Molecular mechanisms underlying HIV & Cocaine-mediated microglial activation: Targeting NLRP3 inflammasome

HIV的分子机制

• 批准号: 10665604
• 负责人: Shilpa J. Buch
• 金额: $ 51.47万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-30 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10665604
• 关键词: Affect; Astrocytes; Behavior assessment; Binding; Blood - brain barrier anatomy; Brain; CASP1 gene; Caspase; Cells; Cocaine; Cocaine Abuse; Complex; Data; Development; Disease; Down-Regulation; Drug abuse; Enzyme-Linked Immunosorbent Assay; Exposure to; Future; Gene Silencing; Genetic Transcription; HIV; HIV-associated neurocognitive disorder; Human; IL18 gene; Immune; In Vitro; Individual; Inflammasome; Inflammatory; Inflammatory Response; Interleukin-1 beta; Leucine-Rich Repeat; Mediating; Memory; Messenger RNA; MicroRNAs; Microglia; Molecular; Motor Activity; Multiprotein Complexes; Mus; Neurocognitive; Neurocognitive Deficit; Neuroglia; Nuclear Translocation; Nucleotides; Pathway interactions; Phosphorylation; Prevalence; Process; Proteins; Quality of life; Reactive Oxygen Species; Rodent; Rodent Model; Role; Signal Pathway; Signal Transduction; Site; Testing; Time; Trans-Activators; Transcription Coactivator; Transgenic Organisms; Up-Regulation; Western Blotting; antiretroviral therapy; combinatorial; comorbidity; cytokine; cytotoxic; efficacy testing; glial activation; in vivo; inhibitor; neuroinflammation; novel therapeutics; pharmacologic; response; symptomatology

In the current era of combination antiretroviral therapy (cART), there is increased prevalence of HIV-associated neurocognitive disorders (HAND) with about 30-50% of HIV-infected individuals afflicted with varying degrees of neurocognitive impairments, substantially affecting their quality of life. It is well-recognized that despite cART, there is unabated persistence & presence of early HIV protein(s) such as the cytotoxic transactivator of transcription (TAT), that contributes to neuroinflammation. Adding fuel to the mix is the comorbidity of drug abuse in HIV-infected individuals, which further exacerbates microglial activation & thus symptomatology of HAND. Until recently, CNS was considered as an immune-privileged site, however, a recent paradigm shift in our understanding has revealed the role of cytosolic, multiprotein complexes, belonging to the nucleotide-binding domain leucine-rich repeat containing (NLR) protein superfamily, termed as “inflammasomes,” in various neuroinflammatory diseases. Assembly of these multiprotein complexes activates the proinflammatory caspases (specifically caspase 1), leading, in turn, to the cleavage & release of IL1β & 1L18, consequently resulting in a potent inflammatory response. Among the various NLRs, NLRC5 & NLRP3 are highly expressed in microglia & differentially regulate inflammatory responses. In our efforts to understand the combinatorial effects of HIV Tat & cocaine on Mg activation, we have made several exciting preliminary observations: 1) Exposure of rodent primary Mg to both HIV Tat & cocaine demonstrated significantly downregulated expression of NLRC5, with a concomitant upregulation of NLRP3 and, exacerbated Mg activation compared to cells exposed to either agent alone; 2) Rodent Mg exposed to HIV TAT demonstrated time-dependent upregulation of microRNA (miR)-34a, leading, in turn, to downregulation of its target - NLRC5; 3) Cocaine-mediated activation of Mg involved induction of reactive oxygen species (ROS), that was accompanied with defective mitophagy & subsequent oligomerization of NLRP3 inflammasome complex, leading to induced expression of mature (m)IL1β & IL18; 4) NLRP3 inhibitor - MCC950 mitigated cocaine-mediated activation of Mg. Based on these observations, we hypothesize that the co-operative effects of HIV TAT & cocaine on Mg activation involve two processes: a) downregulated expression of NLRC5 resulting in activation of NFκB (signal 1), leading, in turn, to transcriptional upregulation of NLRP3, pro IL1β & IL18, & b) induction of ROS-mediated defective mitophagy (signal 2), which, in turn, induces activation of NLRP3 inflammasome, resulting in increased cleavage & secretion of mIL1β & IL18. The hypothesis will be tested in two SA: 1) Explore the molecular mechanism(s) underlying HIV & cocaine- mediated induction of the NLRP3 inflammasome leading to Mg activation and 2) To validate in vivo the role of NLRC5/NFκB & ROS/defective mitophagy/NLRP3 axes underlying HIV Tat & cocaine-mediated activation of Mg. Understanding the mechanisms responsible for microglial activation induced by HIV & cocaine will set the stage for the future development of novel therapeutics aimed at dampening the neuroinflammatory responses.

在当前的联合抗逆转录病毒疗法(CART)时代，艾滋病毒相关疾病的流行率有所增加 神经认知障碍(手)，约30%-50%的艾滋病毒感染者患有不同程度的 神经认知障碍，严重影响他们的生活质量。这是众所周知的，尽管Cart， 存在有增无减的早期艾滋病毒蛋白(S)，如细胞毒性反式激活因子 转录(TAT)，这有助于神经炎症。在混合物中添加燃料是药物滥用的共病 在感染艾滋病毒的人中，这进一步加剧了小胶质细胞的激活&从而导致手部症状。 直到最近，中枢神经系统被认为是一个免疫特权站点，然而，最近我们的 了解到胞浆中的作用，多蛋白复合体，属于核苷酸结合 富含亮氨酸重复序列(NLR)的蛋白超家族，被称为“炎性小体”，在不同的 神经炎症性疾病。这些多蛋白复合体的组装激活了促炎症的caspase (特别是caspase1)，进而导致IL1β&1L18的切割和释放，从而导致 强烈的炎症反应。在多种NLRs中，NLRC5和NLRP3在小胶质细胞中高表达，对炎症反应起不同的调控作用。在我们努力了解hiv、tat和可卡因对镁激活的组合影响的过程中，我们做了几个令人兴奋的初步观察：1)接触啮齿动物。 HIVTAT和可卡因的原始镁均显著下调NLRC5的表达，并具有 与任一种药物暴露的细胞相比，伴随的NLRP3和，加剧了镁的激活 2)暴露于HIV TAT的啮齿动物镁表现出时间依赖性的microRNA(MiR)-34a上调， 进而导致其靶标NLRC5的下调；3)可卡因介导的镁的激活涉及诱导 活性氧物种(ROS)，伴随着有丝分裂缺陷&随后 NLRP3炎症体复合体的寡聚，诱导成熟的(M)IL-1β和IL-18的表达 NLRP3抑制剂MCC950可减轻可卡因介导的MG的激活。基于这些观察，我们 假设HIV、TAT和可卡因对镁激活的协同作用涉及两个过程：a) NLRC5表达下调导致核因子κB(信号1)激活，进而导致转录 上调NLRP3、ProIL1β和IL18，&b)诱导ROS介导的有丝分裂缺陷(信号2)， 进而诱导NLRP3炎性小体的激活，导致mIL1β和IL18的切割和分泌增加。 这一假说将在两个SA中得到验证：1)探索艾滋病毒和可卡因的分子机制(S)- 介导诱导NLRP3炎症体导致镁激活和2)在体内验证 NLRC5/NFκB&ROS/有丝分裂缺陷/NLRP3轴潜在的艾滋病毒TAT和可卡因介导的镁的激活。 了解艾滋病毒和可卡因诱导小胶质细胞激活的机制将奠定基础 为未来旨在抑制神经炎性反应的新疗法的开发奠定基础。