Mechanisms of HIV Tat neurotoxicity and CCL2 protection

HIV Tat神经毒性和CCL2保护机制

• 批准号: 7031511
• 负责人: Joan Weinberger Berman
• 金额: $ 5.6万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-02-17 至 2009-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7031511
• 关键词: AIDS /HIV neuropathy; HIV infections; apoptosis; biological signal transduction; chemokine; clinical research; endocytosis; excitatory aminoacid; human fetus tissue; human immunodeficiency virus; inflammation; intermolecular interaction; intracellular; neurons; neuropathology; neuroprotectants; neurotoxicology; neurotoxins; nitric oxide synthase; protein protein interaction; tissue /cell culture; transcription factor; virus cytopathogenic effect; virus protein

DESCRIPTION (provided by applicant): HIV infection of the CNS can result in neurologic dysfunction that has devastating consequences in a significant number of individuals with AIDS, including motor impairment and cognitive deficits. NeuroAIDS is characterized by neuronal injury and loss, yet there is no evidence of HIV positive neurons. Therefore, the altered function and apoptosis seen in neurons must be due to indirect effects of HIV infection of other CNS cells, including the elaboration of neurotoxic viral proteins, such as tat. Others and we demonstrated that HIV tat is toxic to human neurons. The first goal of this proposal is to identify the signaling pathways that mediate tat-induced apoptosis. Our data suggest that the mechanisms that mediate tat induced apoptosis can be divided into early and late stages. Our hypothesis is that the early events leading to apoptosis involve LRP-mediated tat endocytosis and immediate activation of NMDA channels. These activities are not sufficient to induce cell death. These early events subsequently result in later activation of signaling pathways and additional NMDA channels, leading to the dysregulation of glutamate and NO production, resulting in neuronal apoptosis. Chemokines play an important role in CNS inflammation. While their function in chemotaxis is essential, we identified a novel role for one chemokine, CCL2 (MCP-1), demonstrating that it protects human mixed cultures of neurons and astrocytes from tat-induced apoptosis. Our second goal is to determine the mechanisms by which CCL2 mediates its protection. Our data indicate that CCL2 reduces tat internalization into neurons. While cotreatment of cultures with tat and CCL2 inhibits apoptosis, addition of CCL2 2h after tat treatment does not abrogate tat induced cell death. We propose that CCL2 acts on the early phase of tat-induced apoptosis by altering endocytosis of tat and the intracellular signaling pathways activated during tat internalization. We will: 1. Determine the mechanisms that mediate tat internalization into neurons and the immediate early consequences of this uptake, including changes in protein-protein interactions involved in tat internalization (LRP, PSD-95), and the activation of signaling molecules (PSD-95, Fyn, Pyk2) and effector proteins (NMDA channels, nNOS) that may be involved in regulation of tat induced neuronal apoptosis. 2.Determine the mechanisms by which CCL2 alters tat endocytosis and the signaling pathways responsible for tat-induced apoptosis. 3. Analyze the late pathways involved in tat-induced apoptosis, including expression and activity of NMDA channels, dysregulation of extracellular glutamate and production of NO.

描述（由申请人提供）：中枢神经系统的 HIV 感染可导致神经功能障碍，对大量艾滋病患者造成毁灭性后果，包括运动障碍和认知缺陷。 NeuroAIDS 的特点是神经元损伤和丧失，但没有证据表明 HIV 阳性神经元。因此，神经元中所见的功能改变和细胞凋亡必定是由于 HIV 感染对其他中枢神经系统细胞的间接影响，包括神经毒性病毒蛋白的产生，例如 tat。其他人和我们都证明艾滋病毒对人类神经元有毒。该提案的第一个目标是确定介导 tat 诱导的细胞凋亡的信号通路。我们的数据表明介导tat诱导细胞凋亡的机制可分为早期和晚期。我们的假设是，导致细胞凋亡的早期事件涉及 LRP 介导的 tat 内吞作用和 NMDA 通道的立即激活。这些活性不足以诱导细胞死亡。这些早期事件随后导致信号通路和其他 NMDA 通道的激活，导致谷氨酸和 NO 产生失调，从而导致神经元凋亡。趋化因子在中枢神经系统炎症中发挥重要作用。虽然它们在趋化作用中的功能至关重要，但我们发现了一种趋化因子 CCL2 (MCP-1) 的新作用，证明它可以保护人类神经元和星形胶质细胞混合培养物免受 tat 诱导的细胞凋亡。我们的第二个目标是确定 CCL2 介导其保护的机制。我们的数据表明 CCL2 减少了 tat 内化到神经元中。虽然用tat和CCL2共同处理培养物会抑制细胞凋亡，但在tat处理后2小时添加CCL2并不能消除tat诱导的细胞死亡。我们认为CCL2通过改变tat的内吞作用和tat内化过程中激活的细胞内信号通路来作用于tat诱导的细胞凋亡的早期阶段。我们将： 1. 确定介导 tat 内化进入神经元的机制以及这种摄取的直接早期后果，包括涉及 tat 内化的蛋白质-蛋白质相互作用的变化（LRP、PSD-95）以及信号分子（PSD）的激活-95、Fyn、Pyk2）和效应蛋白（NMDA 通道、nNOS）可能参与调节 tat 诱导的神经元凋亡。 2.确定CCL2改变tat内吞作用的机制以及负责tat诱导细胞凋亡的信号通路。 3. 分析tat诱导细胞凋亡的晚期通路，包括NMDA通道的表达和活性、细胞外谷氨酸的失调和NO的产生。