INTERACTIONS OF HIV-1GP120 WITH BRAIN CELLS: ROLE OF PAG

描述（摘自申请书）：HIV感染患者出现 痴呆症，其发病机制仅部分了解。的 病毒感染小胶质细胞和星形胶质细胞， 损失，虽然神经元本身很少被感染。胶质细胞 类型可能潜在地充当病毒的储存库， 调查人员担心，即使我们成功地 将病毒从周围清除，隔离在这些神经胶质细胞中的病毒 细胞可能会重新植入淋巴组织。虽然显著 我们对病毒进入细胞的机制的理解已经取得了进展， 小胶质细胞，病毒进入星形胶质细胞的机制仍然存在 不确定。进一步了解HIV-星形胶质细胞的重要一步 相互作用将是表征细胞表面分子上的 与HIV包膜相互作用的星形胶质细胞表面。的相互作用 gp 120与星形胶质细胞的结合不仅对病毒的进入很重要， 引起星形胶质细胞的功能变化，从而间接引起神经元的功能变化。它有 已经表明，病毒蛋白，如gp 120，是由 HIV感染的细胞可以作用于星形胶质细胞， 细胞内钙和细胞外谷氨酸样水平增加 可能导致神经毒性的物质。我们在这份提案中提出， 初步数据显示，我们已经确定了一个独特的受体（Pag）上的 星形胶质细胞表面与gp 120相互作用， 细胞内钙和神经元细胞死亡。我们还开发了一个小组 这种蛋白质的单克隆抗体。我们建议建立一个cDNA文库 使用哺乳动物表达载体从人胎儿星形胶质细胞中分离，并使用 抗血清筛选文库。经过几轮免疫选择后， 将分离的克隆测序并转染到Pag阴性细胞中 线将筛选转染细胞的gp 120结合特性， GP 120诱导的细胞内钙变化。Pag在病毒感染中的作用 单独进入或与其他艾滋病毒共受体一起进入也将是 测定

Description (taken from application): Patients with HIV infection develop a dementing illness, the pathogenesis of which is only partially understood. The virus infects microglia and astrocytes and is accompanied by neuronal cell loss, although the neurons themselves are only rarely infected. Both glial cell types may potentially act as a reservoir for the virus and several investigators have raised concern that even if we were to successfully eradicate the virus from the periphery, virus sequestered within these glial cells could potentially re-seed the lymphoid tissue. Although significant progress has been made in our understanding of the mechanism of viral entry in microglial cells, the mechanism of viral entry into astrocytes remains undetermined. An important step to further our understanding of HIV-astrocyte interactions would be to characterize the cell surface molecules on the astrocyte cell surface that interact with the HIV envelope. The interactions of gp120 with astrocytes may be important not only for viral entry, but also for causing functional changes in astrocytes and thus indirectly in neurons. It has been shown that viral proteins, such as gp120, which are released by HIV-infected cells can act on astrocytes to produce large increases in intracellular calcium and increase extracellular levels of glutamate-like substances which can cause neurotoxicity. We present in this proposal, preliminary data showing that we have identified a unique receptor (Pag) on the surface of astrocytes that interacts with gp120 to cause changes in intracellular calcium and neuronal cell death. We have also developed a panel of monoclonal antibodies to this protein. We propose to develop a cDNA library from human fetal astrocytes using a mammalian expression vector and use the antisera to screen the library. Following several rounds of immunoselection, the isolated clones will be sequenced and transfected in Pag negative cell lines. The transfected cells will be screened for gp120 binding properties and for gp120-induced changes in intracellular calcium. The role of Pag in viral entry by itself or in conjunction with other HIV-coreceptors will also be determined.