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.

项目摘要 角质形成细胞是表皮中含量最丰富的一种细胞类型，通过检测和检测细胞中的角质形成细胞来调节正常的触觉。 将触觉信息编码到感觉神经元。然而，目前尚不清楚角质形成细胞是否通过机械转导 会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形成细胞 从神经损伤的动物中分离出来的动物对机械刺激敏感。考虑到这一发现，我建议 探讨损伤诱导的角质形成细胞机械转导敏化是否参与了角质形成细胞的发生。 触摸会引起神经病理性疼痛。我将具体检查这种敏感化是否由机械地 门控离子通道PIEZO1，我的初步数据表明，它是角质形成细胞的主要机械换能器。 在目标1中，我假设角质形成细胞PIEZO1的敏化有助于慢性 化疗诱导的周围神经病变(CIPN)小鼠模型中的机械性疼痛。我会用表皮 特异性PIEZO1基因敲除小鼠和确定PIEZO1基因缺失是否具有保护作用的机械行为分析 抗CIPN机械性疼痛(目标1A)。此外，我将利用体外皮肤神经(目标1B)和全细胞 膜片钳(AIM 1C)记录以确定角质形成细胞PIEZO1的丢失是否减少CIPN感觉神经和 角质形成细胞机械性超敏反应。在目标2中，我假设PIEZO1介导人角质形成细胞 机械转导。我将利用全细胞膜片电生理学和PIEZO1靶向siRNA来确定 如果敲除PIEZO1可降低人角质形成细胞的机械敏感性(目标2A)。接下来，我将确定是否 与化疗药物紫杉醇孵育使人角质形成细胞对机械刺激敏感 PIEZO1基因敲除可降低这种过敏性(目标2B)。最后，我将使用质谱学来研究 PIEZO1在幼稚及以下条件下激活人角质形成细胞释放信号因子 紫杉醇治疗(目标2C)。这些目标将共同决定角质形成细胞是否会增强诱发的机械 神经性损伤后的疼痛以及表皮PIEZO1是否可能成为缓解神经病理性损伤的潜在靶点 疼痛。