COMPARISON OF ANTI-GBM ANTIBODIES IN SERA WITH OR WITHOUT ANCA

An appreciable percentage of patients with serum anti-glomerular basem ent membrane (anti-GEM) antibodies also have antineutrophil cytoplasmi c antibodies (ANCA), against either myeloperoxidase (MPO-ANCA), or pro teinase 3 (PR3-ANCA). In sera without ANCA, the anti-GEM antibodies ha ve been shown to react mainly with the noncollagenous domain (NC1) of Type IV collagen, and especially with its alpha 3 chain, alpha 3(IV)NC 1. In most sera, the antibodies can be partially blocked by a monoclon al antibody (Mab 17) against alpha 3(IV)NC1, suggesting that a limited region is recognized. Although there is evidence that some anti-GEM a ntibodies that coexist with ANCA react with alpha 3(IV)NC1, extensive analysis of the specificity of such anti-GEM antibodies has not been r eported. In the study presented here, sera were analyzed from 332 pati ents tested both for anti-GEM antibodies and ANCA (MPO or PR3-ANCA) an d found to have one or more positive tests. Of the 100 sera with anti- GEM antibodies, 38 also had ANCA-25 with MPO-ANCA (66%), 12 with PR3-A NCA (32%), and one with both (2%). Of the 232 sera with ANCA only, 153 had MPO-ANCA (66%), 75 had PR3-ANCA (32%), and four had both (2%). Se ra was also analyzed from 259 other patients who had positive ANCA tes ts and were not tested for anti-GEM antibodies: 138 had MPO-ANCA (54%) , and 121 had PR3-ANCA (46%). The relative frequencies of MPO or PR3-A NCA in patients with coexisting anti-GEM antibodies did not differ sig nificantly from those in all patients with ANCA (P = 0.35). Seventeen sera with anti-GEM antibodies only and 16 sera with anti-GEM antibodie s plus ANCA were selected for further studies to compare the specifici ty of anti-GEM antibodies in sera with or without ANCA. Using enzyme-l inked immunosorbent assays (ELISA), all sera in both groups were found to react with the NC1 domain (as a hexamer) of bovine Type IV collage n and with alpha 3(IV)NC1 monomers. Furthermore, all but six sera also reacted with one or more of the alpha 1, 2, and 4 (IV)NC1 monomers, g enerally with considerably lower titers. Reactivity to alpha 3(IV)NC1 was partially blocked by Mab17, with comparable degrees of inhibition in both groups. Western blot analysis with the human NC1 domains revea led no differences in reactivity between the two groups. Thus, differe nces in antigen specificities of anti-GEM antibodies in sera with or w ithout ANCA were not detected. The anti-GEM response in both situation s is hypothesized to be driven by the same immunogen, which is probabl y derived from NC1 domains of endogenous Type IV collagen.