Dataset Information

Revisiting the Experimental Methods for Human Skin T-Cell Analysis.

ABSTRACT: Tissue-resident memory T cells exist in both the epidermis and the dermis in human skin. To analyze these cells, the skin needs to be incubated with dispase II to separate the two layers, that is, the epidermis and the dermis. The next step varies among researchers; the subsequent enzymatic digestion of the two layers is popular, whereas the spontaneous migration method can also be done. Scraping of these layers to yield skin T cells may reduce antigen modulation. This study aimed to determine each method's limitations. Dispase II incubation itself cleaves T-cell antigens. Therefore, further enzymatic digestion with collagenases strongly cleaves antigens. The scraping method yields skin T cells that are affected by dispase II as it is. However, skin T-cell yield is low. The spontaneous migration method recovers and/or upregulates antigens with T-cell activation and loses ∼20% of T cells in the floating sheets. However, there was no prominent bias regarding CD103 expression between emigrants and the remaining T cells in the sheets. There were 10⁴ and 10⁵ CD3⁺ T cells per 1 cm² of the epidermis and upper dermis, respectively. Collectively, each method has strengths and limitations to analyze both the epidermal and dermal T cells.

SUBMITTER: Sato T

PROVIDER: S-EPMC9127406 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature

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Revisiting the Experimental Methods for Human Skin T-Cell Analysis.

Sato Takuya T Ogawa Youichi Y Ishikawa Aoha A Nagasaka Yuka Y Kinoshita Manao M Shiokawa Ichiro I Shimada Shinji S Momosawa Akira A Kawamura Tatsuyoshi T

JID innovations : skin science from molecules to population health 20220323 4

Tissue-resident memory T cells exist in both the epidermis and the dermis in human skin. To analyze these cells, the skin needs to be incubated with dispase II to separate the two layers, that is, the epidermis and the dermis. The next step varies among researchers; the subsequent enzymatic digestion of the two layers is popular, whereas the spontaneous migration method can also be done. Scraping of these layers to yield skin T cells may reduce antigen modulation. This study aimed to determine e ...[more]

PMID: 35620704

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Project description:The evolutionarily ancient methoxyindoleamine, melatonin, has long perplexed investigators by its versatility of functions and mechanisms of action, which include the regulation of vertebrate pigmentation. Although first discovered through its potent skin-lightening effects in amphibians, melatonin's role in human skin and hair follicle pigmentation and its impact on melanocyte physiology remain unclear. Synthesizing our limited current understanding of this role, we specifically examine its impact on melanogenesis, oxidative biology, mitochondrial function, melanocyte senescence, and pigmentation-related clock gene activity, with emphasis on human skin, yet without ignoring instructive pointers from nonhuman species. Given the strict dependence of melanocyte functions on the epithelial microenvironment, we underscore that melanocyte responses to melatonin are best interrogated in a physiological tissue context. Current evidence suggests that melatonin and some of its metabolites inhibit both, melanogenesis (via reducing tyrosinase activity) and melanocyte proliferation by stimulating melatonin membrane receptors (MT1, MT2). We discuss whether putative melanogenesis-inhibitory effects of melatonin may occur via activation of Nrf2-mediated PI3K/AKT signaling, estrogen receptor-mediated and/or melanocortin-1 receptor- and cAMP-dependent signaling, and/or via melatonin-regulated changes in peripheral clock genes that regulate human melanogenesis, namely Bmal1 and Per1. Melatonin and its metabolites also accumulate in melanocytes where they exert net cyto- and senescence-protective as well as antioxidative effects by operating as free radical scavengers, stimulating the synthesis and activity of ROS scavenging enzymes and other antioxidants, promoting DNA repair, and enhancing mitochondrial function. We argue that it is clinically and biologically important to definitively clarify whether melanocyte cell culture-based observations translate into melatonin-induced pigmentary changes in a physiological tissue context, that is, in human epidermis and hair follicles ex vivo, and are confirmed by clinical trial results. After defining major open questions in this field, we close by suggesting how to begin answering them in clinically relevant, currently available preclinical in situ research models.

Dataset Information

Revisiting the Experimental Methods for Human Skin T-Cell Analysis.

Publications

Revisiting the Experimental Methods for Human Skin T-Cell Analysis.

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