Mechanistic studies on analgesic effects of terpene enriched extracts from hops

摘要 慢性疼痛影响着世界人口的1/5。这是一种令人衰弱的健康状况，具有显著的社会- 经济成本。慢性疼痛的管理仍然存在治疗障碍，阿片类药物的效用 随着越来越多的人成为鸦片成瘾的受害者， 鸦片危机大麻科植物，如大麻（Cannabis sativa）和葎草（Humulus） 蛇麻草（啤酒花）在缓解疼痛和炎症方面具有悠久的传统使用历史， 这些活动的机理基础和负责的化合物的身份还没有得到很好的理解。 在拟议的工作中，我们将采取多学科的方法来调查萜烯的镇痛作用 来自大麻科。而不是使用大麻，其研究和产品市场因 冲突的联邦和州的法律，我们将集中在萜烯发现啤酒花，这是一个相关的物种， 有着非常相似的萜烯特征我们将获得植物认证的啤酒花锥材料， 通过超临界CO2方法提取以产生用于研究的萜烯富集和耗尽的啤酒花提取物， 从而能够评估萜烯之间的协同作用。我们还将使用个别萜烯标准在我们的 建议的研究。将通过GC-MS和NMR对浸提液进行化学表征，并评估其生物活性 使用体外模型评估影响培养的DRG伤害感受性神经元的兴奋性和致敏性的能力 神经元我们将分析啤酒花提取物或候选萜烯调节伤害感受器TRPV 1的能力 使用钙成像的炎症刺激致敏，并检查萜烯调节 伤害感受器兴奋性采用全细胞膜片钳电生理学方法。PCA和化合物活性映射 对观察到的化学特征和生物活性的分析将指导最有效成分的选择， 包括考虑萜烯的协同组合。在项目的第二部分，我们将 评估最有希望的啤酒花富含萜烯的提取物或分离的萜烯减轻疼痛的功效 和免疫激活的研究。血浆和 还将通过靶向的方法评估小鼠组织中特定萜烯及其代谢物的水平。 高分辨率代谢组学方法除了行为分析，我们还将分析足底和DRG 免疫激活，以及脊髓中的小胶质细胞和星形胶质细胞激活。这项研究是响应 RFA-AT-19-009的目的是1）研究萜烯类可能影响疼痛通路的机制， 包括上行和/或下行神经通路，细胞和分子信号通路， 神经免疫相互作用或其他与疼痛相关的创新调节途径;以及2）探索镇痛 萜类化合物对不同疼痛模式的潜力。这个跨学科的项目利用互补的 Quave实验室在民族植物学和药物化学方面的专业知识， 神经免疫学和疼痛

ABSTRACT Chronic pain affects up to 1/5 of the world population. It is a debilitating health condition with significant socio- economic costs. Management of chronic pain continues to present therapeutic hurdles, and the utility of opiate analgesics has reached a plateau as more and more people have fallen victim to opiate addiction, contributing to the opiate crisis. Plants of the Cannabaceae family, such as Cannabis sativa (cannabis) and Humulus lupulus (hops), have a long history of traditional use in the mitigation of pain and inflammation, yet the mechanistic basis for these activities and the identities of the compounds responsible are not well understood. In the proposed work, we will take a multidisciplinary approach to investigate the analgesic effects of terpenes from the Cannabaceae family. Rather than use cannabis, whose study and product market is complicated by conflicting federal and state laws, we will focus on terpenes found in hops, which is a related species and shares a very similar terpene profile. We will acquire botanically authenticated hops cones materials for extraction by supercritical CO2 methods to create terpene enriched and depleted hops extracts for study, enabling assessment of synergy between terpenes. We will also use individual terpene standards in our proposed studies. Extracts will be chemically characterized by GC-MS and NMR and assessed for bioactivity using in vitro models to assess the capacity to affect excitability and sensitization of cultured DRG nociceptive neurons. We will analyze the ability of Hops extracts or candidate terpenes to modulate nociceptor TRPV1 sensitization by inflammatory stimuli using calcium imaging, and examine the capacity of terpenes to modulate nociceptor excitability use whole cell patch clamp electrophysiology. PCA and Compound Activity Mapping analyses on chemical features and bioactivities observed will guide selection of the most active constituents, including consideration of synergistic combinations of terpenes. In the second part of our project, we will assess the efficacy of the most promising hops terpene-enriched extracts or isolated terpenes to mitigate pain and immune activation in animal models of inflammatory (CFA) and neuropathic (SNI) pain. Plasma and tissues from mice will also be assessed for the levels of specific terpenes and their metabolites via a targeted high-resolution metabolomics approach. In addition to behavioral analysis, we will analyze plantar and DRG immune activation, as well as microglia and astrocyte activation in the spinal cord. This study is responsive to the RFA-AT-19-009 objectives to 1) investigate the mechanisms by which terpenes may affect pain pathways, including ascending and/or descending neural pathways, cellular and molecular signaling pathways, neuroimmune interactions, or other innovative regulatory pathways related to pain; and to 2) explore analgesic potential of terpenes for different pain modalities. This interdisciplinary project leverages the complementary expertise of the Quave laboratory in ethnobotany and medicinal chemistry, and the Chiu laboratory in neuroimmunology and pain.