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pubmed:abstractText |
The nephritogenic antigen of Heymann's nephritis (HN), gp330, was previously demonstrated (4-9) to be a resident glycoprotein of coated pits in the glomerular and proximal tubule epithelium of rats, and anti-gp330 IgG given intravenously was found to form IDs in glomeruli (passive HN). The purpose of this study was to investigate the detailed events that occur in the formation of IDs in passive HN. HN was induced by the injection of either 125I-labeled or unlabeled anti-gp330 IgG. At various times after injection (15 min to 8 d) the kidneys of some of the injected rats were fixed by perfusion, and the distribution of the rabbit IgG was determined by immunofluorescence and by immunoelectron microscopy. Glomeruli were isolated from the kidneys of injected rats and used for isolation of GBM fractions or for elution of the bound IgG. At 15 min to 1 h after injection, the rabbit IgG was localized by immunocytochemistry exclusively in coated pits along the podocyte plasmalemma facing the GBM. By 1-8 d, anti-gp330 IgG was detected in larger electron-dense IDs often located under the slit diaphragms. Serial sectioning revealed that each of the IDs maintained contact with a coated pit at some level. When GBMs isolated from rats given radiolabeled anti-gp330 IgG were examined by electron microscopy, the IDs were found to remain attached to the GBMs as early as 15 min after injection and coisolated with them at all time points. By double-immunolabeling of the isolated GBMs with two sizes of gold particles, both the antigen (gp330) and the anti-gp330 IgG could be demonstrated in IDs at all time points. When the amount of radiolabeled anti-gp330 bound to GBM fractions was compared with that of isolated glomeruli, it was found that 20% of the radiolabel remained bound to the purified GBMs at 15 min after injection, and 90% at 3 d. The bound IgG was released only by treatments that disrupt antibody-antigen complexes (high and low pH), but not by the other treatments we tried (detergent, high salt, heparinase, or collagenase digestion). When the IgG bound to glomeruli was eluted with acid citrate buffer 3 d after injection, it was found to specifically immunoprecipitate only gp330 from detergent-solubilized 125I-labeled kidney microvillar vesicles. By isoelectric focusing the eluate was found to be enriched in IgGs with acidic isoelectric points.(ABSTRACT TRUNCATED AT 400 WORDS)
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