Endoplasmic Reticulum Stress Responses in Pain

疼痛中的内质网应激反应

基本信息

批准号：
10337200
负责人：
Mario Danilo Boada
金额：
$ 40.42万
依托单位：
WAKE FOREST UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-15 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10337200
关键词：
Acetic Acids Acute Pain Address Affect Behavior Behavioral Bioinformatics Biological Assay Biology Cells Cluster Analysis Communities Development Dinoprostone Electrophysiology (science)Endoplasmic Reticulum Exhibits Foundations Gene Expression Genes Genetic Transcription Human Hypersensitivity Immune Immunology In Situ Incidence Individual Infiltration Inflammation Inflammatory Injury Knockout Mice Label Leukocytes Ligation Medical Modeling Molecular Mus Nanotechnology Nerve Degeneration Neuroimmune Neurons Nociceptors Operative Surgical Procedures Outcome Pain Patients Pattern Pattern recognition receptor Peripheral Peripheral nerve injury Pharmacology Phenotype Phosphotransferases Positioning Attribute Preventive Process Production Prostaglandins Protein Biosynthesis Proteins Quality of life Recovery Recovery of Function Reflex action Regulation Resolution Role Signal Transduction Social Impacts Surgical incisions Technology Testing Therapeutic Tissues Toll-like receptors Tumor-infiltrating immune cells XBP1 gene arm base biological adaptation to stress chronic inflammatory disease chronic neuropathic pain chronic pain chronic pain management clinically relevant conditional knockout cytokine economic impact effective therapy endoplasmic reticulum stress genetic signature genome-wide immune activation improved in vivo inflammatory pain inhibitor injured innovation loss of function misfolded protein mouse model nerve injury neurobiotin novel painful neuropathy partial recovery prevent programs recruit response sciatic nerve single-cell RNA sequencing small molecule tool transcriptomics

项目摘要

Summary Peripheral nerve injury activates pattern recognition receptors (i.e. Toll-like receptors) in immune cells, thus triggering and maintaining inflammation, and ultimately determining the perpetuation of pain. Immune cell activation demands high levels of protein synthesis, folding and secretion, which are regulated by the endoplasmic reticulum (ER). An excessive demand in protein handling can evoke ER stress (accumulation of misfolded proteins) and consequently trigger robust activation of the unfolded protein response (UPR). IRE1α- XBP1 is the most evolutionarily conserved arm of the UPR and can be directly activated via Toll-like receptor engagement to promote the expression of pro-inflammatory factors. We have unveiled that conditional knockout (cKO) mice devoid of IRE1α/XBP1 in immune cells (Ern1/Xbp1f/f-Vav1cre), display decreased PGE2 production in vivo, reduced nociceptor responsiveness, and faster resolution of non-reflexive pain-related behaviors following paw incision. Similarly, these cKO mice exhibit improved recovery after partial sciatic nerve ligation (PSNL). Through unbiased genome-wide transcriptomic analyses we found that IRE1α-XBP1 signaling in leukocytes is critically required for the induction of prostanoids, cytokines and other novel factors such as Nupr1 (associated with chronic inflammatory diseases in humans). Of note, IRE1α-XBP1 overactivation has been correlated with painful or inflammatory conditions in humans. Therefore, we hypothesize that IRE1α-XBP1 signaling in leukocytes governs peripheral neuro-immune interactions and the development of chronic pain. Using cutting-edge experimental approaches, we will accomplish the following specific aims: 1) Determine how IRE1α-XBP1 signaling dictates the dynamics of immune cell infiltration and molecular changes that drive behavioral non-reflexive hypersensitivity following peripheral nerve injury. We postulate that the IRE1α-XBP1 arm operates as a key modulator of pro-algesic factors in immune cells, and that Nupr1 is a novel XBP1- dependent factor (using ChIP PCR) implicated in PSNL. 2) Establish how IRE1α-XBP1 activation governs individualized immune cellular reprogramming and how this drives the cross-talk with nociceptor afferents following peripheral nerve injury. Our hypothesis is that the reduced hypersensitivity observed in our cKO mice following PSNL is determined by discrete gene signatures in specific injury-infiltrating leukocyte subsets (using single cell RNA sequencing), and by the acquisition of a less responsive phenotype in nociceptors (via in vivo intracellular DRG recordings). 3) Define the therapeutic potential of inhibiting IRE1a to accelerate recovery from PSNL. We posit that pharmacological inhibition of IRE1α using MKC8866 (RNAase domain inhibitor) or KIRA6 (kinase domain inhibitor) will prevent or treat chronic neuropathic pain. Our team has expertise in pain biology and neuroimmune interactions (Romero-Sandoval), immunology and ER stress biology (Cubillos-Ruiz), in vivo electrophysiology (Boada), and scRNAseq and bioinformatics (Miller). Thus, we are uniquely positioned to test our innovative hypothesis and contribute to the development of novel non-narcotic treatments for chronic pain.

