Chondroitin sulfate proteoglycan core proteins in the interphotoreceptor matrix: A comparative study using biochemical and immunohistochemical analysis

This study characterizes the core proteins of chondroitin sulfate-type glyc osaminoglycans located in the interphotoreceptor matrix and establishes the tissue distribution of chondroitin immunoreactivity in human, bovine, mous e and rat retinas, Monoclonal antibodies specific to unsulfated (Delta DiOS ), 4-sulfated (Delta Di4S) and 6-sulfated (Delta Di6S) chondroitin were emp loyed. Retinal sections and IPM samples were either (a) digested with chond roitinase ABC to expose antibody specific epitopes, (b) double digested wit h chondroitinase ABC and chondroitinase AC II to remove specific epitopes, or (c) left undigested to evaluate mimotope labeling. In tissue sections fr om each species studied, positive immunoreactivity to the Delta Di6S antibo dy was present in the IPM surrounding both rods and cones. In human and bov ine, Delta Di6S labeling of the cone matrix compartments was more intense t han labeling of the matrix surrounding rods, Intense Delta Di6S immunoreact ivity was present surrounding the foveal cones. In mouse and rat, no differ ences in labeling intensity of IPM surrounding rod and cone photoreceptors were evident. although labeling of the IPM near the apical surface of the r etinal pigment epithelium and around the photoreceptor inner segments was m ore pronounced than that surrounding the outer segments. All Delta Di6S ant ibody labeling was eliminated with chondroitinase AC LI digestion. No IPM i mmunoreactivity in tissue sections was observed when the Delta DiOS or Delt a Di4S antibodies were used. In Western blots of IPM extracts treated with chondroitinase ABC, prominent Delta Di6S immunoreactive bands were present at approximately 230 kD and 150 kD in each species studied, with the except ion of the human, where the 150 kD component is not a chondroitin proteogly can. Each of the prominent Delta Di6S immunoreactive bands showed minor imm unoreactivity to the Delta Di4S antibody. No Delta DiOS immunoreactivity wa s noted in Western blots of IPM samples from any species. All immunoreactiv ity was lost following chondroitinase AC II digestion. These observations d ocument similarities in the electrophoretic mobility of IPM proteoglycan co re proteins released following chondroitinase ABC digestion in the four spe cies studied, but reveal pronounced differences in the tissue distribution. Bovine and human IPM show greater concentrations of Delta Di6S immunoreact ivity surrounding cones than rods, whereas rodent tissues show higher conce ntrations near the retinal pigment epithelium and around the photoreceptor inner segments than around the outer segments. The pattern of distribution of these proteoglycan molecules is highly conserved in these species, sugge sting a common role in IPM structure and function, (C) 1999 Academic Press.