Dataset Information

A Novel Method for Visualizing Melanosome and Melanin Distribution in Human Skin Tissues.

ABSTRACT: Melanin incorporated into keratinocytes plays an important role in photoprotection; however, abnormal melanin accumulation causes hyperpigmentary disorders. To understand the mechanism behind the accumulation of excess melanin in the skin, it is essential to clarify the spatial distribution of melanosomes or melanin in the epidermis. Although several markers have been used to detect melanosomes or melanin, no suitable markers to determine the precise localization of melanin in the epidermis have been reported. In this study, we showed that melanocore-interacting Kif1c-tail (M-INK), a recently developed fluorescent probe for visualizing mature melanosomes, binds to purified melanin in vitro, and applied it for detecting melanin in human skin tissues. Frozen skin sections from different phototypes were co-stained for the hemagglutinin (HA)-tagged M-INK probe and markers of melanocytes or keratinocytes, and a wide distribution of melanin was observed in the epidermis. Analysis of the different skin phototypes indicated that the fluorescent signals of HA-M-INK correlated well with skin color. The reconstruction of three-dimensional images of epidermal sheets enabled us to observe the spatial distribution of melanin in the epidermis. Thus, the HA-M-INK probe is an ideal tool to individually visualize melanin (or melanosome) distribution in melanocytes and in keratinocytes in skin tissues.

SUBMITTER: Yoshikawa-Murakami C

PROVIDER: S-EPMC7697890 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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A Novel Method for Visualizing Melanosome and Melanin Distribution in Human Skin Tissues.

Yoshikawa-Murakami Chikako C Mizutani Yuki Y Ryu Akemi A Naru Eiji E Teramura Takashi T Homma Yuta Y Fukuda Mitsunori M

International journal of molecular sciences 20201112 22

Melanin incorporated into keratinocytes plays an important role in photoprotection; however, abnormal melanin accumulation causes hyperpigmentary disorders. To understand the mechanism behind the accumulation of excess melanin in the skin, it is essential to clarify the spatial distribution of melanosomes or melanin in the epidermis. Although several markers have been used to detect melanosomes or melanin, no suitable markers to determine the precise localization of melanin in the epidermis have ...[more]

PMID: 33198186

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Project description:The functional impact of intratumoral heterogeneity has been difficult to assess in the absence of a means to interrogate dynamic, live-cell biochemical events in the native tissue context of a human tumor. Conventional histological methods can reveal morphology and static biomarker expression patterns but do not provide a means to probe and evaluate tumor functional behavior and live-cell responsiveness to experimentally controlled stimuli. Here, we describe an approach that couples vibratome-mediated viable tissue sectioning with live-cell confocal microscopy imaging to visualize human parathyroid adenoma tumor cell responsiveness to extracellular calcium challenge. Tumor sections prepared as 300 micron-thick tissue slices retain viability throughout a >24 hour observation period and retain the native architecture of the parental tumor. Live-cell observation of biochemical signaling in response to extracellular calcium challenge in the intact tissue slices reveals discrete, heterogeneous kinetic waveform categories of calcium agonist reactivity within each tumor. Plotting the proportion of maximally responsive tumor cells as a function of calcium concentration yields a sigmoid dose-response curve with a calculated calcium EC50 value significantly elevated above published reference values for wild-type calcium-sensing receptor (CASR) sensitivity. Subsequent fixation and immunofluorescence analysis of the functionally evaluated tissue specimens allows alignment and mapping of the physical characteristics of individual cells within the tumor to specific calcium response behaviors. Evaluation of the relative abundance of intracellular PTH in tissue slices challenged with variable calcium concentrations demonstrates that production of the hormone can be dynamically manipulated ex vivo. The capability of visualizing live human tumor tissue behavior in response to experimentally controlled conditions opens a wide range of possibilities for personalized ex vivo therapeutic testing. This highly adaptable system provides a unique platform for live-cell ex vivo provocative testing of human tumor responsiveness to a range of physiological agonists or candidate therapeutic compounds.

Dataset Information

A Novel Method for Visualizing Melanosome and Melanin Distribution in Human Skin Tissues.

Publications

A Novel Method for Visualizing Melanosome and Melanin Distribution in Human Skin Tissues.

Similar Datasets

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