Glycobiological Study of Physiological Roles of Interphotoreceptor Matrix

(A)成功进行视网膜移植的实验条件研究：免疫组织化学研究显示，在持续的荧光光照射下，大鼠视网膜的增殖细胞中存在巨噬细胞。用乳糖柱亲和层析从牛视网膜中分离纯化了两种凝集素。我们用抗凝集素的多克隆抗体筛选牛视网膜cDNA文库，从而分离出凝集素的cDNAs。序列分析表明，15kD和35kD凝集素分别对应于半乳糖凝集素和未知凝集素。我们将用这些视网膜凝集素代替植物凝集素用于视网膜移植的实验。(B)光感受器基质中O型糖偶联物介导的视网膜粘连机制的研究：免疫组织化学研究显示，MUC2不存在于视网膜中。这一结果表明，MUC2与视网膜粘连无关。我们用多克隆抗体筛选牛视网膜c DNA文库，获得了O型糖偶联物核心蛋白的c DNA。序列分析表明，该核心蛋白富含丝氨酸和苏氨酸。我们将利用表达的融合蛋白来研究视网膜粘连。(C)光感受器基质中N型糖偶联物的光反应的研究：光反应是用含有戊二醛的固定剂检测的，而不是用多聚甲醛单独用来灌流固定在光镜和超显微镜下的大鼠。这些结果表明，N型糖偶联物可能在视网膜下间隙中移动，而不需要光感受器间基质的桥接。

(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.