Differentiation of human pluripotent stem cells into highly functional classical brown adipocytes.
We describe a detailed method for directed differentiation of human pluripotent stem cells, including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), into functional classical brown adipocytes (BAs) under serum-free and feeder-free conditions. It is a two-tiered culture system, based on very simple techniques, a floating culture and a subsequent adherent culture. It does not require gene transfer. The entire process can be carried out in about 10 days. The key point is the usage of our special hematopoietic cytokine cocktail. Almost all the differentiated cells express uncoupling protein 1, a BA-selective marker, as determined by immunostaining. The differentiated cells show characteristics of classical BA as assessed by morphology and gene/protein expression. Moreover, the expression of myoblast marker genes is transiently induced during the floating culture step. hESC/hiPSC-derived BAs show significantly higher oxygen consumption rates (OCRs) than white adipocytes generated from human mesenchymal stem cell. They also show responsiveness to adrenergic stimuli, with about twofold upregulation in OCR by β-adrenergic receptor (β-AR) agonist treatments. hESC/hiPSC-derived BAs exert in vivo calorigenic activities in response to β-AR agonist treatments as assessed by thermography. Finally, lipid and glucose metabolisms are significantly improved in hESC/hiPSC-derived BA-transplanted mice. Our system provides a highly feasible way to produce functional classical BA bearing metabolism-improving capacities from hESC/hiPSC under a feeder-free and serum-free condition without gene transfer.
Department of Disease Control, Research Institute, National Center for Global Health and Medicine, Tokyo, Japan.
Miwako
M
Nishio
Department of Disease Control, Research Institute, National Center for Global Health and Medicine, Tokyo, Japan. Electronic address: saeki@ri.ncgm.go.jp.
Kumiko
K
Saeki
D052438:Adipocytes, White / Q000166:cytology; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000235:genetics*; D003470:Culture Media; D053595:Embryonic Stem Cells / Q000166:cytology*; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology*; D051379:Mice
D003470:Culture Media
Feeder-free; Glucose tolerance; Hematopoietic cytokine; Human ES cells; Human iPS cells; Oral fat tolerance test; Oxygen consumption rate; Serum-free; Thermography