ENDOGENOUS NEURONAL REPAIR MECHANISMS IN SIV-INFECTED MACAQUES

SIV 感染猕猴的内源性神经修复机制

• 批准号: 8172896
• 负责人: Susan V. Westmoreland
• 金额: $ 6.32万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8172896
• 关键词: Adult; Alzheimer' s Disease; Animal Model; Animals; Anti-Retroviral Agents; Apoptotic; Astrocytes; Binding; Brain; Cessation of life; Chronic; Communication; Computer Retrieval of Information on Scientific Projects Database; Creatine; Dendritic Spines; Development; Disease; Elements; Encephalitis; EphA4 Receptor; Ephrin-B3; Ephrins; Experimental Autoimmune Encephalomyelitis; Funding; Genes; Grant; Growth Associated Protein 43; Growth Cones; HIV encephalitis; Hippocampus (Brain); Inflammatory; Injury; Institution; Ligands; Macaca; Macaca mulatta; Measures; Mediating; Membrane; Messenger RNA; Metabolic; Metabolism; Microglia; Modeling; Monkeys; Multiple Sclerosis; N-acetylaspartate; Natural regeneration; Nerve Growth Factors; Neuronal Plasticity; Neurons; Neuroprotective Agents; Neurotrophic Tyrosine Kinase Receptor Type 1; Pathogenesis; Peripheral; Peripheral Nervous System Diseases; Personal Communication; Phosphotransferases; Presynaptic Terminals; Process; Proteins; Rattus; Recovery; Regulation; Research; Research Personnel; Resources; Role; SIV; Severities; Signal Pathway; Signal Transduction; Source; Spinal Ganglia; Stroke; Synapses; Synaptic Membranes; Synaptic plasticity; Synaptophysin; T-Lymphocyte; Testing; Therapeutic; Time; United States National Institutes of Health; Up-Regulation; Virus; Virus Diseases; antiretroviral therapy; axon growth; axonal guidance; axonal pathfinding; axonal sprouting; cell type; cohort; extracellular; frontal lobe; inhibitor/antagonist; mRNA Stability; macrophage; monocyte; neurotoxic; neurotransmitter release; presynaptic; protein expression; receptor; repaired

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. GAP-43 (neuromodulin) is a component of the presynaptic membrane that is the major protein of axonal growth cones. Its primary functions include axonal growth, axonal pathfinding, synaptic plasticity, and regulating neurotransmitter release. GAP-43 expression has been shown to be up-regulated during development, neuronal plasticity and neuronal death, and it has been used to measure functional axonal recovery in various chronic inflammatory diseases such as multiple sclerosis via the EAE animal model, Alzheimer's disease, and stroke. GAP-43 expression has also been used to test the pathogenesis of antiretroviral-induced peripheral neuropathy and possible effects of neuroprotective drugs in the dorsal root ganglion in rats. The purpose of this study has been to assess neuronal and synaptic damage associated with simian immunodeficiency virus (SIV) encephalitis using markers to evaluate the pre-synaptic terminals (GAP-43 and SYN) and post-synaptic membranes (MAP-2). SIV has been well established as a model for HIV encephalitis (HIVE). It has also been documented that increased severity of SIVE inversely correlates with decreased synaptophysin and MAP-2 