Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement

The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.

Reactivity with Dichotomous Determinants of Ro60 Stratifies Autoantibody Responses in Lupus and Primary Sjogrens Syndrome

Reed1,  Joanne H., Dudek2,  Nadine L., Osborne1,  Shannon E., Kaufman3,  Kenneth M., Jackson1,  Michael W., Purcell2,  Anthony W., Gordon1,  Thomas P.

Flinders Medical Centre, Bedford Park, Australia,
University of Melbourne, Melbourne, Australia,
Oklahoma Medical Research Foundation, Oklahoma City, OK


Flow cytometric-based analysis of B-cell epitopes of Ro60 exposed on the surface of apoptotic cells (apotopes) and intracellular epitopes provides insight into the structural forms of Ro60 that break immune tolerance. The aim of this study was to fine map a Ro60 apotope that is specific for a subset of systemic lupus erythematosus (SLE) and determine whether differentially expressed forms of this determinant stratify anti-Ro60 responses in SLE and primary Sjögren's syndrome (SS).


Ro60 apotopes were mapped by multi-parameter flow cytometry on early apoptotic cells by inhibition experiments using soluble recombinant fragments of Ro60. Anti-Ro60 IgG from patients with SLE (n=32) and primary SS (n=26) were also assessed for reactivity with the immobilised Ro60 fragments by ELISA or immunoblot. Anti-Ro60 IgG was eluted from early apoptotic cells or recombinant Ro60 immobilised on nitrocellulose and binding to membrane-bound and intracellular forms of Ro60 quantitated by flow cytometry. Three-dimensional modelling of immunoreactive domains was based on the crystal structure of Xenopus laevis Ro60.


A homology model was constructed using the crystal structure of X. laevis Ro60 and the apotope mapped to a prominent helix-loop-helix region or peg-like region on the surface of human Ro60. This immunodominant structure was recognised by patients with SLE and isolated anti-Ro. Immobilisation of the same domain to a solid phase revealed a second determinant (epitope) that was specific for linked anti-Ro and anti-La-responses. Virtually all patients recognising the Ro60 apotope were negative on the epitope and vice versa. Affinity-purification experiments revealed that the apotope and epitope were expressed in a mutually exclusive fashion on membrane-bound and intracellular forms of the autoantigen, respectively.


Dichotomous determinant recognition within a single domain of Ro60 stratifies anti-Ro60 responses in SLE and primary SS and supports a model of anti-Ro60 autoimmunity in which tolerance is broken by different immunogenic forms of the autoantigen.

To cite this abstract, please use the following information:
Reed, Joanne H., Dudek, Nadine L., Osborne, Shannon E., Kaufman, Kenneth M., Jackson, Michael W., Purcell, Anthony W., et al; Reactivity with Dichotomous Determinants of Ro60 Stratifies Autoantibody Responses in Lupus and Primary Sjogrens Syndrome [abstract]. Arthritis Rheum 2009;60 Suppl 10 :1987
DOI: 10.1002/art.27060

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