Fig. 1

Porcine placental extract (PPE) decreased lipid droplet size in adipose-derived mesenchymal stromal/stem cells (ASC)-differentiated adipocytes (A) ASC were plated on a 96-well plate at a density of 1 × 10⁴ cells in complete media. After 24 h, cells were treated with various PPE concentrations (1.0, 0.5, 0.25, 0.125, 0.0625 or 0.03125 mg/mL) for additional 48 h. PPE cytotoxicity against ASC was analyzed using the WST assay (see Additional Materials and Methods). The data represent relative cell viabilities compared to ASC cultured without PPE (Control). Experiments were performed in triplicate, and the data are presented as the mean ± SEM. *p < 0.05, ***p < 0.001 vs. Control. (B) Schematic representation for culture with PPE during ASC differentiation. ASC reaching confluence were cultured with or without PPE for 2 days in 0.5 mM IBMX, 0.25 µM DEX, 5 µg/mL insulin, and 1 µM rosiglitazone. The medium was replaced every 2 days to fresh medium containing 5 µg/mL insulin with or without PPE until day 8, followed by staining with Oil Red O or Lipi-Blue. (C) LD in the ASC cultured for 8 days with or without 1.0 mg/mL PPE were stained with Oil Red O and visualized using bright field microscopy. Stained LD were extracted with isopropanol and quantified at 492 nm. Experiments were performed in triplicate, and the data are presented as the mean ± standard error of the mean (SEM) (***p < 0.001 vs. undifferentiation, ^###p < 0.001 vs. Control). Scale bar: 200 μm. (D) LD in ASC cultured for 8 days with or without 1.0 mg/mL PPE were stained with Lipi-Blue fluorescent probe and visualized using a confocal laser scanning microscope; the area and diameter of cellular LD in 50 cells were quantitatively analyzed using cellSens software ver. 4.1. Each point represents a single lipid droplet, and the data are presented as the mean ± SEM. ***p < 0.001 vs. Control. Scale bar: 50 μm