Adipogenic Differentiation from Human Embryonic Stem Cells Through Formation of Embryoid Bodies.

In the context of adipogenesis, new adipocytes arise from the differentiation of pluripotent stem cells, including embryonic stem cells (hESCs). This process involves two fundamental phases: lineage commitment and terminal differentiation. This chapter elucidates a comprehensive 22-day protocol designed for the differentiation of the hESC cell line H9 toward the adipogenic lineage.During the initial phase, we initiate the differentiation process by forming embryoid bodies (EBs). This is achieved by culturing hESCs in an ultralow attachment plate for a duration of 12 days, during which the EBs are exposed to retinoic acid (RA) for a period of 3 days. Subsequently, adipocyte induction is facilitated by transferring EBs onto Geltrex-coated 6-well plates and subjecting them to adipogenic differentiation medium for an additional 10 days. To assess the successful internalization of lipids within the adipocytes, we employ Oil Red O staining. This protocol provides a robust framework for studying adipogenesis from pluripotent stem cells, offering insights into the intricate process of adipocyte development and serving as a valuable tool for related research endeavors.