Adipogenic Differentiation of Mesenchymal Stem Cells

Cells were plated at a concentration of 3 × 10⁴ cells/cm² and cultured in DMEM-low glucose (Sigma-Aldrich), 10% FBS (HyClone, Thermo Fisher Scientific), 4 mM L-glutamine (Euroclone), and 1% antibiotic-antimycotic mixture (Euroclone), with the addition of the mesenchymal stem cell adipogenesis kit (Millipore) for 21 days, according to the manufacturer's instructions. The adipogenic medium was changed every other day. At day 21, Oil Red O solution (Millipore) was used to stain lipid droplets of derived adipocytes, according to the manufacturer's procedures. All photomicrographs were acquired with an Axiovert 40 microscope (Zeiss) equipped with a Moticam 2300 camera (Motic). The mRNA expression of adipogenic markers including PPAR-γ and LPL were also assessed on days 7 and 21 by real-time PCR, as described above [17 (link)].