Reduced UCP-1 Content in In Vitro Differentiated Beige/Brite Adipocytes Derived from Preadipocytes of Human Subcutaneous White Adipose Tissues in Obesity

Abstract

IntroductionBrown adipose tissue (BAT) is a potential therapeutic target to reverse obesity. The purpose of this study was to determine whether primary precursor cells isolated from human adult subcutaneous white adipose tissue (WAT) can be induced to differentiate in-vitro into adipocytes that express key markers of brown or beige adipose, and whether the expression level of such markers differs between lean and obese young adult males.MethodsAdipogenic precursor cells were isolated from lean and obese individuals from subcutaneous abdominal WAT biopsies. Cells were grown to confluence, differentiated for 2.5 weeks then harvested for measurement of gene expression and UCP1 protein.ResultsThere was no difference between groups with respect to differentiation into adipocytes, as indicated by oil red-O staining, rates of lipolysis, and expression of adipogenic genes (FABP4, PPARG). WAT genes (HOXC9, RB1) were expressed equally in the two groups. Post differentiation, the beige adipose specific genes CITED1 and CD137 were significantly increased in both groups, but classic BAT markers ZIC1 and LHX8 decreased significantly. Cell lines from both groups also equally increased post-differentiation expression of the thermogenic-responsive gene PPARGC1A (PGC-1α). UCP1 gene expression was undetectable prior to differentiation, however after differentiation both gene expression and protein content were increased in both groups and were significantly greater in cultures from lean compared with obese individuals (p<0.05).ConclusionHuman subcutaneous WAT cells can be induced to attain BAT characteristics, but this capacity is reduced in WAT cells from obese individuals.

Pioglitazone reduces cold-induced brown fat glucose uptake despite induction of browning in cultured human adipocytes: a randomised, controlled trial in humans

“…Previous TZD studies in patients with type 2 diabetes [14], obese rats and mice [28] and cultured human subcutaneous adipocytes [13,15] showed that chronic treatment with a TZD increases browning in brown and/or white adipose tissues. Our human cell culture experiments involving pre-adipocytes derived from a region known to contain brown/beige adipocyte precursors corroborated this.…”

“…Within 60 min, the tissue was dissected free from connective tissue, minced into~1 mm pieces and then cells were extracted and cultured as described [15].…”

“…neutralbuffered formalin and then stained with Oil Red O (surrogate indicator of general adipogenesis). The cells were assessed for lipolytic rate (basal and stimulated with 1 μmol/l isoprenaline [known as isoproterenol in the USA] for 2 h to assess general adipose function) or UCP-1 protein content was determined (indicator of cell browning) as described [15]. Treatments included vehicle (DMSO), 1 μmol/l rosiglitazone or 1 μmol/l pioglitazone.…”

“…Treatments included vehicle (DMSO), 1 μmol/l rosiglitazone or 1 μmol/l pioglitazone. TZD concentrations were selected based on studies of single clinical doses of these drugs (15-45 mg) which resulted in comparable plasma concentrations (0.5 μmol, l-2 μmol/l) [22,23], and previous human primary adipocyte experiments [15]. Individual experiments were conducted with three or more technical replicates/treatment condition.…”

“…TZDs also induce browning in human adipocyte culture models [13] and rosiglitazone is used almost universally to promote differentiation and browning of human cultured brown adipocytes [13][14][15][16][17]. Rosiglitazone and, to a lesser extent, pioglitazone were widely prescribed for type 2 diabetes until safety concerns were raised (primarily with rosiglitazone [18]), leaving pioglitazone as the predominantly prescribed TZD worldwide [19].…”