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Identification of MHC Class I H-2 K^b/D^b-Restricted Immunogenic Peptides Derived from Retinal Proteins

¹From the Laboratory of Cellular Immunology, Department of Medical Anatomy, and the ³Institute of Medical Microbiology and Immunology, The Panum Institute, and the ⁴Eye Pathology Institute, University of Copenhagen, Copenhagen, Denmark; and the ²Department of Immunology, College of Basic Medicine, Anhui Medical University, Hefei, China.

PURPOSE. To identify H-2 K^b/D^b-binding immunogenic peptides derived from retinal proteins.

METHODS. Computer-based prediction was used to identify potentially H-2 K^b/D^b-binding peptides derived from the interphotoreceptor retinol-binding protein (IRBP), soluble retinal antigen (S-antigen), recoverin, phosducin, and pigment epithelium-derived factor (PEDF). The affinity of the peptides was analyzed by their abilities to upregulate the expression of major histocompatibility complex (MHC) class I on TAP-deficient cells (RMA-S cells) with flow cytometry. C57BL/6 mice were immunized subcutaneously, with individual peptides in incomplete Freund’s adjuvant (IFA). Eight days after immunization, splenocytes were isolated for cytotoxic T-lymphocyte (CTL) analysis. A ⁵¹chromium-release assay was used to detect specific CTL reactivity generated in the cultures. Eyes were enucleated for histopathological analysis on day 21 after immunization with IRBP or IRBP and the immunogenic peptides.

RESULTS. All the 21 predicted peptides were found to upregulate expression of H-2 K^b/D^b on RMA-S cells. Five peptides, the two IRBP-derived peptides IRBP_89-96 and IRBP_101-108, and the three PEDF-derived peptides, PEDF_389-397, PEDF_139-147, and PEDF_272-279, induced specific CTL responses in vivo, whereas the remaining 16 peptides, including 5 IRBP-derived peptides, 5 S-antigen-derived peptides, 1 recoverin-derived peptide, 1 phosducin-derived peptide, and 4 PEDF-derived peptides, did not induce specific CTL reactivity. The immunogenic peptides alone did not induce inflammation in the eyes, but they could enhance severity of uveitis induced by IRBP.

CONCLUSIONS. Five of 21 H-2 K^b/D^b-binding retinal protein-derived peptides were found to be immunogenic, suggesting that these peptides could function as autoantigenic epitopes in the development of inflammatory eye diseases, such as uveitis.

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