B Cell Trafficking to the Eye

项目摘要 自身免疫性后葡萄膜炎是一种影响后眼的炎症性眼病，治疗困难 并且通常导致视力丧失。我们工作的总体目标是改善临床结果， 有这种情况的病人。后葡萄膜炎的特征是在后葡萄膜内的异质性白细胞浸润。 视网膜及邻近组织。以前有关疾病发病机制的研究集中在CD 4 + T细胞 和单核细胞。涉及B细胞的机制很少受到关注，尽管新的治疗方法 对葡萄膜炎患者有治疗作用。这一探索性项目将启动一个新的生产线， 研究B细胞在自身免疫性后葡萄膜炎中的作用，最初关注B细胞迁移到 视网膜。该建议的中心假设是，自身免疫性后葡萄膜炎中B细胞进入视网膜 受到视网膜血管内皮上不同粘附分子的严格调节。一种新的小鼠B细胞模型 提出了将细胞运输到眼睛的方法，其中有效的B细胞趋化因子CXCL 13将被递送到眼睛。 通过眼内注射编码趋化因子的慢病毒，将测试多种条件， 优化免疫方案以获得最大的B细胞流入。转基因表达将通过 ELISA和免疫组织化学，以及炎症将通过局部内窥镜眼底成像进行监测 和组织病理学。B细胞-视网膜内皮细胞相互作用的平行研究将使用人类 系统，包括：使用原代人内皮细胞的视网膜内皮细胞迁移测定;和 使用完整人视网膜的改良Woodruff-Stamper测定。选择的粘附分子的作用- 细胞间粘附分子（ICAM-1）、血管细胞粘附分子（VCAM-1）与活化白细胞 细胞粘附分子（ALCAM）-在B细胞运输到视网膜中将在鼠模型中进行评价， 基于人的测定，使用适当的方法特异性抑制粘附分子-配体结合。

PROJECT SUMMARY Autoimmune posterior uveitis is an inflammatory eye disease affecting the posterior eye that is difficult to treat and commonly results in vision loss. The overall objective of our work is to improve the clinical outcome for patients with this condition. Posterior uveitis is characterized by a heterogeneous leukocytic infiltrate within the retina and adjacent tissues. Previous research relating to disease pathogenesis has centered on CD4+ T cells and monocytes. Mechanisms involving B cells have received little attention, although new treatments directed against these cells are therapeutic for patients with uveitis. This exploratory project will initiate a new line of research on the role of the B cell in autoimmune posterior uveitis, with initial focus on B cell migration to the retina. The central hypothesis of the proposal is that B cell entry into the retina in autoimmune posterior uveitis is strictly regulated by distinct adhesion molecules on retinal vascular endothelium. A novel murine model of B cell trafficking to the eye is proposed, in which the potent B cell chemokine, CXCL13, will be delivered to the posterior eye by intraocular injection of chemokine-encoding lentivirus. Multiple conditions will be tested to optimize the immunization protocol for maximum influx of B cells. Transgene expression will be monitored by ELISA and immunohistochemistry, and inflammation will be monitored by topical endoscopic fundus imaging and histopathology. Parallel studies of B cell-retinal endothelial interactions will be conducted using human systems, including: a retinal endothelial cell transmigration assay with primary human endothelial cells; and a modified Woodruff-Stamper assay using intact human retina. The role of selected adhesion molecules - intercellular adhesion molecule (ICAM)-1, vascular cell adhesion molecule (VCAM)-1 and activated leukocyte cell adhesion molecule (ALCAM) - in B cell trafficking to retina will be evaluated in the murine model and the human-based assays, using appropriate methods to specifically inhibit adhesion molecule-ligand binding.