概括外周神经损伤激活了免疫细胞中的模式识别受体（即toll样受体），因此触发和维持炎症，并最终确定疼痛的延续。免疫细胞激活需要高水平的蛋白质合成，折叠和分泌，这些蛋白质由内质网（ER）。蛋白质处理的过量需求会引起ER应力（积累错误折叠的蛋白质），因此触发了展开的蛋白质反应（UPR）的强大激活。 ire1α- XBP1是UPR的最进化配置的臂，可以直接通过类似收费的接收器激活参与以促进促炎性因素的表达。我们已经公布了有条件的淘汰赛（CKO）在免疫细胞中没有IRE1α/XBP1的小鼠（ERN1/XBP1F/F-VAV1CRE），显示改善的PGE2产生在体内，降低伤害感受器的反应性，并更快地解决非反射性疼痛相关行为按照爪子切口。同样，这些CKO小鼠在部分坐骨神经结扎后暴露了改善的恢复（PSNL）。通过无偏基因组的转录组分析，我们发现IRE1α-XBP1信号在诱导前列腺素，细胞因子和其他新型因素（例如NUPR1）至关重要的白细胞是必需的（与人类的慢性炎症性疾病有关）。值得注意的是，IRE1α-XBP1过度活化已经与人类的疼痛或炎症状况相关。因此，我们假设IRE1α-XBP1 白细胞中的信号传导控制周围神经免疫相互作用和慢性的发展疼痛。使用尖端的实验方法，我们将完成以下特定目的：1）确定 IRE1α-XBP1信号传导如何决定免疫球浸润和驱动分子变化的动力学周围神经损伤后的行为非反射性超敏反应。我们假设IRE1α-XBP1 ARM是免疫细胞中促脂因素的关键调节剂，NUPR1是一种新型的XBP1-- PSNL中实现的依赖因子（使用芯片PCR）。 2）确定IRE1α-XBP1激活如何控制个性化的免疫细胞重编程以及这如何驱动与伤害感受器的串扰外周神经损伤后。我们的假设是在我们的CKO小鼠中观察到的高敏性降低遵循PSNL是通过特定损伤浸润白细胞亚群中的离散基因特征确定的（使用单细胞RNA测序），通过在伤害感受器中获得响应较低的表型（通过体内）细胞内DRG记录）。 3）定义抑制IRE1A加速恢复的治疗潜力 psnl。我们使用MKC8866（RNAase结构域抑制剂）或KIRA6对IRE1α的药物抑制作用阳性（激酶结构域抑制剂）将预防或治疗慢性神经性疼痛。我们的团队在疼痛生物学方面有专业知识和神经免疫相互作用（Romero-Sandoval），免疫学和ER应激生物学（Cubillos-Ruiz），体内电生理学（BOADA）以及SCRNASEQ和生物信息学（Miller）。那就是我们独特地测试我们的创新假设，有助于开发新的非麻醉性治疗方法，以缓解慢性疼痛。