Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
Plasmin Cleavage of Beta2-Glycoprotein I Promotes Maternal Anti-Ro60 IgG Opsonisation of Apoptotic Cells: A Permissive Factor in Congenital Heart Block?
Reed1, Joanne H., Giannakopoulos2, Bill, Kaufman3, Kenneth M., Krilis2, Steven A., Gordon1, Thomas P.
Flinders Medical Centre, Bedford Park, Australia
University of New South Wales, Sydney, Australia
Oklahoma Medical Research Foundation, Oklahoma City, OK
Purpose:
Maternal anti-Ro60 autoantibodies are thought to initiate tissue damage in congenital heart block (CHB) by binding to apoptotic fetal cardiocytes. The plasma protein beta2-glycoprotein I (b2GPI) binds to Ro60 (amino acid residues 82–244) on the surface of apoptotic cells via its fifth domain and prevents the formation of pathogenic anti-Ro60 IgG-apoptotic cell immune complexes. The current study was initiated to define the precise Ro60-b2GPI binding sites and determine whether plasmin mediated cleavage of b2GPI influences immune complex formation.
Method:
The b2GPI binding site on Ro60 was mapped by ELISA using overlapping soluble recombinant Ro60 subfragments spanning amino acids (aa) 82–244. The Ro60 binding on b2GPI was studied by apoptotic cell flow cytometry inhibition experiments using b2GPI peptides and plasmin-cleaved b2GPI. Binding of anti-Ro60 IgG from mothers of infants with CHB to apoptotic cells was determined by flow cytometry in the presence or absence of plasmin-cleaved b2GPI.
Results:
The Ro60 b2GPI-binding site was mapped to a 64 aa region within the middle third of the Ro60 protein. A peptide representing a charged region within domain V (CKNKEKKC) of b2GPI partially inhibited binding to Ro60 on apoptotic cells. Plasmin cleavage of the b2GPI hydrophobic loop (Lys317-Thr318) abrogated binding of b2GPI to both recombinant and apoptotic cell membrane-bound Ro60 by flow cytometry. Maternal anti-Ro60 IgG opsonised apoptotic cells in the presence of plasmin cleaved b2GPI under conditions where an equivalent concentration of native b2GPI completely inhibited immune complex formation.
Conclusion:
b2GPI interacts with a 64 aa region of Ro60 via its hydrophobic loop and adjacent positively charged lysine rich region in domain V. Stimulation of plasmin production by infection or inflammation in the fetal heart may eliminate the protective effect of b2GPI and open the way for maternal anti-Ro60 autoantibody-mediated tissue damage.
To cite this abstract, please use the following information:
Reed, Joanne H., Giannakopoulos, Bill, Kaufman, Kenneth M., Krilis, Steven A., Gordon, Thomas P.; Plasmin Cleavage of Beta2-Glycoprotein I Promotes Maternal Anti-Ro60 IgG Opsonisation of Apoptotic Cells: A Permissive Factor in Congenital Heart Block? [abstract]. Arthritis Rheum 2009;60 Suppl 10 :678
DOI: 10.1002/art.25758
