Abstract
Aside from mature adipocytes, adipose tissue harbors several distinct cell populations including immune cells, endothelial cells, and adipogenic progenitor cells (AdPCs). AdPCs represent the reservoir of regenerative cells that replenishes adipocytes during normal cellular turnover and during times of increased demand for triglyceride-storage capacity. The worldwide increase in pathologies associated with the metabolic syndrome, such as obesity and type-2 diabetes, has heightened public and scientific interest in adipose tissues and the cell biological processes of adipose tissue formation and function. Two distinct types of fat cells are known: White and brown adipocytes. Especially brown adipose tissue (BAT) has received considerable attention due to its unique capacity for thermogenic energy expenditure and potential role in the treatment of adiposity. Accordingly, the cold-induced conversion of white into brown-like adipocytes has become a feasible approach in humans and a study-subject in rodents to better understand the underlying molecular processes. Fluorescence-activated cell sorting (FACS) provides a method to isolate AdPCs and other cell populations from adipose tissue by using antibodies detecting unique surface markers. We here describe an approach to isolate cells committed to the adipogenic lineage and summarize established protocols to differentiate FACS-purified primary AdPCs into UCP1-expressing brown adipocytes under in vitro conditions.
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World Health Organization (2014) Obesity and overweight fact sheet N°311.http://www.who.int/mediacentre/factsheets/fs311/en/
Bluher M (2014) Adipokines—removing road blocks to obesity and diabetes therapy. Mol Metab 3:230–240
Kershaw EE, Flier JS (2004) Adipose tissue as an endocrine organ. J Clin Endocrinol Metab 89:2548–2556
Zhang Y, Proenca R, Maffei M et al (1994) Positional cloning of the mouse obese gene and its human homologue. Nature 372:425–432
Nedergaard J, Bengtsson T, Cannon B (2007) Unexpected evidence for active brown adipose tissue in adult humans. Am J Physiol Endocrinol Metab 293:E444–E452
Cypess AM, Lehman S, Williams G et al (2009) Identification and importance of brown adipose tissue in adult humans. N Engl J Med 360:1509–1517
Saito M, Okamatsu-Ogura Y, Matsushita M et al (2009) High incidence of metabolically active brown adipose tissue in healthy adult humans: effects of cold exposure and adiposity. Diabetes 58:1526–1531
van Marken Lichtenbelt WD, Vanhommerig JW, Smulders NM et al (2009) Cold-activated brown adipose tissue in healthy men. N Engl J Med 360:1500–1508
Virtanen KA, Lidell ME, Orava J et al (2009) Functional brown adipose tissue in healthy adults. N Engl J Med 360:1518–1525
Zingaretti MC, Crosta F, Vitali A et al (2009) The presence of UCP1 demonstrates that metabolically active adipose tissue in the neck of adult humans truly represents brown adipose tissue. FASEB J 23:3113–3120
Yoneshiro T, Aita S, Matsushita M et al (2013) Recruited brown adipose tissue as an antiobesity agent in humans. J Clin Invest 123:3404–3408
Matsushita M, Yoneshiro T, Aita S et al (2013) Impact of brown adipose tissue on body fatness and glucose metabolism in healthy humans. Int J Obes (Lond) 38:812–817
Bakker LEH, Boon MR, van der Linden RAD et al (2014) Brown adipose tissue volume in healthy lean south Asian adults compared with white Caucasians: a prospective, case-controlled observational study. Lancet Diabetes Endocrinol 2:210–217
Cannon B, Nedergaard J (2004) Brown adipose tissue: function and physiological significance. Physiol Rev 84:277–359
Lichtenbelt WV, Kingma B, van der Lans A et al (2014) Cold exposure—an approach to increasing energy expenditure in humans. Trends Endocrinol Metab 25:165–167
Schulz TJ, Tseng YH (2013) Brown adipose tissue: development, metabolism and beyond. Biochem J 453:167–178
Rosen ED, Spiegelman BM (2014) What we talk about when we talk about fat. Cell 156:20–44
Ishibashi J, Seale P (2010) Medicine. Beige can be slimming. Science 328:1113–1114
Petrovic N, Walden TB, Shabalina IG et al (2010) Chronic peroxisome proliferator-activated receptor gamma (PPAR gamma) activation of epididymally derived white adipocyte cultures reveals a population of thermogenically competent, UCP1-containing adipocytes molecularly distinct from classic brown adipocytes. J Biol Chem 285:7153–7164
