Epidermal keratinocytes mediate mechanical pain following neuropathic injury

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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.