Mechanistic studies on analgesic effects of terpene enriched extracts from hops

摘要 慢性疼痛影响着多达五分之一的世界人口。这是一种令人衰弱的健康状况，具有重大的社会- 经济成本。慢性疼痛的治疗仍然存在治疗障碍，阿片类药物的使用 随着越来越多的人成为阿片成瘾的受害者，止痛药已经达到了平台期，这对 阿片类药物危机。大麻科植物，如大麻属(大麻属)和胡萝卜属 狼尾草(啤酒花)，在缓解疼痛和炎症方面有很长的传统使用历史，但 这些活动的机制基础和负责化合物的身份还没有被很好地理解。 在拟议的工作中，我们将采取多学科的方法来研究萜类化合物的镇痛作用。 从大麻科植物中提取。而不是使用大麻，大麻的研究和产品市场因 相互冲突的联邦和州法律，我们将集中在啤酒花中发现的萜类化合物，这是一个相关的物种和 具有非常相似的萜烯特征。我们将购买经植物鉴定的啤酒花球果材料 用超临界二氧化碳方法提取，以产生富含和贫化的啤酒花提取物以供研究， 能够评估萜类化合物之间的协同作用。我们还将在我们的 建议进行的研究。萃取物将通过GC-MS和核磁共振进行化学表征，并进行生物活性评估 用体外模型评价培养的背根节伤害性感受器对兴奋性和敏感化的影响 神经元。我们将分析啤酒花提取物或候选萜类化合物调节伤害感受器TRPV1的能力 使用钙成像技术对炎症刺激进行增敏，并检测萜类化合物的调节能力。 伤害性感受器兴奋性采用全细胞膜片钳电生理学。主成分分析与化合物活度作图 对观察到的化学特征和生物活性的分析将指导选择最活跃的成分， 包括考虑萜类化合物的协同组合。在我们项目的第二部分，我们将 评估最有希望的啤酒花萜类提取物或分离的萜类化合物的止痛效果 在炎症(CFA)和神经病理性(SNI)疼痛的动物模型中进行免疫激活。血浆和 来自小鼠的组织也将被评估特定的萜类及其代谢物的水平 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背根节 免疫激活，以及脊髓内小胶质细胞和星形胶质细胞的激活。这项研究是对 RFA-AT-19-009的目标是1)研究萜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控途径；以及2)探索止痛药 不同疼痛方式下萜类化合物的潜力。这一跨学科项目充分利用了 夸夫实验室在民族植物学和药物化学方面的专业知识，以及赵实验室在 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.