1,25-Dihydroxyvitamin D regulation of triacylglycerol accumulation in differentiated adipocytes
Obesity is a major public health concern, both in the United States and worldwide. Therefore, identification of measures by which obesity may be prevented and reversed is of high priority. Epidemiological studies consistently demonstrate an inverse relationship between serum 25(OH)D levels, an indicator of vitamin D status, and measures of adiposity. These data suggest that vitamin D may play a role in the prevention of excessive adiposity. However, whether vitamin D impacts lipid storage and metabolism in terminally differentiated adipocytes is not yet known. The purpose of this work was to determine the impact of 1,25-dihydroxyvitamin D (1,25(OH)2D), the bioactive vitamin D metabolite, on triacylglycerol accumulation and lipid and glucose metabolism in differentiated adipocytes. To study this, 3T3-L1 adipocytes were differentiated for 9 days, followed by stimulation with 1,25(OH)2D (10 nM) or vehicle for 1-7 days. Results indicate that 1,25(OH)2D stimulates a 21% reduction in TAG accumulation in differentiated 3T3-L1 adipocytes after 4 days ( P=0.01). This occurs despite a significant increase in fatty acid uptake (P<0.01), assessed using BODIPY FL C16, and with concomitant stimulation of PKA-dependent glycerol release ( P<0.01), which is typically indicative of lipolysis. Additionally, we demonstrate that 1,25(OH)2D stimulates a 2.5-fold increase in complete fatty acid oxidation (P<0.01), assessed by quantifying the production of 14CO2 from [1-14C] palmitic acid. These results suggest a novel mechanism by which 1,25(OH) 2D may be protective against excessive adiposity. In addition to its impact on fatty acid metabolism, the impact of 1,25(OH)2D on glucose metabolism was also examined. Glucose contributes significantly to the intracellular TAG pool, serving as a substrate for both fatty acid synthesis as well as glycerol production to support TAG synthesis. The results indicate that 1,25(OH) 2D reduces the incorporation of D-[U-13C]glucose incorporation into palmitic, palmitoleic, stearic, and oleic acids (P=0.03), determined by liquid chromatography-mass spectrometry (LC-MS). Interestingly, [13C2]acetate incorporation into these fatty acids was reduced by only 10% (p<0.01), suggesting that while de novo lipogenesis is slightly inhibited in response to 1,25(OH)2D, the contribution of glucose specifically as a substrate for fatty acid synthesis is reduced. Study of glucose uptake and disposal as lactate revealed that these two processes are not impacted by 1,25(OH) 2D, suggesting that glucose may instead be used for the synthesis of glycerol. Indeed, inhibition of glycolysis to reduce substrate availability for glycerol synthesis completely prevented 1,25(OH)2D-stimulation of glycerol release. These data suggest that rather than stimulating TAG hydrolysis, 1,25(OH)2D stimulates disposal of glucose as glycerol, while reducing its utilization as a substrate for fatty acid synthesis. While the mRNA expression of pyruvate carboxylase (PC) is reduced by 40% in response to 1,25(OH) 2D (P<0.01), suggesting that 1,25(OH)2D may limit pyruvate entry into the TCA cycle, at this time it is not clear whether this underlies the 1,25(OH)2D-stimulated changes in glucose metabolism and TAG storage. In conclusion, 1,25(OH)2D stimulates fatty acid oxidation in 3T3-L1 adipocytes, and reduces the contribution of glucose to the fatty acid pool, likely by stimulating glucose disposal as glycerol. These results demonstrate that 1,25(OH)2D regulates both fatty acid and glucose metabolism in differentiated 3T3-L1 adipocytes to reduce TAG storage. These novel findings demonstrate a mechanism by with 1,25(OH) 2D may protect against excessive fat mass accumulation, and provide support for the inverse relationship between vitamin D and obesity.^
Dorothy Teegarden, Purdue University.