RO1E OF CELL INTEGRINS AND ADENOVIRAL CONJUNCTIVITIES

腺病毒(Ad)是引起病毒性结膜炎的主要原因 天哪。虽然通常是自我限制的，但眼部感染有很高的发病率 传染性水平和传播率导致显著 因大量工作日损失造成的发病率和社会经济问题 每年。在眼部感染期间发生的免疫反应也可能 导致严重的视觉障碍，包括失明。 旨在阻止病毒进入细胞的抗病毒策略已经被 由于缺乏准确的知识，相对不发达 介导进入事件的病毒和细胞成分。单元格 表面整合素α-v-Beta3和α-v-Beta5介导腺病毒 通过特定的交互作用实现内化，而不是病毒附着 与病毒五元碱基衣壳蛋白中的RGD序列同源。此外， 我们最近的研究表明，整合素α-M-β2和α-L- β2促进腺病毒与人单核细胞和淋巴系的黏附 细胞。这项提议寻求利用这些发现来开发 体内封堵腺病毒眼部感染的合理方法。 由于几种不同的腺病毒血清型使用α-v整合素 感染时，使用这些受体的强效拮抗剂可能会 针对多种病毒血清型的保护。 一系列详细的分子和生化研究将用于 阐明Ad Penton碱基与不同细胞的精确相互作用 整合素，以获得对腺病毒细胞进入的进一步了解。 五酮碱结合的动力学、热力学和化学计量 到细胞的整合素将使用自动生物传感器进行测定 系统。五酮碱基中精确的氨基酸序列 与不同细胞整合素的中介结合将通过以下方式确定 激光分析与固定化整合素结合的多肽 解吸质谱。该提案的第二个目标是 确定β2整合素在腺病毒感染中的整体作用 人类单核/巨噬细胞和淋巴细胞的两种细胞类型 在眼球疾病发病机制中起重要作用。这些细胞也可能是一种 宿主中持续的Ad感染的站点。最后，一个强有力的 细胞整合素或功能阻断的合成肽拮抗剂 识别多发性硬化RGD结构域的单抗 将使用腺病毒血清型来防止腺病毒感染 NZW兔眼模型。 这些研究代表了一种合理的方法来阻止广告眼球 体内感染，也可能导致使用的改进策略 用于眼部基因治疗的复制缺陷型腺病毒载体。

Adenoviruses (Ad) represent a major cause of viral conjunctivitis in man. Although usually self-limiting, Ad ocular infections have a high level of infectivity and transmission rate causing significant morbidity and socioeconomic problems due to numerous lost working days each year. Immune responses occurring during ocular infections can also lead to severe visual disturbances including blindness. Antiviral strategies aimed at blocking virus entry into cells have been relatively underdeveloped due to a lack of knowledge of the precise viral and cellular components that mediate the entry events. The cell surface integrins alpha-v-beta3 and alpha-v-beta5 mediate adenovirus internalization rather than virus attachment via a specific interaction with an RGD sequence in the virus penton base capsid protein. Moreover, our recent studies indicate that integrins alpha-M-beta2 and alpha-L- beta2 promote adenovirus attachment to human monocytic and lymphoid cells. This proposal seeks to capitalize on these findings to develop a rational approach to blocking adenovirus ocular infections in vivo. Since several different adenovirus serotypes use alpha-v integrins for infection, the use of potent antagonists of these receptors may confer protection against multiple virus serotypes. A series of detailed molecular and biochemical studies will be used to elucidate the precise interactions of Ad penton base with distinct cell integrins in order to gain further insights into adenovirus cell entry. The kinetics, thermodynamics and stoichiometry of penton base binding to cell integrins will be determined using an automated biosensor system. The precise amino acid sequences in the penton base that mediate binding to different cell integrins will be identified by analyzing peptide binding to immobilized integrins using laser- desorption mass spectrometry. A second goal of the proposal is to determine the overall role of beta2 integrins in adenovirus infection of human monocyte/macrophages and lymphocytes, two cell types that contribute to Ad ocular pathogenesis. These cells are also a likely site for persistent Ad infection in the host. Finally, a potent synthetic peptide antagonist of cell integrins or a function-blocking monoclonal antibody that recognizes the RGD domain of multiple adenovirus serotypes will be used to prevent adenovirus infection in the NZW rabbit ocular model. These studies represent a rational approach to block Ad ocular infection in vivo and may also lead to improved strategies for using replication-defective adenovirus vectors for ocular gene therapy.