Glycobiological Study of Physiological Roles of Interphotoreceptor Matrix

(a)视网膜移植成功的实验条件研究：免疫组化研究显示，持续荧光照射诱导的大鼠视网膜增殖细胞中存在巨噬细胞。用亲和层析法从牛视网膜中纯化了两种凝集素。利用抗凝集素的多克隆抗体对牛视网膜cDNA文库进行筛选，分离出凝集素的cDNA。序列分析显示15kD-和35kd -凝集素分别对应于凝集素和未知凝集素。我们将使用这些视网膜凝集素代替植物凝集素进行视网膜移植实验。(b)光感受器间基质中o型糖缀合物介导的视网膜粘附机制研究：免疫组织化学研究显示，视网膜中不存在MUC2。提示MUC2与视网膜粘连无关。利用多克隆抗体对牛视网膜cDNA文库进行筛选，分离出o型糖缀合物核心蛋白的cDNA。序列分析表明，该核心蛋白富含丝氨酸和苏氨酸。我们将利用cDNA表达的融合蛋白进行视网膜粘附的研究。(c)光感受器间基质中n型糖缀合物的光反应研究：使用含有戊二醛的固定剂检测光反应，而在光和超显微镜研究中，仅使用多聚甲醛进行大鼠灌注固定时未检测到光反应。这些结果表明，n型糖缀合物可能在视网膜下空间中移动而不桥接光感受器间基质。

(a) Study of experimental conditions for successful retinal transplantation : Immunohistochemical studies revealed that macrophages are present in the proliferating cells of the rat retina induced by constant fluorescence light exposure. Two lectins were purified from bovine retinas by affinity chromatography using a lactose column. We isolated the cDNAs of the lectins by screening bovine retinal cDNA libraly using the polyclonal antibodies against the lectins. The sequence analysis revealed that the 15kD- and 35kD-lectins correspond to galectin and unknown lectin, respectively. We will use these retinal lectins for the experiments of retinal transplantation instead of plant lectins.(b) Study of mechanism of retinal adhesion mediated by the O-type-glycoconjugates in the interphotoreceptor matrix : Immunohistochemical studies revealed that MUC2 is not present in the retina. This result suggests that MUC2 is not related to retinal adhesion. We isolated the, cDNA of a core protein of the O-type-glycoconjugate by screening bovine retinal cDNA libraly using polyclonal antibodies. The sequence analysis revealed that the core protein is rich in serine and threonine. We will use the fusion protein from expression of the cDNA for the study of retinal adhesion.(c) Study of light-response of the N-type-glycoconjugates in the interphotoreceptor matrix : The light- response was detected by use of a fixative containing glutaraldehyde, whereas it was not deteted using paraformaldehyde alone for the perfusion-fixation of rats in light- and ultra-microscopic sudies. These results suggest that the N-type-glycoconjugates may move in the subretinal space without bridging of the interphotoreceptor matrix.