Protective Effects of Cannabinoids in HIV Neuropathogenesis

描述(申请人提供)：HIV-1的神经发病机制涉及病毒蛋白、神经胶质和神经介质以及滥用药物等环境因素之间的复杂相互作用。然而，令人惊讶的是，尽管大麻素受体在纹状体中大量表达，黑质纹状体多巴胺能区对HIV-1相关的神经病理非常敏感，但关于大麻样物质对HIV-1神经发病机制的影响知之甚少。此外，这个脑区的多巴胺能神经元、激活的小胶质细胞、反应性星形胶质细胞和氧化损伤是HAD的显著特征。小胶质细胞是炎症介质的重要来源，包括细胞因子、趋化因子和活性氧物种，所有这些都会导致脑损伤。当受到白介素12的刺激时，星形胶质细胞产生神经毒性的反应性氮物种和趋化因子，如CCL2和CX3CL1，这可能会将巨噬细胞/小胶质细胞招募到纹状体。在病毒相关因子中，病毒毒素HIV-1 gp120蛋白在小胶质细胞和星形胶质细胞中触发多种神经致病反应。基于我们实验室以前的工作证明合成大麻素抑制人类小胶质细胞中HIV-1的表达并抑制人脑星形胶质细胞产生炎症介质，以及来自非人类系统的大量文献证明了大麻素的神经保护作用，本研究项目背后的中心假设是合成大麻素将通过抑制激活的小胶质细胞和星形胶质细胞的机制来保护多巴胺能神经元免受gp120的毒性影响。在认识到gp120对神经元的直接毒性效应的同时，我们打算填补关于大麻类物质对gp120与参与HIV-1神经发病的神经胶质细胞相互作用的影响的重要知识空白。在这个项目中，我们将使用两个新开发的人类细胞培养模型：中脑细胞培养模型和神经前体细胞(NPC)来源的多巴胺能神经元模型。我们将使用这些模型以及高纯度的人小胶质细胞和星形胶质细胞进行实验，通过表征神经胶质细胞产生趋化因子和多巴胺能神经元的功能损伤、凋亡和氧化损伤来研究合成大麻素对gp120诱导的多巴胺能神经元损伤的保护作用，评估大麻素在两种细胞培养模型中减轻gp120介导的多巴胺能神经元损伤和抑制趋化因子产生的能力，并探讨大麻素抑制小胶质细胞介导的多巴胺能神经元损伤和抑制gp120和IL-12介导的星形胶质细胞功能效应的具体机制。通过创新地使用原代人脑细胞培养模型，我们希望促进对合成大麻如何改变gp120对参与HIV-1神经发病机制的脑细胞的毒性效应的理解，并将这些研究的发现转化为HAD的新治疗方法。项目简介尽管抗逆转录病毒疗法(ART)具有治疗效果，但艾滋病毒-1相关性痴呆(HAD)仍然对艾滋病患者构成严重威胁，目前仍没有针对艾滋病毒-1的神经表现的具体治疗方法。基于最近的工作，即黑质纹状体多巴胺能区是HAD中神经元功能障碍和死亡的关键大脑区域，合成大麻素能抑制人小胶质细胞中艾滋病毒-1的表达并抑制人星形胶质细胞中炎症介质的产生，以及大量文献证明大麻素类在其他系统中具有神经保护作用，实验旨在验证这一中心假设，即合成大麻素将通过抑制激活的小胶质细胞和星形胶质细胞的机制来保护多巴胺能神经元免受艾滋病毒蛋白gp120的毒性影响。通过创新地使用原代人脑细胞培养模型，我们希望促进对合成大麻如何改变gp120对参与HIV-1神经发病机制的脑细胞的毒性效应的理解，并将这些研究的发现转化为HAD的新治疗方法。

DESCRIPTION (provided by applicant): The neuropathogenesis of HIV-1 involves a complex interaction between viral proteins, glial and neuronal mediators, and environmental factors, such as drugs of abuse. However, surprisingly little is known about the effects of cannabinoids on HIV-1 neuropathogenesis, despite the facts that cannabinoid receptors are abundantly expressed in the striatum and that the nigrostriatal dopaminergic area is highly susceptible to HIV-1-related neuropathology. In addition, marked depletion of dopaminergic neurons, activated microglia, reactive astrocytes, and oxidative damage within this brain region are salient features of HAD. Microglia are an important source of inflammatory mediators, including cytokines, chemokines, and reactive oxygen species, all of which contribute to brain damage. When stimulated by interleukin (IL)-12, astrocytes produce neurotoxic reactive nitrogen species and chemokines, such as CCL2 and CX3CL1, which may recruit macrophages/microglia to the striatum. Among the viral-related factors, the virotoxin HIV-1 gp120 protein triggers several neuropathogenic responses in microglia and astrocytes. Based upon previous work in our laboratory demonstrating that synthetic cannabinoids inhibit expression of HIV-1 in human microglia and suppress production of inflammatory mediators by human astrocytes, and a substantial body of literature from non-human systems demonstrating neuroprotective properties of cannabinoids, the central hypothesis underlying this research project is that synthetic cannabinoids will protect dopaminergic neurons against the toxic effects of gp120 through a mechanism involving inhibition of activated microglia and astrocytes. While acknowledging the direct toxic effect gp120 can have on neurons, we intend to fill important gaps in knowledge regarding the effects of cannabinoids on the interaction of gp120 with the glial cells involved in HIV-1 neuropathogenesis. For this project, we will use two newly developed human cell culture models: a midbrain cell culture model and a neural precursor cell (NPC)-derived dopaminergic neuron model. We will use these models, as well as highly purified human microglial cells and astrocytes in experiments that investigate the protective effects of synthetic cannabinoids against gp120-induced damage to dopaminergic neurons through characterization of functional impairment, apoptosis, and oxidative damage of dopaminergic neurons and chemokine production by glial cells, assess the ability of cannabinoids to ameliorate gp120-mediated damage to dopaminergic neurons and to inhibit chemokine production in both cell culture models, and investigate the specific mechanisms by which cannabinoids inhibit microglia-mediated damage to dopaminergic neurons and inhibit gp120- and IL-12-mediated functional effects on astrocytes. Through the innovative use of primary human brain cell culture models, we hope to advance the understanding of how synthetic cannabinoids alter the toxic effects of gp120 on the brain cells involved in the neuropathogenesis of HIV-1, with the long term objective of translating discoveries arising from these studies into a new treatment method for HAD. Project Narrative Despite the therapeutic impact of anti-retroviral therapy (ART), HIV-1-associated dementia (HAD) remains a serious threat to AIDS patients, and there currently remains no specific therapy for the neurological manifestations of HIV-1. Based upon recent work that the nigrostriatal dopaminergic area is a critical brain region for the neuronal dysfunction and death seen in HAD, that synthetic cannabinoids inhibit HIV-1 expression in human microglia and suppress production of inflammatory mediators in human astrocytes, as well as a substantial literature demonstrating neuroprotective properties of cannabinoids in other systems, experiments have been designed to test the central hypothesis that synthetic cannabinoids will protect dopaminergic neurons against the toxic effects of the HIV protein gp120 through a mechanism involving inhibition of activated microglia and astrocytes. Through the innovative use of primary human brain cell culture models, we hope to advance the understanding of how synthetic cannabinoids alter the toxic effects of gp120 on the brain cells involved in the neuropathogenesis of HIV-1, with the long term objective of translating discoveries arising from these studies into a new treatment method for HAD.