A new member of the uncoupling protein (UCP) family called UCP3 has recently been cloned and shown to be highly expressed in skeletal muscle of rodents and humans. In the present study, UCP3 was overexpressed in C 2 C 12 myoblasts where it acts as an uncoupling protein.Changes in UCP3 mRNA expression were examined in rodent muscles under conditions known to modulate thermogenesis in brown adipose tissue. In skeletal muscle, UCP3 expression did not change in response to 48 h of cold exposure (6°C), whereas it was decreased by 81% or increased 5.6-fold by 1 week of 50% food restriction or fasting, respectively. It was also decreased by 36% in soleus muscle of obese (fa/fa) as compared with lean Zucker rats. The unexpected rise of UCP3 mRNA level induced by fasting did not change in vitro muscle basal heat production rate but decreased by 31% the capacity to produce heat in response to the uncoupler carbonylcyanide p-trifluoromethoxyphenylhydrazone. This decrease may reflect underlying uncoupling by UCP3. Upregulation of UCP3 mRNA after a 24-h fast was still observed in mice exposed at thermoneutrality. These results show that the increase in UCP3 expression induced by fasting is associated with the maintenance of thermogenesis measured in muscle in vitro and is not modulated by environmental temperature. The notion that UCP3 expression is modulated by food intake is of importance to better understand the pathophysiology of obesity in humans.

The mechanism by which long-term exposure of the ␤-cell to elevated concentrations of fatty acid alters glucose-induced insulin secretion has been examined. Exposure of INS-1 ␤-cells to 0.4 mmol/l oleate for 72 h increased basal insulin secretion and decreased insulin release in response to high glucose, but not in response to agents acting at the level of the K ATP channel (tolbutamide) or beyond (elevated KCl). This also suppressed the glucose-induced increase in the cellular ATP-to-ADP ratio. The depolarization of the plasma membrane promoted by glucose was decreased after oleate exposure, whereas the response to KCl was unchanged. Cells exposed to free fatty acids displayed a lower mitochondrial membrane potential and a decreased glucose-induced hyperpolarization. The possible implication of uncoupling protein (UCP)-2 in the altered secretory response was examined by measuring UCP2 gene expression after chronic exposure of the cells to fatty acids. UCP2 mRNA and protein were increased twofold by oleate. Palmitate and the nonoxidizable fatty acid bromopalmitate had similar effects on UCP2 mRNA, suggesting that UCP2 gene induction by fatty acids does not require their metabolism. The data are compatible with a role of UCP2 and partial mitochondrial uncoupling in the decreased secretory response to glucose observed after chronic exposure of the ␤-cell to elevated fatty acids, and suggest that the expression and/or activity of the protein may modulate insulin secretion in response to glucose. Diabetes 50:803-809, 2001

To better understand the action of glucose on fatty acid metabolism in the ␤-cell and the link between chronically elevated glucose or fatty acids and ␤-cell decompensation in adipogenic diabetes, we investigated whether glucose regulates peroxisomal proliferator-activated receptor (PPAR) gene expression in the ␤-cell. Islets or INS(832/13) ␤-cells exposed to high glucose show a 60 -80% reduction in PPAR␣ mRNA expression. Oleate, either in the absence or presence of glucose, has no effect. The action of glucose is dose-dependent in the 6 -20 mM range and maximal after 6 h. Glucose also causes quantitatively similar reductions in PPAR␣ protein and DNA binding activity of this transcription factor. The effect of glucose is blocked by the glucokinase inhibitor mannoheptulose, is partially mimicked by 2-deoxyglucose, and is not blocked by the 3-O-methyl or the 6-deoxy analogues of the sugar that are not phosphorylated. Chronic elevated glucose reduces the expression levels of the PPAR target genes, uncoupling protein 2 and acyl-CoA oxidase, which are involved in fat oxidation and lipid detoxification. A 3-day exposure of INS-1 cells to elevated glucose results in a permanent rise in malonyl-CoA, the inhibition of fat oxidation, and the promotion of fatty acid esterification processes and causes elevated insulin secretion at low glucose. The results suggest that a reduction in PPAR␣ gene expression together with a rise in malonyl-CoA plays a role in the coordinated adaptation of ␤-cell glucose and lipid metabolism to hyperglycemia and may be implicated in the mechanism of ␤-cell "glucolipotoxicity."

The ob gene mRNA expression in rat brown adipose tissue (BAT) and epididymal white adipose tissue (WAT) was measured on Northern blots hybridized with a rat ob gene probe. The level of ob gene mRNA in BAT was about 40% of that in WAT. Fasting (36 h) or semi-starvation (10 days) decreased the ob gene mRNA level in both tissues by 62-68%, and cold exposure at 6°C (24 h) decreased it in BAT (-84%) but not in WAT. Acute administration of the /3a-adrenergic agonist Ro 16-8714 decreased the ob gene mRNA level in BAT (-51%) and WAT (-28%) of lean Zucker rats and only in BAT (-74%) of obese fa/fa rats. This study demonstrates that, in the rat, the ob gene is not only expressed in WAT but also in BAT, and suggests that in these two tissues, the modulation of the ob gene expression might be more closely associated with known alterations in cell lipid content than with changes in sympathetic activity.Key words." ob Gene; Fast; Cold; Rat brown and white adipose tissue db/db mouse and that would account for the up-regulation of the ob gene expression in the WAT of these animals.To date, the ob gene expression has been demonstrated in WAT [1,2] and also detected in brown adipose tissue (BAT) [3]. Since BAT is known to play an important role in the regulation of energy balance in rodents [4], the expression of the ob gene was examined in this tissue. The aims of this study therefore were: (i) to quantify the ob gene expression in rat interscapular BAT and to compare its level to that measured in epididymal WAT; and (ii) to investigate if the ob gene level is modulated by conditions which are known to reduce or increase BAT sympathetic and thermogenic activity, namely caloric restriction [5,6], cold exposure [6,7], or the administration of a fl3-adrenoceptor agonist [8]. Under each of these conditions, possible changes in the ob gene expression in epididymal WAT were also studied in parallel. Materials and methods

Chronic exposure of pancreatic beta-cells to high glucose has pleiotropic action on beta-cell function. In particular, it induces key glycolytic genes, promotes glycogen deposition, and causes beta-cell proliferation and altered insulin secretion characterized by sensitization to low glucose. Postglycolytic events, in particular, anaplerosis and lipid signaling, are thought to be implicated in beta-cell activation by glucose. To understand the biochemical nature of the beta-cell adaptive process to hyperglycemia, we studied the regulation by glucose of lipogenic genes in the beta-cell line INS-1. A 3-day exposure of cells to elevated glucose (5-25 mmol/l) increased the enzymatic activities of fatty acid synthase 3-fold, acetyl-CoA carboxylase 30-fold, and malic enzyme 1.3-fold. Pyruvate carboxylase and citrate lyase expression remained constant. Similar observations were made at the protein and mRNA levels except for malic enzyme mRNA, which did not vary. Metabolic gene expression changes were associated with chronically elevated levels of citrate, malate, malonyl-CoA, and conversion of glucose carbon into lipids, even in cells that were subsequently exposed to low glucose. Similarly, fatty acid oxidation was suppressed and phospholipid and triglyceride synthesis was enhanced independently of the external glucose concentration in cells preexposed to high glucose. The results suggest that a coordinated induction of glycolytic and lipogenic genes in conjunction with glycogen and triglyceride deposition, as well as increased anaplerosis and altered lipid partitioning, contribute to the adaptive process to hyperglycemia and glucose sensitization of the beta-cell.