Amber Extract Reduces Lipid Content in Mature 3T3-L1 Adipocytes by Activating the Lipolysis Pathway

Amber—the fossilized resin of trees—is rich in terpenoids and rosin acids. The physiological effects, such as antipyretic, sedative, and anti-inflammatory, were used in traditional medicine. This study aims to clarify the physiological effects of amber extract on lipid metabolism in mouse 3T3-L1 cells. Mature adipocytes are used to evaluate the effect of amber extract on lipolysis by measuring the triglyceride content, glucose uptake, glycerol release, and lipolysis-related gene expression. Our results show that the amount of triacylglycerol, which is stored in lipid droplets in mature adipocytes, decreases following 96 h of treatment with different concentrations of amber extract. Amber extract treatment also decreases glucose uptake and increases the release of glycerol from the cells. Moreover, amber extract increases the expression of lipolysis-related genes encoding perilipin and hormone-sensitive lipase (HSL) and promotes the activity of HSL (by increasing HSL phosphorylation). Amber extract treatment also regulates the expression of other adipocytokines in mature adipocytes, such as adiponectin and leptin. Overall, our results indicate that amber extract increases the expression of lipolysis-related genes to induce lipolysis in 3T3-L1 cells, highlighting its potential for treating various obesity-related diseases.

Lugol Increases Lipolysis through Upregulation of PPAR-Gamma and Downregulation of C/EBP-Alpha in Mature 3T3-L1 Adipocytes

Overweight and obesity are defined as excessive and abnormal fat accumulation that is harmful to health. This study analyzes the effect of different concentrations of the lugol solution (molecular iodine dissolved in potassium iodide) on lipolysis in cultured 3T3-L1-differentiated adipocytes. The mature adipocytes were treated with doses from 1 to 100 µm of lugol for 0.5, 6, and 24 h. The results showed that mature adipocytes exposed to lugol decrease their viability and increase caspase-3 activity with a lethal dose (LD50) of 473 µm. In mature adipocytes, lugol decreased the total intracellular lipid content, being significant at doses of 10 and 100 µm after 6 and 24 h of treatment (P<0.01), and the accumulation of intracellular triglycerides decreased after 24 h of exposure to lugol (P<0.05). Lugol treatment significantly increases the release of glycerol to the culture medium (P<0.05). The levels of adipocyte-specific transcription factors C/EBP-α were downregulated and PPAR-γ upregulated after 30 min with lugol. These results indicate a lipolytic effect of lugol dependent on PPAR-γ and C/EBP-α expression in mature 3T3-L1 adipocytes.

Insulin Controls Triacylglycerol Synthesis through Control of Glycerol Metabolism and Despite Increased Lipogenesis

Under normoxic conditions, adipocytes in primary culture convert huge amounts of glucose to lactate and glycerol. This “wasting” of glucose may help to diminish hyperglycemia. Given the importance of insulin in the metabolism, we have studied how it affects adipocyte response to varying glucose levels, and whether the high basal conversion of glucose to 3-carbon fragments is affected by insulin. Rat fat cells were incubated for 24 h in the presence or absence of 175 nM insulin and 3.5, 7, or 14 mM glucose; half of the wells contained 14C-glucose. We analyzed glucose label fate, medium metabolites, and the expression of key genes controlling glucose and lipid metabolism. Insulin increased both glucose uptake and the flow of carbon through glycolysis and lipogenesis. Lactate excretion was related to medium glucose levels, which agrees with the purported role of disposing excess (circulating) glucose. When medium glucose was low, most basal glycerol came from lipolysis, but when glucose was high, release of glycerol via breakup of glycerol-3P was predominant. Although insulin promotes lipogenesis, it also limited the synthesis of glycerol-3P from glucose and its incorporation into acyl-glycerols. We assume that this is a mechanism of adipose tissue defense to avoid crippling fat accumulation which has not yet been described.

Homocysteine suppresses lipolysis in adipocytes by activating the AMPK pathway

Hyperhomocysteinemia (HHcy) is an independent risk factor for coronary artery disease. Emerging evidence suggests that HHcy is also associated with adipocyte tissue dysfunction. One of the principal functions of adipose tissue is to provide energy substrate via lipolysis. In the present study, we investigated the effects of homocysteine (Hcy) on lipolysis in adipocytes. We found that Hcy inhibited release of glycerol and fatty acids, two typical indicators of the lipolytic response, in primary adipocytes and fully differentiated 3T3-L1 adipocytes in a dose-dependent manner under both basal and isoproterenol-stimulated conditions. In differentiated 3T3-L1 adipocytes, decreased glycerol and free fatty acid (FFA) release was associated with elevation of intracellular TG content. Further studies showed that Hcy-mediated antilipolytic responses were independent of the cyclic AMP-PKA and MEK-ERK1/2 pathways. However, Hcy increased phosphorylation levels of AMP-activated protein kinase (AMPK) and its downstream enzyme acetyl-CoA carboxylase. Compound C, an AMPK inhibitor, abolished Hcy-induced reduction of glycerol and FFA release under both basal and isoproterenol-stimulated conditions. Furthermore, AMPKα1 siRNA reversed Hcy-inhibited glycerol release. Supplementation of exogenous Hcy in the diet for 2 wk lowered circulating glycerol and FFA levels. Moreover, Hcy supplementation was associated with elevated leptin levels and reduced adiponectin levels in plasma. These results show that Hcy inhibits lipolysis through a pathway that involves AMPK activation.