expression and with decreased N-acetylaspartate/creatine ratio (NAA/Cr), a marker of neuronal metabolism. Immunohistochemical examination of GAP-43 protein expression in frontal cortex (FC) and hippocampus (HI) was undertaken was evaluated in FC and HI from a cohort of CD8-depleted SIVmac251-infected rhesus with and without combined antiretroviral therapy (CART). We had previously demonstrated that NAA/Cr decline in CD8-depleted SIV-infected macaques is reversed after 28 days of antiretroviral therapy associated with near complete clearance of CD68+ SIV-infected perivascular macrophages from the brain. We hypothesized that part of the cause of the normalization of NAA in treated macaques was reversal of presynaptic membrane loss that would be represented by increased synaptophysin. We also hypothesized that GAP-43 would be upregulated as a repair mechanism with CART. We demonstrate here that although synaptophysin loss is not reversed in this time frame with CART, GAP-43 is upreglulated, but the upregulation occurs with initial SIV infection and is not augmented with CART. Increased GAP-43 represents an endogenous repair mechanism that occurs with initial injury and may serve as a potential marker of repair in the SIV model. We have also demonstrated that microglia and infiltrating macrophages activated during SIV-infection express the axonal guidance molecule ephrin B3. Ephrin B3 is a ligand for EphA4 and EphB3 and is involved in synapse and dendritic spine formation. It mediates demonstrated anti-apoptotic activity in neurons, but also functions as an inhibitor of axonal growth, a mechanism to maintain axonal stability. Although ephrin B3 can regulate axonal sprouting through repulsion, its action has been associated with induction of GAP-43. The discovery of ephrin B3 in macrophages and microglia in the brain was not unanticipated. Ephrin B3 and its receptors are expressed in peripheral T cells and monocytes/macrophages. This may provide a mechanism whereby macrophages and microglia could modulate neuron and astrocyte function, as both cell types express the ephrin B3 receptors, EphA4 and EphB3. Despite the documented significant improvement in neuronal metabolism with short-term CART evidenced by normalization of NAA/Cr in the treated animals, there was no significant difference in GAP-43 expression between SIV-infected untreated monkeys and those that received CART. We speculate that there is ongoing injury (and attempts at repair) that continues to induce GAP-43 expression despite the marked reduction in brain virus burden that is observed in the animals that received CART in this study. We observe a mismatch in marked reduction of brain virus burden with CART, but persistent levels of microglial activation and TNFa, a neurotoxic factor, in the treated animals (L. Annamalai personal communication). The mismatch in parameters of improvement, neuronal metabolism and GAP-43/ephrin B3, may also reflect differences in the time course of metabolic improvement, compared with the time course of induction and