ENDOTHELIAL-BINDING LECTINS OF LYMPHOID MALIGNANCIES

本项目主要研究人淋巴细胞与淋巴细胞之间的粘附相互作用 细胞和外周淋巴结高内皮微静脉（PNHEV） 其启动循环细胞迁移到组织中。在啮齿类动物中， 这一过程有助于正常淋巴细胞的运输， 淋巴恶性肿瘤的扩散。首席调查员 提出淋巴细胞表面的凝集素样结构， 磷酸甘露聚糖结合活性（PMRad）介导这种相互作用。在 鼠系统，PMRad和mel 14抗原，一个独立表征的 90 kd的粘附结构，或者紧密相连或者相同。在 人PMRad与爱马仕抗原的关系，85-95 kd Mel 14抗原的类似物是未知的。此外， PMRad与充分表征的磷酸甘露聚糖受体介导的递送 酸性水解酶对溶酶体的作用（PMRup）尚不清楚， 确立了习这些识别结构的比较分析是 由于无法获得纯化的PMRad和缺乏抗体， 对关键的配体结合域具有特异性。私家侦探。因此，在本发明中， 建议开发特异性针对 淋巴细胞凝集素的碳水化合物结合域。初步研究表明， 鉴定了一系列培养的T淋巴母细胞恶性肿瘤， PMRad、PMRup的组成型或诱导型表达以及与PNHEV的结合。 这些细胞系将用于：（1）产生阻断抗体连接的Mab。 将磷甘露聚糖衍生的荧光珠与PMRad或PMRup连接;（2）产生 在宽密度范围内表达表面凝集素的克隆;（3） 确定完整的PMRad和PMRup之间的功能关系 细胞使用多参数荧光细胞仪和直接 定量与PNHEV的结合;（4）提取并比较凝集素 结构;（5）评估PMRad、爱马仕 抗原和完整细胞中的Mel 14交叉反应物质，和（6）提取物 并在结构上比较这些独立表征的PNHEV粘附性 结构.研究（5）和（6）也将在白血病中进行 标本，以确定是否新鲜分离造血恶性肿瘤 利用与正常细胞和培养细胞相同的PNHEV粘附受体， 建立表面标志物预测和定量的能力 对PNHEV的亲和力，并提高检测 粘合结构之间的功能或结构连接。

This project focuses on the adhesive interaction between human lymphoid cells and the high endothelial venules of peripheral lymph nodes (PNHEV) which initiates the migration of circulating cells into tissue. In rodents, this process contributes to the trafficking of normal lymphocytes and the dissemination of lymphoid malignancies. The principle investigator has proposed that a lectin-like structure at the lymphocyte surface with phosphomannan-binding activity (PMRad) mediates this interaction. In the murine system, PMRad and the mel 14 antigen, an independently characterized 90 kd adhesion structure, are either closely linked or identical. In humans, the relationship between PMRad and the hermes antigen, the 85-95 kd analog of the mel 14 antigen, is unknown. Furthermore, the relationship of PMRad to the well characterized phosphomannan receptors mediating delivery of acid hydrolases to the lysosome (PMRup) has not been clearly established. Comparative analyses of these recognition structures are hampered by the unavailability of purified PMRad and the lack of antibodies specific for the crucial ligand-binding domains. The PI. therefore, proposes development of monoclonal antibodies (Mabs) specific for the carbohydrate-binding domains of lymphocytic lectins. Initial studies have identified a series of cultured T-lymphoblastic malignancies with either constitutive or inducible expression of PMRad, PMRup and binding to PNHEV. These lines will be used to: (1) generate Mabs which block attachment of phosphomannan-derivatized fluorescent beads to PMRad or PMRup; (2) generate clones expressing the surface lectins over a broad range of densities; (3) determine the functional relationships between PMRad and PMRup in intact cells using multiparameter fluorescence cytometry and the direct quantitation of binding to PNHEV; (4) extract and compare the lectins structurally; (5) assess the functional linkages between PMRad, the Hermes antigen and Mel14 cross-reactive material in intact cells and (6) extract and compare structurally these independently characterized PNHEV adhesion structures. Studies (5) and (6) will also be conducted in leukemia specimens to determine whether freshly isolate hematopoetic malignancies utilize the same PNHEV adhesion receptors as normal and cultured cells, to establish the capacities of surface markers to predict and quantitate affinity for PNHEV and to enhance the likelihood of detecting either functional or structural linkages between adhesion structures.