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Lauren Drowley, High Content Biology, AstraZeneca, Patrick O'Shea, New Opportunities, AstraZeneca, Xiao-Rong Peng, Diabetes and Obesity, AstraZeneca, Mike Sullivan, High Content Biology, AstraZeneca

Due to the increasing epidemic of obesity, there has been significant interest in adipose tissue. There are two types of fat cells, each with distinct properties. White fat cells store energy whereas brown fat cells use energy in a process known as thermogenesis. Uncoupling protein 1 (UCP-1) is expressed in the mitochondrial membranes of brown fat and provides a major means of heat generation. Brown adipose tissue is more prevalent in infants, though recently studies have shown brown fat depots in adults. Increasing differentiation towards brown fat is a potential target of therapeutic treatments to decrease obesity and metabolic diseases. We describe the characterization and development of cellular models of adipose differentiation. There are two types of brown fat cells, which appear to arise from different developmental origins. One subset is derived from an adipogenic precursor and is responsive to β-adrenergic stimuli. The other type of cell arises from a myf5+ myogenic precursor and appears to be dependent on the transcriptional regulator PRDM16.

Adipose derived stem cells (ADSCs) or myoblasts were plated in 96-well plates and maintained in differentiation media for 1, 2, or 3 weeks in the presence or absence of the thiazolidinedione PPAR-agonist pioglitazone, a compound targeting a known pathway involved in differentiation. The cells were stained with adiponectin, a general adipose marker, and UCP-1, with cell nuclei counterstained with Hoescht 33342.

In ADSCs, we demonstrated a significant increase in UCP-1 expression in the presence of pioglitazone compared to cells treated with differentiation media alone. In contrast, ADSCs not exposed to differentiation media showed no increase in UCP-1 expression. In skeletal myoblasts, pioglitazone also induced expression of UCP-1, though not to the same levels as in ADSCs.

We demonstrate the application of multiparametric high-content technology to monitor differentiation of cells to brown fat following small molecule intervention. Furthermore, we apply this approach to cells from distinct developmental origins in order that compounds with selective effects are identified. These assays have potential to be used as the basis for regenerative medicine screening assays looking to identify modulators of adipose tissue for indications such as obesity and diabetes.

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