BetaB1-crystallin: identification of a candidate ciliary body uveitis antigen.
OBJECTIVE
Perineuclear anti-neutrophil cytoplasmic antibody (pANCA), a marker antibody present in 12% of patients with anterior uveitis, recognizes cytoplasmic antigens in the nonpigmented ciliary body epithelium, a probable site of immunologic reactivity in this inflammatory disease. In this study, a recombinantly isolated pANCA monoclonal antibody was used to identify the corresponding antigenic target(s) in the ciliary body.


METHODS
Proteins from microdissected eye bank ocular ciliary body tissue were used to identify the corresponding ANCA antigen. Parallel two-dimensional protein gels were used for simultaneous identification of candidate antigenic protein spots by Western blot analysis and as a source of material for proteomic analysis. Multiple independent methods including Western blot analysis, confocal microscopy, and RT-PCR were used to provide additional characterization of the candidate protein.


RESULTS
Proteomic analysis suggested that beta B1 (betaB1)-crystallin is the primary ciliary body antigen. The presence of betaB1-crystallin in the human ciliary body was confirmed by Western blot with a betaB1 specific anti-peptide antibody. Confocal microscopy revealed colocalization of the antigenic reactivity of both anti-betaB1 antibody and monoclonal pANCA. RT-PCR confirmed the presence of betaB1-crystallin RNA in the ciliary body tissues.


CONCLUSIONS
This study identified betaB1-crystallin as a new cytoplasmic ciliary body antigenic target of a marker autoantibody associated with uveitis. This characterization of betaB1-crystallin outside the lens raises questions about its extralenticular expression, intracellular role, and potential target of inflammation in uveitis.
Stempel
David
D
Department of Ophthalmology, University of California-Los Angeles, Jules Stein Eye Institute, 100 Stein Plaza, Los Angeles, CA 90095, USA.
Sandusky
Hallie
H
Lampi
Kirsten
K
Cilluffo
Marianne
M
Horwitz
Joe
J
Braun
Jonathan
J
Goodglick
Lee
L
Gordon
Lynn K
LK
D000595:Amino Acid Sequence; D000818:Animals; D019268:Antibodies, Antineutrophil Cytoplasmic / Q000276:immunology; D000911:Antibodies, Monoclonal; D001323:Autoantibodies / Q000276:immunology; D001324:Autoantigens / Q000276:immunology*; D015153:Blotting, Western; D002417:Cattle; D002924:Ciliary Body / Q000276:immunology*; D006801:Humans; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014606:Uveitis, Anterior / Q000276:immunology*; D038206:beta-Crystallin B Chain / Q000235:genetics / Q000276:immunology*
D019268:Antibodies, Antineutrophil Cytoplasmic; D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D001324:Autoantigens; D012333:RNA, Messenger; D038206:beta-Crystallin B Chain