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Identification of a clinically relevant immunodominant region out lung hemorrhage. The pathogenic process is driven of type IV collagen in Goodpasture disease. by antibodies to type IV collagen, one of the main struc- Background. The characteristic feature of Goodpasture dis- tural components of basement membranes. These auto- ease is the occurrence of an autoantibody response to the antibodies can be detected as circulating antibodies or noncollagenous domain of the a3 chain of type IV collagen as linear deposits of IgG in the affected basement mem- (a3(IV)NC1) in the alveolar and glomerular basement mem- brane. These antibodies are associated with the development branes. Studies have revealed that the major antigenic of a rapidly progressive glomerulonephritis, with or without epitopes are located in the C-terminal region of the lung hemorrhage, whereas autoantibodies specific for the other a3(IV) chain (1, 2), one of the six known a(IV) chains a chains of the heterotrimeric type IV collagen probably do (3). In this region, each chain is folded into a globular not cause disease. In this study, we have investigated whether domain (4), called the NC1 domain. The limited distribu- differences in fine specificity of autoimmune recognition of the tion of the a3(IV) chain (5), mainly in glomerular and a3(IV)NC1 correlate with clinical outcome. Methods. For mapping of antibody binding to type IV colla- alveolar basement membranes, correlates well with the gen, chimeric collagen constructs were generated in which parts organ involvement in Goodpasture disease. of the a3(IV)NC1 domain were replaced by the corresponding There are several reports discussing specificity of the sequences of homologous nonreactive a1(IV). The different Goodpasture autoantibodies and their epitopes. The epi- recombinant collagen chimeras allowed the analysis of anti- tope has been shown to be a conformational epitope

Circulating anti-entactin antibodies in patients with glomerulonephritis. Sera from 305 consecutive patients in a renal biopsy series were analyzed for the presence of anti-entactin antibodies by ELISA. Of these patients, 59% had primary glomerulonephritis, 21% had secondary glomerulonephritis, while 20% had other nephropathies (noninflammatory conditions like amyloidosis, diabetic nephropathy, nephrosclerosis, etc.). Forty-one of these patients (13.4%) were positive for IgG/IgM antibodies against entactin; 60% of them had primary glomerulonephritis, 35% had secondary glomerulonephritis, while the remaining 3 patients had other nephropathies. Fifteen (70%) of the 23 patients with primary glomerulonephritis had proliferative glomerulonephritis (PGN), whereas 13 (87%) of the 15 patients with secondary glomerulonephritis were due to systemic connective tissue diseases (SCTD); 7 due to SLE, 4 due to SLE like SCTD and two due to other SCTD. There was a peak of incidence corresponding to the group aged 18 to 30 years. A majority of these patients (12 of the total 17) had primary glomerulonephritis and were associated with nephrotic or subnephrotic grade proteinuria, poorly or nonresponsive to immunosuppressive treatment and associated, in several cases, with progressive deterioration of renal function. In addition, there was a tendency to another peak in the age group 51 to 60 years. Most of these patients (6 of the total 8) had glomerulonephritis secondary, mainly, to SLE or SLE like SCTD with milder degree of proteinuria and better preserved renal functions Anti-entactin antibodies were not found in certain glomeruionephritides like IgA nephropathy and those secondary to systemic vasculitides and in control subjects (healthy subjects, and patients with a variety of non-renal disorders including inflammatory diseases). There was a significant correlation between the presence of circulating anti-entactin antibodies and the deposition of corresponding class of immunoglobulin along the glomerular basement membrane. In addition, a significant correlation between the absence of immune deposits and negative results for anti-entactin antibodies by ELISA was also observed. Our observations suggest that anti-entactin antibodies are associated with certain definite categories of glomeruionephritides in human beings and may well be involved in their immunopathogenesis.

The PiZ-gene of α1-antitrypsin as a determinant of the outcome in PR3-ANCA-positive vasculitis. We have previously demonstrated that a strong correlation exists between systemic vasculitis with proteinase 3-anti-neutrophil cytoplasm antibodies (PR3-ANCA) and heterozygosity for α1-antitrypsin (α1-AT) deficiency, PiZ. In the present study we characterized the PiZ-positive subgroup by laboratory findings, clinical features and outcome. The series studied consisted of 18 PiZ-positive and 81 PiZ-negative PR3-ANCA patients, comparable in sex ratio, age, C-reactive protein concentrations and renal function at diagnosis, and treatment. PiZ-positive patients had a more disseminated disease as reflected by the number of affected organs (P < 0.01). We found no group differences in relapse tendency. Overall mortality was 39% (7 of 18) in the PiZ-positive and 16% (13 of 81) in the non-PiZ group (P = 0.048). When survival analysis was restricted to 66 patients included in the study at disease onset, the group difference was significant (P = 0.016). The results suggest that the subnormal response of plasma α1-AT seen in PiZ-heterozygotes enhances the risk of dissemination of the vasculitic process and the risk of a fatal outcome. We consider α1-AT phenotyping to be justified in cases of PR3-ANCA-positive vasculitis. Treatments decreasing plasma α1-AT (such as plasmapheresis without plasma replacement) may be deleterious.