Effects of adenosine A1 receptor antagonism on lipogenesis and lipolysis in isolated rat adipocytes

Adenosine is secreted from adipocytes, binds to adenosine A1 receptor and modulates various functions of these cells. In the present study, the effects of an adenosine A1 receptor antagonist (DPCPX; 0.01, 0.1 and 1 μM) on lipogenesis, glucose transport, lipolysis and the antilipolytic action of insulin were tested in rat adipocytes. DPCPX had a very weak effect on lipogenesis and did not significantly affect glucose uptake. In adipocytes incubated with 1 μM DPCPX, lipolysis increased. This effect was blunted by insulin and by a direct inhibitor of protein kinase A. Moreover, 0.1 μM DPCPX substantially enhanced the lipolytic response to epinephrine and increased cAMP in adipocytes. However, DPCPX was ineffective when lipolysis was stimulated by direct activation of protein kinase A. Adipocyte exposure to epinephrine and insulin with or without 0.1 μM DPCPX demonstrated that this antagonist increased the release of glycerol. However, despite the presence of DPCPX, insulin was able to reduce lipolysis. It is concluded that DPCPX had a weak effect on lipogenesis, whereas lipolysis was significantly affected. The partial antagonism of adenosine A1 receptor increased lipolysis in cells incubated with epinephrine alone and epinephrine with insulin due to the synergistic action of 0.1 μM DPCPX and epinephrine.

Peroxisome proliferator-activated receptor α activation regulates lipid metabolism in the feto-placental unit from diabetic rats

Maternal diabetes promotes an overaccumulation of lipids in the feto-placental unit and impairs feto-placental development and growth. Here, we investigated the role played by the nuclear receptor peroxisome proliferator-activated receptor (PPAR)α in lipid metabolism in fetuses and placentas from control and neonatal streptozotocin-induced diabetic rats. Placentas and fetuses were studied on day 13.5 of gestation. The concentrations of PPARα (by Western blot) and its endogenous agonist leukotriene B4 (LTB4) (by enzyme immunoassay) were analysed. Placental explants and fetuses were cultured with LTB4 or clofibrate, and then lipid metabolism analysed (concentrations and synthesis from 14C-acetate of triglycerides, phospholipids, cholesterol and cholesteryl esters; release of glycerol and free fatty acids (FFAs)). We found that maternal diabetes led to increases in placental concentrations of triglycerides and cholesteryl esters, and fetal concentrations of phospholipids. PPARα agonists downregulated fetal and placental lipid concentrations in control and diabetic rats. The synthesis of lipids was reduced in the diabetic placenta but increased in fetuses from diabetic animals. PPARα agonists reduced the synthesis of lipids in control placenta and in the fetuses from control and diabetic rats. Glycerol and FFA release was enhanced in the diabetic placenta and in control placenta cultured with PPARα agonists. Maternal diabetes led to reductions in fetal and placental LTB4 concentrations and to increases in placental PPARα concentrations. Overall, these data support a novel role of PPARα as a regulator of lipid metabolism in the feto-placental unit, relevant in maternal diabetes where fetal and placental PPARα, LTB4 and lipid concentrations are altered.

Effect of training on insulin-mediated glucose uptake in human muscle

During insulin stimulation whole body glucose uptake is increased in trained compared with untrained humans. However, it is not known which tissue is responsible. Seven young male subjects bicycle trained one leg for 10 wk at 70% of maximal O2 consumption (VO2max). Sixteen hours after last exercise bout, a three-step euglycemic hyperinsulinemic clamp (clamp 1) was performed (insulin levels, means +/- SE: 9 +/- 1, 53 +/- 3, 174 +/- 5, and 2,323 +/- 80 was microU/ml), with measurement of arteriovenous differences and blood flow in both legs. After 6 days of detraining subjects were restudied, having exercised the untrained leg 16 h before. VO2max for trained (T) and untrained (UT) legs was 52 +/- 2 vs. 44 +/- 2 ml.min-1.kg-1 (P < 0.05). In clamp 1 glucose uptake in T and UT legs was 1.0 +/- 0.2 vs. 0.5 +/- 0.1 mg.min-1.kg-1 (basal), 9.7 +/- 2.3 vs. 6.7 +/- 1.7 (P < 0.05) (step I), 19.2 +/- 2.8 vs. 14.3 +/- 2.0 (P < 0.05) (step II), and 22.8 +/- 2.3 vs. 18.6 +/- 2.2 (P < 0.05) (step III). During insulin infusion lactate release (P < 0.05) [8.9 +/- 1.8 vs. 2.9 +/- 0.9 mumol.min-1.kg-1 (step I), 24.6 +/- 3.1 vs. 12.5 +/- 2.6 (step III)] and glycogen storage (P < 0.1) calculated by indirect calorimetry [6.7 +/- 2.3 vs. 5.0 +/- 1.7 mg.min-1.kg-1 (step I), 16.8 +/- 2.1 vs. 14.1 +/- 1.8 (step III)] were always higher in T than in UT legs. Release of glycerol, free fatty acids, and tyrosine and clearance of insulin were not influenced by training. Insulin-mediated glucose uptake was not increased after detraining or a single bout of exercise. In conclusion, training increases sensitivity and responsiveness of insulin-mediated glucose uptake in human muscle by local mechanisms. Glycolysis and glycogen storage are equally enhanced. The training effect represents a genuine adaptation to repeated exercise but is short lived. Insulin clearance in muscle is not influenced by training.