Chronic low level complement activation within the eye is controlled by intraocular complement regulatory proteins

PURPOSE. TO explore the role of the complement system and complement regula tory proteins in an immune-privileged organ, the rye. METHODS. Eyes Of normal Lewis rats were analyzed for the expression of comp lement regulatory proteins, membrane cofactor protein (MCP), decay-accelera tion factor (DAF), membrane inhibitor of reactive lysis (MIRL, CD59), and c ell surface regulator of complement (Crry) using immunohistochemistry, West ern blot analysis, and reverse transcription-polymerase chain reaction (RT- PCR). Zymosan, a known activator of the alternative pathway of complement s ystem was injected into the anterior chamber of the eye of Lewis rats. Anim als were also injected intracamerally with 5 mu l (25 mu g) Of neutralizing monoclonal antibody (mAb) against rat Crry (512) or CD59 (6D1) in an attem pt to develop antibody induced anterior uveitis; control animals received 5 mu l of sterile phosphate-buffered saline (PBS), OX-18 (25 mu g), G-16-510 E3 (25 mu g), or MOPC-21 (25 mu g). The role of complement system in antibo dy-induced uveitis was explored by intraperitoneal injection of 35 U cobra venom factor(CVF), 24 hours before antibody injection. Immunohistochemical staining and sodium dodecyl sulfate-poly acrylamide gel electrophoresis (SD S-PAGE) with Western blot analysis were used to detect the presence of memb rane attack complex (MAC) and C3 activation products, respectively, in norm al and antibody-injected rat eyes. RESULTS. Complement activation product MAC was present in the normal rat ey e, and intraocular injection of zymosan induced severe anterior uveitis. Th e complement regulatory proteins, MCP, DAF, CD59, and Crry, were identified in the normal rat eye. Soluble forms of Crry and CD59 were also detected i n normal rat aqueous humor. Severe anterior uveitis developed in Lewis rats injected with a neutralizing mAb against Crry, with increased formation of C3 split products. Systemic complement depletion by CVF prevented the indu ction of anterior uveitis by anti-Crry mAb. Intracameral injection of anti- rat CD59 (6D1), anti-rat MHC class I antigen (OX-18), anti-rat ig (G-16-510 E3), or MOPC-21 caused no inflammatory reaction. CONCLUSIONS. The results suggest that the complement system is continuously active at a low level in the normal eye and is tightly regulated by intrao cular complement regulatory proteins.