Shabalina IG, Petrovic N, de Jong JM et al (2013) UCP1 in brite/beige adipose tissue mitochondria is functionally thermogenic. Cell Rep 5:1196–1203
Kajimura S, Seale P, Spiegelman BM (2010) Transcriptional control of brown fat development. Cell Metab 11:257–262
Kajimura S, Seale P, Kubota K et al (2009) Initiation of myoblast to brown fat switch by a PRDM16-C/EBP-beta transcriptional complex. Nature 460:1154–1158
Seale P, Bjork B, Yang W et al (2008) PRDM16 controls a brown fat/skeletal muscle switch. Nature 454:961–967
Seale P, Conroe HM, Estall J et al (2011) Prdm16 determines the thermogenic program of subcutaneous white adipose tissue in mice. J Clin Invest 121:96–105
Vitali A, Murano I, Zingaretti MC et al (2012) The adipose organ of obesity-prone C57BL/6J mice is composed of mixed white and brown adipocytes. J Lipid Res 53:619–629
Himms-Hagen J, Melnyk A, Zingaretti MC et al (2000) Multilocular fat cells in WAT of CL-316243-treated rats derive directly from white adipocytes. Am J Physiol Cell Physiol 279:C670–C681
Barbatelli G, Murano I, Madsen L et al (2010) The emergence of cold-induced brown adipocytes in mouse white fat depots is determined predominantly by white to brown adipocyte transdifferentiation. Am J Physiol Endocrinol Metab 298:E1244–E1253
Schulz TJ, Huang P, Huang TL et al (2013) Brown-fat paucity due to impaired BMP signalling induces compensatory browning of white fat. Nature 495:379–383
Weisberg SP, McCann D, Desai M et al (2003) Obesity is associated with macrophage accumulation in adipose tissue. J Clin Invest 112:1796–1808
Rodeheffer MS, Birsoy K, Friedman JM (2008) Identification of white adipocyte progenitor cells in vivo. Cell 135:240–249
Schulz TJ, Huang TL, Tran TT et al (2011) Identification of inducible brown adipocyte progenitors residing in skeletal muscle and white fat. Proc Natl Acad Sci USA 108:143–148
Tang W, Zeve D, Suh JM et al (2008) White fat progenitor cells reside in the adipose vasculature. Science 322:583–586
Berry DC, Stenesen D, Zeve D et al (2013) The developmental origins of adipose tissue. Development 140:3939–3949
Berry R, Rodeheffer MS (2013) Characterization of the adipocyte cellular lineage in vivo. Nat Cell Biol 15:302–308
Tseng YH, Kokkotou E, Schulz TJ et al (2008) New role of bone morphogenetic protein 7 in brown adipogenesis and energy expenditure. Nature 454:1000–1004
Vegiopoulos A, Muller-Decker K, Strzoda D et al (2010) Cyclooxygenase-2 controls energy homeostasis in mice by de novo recruitment of brown adipocytes. Science 328:1158–1161
Steenhuis P, Pettway GJ, Ignelzi MA Jr (2008) Cell surface expression of stem cell antigen-1 (Sca-1) distinguishes osteo-, chondro-, and adipoprogenitors in fetal mouse calvaria. Calcif Tissue Int 82:44–56
Himms-Hagen J, Cui J, Danforth E Jr et al (1994) Effect of CL-316,243, a thermogenic beta 3-agonist, on energy balance and brown and white adipose tissues in rats. Am J Physiol 266:R1371–R1382
Klaus S, Choy L, Champigny O et al (1994) Characterization of the novel brown adipocyte cell line HIB 1B. Adrenergic pathways involved in regulation of uncoupling protein gene expression. J Cell Sci 107:313–319
Lehr L, Canola K, Asensio C et al (2006) The control of UCP1 is dissociated from that of PGC-1alpha or of mitochondriogenesis as revealed by a study using beta-less mouse brown adipocytes in culture. FEBS Lett 580:4661–4666
Orr JS, Kennedy AJ, Hasty AH (2013) Isolation of adipose tissue immune cells. J Vis Exp 22(75):e50707
Acknowledgments
This work was supported by grants from the German Research Foundation (DFG; grant # SCHU 2445/2-1) and the European Research Council (grant # ERC-StG-2012-311082) to T.J.S. The authors gratefully acknowledge support from the German Center for Diabetes Research (DZD).
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Steinbring, J., Graja, A., Jank, AM., Schulz, T.J. (2017). Flow Cytometric Isolation and Differentiation of Adipogenic Progenitor Cells into Brown and Brite/Beige Adipocytes. In: Wu, J. (eds) Thermogenic Fat. Methods in Molecular Biology, vol 1566. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6820-6_4
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DOI: https://doi.org/10.1007/978-1-4939-6820-6_4
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