FIBRONECTIN-BUCCAL EPITHELIAL CELL INTERACTIONS

本研究项目的总体目标是确定 纤维连接蛋白（FN）在细菌附着到粘膜表面的作用 口咽 为了逻辑地进行这些研究， 在该第一申请中提出的方法集中于表征 FN与人颊上皮细胞的相互作用。 具体努力 将针对：（1）检查颊细胞的内容物， FN的分布结合到他们的表面自然和实验后， 治疗，（2）研究结合的具体特点， 外源性FN的颊细胞，和（3）确定主要功能 介导完整分子与颊粘膜结合的血浆FN结构域 上皮细胞 颊上皮细胞的增殖将在 测定与细胞结合的标记抗FN抗体的量， 与细胞结合的标记FN的量和形态学特征 与细胞结合的抗FN抗体的分布。 几 将使用不同的化学和酶处理来操纵FN 这些细胞的水平。 将进行结合研究，以确定 颊上皮FN结合位点的特异性、亲和力和数量 细胞 我们将特别注意pH值、离子 强度和二价阳离子组成和浓度。 明确定义 FN片段和已知与FN相互作用的其他特异性配体将被 测试了它们抑制完整FN与颊粘膜结合的能力， 细胞 确定的FN片段直接结合细胞的能力 也将受到考验。 此外，将检测定义的单克隆抗体 因为它们抑制FN与细胞结合的能力。 我们认为 提出的FN-颊上皮细胞表征实验 互动将是我们努力了解 FN在细菌粘附中的作用。

The overall goal of this research project is to determine the role of fibronectin (FN) in the attachment of bacteria to the mucosal surface of the oropharynx. To pursue these studies logically, the experiments proposed in this first application are focused on characterizing the interaction of FN with human buccal epithelial cells. Specific efforts will be directed toward: (1) examining buccal cells for content and distribution of FN bound to their surfaces naturally and after experimental treatment, (2) studying the specific characteristics of the binding of exogenous FN to the buccal cells, and (3) determining the major functional domain of plasma FN that mediates binding of the intact molecule to buccal epithelial cells. Pools of buccal epithelial cells will be examined in assays of the amount of labeled anti-FN antibody that binds to the cells, the amount of labeled FN that binds to the cells and the morphological distribution of anti-FN antibodies that bind to the cells. Several different chemical and enzymatic treatments will be used to manipulate FN levels on these cells. Binding studies will be performed to determine the specificity, affinity and number of FN binding sites on buccal epithelial cells. We will pay particular attention to the effects of pH, ionic strength and divalent cation composition and concentration. Well defined FN fragments, and other specific ligands known to interact with FN, will be tested for their ability to inhibit the binding of intact FN to buccal cells. The ability of defined FN fragments to bind to the cells directly will also be tested. Further, defined monoclonal antibodies will be tested for their ability to inhibit the binding of FN to the cells. We believe the proposed experiments on characterization of FN-buccal epithelial cell interactions will be an important step in our efforts to understand the role of FN in bacterial adherence.