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利用抑制脂肪来源干细胞的脂肪生成进行毒性预测。

Using inhibition of the adipogenesis of adipose-derived stem cells for toxicity prediction.

作者信息

Abud Ana Paula Ressetti, Paschoal Ariane Caroline Campos, Kuligovski Crisciele, Caruso Rodrigo Rêgo Barros, Dallagiovanna Bruno, de Aguiar Alessandra Melo

机构信息

Rede de Plataformas Tecnológicas FIOCRUZ - Bioensaios com Métodos alternativos em Citotoxicidade, Rua Professor Algacyr Munhoz Mader, 3775, Instituto Carlos Chagas, FIOCRUZ Paraná, Curitiba, PR 81350-010, Brazil.

Laboratório de Biologia Básica de Células-Tronco, Rua Professor Algacyr Munhoz Mader, 3775, Instituto Carlos Chagas, FIOCRUZ Paraná, Curitiba, PR 81350-010, Brazil.

出版信息

MethodsX. 2021 Sep 14;8:101515. doi: 10.1016/j.mex.2021.101515. eCollection 2021.

DOI:10.1016/j.mex.2021.101515
PMID:34754786
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8564732/
Abstract

In vitro stem cell models are used as alternatives to animal models and are important tools for cytotoxicity studies. Researchers can determine the effects of test substances on human cells by evaluating cell viability and differentiation. Here, we describe an in vitro model to quantify adipogenesis based on the Nile red staining of specific lipid droplets and the emission of basic lipids from human adipose tissue-derived mesenchymal stromal cells (AD-MSCs) in the presence of test substances. This assay allows for the prediction of toxicity based on the inhibition of adipogenesis in vitro in a 96-well format. The differentiation of a progenitor cell into a specialized cell, the adipocyte, is easy to monitor and quantify, making this a simple assay. The fluorescence staining of nuclei and lipid droplets is measured after 14 days of cell differentiation to determine cell number and assess cell differentiation using high-content imaging analysis, thus allowing for the identification of chemicals that impact differentiation. We also describe a protocol to assess adipocyte differentiation by fluorescence intensity using a multiplate reader.•Researchers can utilize the protocol described here for many purposes to evaluate in vitro adipogenesis.•

摘要

体外干细胞模型被用作动物模型的替代物，是细胞毒性研究的重要工具。研究人员可以通过评估细胞活力和分化来确定测试物质对人类细胞的影响。在此，我们描述了一种体外模型，该模型基于特定脂滴的尼罗红染色以及在测试物质存在下从人脂肪组织来源的间充质基质细胞（AD-MSCs）中碱性脂质的发射来量化脂肪生成。该测定法允许以96孔板形式基于体外脂肪生成的抑制来预测毒性。祖细胞向特化细胞脂肪细胞的分化易于监测和量化，使其成为一种简单的测定法。在细胞分化14天后测量细胞核和脂滴的荧光染色，以确定细胞数量并使用高内涵成像分析评估细胞分化，从而能够鉴定影响分化的化学物质。我们还描述了一种使用多板读数器通过荧光强度评估脂肪细胞分化的方案。•研究人员可以将此处描述的方案用于许多目的，以评估体外脂肪生成。•

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/4ae91d2bf30a/gr9.jpg
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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/4ae91d2bf30a/gr9.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/d28c11195962/ga1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/adb31ef2327d/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/47434f8b868a/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/111095696b61/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/b10d68605d63/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/41e1d6852fe4/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/102ac3627a75/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/ec5d829bf26d/gr7.jpg
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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3353/8564732/4ae91d2bf30a/gr9.jpg

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