RPE SECRETED PROTEINS AND ANTIBODY INFLUENCE PHOTORECEPTOR CELL-SURVIVAL AND MATURATION

Proteins in media conditioned by retinal pigment epithelial cells (RPE -CM) and an antibody against these proteins (RPE-SP) were tested for t heir respective effects on rat retinal development in vitro and in viv o. Proteins of RPE-CM were separated in denaturing gels and evaluated by Western blot analysis. Retinal explants from postnatal day 2 (P2) r ats were cultured in RPE-CM only or CM diluted with the RPE-SP antibod y and, after 7 days, the explants were dissociated into single cells t hat were immunostained for opsin. RPE-CM or antibody was also injected into the vitreous of postnatal day 7 (P7) Long-Evans rats and analyze d 7 and 21 days later. Electrophoretic analysis of RPE-CM predominantl y showed 60-70 kDa proteins; when these proteins were probed with RPE- SP antibody by Western blot, immunoreactive proteins were restricted t o this narrow molecular weight range. In P2 retinal explant cultures s upplemented with RPE-CM, long ganglion cell-like neurites were detecte d in 3 days. This activity was nullified in explant cultures grown in RPE-CM titrated with antibody, and these explants appeared to degenera te within 5 days. Over 80% of dissociated retinal cells from explants 7 days after treatment with RPE-CM expressed opsin, compared to only 2 0% of cells from explants grown in defined medium or serum. Retinas of P14 rats injected intravitreally with RPE-CM at P7 had increased numb ers of ectopic photoreceptor cells within the inner nuclear layer when compared to retinas of sham-injected eyes. In contrast, retinas of ey es injected intravitreally with RPE-SP antibody exhibited shorter oute r (OS) and inner (IS) segments and thinner outer nuclear (ONL) and out er plexiform (OPL) layers than retinas of sham-injected eyes. In concl usion, proteins in RPE-CM appeared to accelerate and maximize the deve lopment of rat photoreceptor cells in vitro, while intravitreal inject ions of its antibody caused an apparent retardation of outer segment m aturation. These results suggest that a protein(s) secreted by RPE pla ys a key role in normal retinal development, particularly in photorece ptor cell survival and outer segment maturation. (C) 1998 Elsevier Sci ence B.V.