Moleclar Regulation of HIV-1 Nef Neuropathogenesis by Transforming Growth Factor Beta

描述(由申请人提供)：神经病理学对HIV-1患者来说是一个问题，不仅是由于精神和身体上的限制，而且还因为对治疗坚持的干扰。在一项队列研究中，观察到神经损伤不是药物毒性或衰老的结果，而是表明大脑中一定有其他因素引起了这些较轻微的损伤[14]。HIV-1患者死后脑组织中的星形胶质细胞大量表达HIV-1辅助蛋白阴性因子(Nef)[15]。NEF是HIV最佳复制所需的早期病毒蛋白[16，17]。研究表明，Nef诱导星形胶质细胞表达CCL2/MCP-1有助于单核细胞向脑内的渗透，从而促进HAD的进展[7]。众所周知，Nef在持续感染的星形胶质细胞中存在[19]，但它是否存在并与大脑中的其他细胞相互作用仍是一个新兴的研究领域。 我们的长期目标是更好地了解这种持续损害的基础，并帮助改善HIV-1患者的健康，这些患者寿命更长，但仍面临认知障碍，限制了他们的日常任务。本研究的目的是探讨Nef致神经元损伤的机制，了解转化生长因子β级联通路在炎症反应中的作用。由于转化生长因子β促进脑损伤后的星形胶质细胞增生[20]，我们想要分析它在Nef存在的情况下的行为。我们的中心假设是，HIV-1Nef蛋白改变了转化生长因子β途径，改变了SMADS信号，同时导致炎症反应增加，最终导致神经元损伤。我们的假设得到了初步数据和现有文献的支持，这些文献描述了Nef在大脑中的作用，以及转化生长因子β在其他神经病理中的作用。我们设计了两个目的，以便在Nef存在的情况下，通过转化生长因子β来研究和理解神经元的命运。在AIM 1中，我们将量化转化生长因子β信号和随后的促炎细胞因子在与Nef基因转染的星形胶质细胞共培养的神经元中的表达和活性。在AIM2中，我们将研究在转化生长因子β存在和不存在的情况下，HIV-1Nef对星形胶质细胞和树突生长的影响。这项研究具有重要意义，因为它旨在解决一些需要填补的空白，以便了解Nef的作用及其在学习障碍中的意义。该项目的创新将有助于理解Nef是如何通过改变转化生长因子β信号通路而导致认知障碍的。深入了解SMAD蛋白在暴露于Nef时的功能将有助于阐明该途径在HIV-1患者中的行为，即使他们的病毒载量低于可检测的水平。

DESCRIPTION (provided by applicant): Neuropathologies represent a problem for HIV-1 patients, not only due to the mental and physical limitations but also because of interference with therapy adherence. In a cohort study it was observed that the neurological impairments are not a result of drug toxicity or aging but rather suggests that there must be something else acting in the brain that provokes these milder forms of impairments [14]. The HIV-1 accessory protein negative factor (Nef) has been found to be abundantly expressed by astrocytes in postmortem brain tissue from HIV-1 patients [15]. Nef is an early viral protein required for optimal HIV replication [16, 17]. Studies suggest that Nef-induced CCL2/MCP-1 expression in astrocytes contributes to infiltration of monocytes into the brain, thereby to progression of HAD [7]. It is known that Nef is constitutively present in persistently infected astrocytes [19], but hw it exits and interacts with other cells in the brain is still an emerging area of investigation. Our long term goal is to better understand the basis of this ongoing impairment and help improve the health of HIV-1 patients that are living longer but are still confronting cognitive disorders that limit their daily tasks. The objective of this proposal is to investigate the mechanism through which Nef causes neuronal damage and understand the participation of TGFβ cascade in the inflammatory response. Since TGFβ promotes astrogliosis after brain injury [20], we want to analyze its behavior in the presence of Nef. Our central hypothesis is that HIV-1 Nef protein alters TGFβ pathway, modifying SMADs signaling while leading to increased inflammatory response and eventually neuronal damage. Our hypothesis is supported by our preliminary data and by the available literature describing the effects of Nef in the brain as well as the role of TGFβ in other neuropathologies. We have designed two aims in order to study and understand neuronal fate by TGFβ in the presence of Nef. In aim1 we will quantify the expression and activity of TGFβ signaling and consequent pro-inflammatory cytokines in neurons co-cultured with astrocytes transfected with Nef. In aim2 we will study the effects of HIV-1 Nef on astrogliosis and dendritic growth in the presence and absence of TGFβ. This study is significant since it aims to address some of those gaps that need to be filled in order to understand the action of Nef and its implication in learning impairment. The innovation of this project will provie understanding of how Nef causes cognitive disorders by altering TGFβ signaling pathway. Insight in how SMAD proteins function when exposed to Nef will help clarify how the pathway behaves in HIV-1 patients even when they have viral loads under detectable levels.