Myocardin-Related Transcription Factor A (MRTF-A) Regulates the Balance between Adipogenesis and Osteogenesis of Human Adipose Stem Cells
Adipogenesis of hASCs treated with MRTF-A inhibitors at 21 d. hASCs were cultured 21 d in BM or AM conditions supplemented with 15 or 20 μM CCG-1423 or 10 or 12 μM CCG-100602 inhibitors and stained with ORO method. (a) Representative fluorescence images of hASCs stained with ORO for intracellular lipid accumulation followed by nuclei staining with DAPI. Fluorescence images were taken with Alexa 546 for ORO (red) and DAPI (blue) filters, and 20x magnification. Scale bars 100 μm. (b) Lipid droplet areas were quantified from the ORO-stained fluorescence images with a custom analysis pipeline designed for CellProfiler. The graphs present the total ORO-stained area or lipid droplet clusters with a diameter of over 5 μm as percentages of the total image area. for BM +20 μM CCG-1423 and AM +20 μM CCG-1423, other conditions images (15 independent biological replicates from 4 donors). (c) Representative Plin1- and DAPI-stained samples of MTRF-A inhibitor-treated hASCs at 21 d imaged with a fluorescence microscope using Alexa 488 for Plin1 (green) and DAPI (blue) filters, and 40x magnification. The lowest row represents 4x digital zoom of the white rectangles in AM images. Scale bars 100 μm. BM: basic medium; AM: adipogenic medium; ORO: Oil Red O.
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