Epidermal keratinocytes mediate mechanical pain following neuropathic injury

表皮角质形成细胞介导神经性损伤后的机械性疼痛

• 批准号: 10576798
• 负责人: Alexander Mikesell
• 金额: $ 4.77万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10576798
• 关键词: Afferent Neurons; Agonist; Animals; Antineoplastic Agents; Behavioral; Behavioral Assay; Cells; Central Nervous System; Chemicals; Chemotherapy-induced peripheral neuropathy; Chronic; Data; Development; Electrophysiology (science); Epidermis; Exhibits; Fiber; Goals; Growth; Human; Hyperalgesia; Hypersensitivity; Incubated; Injury; Ion Channel; Ion Channel Gating; Knockout Mice; Mass Spectrum Analysis; Mechanical Stimulation; Mechanics; Mediating; Molecular Biology; Mus; Nerve; Neurons; Neuropathy; Paclitaxel; Pain; Patients; Peripheral; Peripheral Nervous System Diseases; Piezo 1 ion channel; Pre-Clinical Model; Property; Role; Signal Transduction; Skin; Skin Tissue; Small Interfering RNA; Sodium Channel; Tactile; Testing; Therapeutic; Tissue Donors; Touch sensation; Transgenic Mice; Work; afferent nerve; behavioral response; biomedical scientist; career; cell type; chronic neuropathic pain; chronic pain; human model; inflammatory pain; injured; keratinocyte; knock-down; mechanical allodynia; mechanical stimulus; mechanotransduction; mouse model; nerve injury; painful neuropathy; patch clamp; release factor; sensory neuropathy; side effect; skills; tissue injury; voltage

Project Summary Keratinocytes, the most abundant cell type in the epidermis, mediate normal touch sensation by detecting and encoding tactile information to sensory neurons. However, it is unknown if keratinocyte mechanotransduction contributes to chronic mechanical pain following tissue injury. My preliminary data suggest that keratinocytes isolated from nerve injured animals are sensitized to mechanical stimulation. Considering this finding, I propose to investigate if injury induced sensitization of keratinocyte mechanotransduction contributes to the development of touch evoked neuropathic pain. I will specifically examine if this sensitization is mediated by the mechanically gated ion channel PIEZO1, which my preliminary data suggests is a major keratinocyte mechanotransducer. Here in Aim 1, I hypothesize that sensitization of keratinocyte PIEZO1 contributes to the development of chronic mechanical pain in a mouse model chemotherapy-induced peripheral neuropathy (CIPN). I will utilize epidermal specific PIEZO1 knockout mice and mechanical behavioral assays to determine if loss of PIEZO1 protects against CIPN mechanical pain (Aim 1A). Additionally, I will utilize ex vivo skin nerve (Aim 1B) and whole-cell patch clamp (Aim 1C) recordings to determine if loss of keratinocyte PIEZO1 reduces CIPN sensory nerve and keratinocyte mechanical hypersensitivity. In Aim 2, I hypothesize that PIEZO1 mediates human keratinocyte mechanotransduction. I will utilize whole-cell patch electrophysiology and PIEZO1 targeted siRNA to determine if knockdown of PIEZO1 reduces human keratinocyte mechanical sensitivity (Aim 2A). Next, I will determine if incubation with the chemotherapeutic paclitaxel sensitizes human keratinocytes to mechanical stimulation and if PIEZO1 knockdown reduces this hypersensitivity (Aim 2B). Finally, I will use mass spectrometry to investigate signaling factors released from human keratinocytes by PIEZO1 activation under naïve conditions and following paclitaxel treatment (Aim 2C). Together these aims will determine if keratinocytes enhance evoked mechanical pain following neuropathic injury and if epidermal PIEZO1 may be potentially targeted to relieve neuropathic pain.

项目摘要 角质形成细胞是表皮中最丰富的细胞类型，通过检测和调节正常的触觉， 将触觉信息编码到感觉神经元。然而，角质形成细胞的机械转导是否 导致组织损伤后的慢性机械性疼痛。我的初步数据显示角质细胞 从神经损伤的动物中分离的细胞对机械刺激敏感。考虑到这一发现，我建议 研究损伤诱导的角质形成细胞机械转导的致敏是否有助于发展 触摸引起的神经性疼痛。我将特别检查这种致敏是否是由机械介导的， 门控离子通道PIEZO 1，我的初步数据表明，这是一个主要的角质形成细胞mechanotransducer。 在目的1中，我假设角质形成细胞PIEZO 1的致敏有助于慢性炎症的发展。 化学疗法诱导的周围神经病变（CIPN）小鼠模型中的机械性疼痛。我会利用表皮细胞 特异性PIEZO 1敲除小鼠和机械行为测定，以确定PIEZO 1的缺失是否保护 对抗CIPN机械性疼痛（Aim 1A）。此外，我将利用离体皮肤神经（Aim 1B）和全细胞 膜片钳（Aim 1C）记录以确定角质形成细胞PIEZO 1的损失是否减少CIPN感觉神经， 角质形成细胞机械过敏。在目的2中，我假设PIEZO 1介导人角质形成细胞 机械传导我将利用全细胞贴片电生理学和PIEZO 1靶向siRNA来确定 如果PIEZO 1的敲低降低人角质形成细胞的机械敏感性（Aim 2A）。接下来，我将确定 与化疗紫杉醇一起孵育使人角质形成细胞对机械刺激敏感， PIEZO 1敲低降低了这种超敏反应（目的2B）。最后，我会用质谱仪来研究 在初始条件下和之后通过PIEZO 1活化从人角质形成细胞释放的信号传导因子 紫杉醇治疗（Aim 2C）。这些目标将共同确定角质形成细胞是否增强诱发的机械 神经性损伤后的疼痛，以及如果表皮PIEZO 1可以潜在地靶向缓解神经性损伤， 痛苦