Incorporation of [1-¹³C]oleate into cellular triglycerides in differentiating 3T3L1 cells

Oleate is one of the most abundant dietary fatty acids, and much remains to be learned about its metabolism in fat cells. We studied the incorporation of exogenous [1-¹³C]oleate into triglycerides (TG) in differentiating 3T3L1 preadipocytes using¹³C NMR spectroscopy. The quantity of oleate incorporated into TG was found to increase as preadipocytes differentiated into fat cells. The ratio of unesterified [1-¹³C]oleate to total stored fatty acids was higher in less differentiated cells, and declined at later stages of differentiation as cells accumulated fatty acids through de novo synthesis. When added as the only exogenous fatty acid, oleate was largely esterified at the sn-2 position. When equimolar unlabeled linoleate was co- provided at the same time, the ratio of [1-¹³C]oleate esterified at the sn- 1,3 position increased, implying competition between linoleate and oleate for esterification, especially at the sn-2 position. When cells pre-enriched with [1-¹³C]oleate (esterified to TG) were treated with isoproterenol, a lipolytic agent, most of the [1-¹³C]oleate was still found in TG, despite a high rate of lipolysis determined by measuring glycerol release. This implies extensive re-esterification of the oleate released by lipolysis.