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: 10387370
• 负责人: Alexander Mikesell
• 金额: $ 4.68万
• 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/10387370
• 关键词: Afferent Neurons; Agonist; Animals; Antineoplastic Agents; Behavioral; Behavioral Assay; Cells; Chemicals; Chemotherapy-induced peripheral neuropathy; Chronic; Data; Development; Electrophysiology (science); Epidermis; Exhibits; Fiber; Goals; Growth; Human; Hyperalgesia; Hypersensitivity; Injury; Ion Channel; Ion Channel Gating; Knockout Mice; Mass Spectrum Analysis; Mechanical Stimulation; Mechanics; Mediating; Molecular Biology; Mus; Nerve; Neuraxis; Neurons; Neuropathy; Paclitaxel; Pain; Patients; Peripheral; Peripheral Nervous System Diseases; Property; Role; Signal Transduction; Skin; Skin Tissue; Small Interfering RNA; Sodium; Tactile; Testing; Therapeutic; Tissue Donors; Touch sensation; Transgenic Mice; Work; afferent nerve; base; behavioral response; biomedical scientist; career; cell type; chronic neuropathic pain; chronic pain; inflammatory pain; injured; keratinocyte; knock-down; mechanical allodynia; mechanical stimulus; mechanotransduction; mouse model; nerve injury; painful neuropathy; patch clamp; pre-clinical; 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.

项目概要 角质形成细胞是表皮中最丰富的细胞类型，通过检测和介导正常的触觉 将触觉信息编码到感觉神经元。然而，尚不清楚角质形成细胞机械转导是否 导致组织损伤后的慢性机械性疼痛。我的初步数据表明角质形成细胞 从神经受损的动物中分离出来的对机械刺激敏感。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敏化是否有助于发育 触摸引起神经性疼痛。我将专门检查这种敏化是否是由机械介导的 门控离子通道 PIEZO1，我的初步数据表明它是主要的角质形成细胞机械转导器。 在目标 1 中，我假设角质形成细胞 PIEZO1 的敏化有助于慢性疾病的发展 化疗引起的周围神经病变（CIPN）小鼠模型中的机械性疼痛。我会利用表皮 特定 PIEZO1 敲除小鼠和机械行为测定以确定 PIEZO1 缺失是否能提供保护 对抗 CIPN 机械性疼痛（目标 1A）。此外，我将利用离体皮肤神经（Aim 1B）和全细胞 膜片钳（目标 1C）记录以确定角质形成细胞 PIEZO1 的丢失是否会降低 CIPN 感觉神经和 角质形成细胞机械超敏反应。在目标 2 中，我假设 PIEZO1 介导人类角质形成细胞 力传导。我将利用全细胞贴片电生理学和 PIEZO1 靶向 siRNA 来确定 如果 PIEZO1 的敲低会降低人类角质形成细胞的机械敏感性（目标 2A）。接下来，我将确定是否 与化疗紫杉醇一起孵育会使人角质形成细胞对机械刺激敏感，如果 PIEZO1 敲低可降低这种超敏反应（目标 2B）。最后，我将使用质谱来研究 在初始条件下和以下条件下通过 PIEZO1 激活从人角质形成细胞释放信号因子 紫杉醇治疗（目标 2C）。这些目标共同将决定角质形成细胞是否增强诱发机械力 神经性损伤后的疼痛，如果表皮 PIEZO1 可能有潜在的靶点来缓解神经性损伤 疼痛。