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

描述(摘自申请)：感染艾滋病毒的患者会发展为 痴呆性疾病，其发病机制尚不完全清楚。这个 病毒感染小胶质细胞和星形胶质细胞，并伴有神经细胞 损失，尽管神经元本身很少被感染。两者都是神经胶质细胞 类型可能潜在地作为病毒和几个 调查人员担心，即使我们成功地 从周围清除病毒，病毒隔离在这些神经胶质细胞内 细胞可能会重新种植淋巴组织。虽然意义重大 在我们对病毒进入的机制的理解上取得了进展 小胶质细胞，病毒进入星形胶质细胞的机制仍然存在 未定。进一步了解艾滋病毒的重要一步--星形胶质细胞 相互作用将是表征细胞表面分子在 与HIV包膜相互作用的星形胶质细胞表面。的相互作用 带有星形胶质细胞的gp120可能不仅对病毒的进入很重要，而且对 导致星形胶质细胞的功能改变，从而间接改变神经元的功能。它有 已经证明，病毒蛋白，如gp120，由 HIV感染的细胞可以作用于星形胶质细胞，使其产生大量的 细胞内钙离子和细胞外类谷氨酸水平升高 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，我们已经确定了一种独特的受体(PAG)在 星形胶质细胞表面与gp120相互作用引起 细胞内钙离子和神经细胞死亡。我们还开发了一个面板 这种蛋白质的单抗。我们建议开发一个cdna文库 使用哺乳动物表达载体从人胚胎星形胶质细胞中分离出 用抗血清筛选文库。在几轮免疫选择之后， 将分离的克隆进行测序，并在PAg阴性细胞中进行转染 台词。将对转基因细胞进行gp120结合特性的筛选和 用于gp120诱导的细胞内钙变化。PAG在病毒中的作用 单独或与其他艾滋病毒辅助受体一起进入也将是 下定决心。

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.