suppression of factors that promote neuroadaptive processes. GAP-43 mRNA is induced by nerve growth factor (NGF), which is dependent on Mst3b, a neuron-specific kinase central to regulation of axonal outgrowth. The role of ephrins as factors that can promote or hinder neuronal repair mechanisms is controversial, but it is clear that ephrins are involved in the intricate communication of neurons with astrocytes and microglia. We propose a model in which microglia activated by virus infection (and ephrin B3 expression) stimulate astrocytes to secrete NGF, which binds TrkA receptors on neurons, thus inducing neuroprotective signaling cascades that result in the induction and increased mRNA stability of GAP-43. Other ephrins in addition to ephrin B3 and their receptors are likely to be involved in this multicellular activation and signaling pathway. Therapeutic modulation of these extracellular axonal growth molecules, their receptors, or downstream signaling elements may augment axonal sprouting and regeneration in the adult CNS. The adult SIV-infected macaque model will be useful for exploring additional genes and ephrins that are associated with synaptic plasticity as well as for further characterization of endogenous neuroregenerative potential.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 GAP-43（神经调节素）是突触前膜的一种成分，是轴突生长锥的主要蛋白质。其主要功能包括轴突生长、轴突寻路、突触可塑性和调节神经递质释放。GAP-43表达已显示在发育、神经元可塑性和神经元死亡期间上调，并且其已用于测量各种慢性炎性疾病（例如通过EAE动物模型的多发性硬化、阿尔茨海默病和中风）中的功能性轴突恢复。GAP-43的表达也被用来测试抗逆转录病毒诱导的周围神经病变的发病机制和神经保护药物在大鼠背根神经节中的可能作用。 本研究的目的是评估与猴免疫缺陷病毒（SIV）脑炎相关的神经元和突触损伤，使用标记物评估突触前末梢（GAP-43和SYN）和突触后膜（MAP-2）。SIV已被很好地确立为HIV脑炎（HIVE）的模型。也已经证明SIVE的严重程度增加与突触素和MAP-2表达的降低以及N-乙酰天冬氨酸/肌酸比率（NAA/Cr）（神经元代谢的标志物）的降低负相关。在有和没有联合抗逆转录病毒治疗（CART）的CD 8-耗尽的SIVmac 251感染的恒河猴的队列中，在FC和HI中评价了额叶皮质（FC）和海马（HI）中GAP-43蛋白表达的免疫组织化学检查。 我们以前已经证明，NAA/Cr下降CD 8-耗尽SIV感染的猕猴逆转后，28天的抗逆转录病毒治疗与CD 68 + SIV感染的血管周围巨噬细胞从大脑中几乎完全清除。 我们假设，在治疗猕猴的NAA正常化的原因的一部分是逆转突触前膜的损失，将由突触素增加代表。我们还假设GAP-43将作为CART的修复机制上调。 我们在这里证明，虽然突触体蛋白的损失是不逆转的，在这个时间范围内与CART，GAP-43是上调，但上调发生在初始SIV感染，并没有增强与CART。 增加的GAP-43代表了一种内源性修复机制，其在初始损伤时发生，并可作为SIV模型中修复的潜在标志物。 我们还证明了在SIV感染过程中激活的小胶质细胞和浸润性巨噬细胞表达轴突导向分子ephrin B3。 Ephrin B3是EphA 4和EphB 3的配体，参与突触和树突棘的形成。 它介导神经元中的抗凋亡活性，但也作为轴突生长的抑制剂发挥作用，这是维持轴突稳定性的机制。虽然ephrin B3可以通过排斥调节轴突发芽，但其作用与GAP-43的诱导有关。 在大脑中的巨噬细胞和小胶质细胞中发现肝配蛋白B3并非意料之外。 肝配蛋白B3及其受体在外周T细胞和单核细胞/巨噬细胞中表达。这可能提供了巨噬细胞和小胶质细胞可以调节神经元和星形胶质细胞功能的机制，因为这两种细胞类型都表达肝配蛋白B3受体EphA 4和EphB 3。 尽管短期CART治疗的动物中NAA/Cr的正常化证明了神经元代谢的显着改善，但SIV感染的未治疗猴和接受CART的猴之间的GAP-43表达没有显着差异。 我们推测，尽管在本研究中接受CART的动物中观察到脑病毒负荷显著降低，但仍存在持续的损伤（并尝试修复），继续诱导GAP-43表达。 我们观察到在用CART显著降低脑病毒负荷方面的不匹配，但在治疗的动物中持续存在小胶质细胞活化和TNFa（一种神经毒性因子）水平（L. Annamalai个人通信）。 与促进神经适应性过程的因子的诱导和抑制的时程相比，改善参数、神经元代谢和GAP-43/ephrin B3的不匹配也可能反映了代谢改善时程的差异。 GAP-43 mRNA由神经生长因子（NGF）诱导，其依赖于Mst 3b，Mst 3b是调节轴突生长的神经元特异性激酶。 肝配蛋白作为促进或阻碍神经元修复机制的因子的作用是有争议的，但很明显，肝配蛋白参与神经元与星形胶质细胞和小胶质细胞的复杂通信。 我们提出了一种模型，其中通过病毒感染（和肝配蛋白B3表达）激活的小胶质细胞刺激星形胶质细胞分泌NGF，所述NGF结合神经元上的TrkA受体，从而诱导神经保护性信号传导级联，导致GAP-43的诱导和增加的mRNA稳定性。 除了肝配蛋白B3及其受体之外，其他肝配蛋白可能参与这种多细胞活化和信号传导途径。 这些细胞外轴突生长分子、其受体或下游信号传导元件的治疗性调节可增强成年CNS中的轴突发芽和再生。 成年SIV感染的猕猴模型将有助于探索与突触可塑性相关的其他基因和肝配蛋白，以及进一步表征内源性神经再生潜力。