Glomerulonephritis induced in sheep by immunization with human glomerular basement membrane. The specificity of the anti-glomerular basement membrane (GBM) antibodies in experimental nephritis in sheep (Steblay's nephritis) was studied and compared with the specifity of antibodies in human anti-GBM nephritis (Goodpasture's syndrome). Sheep were injected monthly with isolated human GBM and antibody reactivities with isolated human and sheep GBM proteins were quantified with ELISA. Expectedly, the sheep had high titers of antibodies against several human GBM antigens. These antibodies remained for the most part in the circulation. In contrast, circulating antibody levels against sheep GBM antigens remained low for a long period of time, but a significant and progressive increase coincided with the development of acute nephritis. These antibodies accumulated in the kidneys of the nephritic sheep and could be eluted from diseased kidneys. They represent auto-antibodies immunologically cross-reacting with antigens of both sheep and human GBM. The specificity of auto-antibodies eluted from the kidneys was analyzed by immunoblotting and ELISA. The major populations reacted with one subunit, termed M2, of the globular domain of collagen IV. The same subunit contains the major antigen in Goodpasture's syndrome. It is concluded that the M2 subunit of the globular domain of collagen IV is recognized by IgG antibodies that primarily bind to the glomerular basement membrane in both Steblay's nephritis and Goodpasture's syndrome, indicating that it is a main nephritogen in both diseases.

Characterization of anti-GBM antibodies involved in Goodpasture's syndrome. Goodpasture's syndrome is a life threatening autoimmune kidney disease. The patients have autoantibodies to the glomerular basement membrane, which are specific for the C-terminal domain of type IV collagen (NC1). The major antigen has been localized to the α3(IV)-chain. We have investigated sera from 44 patients with anti-NC1 antibodies. The quantity of antibodies to four different α(IV)-chains of type IV collagen was measured with direct ELISA. We used affinity chromatography to separate the antibodies and their specificities were studied with ELISA. The results show that about 1% of the patients total IgG are anti-NC1 antibodies and that 90% of these antibodies are specific for the α3(IV)-chain. Antibodies to the other α(IV)-chains were found in 80% of the patients. Furthermore, affinity purified anti-α3(IV) antibodies from one patient were inhibited by antibodies from the other patients, from 4 to 72%. The antibodies, from 39 of the patients, were inhibited by a monoclonal antibody against the α3(IV)-chain. The results indicate that patients with Goodpasture's syndrome can have antibodies to most of the α(IV)-chains, while the majority of anti-NC1 antibodies are restricted to the α3(IV)-chain. Moreover the number of epitopes seems to be limited and the majority of the antibodies from most patients are against one single epitope on the α3(IV)-chain.

Antiglomerular basement membrane antibody: Antibody specificity in different forms of glomerulonephritis. Components were solubilized from human glomerular basement membrane by digestion with collage-nase and pepsin or by extraction with guanidine-HCl either directly or after previous digestion with the enzyme. The diverse preparations were used as antigens in the enzyme-linked immunosorbent assay (ELISA) of antibody titers in sera from patients with Goodpasture syndrome and patients with other forms of glomerulonephritis, that is, systemic lupus erythematosus, periarteritis nodosa, and IgA-related nephropathy. Patients with Goodpasture syndrome had high titers of IgG antibodies reacting most strongly with collagenase digests. The antigen(s) was only partly solubilized by guanidine-HCl extraction, was destroyed by pepsin digestion as well as reduction, and partly destroyed by trypsin digestion. The antigen(s) is most likely noncollagneous protein. Antibodies from patients with other forms of nephritis were directed primarily against antigens in guanidine-HCl extracts, while the antigen(s) was not solubilized by collagenase digestion. Pepsin digestion destroyed the antigen(s). The antibodies were of a different class, that is, the patients with systemic lupus erythematosus had IgG and IgA as well as IgM antibodies; the patients with periarteritis nodosa had IgM or IgG and IgA antibodies, while the patients with IgA-related nephritis had the highest recorded titers of IgA but also had IgG as well as IgM antibodies. None of the patients had antibodies directed against triple helical collagen. The antibody response in anti-GBM antibody-related nephritis, then, is different both with respect to antigen and antibody class and depends on the underlying disease syndrome. Anticorps anti-membrane basale glomerulaire: Specificite des anticorps dans differentes formes de glomerulonephrites. Les constituants de membranes basales glomerulaires humaines ont ete solubilises par digestion avec de la collagenase et de la pepsine ou par extraction avec de la guanidine-HCl directement ou apres digestion prealable par l'enzyme. Ces diverses preparations ont ete utilisees comme antigene dans la titration d'anticorps par essai immunosorbant relies par enzyme (ELISA) dans des serums de malades ayant un syndrome de Goodpasture, et de malades avec d'autres formes de glomerulonephrites, notamment lupus erythemateux dissemine, periarterite noueuse, et nephropathie a IgA. Les malades ayant un syndrome de Goodpasture avaient des titres eleves d'anticorps IgG reagissant tres fortement avec des produits de digestion de collagenase. L'antigene(s) etait seulement partiellement solubilise par extraction a la guanidine-HCl, etait detruit par la digestion par la pepsine et par reduction, et etait partiellement detruit par digestion a la trypsine. L'antigene(s) est probablement une proteine noncollagene. Les anticorps provenant de malades ayant d'autres formes de nephrites etaient essentiellement diriges contre des antigenes provenant d'extraits guanidine-HCl, tandis que l'antigene(s) n'etait pas solubilise par digestion a la collagenase. La digestion par la pepsine a detruit l'antigene(s). Les anticorps etaient de differentes classes, les malades ayant un lupus erythemateux dissemine avaient des IgG, des IgA, et des IgM comme anticorps; les malades ayant une periarterite noueuse avaient des anticorps IgM, IgG, IgA, alors que les malades ayant une nephrite a IgA avaient les titres les plus eleves d'anticorps IgA, mais avaient egalement des IgG et des IgM. Aucun des malades n'avait d'anticorps diriges contre du collagene a triple helice. La reponse anticorps dans la nephrite liee a un anticorps anti-GBM est donc differente par la classe d'antigenes ou d'anticorps en fonction du syndrome sous-jacent.