Endoplasmic Reticulum Stress Responses in Pain

疼痛中的内质网应激反应

• 批准号: 10094265
• 负责人: Mario Danilo Boada
• 金额: $ 39.77万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 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/10094265
• 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; clinically relevant; conditional knockout; cytokine; economic impact; effective therapy; endoplasmic reticulum stress; genetic signature; genome-wide; immune activation; improved; in vivo; inflammatory pain; inhibitor/antagonist; 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

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.

总结 外周神经损伤激活免疫细胞中的模式识别受体（即Toll样受体）， 引发和维持炎症，并最终决定疼痛的持续。免疫细胞 激活需要高水平的蛋白质合成，折叠和分泌，这是由蛋白质合成，折叠和分泌调节。 内质网（ER）。对蛋白质处理的过度需求可以引起ER应激（ER的积累）。 错误折叠的蛋白质），并因此触发未折叠蛋白质反应（UPR）的强烈激活。IRE1α- XBP 1是UPR中进化上最保守的一条臂，可通过Toll样受体直接激活 参与促进促炎因子的表达。我们已经公布了条件性击倒 (cKO)在免疫细胞中缺乏IRE 1 α/XBP 1的小鼠（Ern 1/Xbp 1f/f-Vav 1cre）显示PGE 2产生减少 在体内，伤害感受器反应性降低，非反射性疼痛相关行为的消退更快 在爪切口之后。类似地，这些cKO小鼠在部分坐骨神经结扎后表现出改善的恢复 （PSNL）。通过无偏的全基因组转录组分析，我们发现，IRE 1 α-XBP 1信号转导在 白细胞是诱导前列腺素类、细胞因子和其他新因子（如Nupr 1）所必需的 （与人类慢性炎症性疾病有关）。值得注意的是，IRE 1 α-XBP 1的过度激活已经被证实是一种基因突变。 与人类的疼痛或炎症有关。因此，我们假设IRE 1 α-XBP 1 白细胞中的信号传导控制外周神经免疫相互作用和慢性 痛苦使用尖端的实验方法，我们将实现以下具体目标：1）确定 IRE 1 α-XBP 1信号传导如何决定免疫细胞浸润的动力学和驱动免疫细胞浸润的分子变化 周围神经损伤后的行为非反射性超敏反应我们假设IRE 1 α-XBP 1 手臂作为免疫细胞中促痛觉因子的关键调节剂，Nupr 1是一种新的XBP 1- 依赖因子（使用ChIP PCR）参与PSNL。2)确定IRE 1 α-XBP 1激活如何支配 个体化免疫细胞重编程以及这如何驱动与伤害感受器传入的串扰 周围神经损伤后。我们的假设是，在我们的cKO小鼠中观察到的超敏反应降低 随后的PSNL是通过特定损伤浸润性白细胞亚群中的离散基因标签确定的（使用 单细胞RNA测序），以及通过在伤害感受器中获得反应性较低的表型（通过体内 细胞内DRG记录）。3)确定抑制IRE 1a以加速从 PSNL。我们证实，使用MKC 8866（RNA酶结构域抑制剂）或KIRA 6 （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长 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）和scRNAseq和生物信息学（米勒）。因此，我们处于独特的地位， 我们的创新假设，并有助于开发新的非麻醉性治疗慢性疼痛。