SBM induces apoptosis in mature adipocytes.

<p>Effect of SBM on the viability of, (A) 3T3L-1 preadipocyte and (B) mature adipocytes (8 days), as determined by MTT assay. Values are expressed as a percentage survival as compared to control after a 24 h incubation. (C) SBM-induced apoptosis in mature adipocytes evaluated by annexin-V/PI-FACS analysis. Annexin: FL1-H; PI: FL2-H. (D) Quantification of annexin V positive cells by imaging. At least 500 cells were analyzed. (E) Control (vehicle-treated) and SBM-treated mature adipocytes were either fixed and stained with DAPI and analyzed by fluorescence microscopy, or analyzed for DNA fragmentation by TUNEL. Cells labeled with the Fluorescein-FragEL were analysed by FACS shown, cell only (green) and cell treated with 100 µg/ml SBM (red), 1 µg/ml Vinblastine (purple) (left panel). Quantification of SBM-induced apoptosis in mature adipocytes by TUNEL assay (right panel). Micrographs of control and SBM-treated cells stained with DAPI are shown (bottom panel). (F) Demonstration of apoptosis by gel electrophoresis. Mature adipocytes were incubated with SBM at various concentrations for 24 h. Lane 1, DNA marker; lane 2, vehicle; lane 7, 1μM vinblastine positive control. Data are expressed as the mean ± SD. *<i>P</i><0.01, **<i>P</i><0.05 vs. controls. Results were verified by three repetitions of the experiments, which were each conducted in triplicate.