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F Gonzalez-Fernandez, M Bussard, SS Agersborg, KS K Tung; Epitope Spreading and Immune Complex Deposition Following IRBP Peptide Immunization . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1531.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To evaluate the antibody response and eye immunopathology in mice with autoimmune uveitis induced by immunization with human IRBP peptide 161-180 (pIRBP). Methods: B10RIII mice were immunized with CFA only, CFA/PTX, CFA/ human pIRBP161-180, or CFA/ human pIRBP161-180/PTX, and sacrificed 21 days later. Sera and eyes were evaluated using western blot analysis, light and electron microscopy, direct IF for IgG1, IgG2a, IgG2b, IgG3, IgM, IgA and C3, and indirect IF and ELISA assay for antibody to IRBP. Results: Autoimmune uveitis was induced in mice injected with pIRBP in CFA and PTX but not with pIRBP in CFA. However, both immunization protocols elicited serum antibody to mouse IRBP as demonstrable by indirect IF on normal eyes and eyes from IRBP -/- mice used as controls. Preadsorption of the mouse sera with IPM from IRBP +/+ mice or native bovine IRBP, but not IPM from IRBP -/- mice or pIRBP blocked binding of serum IRBP antibodies to the retina by IF or ELISA. However, no significant reduction in signal was apparent using pIRBP at even molar equivalents 10,000 fold higher than that at which mouse or bovine IRBP demonstrated ∼50% reduction in signal. Direct IF and EM demonstrated that animals immunized with CFA/pIRBP with or without PTX had extensive immune complex depositions (granular IgG1, IgG2a, IgG2b, C3) in Bruch's membrane, the corneal endothelium, trabecular meshwork, and ciliary body. Conclusion: Immunization with a restricted peptide domain of IRBP elicits autoantibody response to epitopes of the IRBP distant from the immunizing peptide. Such immunization induces extensive deposition of immune complexes in portions of the eye where IRBP is not usually present. The eye immune complexes are detected in mice with or without uveitis. Therefore, IRBP may not be sequestered in the eye.
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