL-4 and/or IL-5 to see if the IgA enhancing activity was associated with these molecules.

Immunopathological study of glomerular IgA deposition in ddY mice.

Serial renal biopsies were performed to observe in detail the age-related glomerular IgA deposition in ddY mice. Moreover, lymphocytes from spleens and Peyer's patches (PP) were immunologically examined in the mice at 10 months of age. The incidence of IgA deposition in the glomeruli was 0% at 3 months, 33% at 7 months, 60% at 10 months and 100% at 13 months, but IgA deposits were less intense than IgG and IgM deposits. Serum levels of IgA and polyclonal IgA responses of PP cells were significantly higher in the mice with glomerular IgA deposits. However, neither the Lyt-1+/Lyt-2+ cell ratio nor the polyclonal IgA responses of spleen cells were related to IgA deposition. Therefore, gut-associated lymphoreticular tissue (GALT) probably plays an important role in the pathogenesis of the high IgA response and glomerular IgA deposition observed in this strain. These results suggest that ddY mice would be valuable as a model of spontaneous glomerular IgA deposition, as well as being useful for studying the possible relationship between the mucosal immune system and glomerular IgA deposition.

Activation of the alternative pathway of complement by human serum IgA.

In order to study the activation of complement by soluble aggregates of human polyclonal serum IgA, lysis of sheep erythrocytes (E) coated with several IgA preparations was used as a model. A complement nonactivating monoclonal mouse IgG1 against IgA was used to coat the cells. IgA, isolated from normal human serum, was aggregated by either N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP), glutaraldehyde, carbodiimide or heating. Depending on the size of the aggregates, and on the method of aggregation, E coated with aggregated IgA (E gamma 1.AIgA) could be lysed. The alternative pathway of complement appeared to mediate the lysis because the latter was observed in the presence of EGTA containing 5 mM Mg2+ (MgEGTA) and properdin (P) was deposited on the cells. Furthermore, no lysis was observed in C3-deficient serum. In the absence of AIgA the cells were not lysed, and no P deposition was observed. In another set of experiments E gamma 1.AIgA were first reacted with purified C3, B, D and P for 30 min at 30 degrees C, and subsequently in rat serum EDTA at 37 degrees C. Lysis occurred when E gamma 1.AIgA were prepared using SPDP-, glutaraldehyde- or carbodiimide-AIgA. Incubation of 100 micrograms/ml SPDP-AIgA with normal human serum for 30 min at 37 degrees C in the presence or absence of MgEGTA also induced consumption of total complement. The other soluble AIgA preparations were less effective in activating complement. These results suggest that polymeric serum IgA is capable of activating the alternative pathway of complement.