The Journal of Experimental Medicine
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© The Rockefeller University Press, 0022-1007/1996/12/2231/ $5.00
The Journal of Experimental Medicine, Volume 184, Number 6, December 1, 1996 2231-2242

Articles

Antineutrophil Cytoplasmic Autoantibodies Interact with Primary Granule Constituents on the Surface of Apoptotic Neutrophils in the Absence of Neutrophil Priming

Hannah M. Gilligan*,{ddagger}, Brunel Bredy*,{ddagger}, Hugh R. Brady||, Marie-Josée Hébert, Henry S. Slayter**, Yuhui Xu**, Joyce Rauch{dagger}{dagger}, Michael A. Shia*,§, Jason S. Koh*,{ddagger}, and Jerrold S. Levine*,{ddagger}

From the * Renal Section, Evans Memorial Department of Clinical Research, and {ddagger} Department of Medicine and § Department of Biochemistry, Boston University Medical Center Hospital, Boston, Massachusetts 02118; || Department of Medicine and Therapeutics, University College Dublin, Mater Miseracordiae, Dublin 7, Ireland; Renal Division, Department of Medicine, Brigham and Women's Hospital, and the Renal Section, Medical Service, Brockton-West Roxbury Department of Veterans Affairs Medical Center, Harvard Medical School, Boston, Massachusetts 02132; ** Laboratories of Electron Microscopy and Structural Molecular Biology, Dana Farber Cancer Institute, and Department of Cellular and Molecular Physiology, Harvard Medical School, Boston, Massachusetts 02132; and {dagger}{dagger} Division of Rheumatology, Montreal General Hospital Research Institute, McGill University, Montreal, QC, Canada H3G 1A4

The pathogenic role of antineutrophil cytoplasmic autoantibodies (ANCA) remains controversial because of the difficulty in explaining how extracellular ANCA can interact with intracellular primary granule constituents. It has been postulated that cytokine priming of neutrophils (PMN), as may occur during a prodromal infection, is an important trigger for mobilization of granules to the cell surface, where they may interact with ANCA. We show by electron microscopy that apoptosis of unprimed PMN is also associated with the translocation of cytoplasmic granules to the cell surface and alignment just beneath an intact cell membrane. Immunofluorescent microscopy and FACS® analysis demonstrate reactivity of ANCA-positive sera and antimyeloperoxidase antibodies with apoptotic PMN, but not with viable PMN. Moreover, we show that apoptotic PMN may be divided into two subsets, based on the presence or absence of granular translocation, and that surface immunogold labeling of myeloperoxidase occurs only in the subset of PMN showing translocation. These results provide a novel mechanism that is independent of priming, by which ANCA may gain access to PMN granule components during ANCA-associated vasculitis.

Address correspondence to Jerrold S. Levine, Renal Section, E428, Boston University Medical Center Hospital, 88 East Newton Street, Boston, Massachusetts, 02118.

This work was supported by National Institutes of Health grants AR/AI42732 (J.S. Levine), DK44380 (H.R. Brady), and DK45047 (M.A. Shia); a Young Investigator Award from the National Kidney Foundation (J.S. Levine); a Research Unit Grant from the Health Research Board, Ireland (H.R. Brady); The Arthritis Society of Canada (J. Rauch); The Medical Research Council of Canada (J. Rauch); and The Evans Department of Clinical Research, Boston University Medical Center.

1Abbreviations used in this paper: ANCA, antineutrophil cytoplasmic autoantibodies; C-ANCA, ANCA with cytoplasmic staining pattern on indirect immunofluorescence; CHX, cycloheximide; EM, electron microscopy; GBM, glomerular basement membrane; IF, immunofluorescence; MPO, myeloperoxidase; P-ANCA, ANCA with perinuclear staining pattern on indirect immunofluorescence; PI, propidium iodide; PR3, proteinase 3.


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