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A New Antioxidant Prevents Toxicity of HIV Proteins with Methamphetamine

一种新的抗氧化剂可防止 HIV 蛋白与甲基苯丙胺的毒性

基本信息

批准号：
7418174
负责人：
NURAN ERCAL
金额：
$ 22.58万
依托单位：
MISSOURI UNIVERSITY OF SCIENCE & TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-09-30 至 2009-09-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7418174
关键词：
AIDS Dementia Complex Acetylcysteine Acquired Immunodeficiency Syndrome Address Affect Albumins Alcohols Amides Animals Antioxidants Apoptosis Astrocytes Behavioral Biological Assay Biological Availability Blood - brain barrier anatomy Brain Cell membrane Cells Characteristics Charge Cognitive deficits Complication Cysteine Deacetylation Dementia Development Disease Disruption Dose Drug or chemical Tissue Distribution End Point Endocytosis Endothelial Cells Enzymes Equilibrium Event Exposure to Extravasation Failure Free Radical Scavenging Free Radicals Functional disorder Glutathione Glutathione Disulfide Glutathione Reductase Glycoproteins HIV HIV Envelope Protein gp120 HIV Infections HIV Seropositivity HIV-1 High Pressure Liquid Chromatography Human In Vitro Individual Infection Laboratories Lipid Peroxidation Measures Methamphetamine Microglia Modeling Modification Morphine Motor N-Acetylcysteinamide Neurologic Neurons Neuropathogenesis Nitrogen Numbers Oral Oxidative Stress Oxygen Pathway interactions Patients Permeability Pharmaceutical Preparations Play Production Protein Overexpression Proteins Rate Rattus Reactive Oxygen Species Recovery Risk Role Solubility Sulfhydryl Compounds Syndrome Techniques Testing Therapeutic Agents Time Tissues Toxic effect Transgenic Animals Transgenic Mice Transgenic Organisms Wild Type Mouse Work antiretroviral therapy carboxyl group caspase-3 catalase cytokine design ecstasy glutathione peroxidase gp-120 Antigen human tissue in vitro Model in vivo in vivo Model neurotoxicity prevent research study response stress related disorder tat Protein transcytosis

项目摘要

DESCRIPTION (provided by applicant): A major complication of HIV-1 infection is the development of HIV-1-associated dementia (HAD). The mechanisms behind HAD are not yet known. Certain studies have indicated that the HIV-1 envelope glycoprotein (gp120) and transregulatory protein (Tat) play a role in the development of HAD. In particular, these proteins affect the integrity of the blood- brain barrier (BBB) by crossing the BBB, altering BBB transporters, and disrupting the BBB. Our studies show that these proteins also induce oxidative stress in RBE4 cells, an in vitro BBB model. We determined that gp120 and Tat induced oxidative stress in RBE4 cells by measuring selected oxidative stress parameters. Subsequently, the levels of glutathione (GSH), the principal intracellular thiol responsible for maintaining the oxidative balance in cells, significantly decreased, indicating that the cells were undergoing oxidative stress. Antioxidants are becoming increasingly popular in oxidative stress-related disorders and hold promise as therapeutic agents. We have also found that N-acetylcysteine amide (NACA), a new thiol antioxidant, significantly increased the levels of GSH in gp120 and Tat-exposed RBE4 cells. Furthermore, many AIDS/HIV-positive patients use addictive drugs, including methamphetamines (METH). Since METH induces oxidative stress as well, drug abusing patients might be at risk of a synergistic effect and increased damage. Our preliminary results showed that METH does indeed work synergistically with gp120 and Tat to induce oxidative stress in RBE4 cells. Since work with RBE4 cells is open to the criticism, discussed under the "Specific Aims" section, we plan to use a more appropriate in vitro BBB model. Human brain microvascular endothelial cells (HBMVECs) will be used to evaluate the synergistic effect of METH and the two HIV proteins in inducing oxidative stress. Transgenic mice overexpressing gp120 or Tat will serve as our in vivo model. Permeability experiments in BBB models (both in HBMVECs and isolated brain microvascular endothelial cells from the transgenic animals) will also be performed to serve as functional endpoints. Therefore, we propose to determine whether the potent antioxidant NACA protects the BBB from gp120, Tat, or METH (alone and/or in combination).Despite the development and use of effective antiretroviral therapy, HIV-1 associated dementia (HAD) still persists. Certain studies have indicated that the HIV-1 envelope glycoprotein (gp120) and transregulatory protein (Tat) play a role in the development of HAD. One proposed mechanism is that HIV-1 induced neurotoxicity may be due to an increased production of reactive oxygen species (ROS) by HIV-1 proteins (gp120 and Tat). Furthermore, many AIDs/HIV-positive patients use addictive drugs such as methamphetamine (METH). Among these addictive drugs, METH, in particular, induces free radicals in various cells. If neuropathogenesis of HIV infection is found to be caused by a significant increase in oxidative damage by METH and gp120 and/or Tat, antioxidants should be included in the treatment to prevent HIV-induced dementia. Glutathione (GSH) is the principal intracellular thiol responsible for scavenging reactive oxygen species (ROS) and maintaining the oxidative balance in tissues. Cysteine and glutathione delivery compounds protect cells from free radical damage. For example, N-acetylcysteine (NAC) indirectly replenishes GSH through deacetylation to cysteine and prevents oxidative stress. However, NAC has a very low oral bioavailability (30%) and its efficacy is only significant when given intravenously. NAC's failure to provide significant antioxidant effects in vivo might originate from its low solubility and tissue distribution. This inability to provide antioxidant defense may result from NAC's negatively charged carboxyl group. Recently, an amide form of NAC has been synthesized and our laboratory has tested this new compound in various oxidative stress models. Because the carboxylic group in NACA is neutralized, NACA is more lipophilic than NAC and can easily cross the cell membranes. Therefore, we hypothesize that addictive drugs, as exemplified by METH, potentiate the oxidative stress induced by gp120 and Tat at the blood brain barrier (BBB) and that NACA blocks oxidative stress and protects the BBB.

