Flow Cytometric Isolation and Differentiation of Adipogenic Progenitor Cells into Brown and Brite/Beige Adipocytes

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.

Seasonal variations of adipose tissue in Tupaia belangeri (Mammalia: Scandentia: Tupaiidae)

“…PPARα is a unique link between lipid metabolism and immune function, which plays an important role in inflammatory reaction (Hines et al 2018). Therefore, some effector molecules produced by immune cells in WAT and BAT and their regulation of the development and proliferation of beige and brown adipocytes (Brestoff & Artis 2015;DiSpirito & Mathis 2015), and activation of beige adipocytes in vitro, have become current research directions (Steinbring et al 2017). Expression of UCP1 in WAT is dependent on the activation of COX-2 (Lau et al 2013).…”

Loss of periostin occurs in aging adipose tissue of mice and its genetic ablation impairs adipose tissue lipid metabolism

“…AT‐derived progenitor cells were isolated as described (Schulz et al, 2011; Steinbring, Graja, Jank, & Schulz, 2017). A detailed description of the procedures is supplied in the Supporting Information.…”

Identification of functional lipid metabolism biomarkers of brown adipose tissue aging

“…AT-derived progenitor cells were isolated from the BAT and iWAT of mice according to published procedures [21], [22]. In brief, cells stained positive for surface antigen Stem cell antigen 1 (Sca1, #17-5981-82, Thermo Fisher Scientific, Dreieich, Germany) and negative for the hematopoietic marker antigen CD45 (#11-0451-82, Thermo Fisher Scientific, Dreieich, Germany) and the endothelial marker antigen CD31 (#11-0311-82, Thermo Fisher Scientific, Dreieich, Germany) were purified by fluorescence-activated cell sorting (FACS, BD FACSAria™ III, BD Biosciences, San Jose, CA, USA).…”

“…Primary, FACS-isolated APCs were differentiated as described before with minor modifications [22]. In brief, after seeding cells were grown to confluence for three days with 1 μM rosiglitazone (Sigma Aldrich, Taufkirchen, Germany).…”