Exposure and binding of selected immunodominant La/SSB epitopes on human apoptotic cells.
OBJECTIVE
Opsonization of apoptotic cells by autoantibodies bound to surface membrane-translocated La/SSB antigens may initiate tissue damage in the setting of congenital heart block. By injecting pregnant mice with human anti-La antibodies, we previously demonstrated the formation of IgG-apoptotic cell complexes in the developing mouse fetus; however, the binding of anti-La antibodies to human-specific epitopes could not be addressed. Accordingly, the objective of the current study was to delineate the epitope specificity of human La antibodies that are exposed on the surface of apoptotic cells.


METHODS
We used fluorescence microscopy and flow cytometry to assess the binding of human anti-La antibodies affinity purified against immunodominant epitopes of La to human cells undergoing spontaneous apoptosis, in a murine xenograft model in vivo and in cultured human fetal cardiocytes rendered apoptotic in vitro, respectively.


RESULTS
Anti-La antibodies bound to immunodominant epitopes of La within the NH(2)-terminus and the RNA recognition motif (RRM) region of apoptotic human cells, in both xenografts and fetal cardiocytes. In contrast, human antibodies affinity purified against the COOH-terminal La epitope did not bind apoptotic cells in either model. This defines the topology of redistributed La during apoptosis, with surface exposure of the NH(2)-terminus and RRM regions. The potential importance of anti-La NH(2)-terminal and anti-La RRM specificity was confirmed by detection of this reactivity in mothers of children with congenital heart block.


CONCLUSIONS
These findings provide insight into both the molecular modification of the La autoantigen during apoptosis and the specificity of antibodies capable of binding to surface-exposed La. Subsequent formation of surface immune complexes may lead to tissue injury in patients with autoimmune diseases such as congenital heart block.
Neufing
Petra J
PJ
Department of Immunology, Allergy and Arthritis, Flinders University, Adelaide, South Australia, Australia. petra.neufing@fmc.sa.gov.au
Clancy
Robert M
RM
Jackson
Michael W
MW
Tran
Hai Bac
HB
Buyon
Jill P
JP
Gordon
Tom P
TP
D000818:Animals; D000974:Antibodies, Antinuclear / Q000276:immunology; D017209:Apoptosis / Q000276:immunology*; D001324:Autoantigens / Q000737:chemistry / Q000276:immunology*; D002478:Cells, Cultured; D000939:Epitopes / Q000276:immunology*; D005260:Female; D006327:Heart Block / Q000151:congenital / Q000276:immunology*; D006801:Humans; D016056:Immunodominant Epitopes / Q000276:immunology*; D020287:In Situ Nick-End Labeling; D008180:Lupus Erythematosus, Systemic / Q000276:immunology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D032383:Myocytes, Cardiac / Q000166:cytology / Q000276:immunology* / Q000637:transplantation; D017434:Protein Structure, Tertiary; D012261:Ribonucleoproteins / Q000737:chemistry / Q000276:immunology*; D012859:Sjogren's Syndrome / Q000276:immunology; D014183:Transplantation, Heterologous; D000097585:SS-B Antigen
D000974:Antibodies, Antinuclear; D001324:Autoantigens; D000939:Epitopes; D016056:Immunodominant Epitopes; D012261:Ribonucleoproteins; D000097585:SS-B Antigen; C035357:SS-B antibodies