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利用 FLIM 评估 iPS 细胞来源的真皮乳头细胞与分化细胞之间的代谢状态差异。

FLIM for Evaluation of Difference in Metabolic Status between Native and Differentiated from iPSCs Dermal Papilla Cells.

机构信息

Institute of Experimental Oncology and Biomedical Technologies, Privolzhsky Research Medical University, 603005 Nizhny Novgorod, Russia.

Koltzov Institute of Developmental Biology, Russian Academy of Sciences, 119334 Moscow, Russia.

出版信息

Cells. 2022 Sep 1;11(17):2730. doi: 10.3390/cells11172730.

DOI:10.3390/cells11172730
PMID:36078136
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC9454864/
Abstract

iPSCs and their derivatives are the most promising cell sources for creating skin equivalents. However, their properties are not fully understood. In addition, new approaches and parameters are needed for studying cells in 3D models without destroying their organization. Thus, the aim of our work was to study and compare the metabolic status and pH of dermal spheroids created from dermal papilla cells differentiated from pluripotent stem cells (iDP) and native dermal papilla cells (hDP) using fluorescence microscopy and fluorescence lifetime imaging microscopy (FLIM). For this purpose, fluorescence intensities of NAD(P)H and FAD, fluorescence lifetimes, and the contributions of NAD(P)H, as well as the fluorescence intensities of SypHer-2 and BCECF were measured. iDP in spheroids were characterized by a more glycolytic phenotype and alkaline intra-cellular pH in comparison with hDP cells. Moreover, the metabolic activity of iDP in spheroids depends on the source of stem cells from which they were obtained. So, less differentiated and condensed spheroids from iDP-iPSDP and iDP-iPSKYOU are characterized by a more glycolytic phenotype compared to dense spheroids from iDP-DYP0730 and iDP-hES. FLIM and fluorescent microscopy in combination with the metabolism and pH are promising tools for minimally invasive and long-term analyses of 3D models based on stem cells.

摘要

iPSCs 及其衍生物是用于创建皮肤等效物的最有前途的细胞来源。然而，它们的特性尚未完全了解。此外，还需要新的方法和参数来研究 3D 模型中的细胞，而不会破坏其组织。因此，我们的工作旨在使用荧光显微镜和荧光寿命成像显微镜 (FLIM) 研究和比较从多能干细胞 (iDP) 和天然真皮乳头细胞 (hDP) 分化而来的真皮乳头细胞 (iDP) 形成的真皮球体的代谢状态和 pH 值。为此，测量了 NAD(P)H 和 FAD 的荧光强度、荧光寿命以及 NAD(P)H 的贡献，以及 SypHer-2 和 BCECF 的荧光强度。与 hDP 细胞相比，iDP 球体中的细胞表现出更具糖酵解表型和碱性细胞内 pH 值。此外，iDP 球体中的代谢活性取决于它们获得的干细胞来源。因此，与 iDP-DYP0730 和 iDP-hES 的致密球体相比，来自 iDP-iPSDP 和 iDP-iPSKYOU 的分化程度较低且较密集的球体表现出更具糖酵解表型。FLIM 和荧光显微镜结合代谢和 pH 值是基于干细胞的 3D 模型进行微创和长期分析的有前途的工具。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/96b8/9454864/ad3334cbec57/cells-11-02730-g011.jpg
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