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*Lupus
The Journal of Immunology, 2002, 169: 4046-4053.
Copyright © 2002 by The American Association of Immunologists

Orderly Pattern of Development of the Autoantibody Response in (New Zealand White x BXSB)F1 Lupus Mice: Characterization of Target Antigens and Antigen Spreading by Two-Dimensional Gel Electrophoresis and Mass Spectrometry1

Sandrine Thébault*, Danièle Gilbert*, Marie Hubert{dagger}, Laurent Drouot*, Nadine Machour*, Catherine Lange{dagger}, Roland Charlionet* and François Tron2,*

* Institut de la Santé et de la Recherche Médicale Unité 519, Faculté de Médecine et de Pharmacie, Hôpital Charles-Nicolle, and {dagger} Laboratoire de Spectroscopie, Institut Fédératif de Recherches Multidisciplinaires sur les Peptides, Institut de Rechereche en Chimie Organique Fine, Rouen, France.

Immunoblots of a two-dimensional PAGE-separated HL-60 cell proteomic map and mass spectrometry were combined to characterize proteins targeted by autoantibodies produced by male (New Zealand White x BXSB)F1 (WB) mice that develop lupus and anti-phospholipid syndrome. Analysis of sera sequentially obtained from seven individual mice at different ages showed that six proteins, vimentin, heat shock protein 60, UV excision-repair protein RAD23, {alpha}-enolase, heterogeneous nuclear ribonucleoprotein L, and nucleophosmin, were the targets of the B cell autoimmune response, and that autoantibodies to them were synthesized sequentially in an orderly pattern that recurred in all the male WB mice analyzed: anti-vimentin first and anti-nucleophosmin last, with anti-RAD23 and anti-heat shock protein 60, then anti-{alpha}-enolase and anti-heterogeneous nuclear ribonucleoprotein L Abs occuring concomitantly. Anti-vimentin reactivity always appeared before anti-cardiolipin and anti-DNA Abs, suggesting that vimentin is the immunogen initiating the autoimmune process. The pattern of HL-60 proteins recognized by female WB sera differed from that of male sera, indicating that the Y chromosome-linked autoimmune acceleration gene is not an accelerator but a strong modifier of the autoimmune response. Thus, 1) combining two-dimensional PAGE and mass spectrometry constitutes a powerful tool to identify the set of Ags bound by autoantibodies present in a single serum and the whole autoantibody pattern of an autoimmune disease; 2) the diversification of the autoimmune response in male WB mice occurs in a predetermined pattern consistent with Ag spreading, and thus provides a useful model to further our understanding of the development of the autoantibody response in lupus.




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