Using inhibition of the adipogenesis of adipose-derived stem cells in vitro for toxicity prediction
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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.• With this method, it is possible to reduce the use of animals. Graphical abstract AD-MSCs adipogenesis-inhibition assay in culture. Schematic diagram showing the adipogenic inhibition assay. In this method, AD-MSCs are induced to differentiate and are maintained for 14 days in the presence of different concentrations of test chemicals. Therefore, it is possible to visualize the effects of the substances during the cell differentiation process. The results can be obtained by a high-content imaging system or microplate reader.Image, graphical abstract
SUBMITTER: Abud A
PROVIDER: S-EPMC8564732 | biostudies-literature |
REPOSITORIES: biostudies-literature
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