University of Padua

Calcium signaling in the photodamaged skin

Donati, Viola and Peres, Chiara and Nardin, Chiara and Scavizzi, Ferdinando and Raspa, Marcello and Ciubotaru, Catalin D. and Bortolozzi, Mario and Mammano, Fabio (2021) Calcium signaling in the photodamaged skin. [Data Collection]

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BACKGROUND The mammalian skin, the body’s largest single organ, is a highly organized tissue that forms an essential barrier against dehydration, pathogens, light and mechanical injury. Damage triggers perturbations of the cytosolic free Ca2+ concentration ([Ca2+]c) that spread from cell to cell (known as intercellular Ca2+ waves) in different epithelia, including epidermis. Ca2+ waves are considered a fundamental mechanism for coordinating multicellular responses, however the mechanisms underlying their propagation in the damaged epidermis are incompletely understood. AIM OF THE PROJECT To dissect the molecular components contributing to Ca2+ wave propagation in murine model of epidermal photodamage. METHODS To trigger Ca2+ waves, we used intense and focused pulsed laser radiation and targeted a single keratinocyte of the epidermal basal layer in the earlobe skin of live anesthetized mice. To track photodamage-evoked Ca2+ waves, we performed intravital multiphoton microscopy in transgenic mice with ubiquitous expression of the sensitive and selective Ca2+ biosensor GCaMP6s. To dissect the molecular components contributing to Ca2+ wave propagation, we performed in vivo pharmacological interference experiments by intradermal microinjection of different drugs. EXPERIMENTAL RESULTS The major effects of drugs that interfere with degradation of extracellular ATP or P2 purinoceptors suggest that Ca2+ waves in the photodamaged epidermis are primarily due to release of ATP from the target cell, whose plasma membrane integrity was compromised by laser irradiation. The limited effect of the Connexin 43 (Cx43) selective inhibitor TAT-Gap19 suggests ATP-dependent ATP release though connexin hemichannels (HCs) plays a minor role, affecting Ca2+ wave propagation only at larger distances, where the concentration of ATP released from the photodamaged cell was reduced by the combined effect of passive diffusion and hydrolysis due to the action of ectonucleotidases. The ineffectiveness of probenecid suggests pannexin channels have no role. As GCaMP6s signals in bystander keratinocytes were augmented by exposure to the Ca2+ chelator EGTA in the extracellular medium, the corresponding transient increments of the [Ca2+]c should be ascribed primarily to Ca2+ release from the ER, downstream of ATP binding to P2Y purinoceptors, with Ca2+ entry through plasma membrane channels playing a comparatively negligible role. The effect of thapsigargin (a well-known inhibitor of SERCA pumps) and carbenoxolone (a recently recognized inhibitor of Ca2+ release through IP3 receptors) support this conclusion. CONCLUSIONS The one presented here is an experimental model for accidental skin injury that may also shed light on the widespread medical practice of laser skin resurfacing, used to treat a range of pathologies from photodamage and acne scars to hidradenitis suppurativa and posttraumatic scarring from basal cell carcinoma excision. The results of our experiments support the notion that Ca2+ waves reflect chiefly the sequential activation of bystander keratinocytes by the ATP released through the compromised plasma membrane of the cell hit by laser radiation. We attributed the observed increments of the [Ca2+]c chiefly to signal transduction through purinergic P2Y receptors. Several studies have highlighted fundamental roles of P2Y receptors during inflammatory and infectious diseases, and the initial phase of wound healing involves acute inflammation. In addition, hyaluronan is a major component of the extracellular matrix and its synthesis is rapidly upregulated after tissue wounding via P2Y receptor activation. It is tempting to speculate that response coordination after injury in the epidermis occurs via propagation of the ATP-dependent intercellular Ca2+ waves described in this work.

DOI: 10.25430/
Keywords: epidermis, calcium, purinergic signaling, biosensors, skin, laser-induced intra-epidermal photodamage, wound healing, connexins, pannexins
Subjects: Physical:Sciences and Engineering > Condensed Matter Physics: Structure, electronic properties, fluids, nanosciences, biological physics > Physics of biological systems
Department: Departments > Dipartimento di Fisica e Astronomia "Galileo Galilei" (DFA)
Depositing User: Viola Donati
Date Deposited: 18 Oct 2021 07:24
Last Modified: 18 Oct 2021 07:24
Donati, Violaviola.donati@ibbc.cnr.itUNSPECIFIED
Type of data: Database
Research funder: University of Padova, Fondazione Telethon
Grant number: Grant No. BIRD187130, Grant No. GGP19148
Collection period:
September 2020October 2021
Resource language: English
Metadata language: English
Publisher: Research Data Unipd
Related publications:
Date: 15 October 2021
Copyright holders: The Author

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