描述（由申请人提供）：HIV-1感染的一个主要并发症是HIV-1相关性痴呆（HAD）的发展。HAD背后的机制尚不清楚。某些研究表明HIV-1包膜糖蛋白（gp 120）和反式调节蛋白（达特）在HAD的发展中起作用。特别地，这些蛋白质通过穿过血脑屏障（BBB）、改变BBB转运蛋白和破坏BBB来影响血脑屏障（BBB）的完整性。我们的研究表明，这些蛋白质也诱导氧化应激RBE 4细胞，在体外血脑屏障模型。我们通过测量选择的氧化应激参数确定了gp 120和达特诱导RBE 4细胞的氧化应激。随后，谷胱甘肽（GSH）（负责维持细胞内氧化平衡的主要细胞内硫醇）的水平显著降低，表明细胞正在经历氧化应激。抗氧化剂在氧化应激相关疾病中越来越受欢迎，并有望成为治疗剂。我们还发现，N-乙酰半胱氨酸酰胺（NACA），一种新的巯基抗氧化剂，显着增加GSH的水平在gp 120和Tat暴露的RBE 4细胞。此外，许多艾滋病/艾滋病毒阳性患者使用成瘾药物，包括甲基苯丙胺（METH）。由于METH也诱导氧化应激，药物滥用患者可能有协同效应和增加损害的风险。我们的初步结果表明，METH确实与gp 120和达特协同作用，诱导RBE 4细胞中的氧化应激。由于使用RBE 4细胞的工作受到批评，在“特定目的”部分讨论，我们计划使用更合适的体外BBB模型。人脑微血管内皮细胞（HBMVEC）将用于评估METH和两种HIV蛋白在诱导氧化应激中的协同作用。过表达gp 120或达特的转基因小鼠将作为我们的体内模型。还将在BBB模型（HBMVEC和转基因动物分离的脑微血管内皮细胞）中进行渗透性实验，作为功能终点。因此，我们建议，以确定是否有效的抗氧化剂NACA保护BBB从gp 120，达特，或METH（单独和/或组合）。尽管开发和使用有效的抗逆转录病毒疗法，HIV-1相关的痴呆症（HAD）仍然存在。某些研究表明HIV-1包膜糖蛋白（gp 120）和反式调节蛋白（达特）在HAD的发展中起作用。一种提出的机制是HIV-1诱导的神经毒性可能是由于HIV-1蛋白（gp 120和达特）产生的活性氧（ROS）增加。此外，许多艾滋病/艾滋病毒阳性患者使用甲基苯丙胺（METH）等成瘾药物。在这些成瘾性药物中，特别是METH在各种细胞中诱导自由基。如果发现HIV感染的神经发病机制是由METH和gp 120和/或达特的氧化损伤显著增加引起的，则应在治疗中包括抗氧化剂以预防HIV诱导的痴呆。谷胱甘肽（GSH）是细胞内主要的巯基，负责清除活性氧（ROS）和维持组织中的氧化平衡。半胱氨酸和谷胱甘肽递送化合物保护细胞免受自由基损伤。例如，N-乙酰半胱氨酸（NAC）通过脱乙酰化为半胱氨酸间接还原GSH并防止氧化应激。然而，NAC具有非常低的口服生物利用度（30%），并且其功效仅在静脉内给予时才显著。NAC在体内不能提供显著的抗氧化作用可能源于其低溶解度和组织分布。这种不能提供抗氧化防御可能是由于NAC的带负电荷的羧基。最近，已经合成了NAC的酰胺形式，我们的实验室已经在各种氧化应激模型中测试了这种新化合物。由于NACA中的羧基被中和，NACA比NAC更亲脂，可以很容易地穿过细胞膜。因此，我们假设成瘾性药物，如METH，加强gp 120和达特诱导的氧化应激在血脑屏障（BBB）和NACA阻断氧化应激和保护BBB。