To identify genes involved in the central regulation of energy balance, we compared hypothalamic mRNA from lean and obese Psammomys obesus, a polygenic model of obesity, using differential display PCR. One mRNA transcript was observed to be elevated in obese, and obese diabetic, P. obesus compared with lean animals and was subsequently found to be increased 4-fold in the hypothalamus of lethal yellow agouti (A(y)/a) mice, a murine model of obesity and diabetes. Intracerebroventricular infusion of antisense oligonucleotide targeted to this transcript selectively suppressed its hypothalamic mRNA levels and resulted in loss of body weight in both P. obesus and Sprague Dawley rats. Reductions in body weight were mediated by profoundly reduced food intake without a concomitant reduction in metabolic rate. Yeast two-hybrid screening, and confirmation in mammalian cells by bioluminescence resonance energy transfer analysis, demonstrated that the protein it encodes interacts with endophilins, mediators of synaptic vesicle recycling and receptor endocytosis in the brain. We therefore named this transcript Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1 (SGIP1). SGIP1 encodes a large proline-rich protein that is expressed predominantly in the brain and is highly conserved between species. Together these data suggest that SGIP1 is an important and novel member of the group of neuronal molecules required for the regulation of energy homeostasis.

Casitas b-lineage lymphoma (c-Cbl) is a multiadaptor protein with E3-ubiquitin ligase activity involved in regulating the degradation of receptor tyrosine kinases. We have recently reported that c-CblϪ/Ϫ mice exhibit a lean phenotype and enhanced peripheral insulin action likely due to elevated energy expenditure. In the study reported here, we examined the effect of a high-fat diet on energy homeostasis and glucose metabolism in these animals. When c-Cbl Ϫ/Ϫ mice were fed a high-fat diet for 4 weeks, they maintained hyperphagia, higher whole-body oxygen consumption (27%), and greater activity (threefold) compared with wild-type animals fed the same diet. In addition, the activity of several enzymes involved in mitochondrial fat oxidation and the phosphorylation of acetyl CoA carboxylase was significantly increased in muscle of high-fat-fed c-Cbl-deficient mice, indicating a greater capacity for fat oxidation in these animals. As a result of these differences, fat-fed c-Cbl Ϫ/Ϫ mice were 30% leaner than wild-type animals and were protected against high-fat diet-induced insulin resistance. These studies are consistent with a role for c-Cbl in regulating nutrient partitioning in skeletal muscle and emphasize the potential of c-Cbl as a therapeutic target in the treatment of obesity and type 2 diabetes. Diabetes 55:708 -715, 2006 T he incidence of obesity and type 2 diabetes is increasing throughout the world. This has been ascribed to changes in food intake combined with a more sedentary lifestyle. Regardless of the cause, this emerging health care problem has sparked renewed interest in the study of insulin action and fuel metabolism. In particular, the identification of genes that regulate energy homeostasis in mammals has become a major research interest. Over the past decade, largely through the use of genetically manipulated animal models, a number of genes that result in lean phenotypes have been described. These genes include those that regulate appetite, food absorption, and increased energy expenditure in either muscle or adipose tissue (1). A major advantage of manipulations that increase energy expenditure is that this depletes fat stores not only in adipose tissue but possibly in other cells that are susceptible to lipotoxic damage, thus providing a protective mechanism against the development of insulin resistance and diabetes (2,3).Genes that are known to regulate whole-body energy expenditure include mitochondrial uncoupling proteins that divert energy stores into heat production (4,5) and lipid handling enzymes, such as acetyl CoA carboxylase (ACC), which regulates the entry of long-chain acyl CoAs into mitochondria (6), and DGAT, a key enzyme in triacylglyceride synthesis (7). Unexpectedly, reduced expression of several molecules that negatively regulate insulin signaling, like the tyrosine phosphatase PTP1b (8), have also been shown to cause a significant increase in whole-body energy expenditure. This provides further evidence for an intimate link between insulin action and energy homeostasis.We...

In the past, it has been noted that experimental tumour cells inoculated into the peritoneal cavity or into the lumen of the bowel will grow at a recently formed colonic anastomosis. However, it has previously been unclear whether the healing process enhances tumour growth or whether the presence of a suture line merely allows the tumour cells to gain access to the tissues. In the present study, using the hooded Lister rat, we have confirmed these findings by showing that growth of the syngeneic MC28 sarcoma and OES5 breast carcinoma occurs preferentially at colonic anastomoses and laparotomy wounds after intraperitoneal injection, and at colonic anastomoses after intraluminal injection. In previous studies using the MC28 sarcoma and the OES5 breast carcinoma injected by the intracardiac route (so that tumour cells reach normal and healing tissues in approximately equal numbers) we have shown that tumour growth is enhanced in healing wounds but not in the surrounding normal tissues when cells reach a healing colonic anastomosis or laparotomy wound within 2 h of its formation. Furthermore, by studying the distribution of radiolabelled tumour cells after intracardiac injection, we have calculated that the probability of a tumour cell leading to a deposit in a healing anastomosis or laparotomy wound is increased 1,000 fold compared to normal tissue. No previous studies have combined the data for intracardiac, intraluminal and intraperitoneal injection of tumour cells using the same animal model. We conclude that the same phenomenon of tumour growth enhancement in colonic anastomoses and laparotomy wounds reported after intracardiac injection of tumour cells may well be enhancing tumour growth after intraperitoneal and intraluminal injection.(ABSTRACT TRUNCATED AT 250 WORDS)

We previously used Gene Expression Signature technology to identify methazolamide (MTZ) and related compounds with insulin sensitizing activity in vitro. The effects of these compounds were investigated in diabetic db/db mice, insulin-resistant diet-induced obese (DIO) mice, and rats with streptozotocin (STZ)-induced diabetes. MTZ reduced fasting blood glucose and HbA1c levels in db/db mice, improved glucose tolerance in DIO mice, and enhanced the glucose-lowering effects of exogenous insulin administration in rats with STZ-induced diabetes. Hyperinsulinemic-euglycemic clamps in DIO mice revealed that MTZ increased glucose infusion rate and suppressed endogenous glucose production. Whole-body or cellular oxygen consumption rate was not altered, suggesting MTZ may inhibit glucose production by different mechanism(s) to metformin. In support of this, MTZ enhanced the glucose-lowering effects of metformin in db/db mice. MTZ is known to be a carbonic anhydrase inhibitor (CAI); however, CAIs acetazolamide, ethoxyzolamide, dichlorphenamide, chlorthalidone, and furosemide were not effective in vivo. Our results demonstrate that MTZ acts as an insulin sensitizer that suppresses hepatic glucose production in vivo. The antidiabetic effect of MTZ does not appear to be a function of its known activity as a CAI. The additive glucose-lowering effect of MTZ together with metformin highlights the potential utility for the management of type 2 diabetes.

Reciprocal regulation of hepatic glycolysis and gluconeogenesis contributes to systemic metabolic homeostasis. Recent evidence from lower order organisms has found that reversible post-translational modification of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), particularly acetylation, contributes to the reciprocal regulation of glycolysis/gluconeogenesis. However, whether this occurs in mammalian hepatocytes or is unknown. Several proteomics studies have identified 4 lysine residues in critical regions of mammalian GAPDH that are altered by multiple post-translational modifications. In FAO hepatoma cells, mutation of all 4 lysine residues (4K-R GAPDH) to mimic their unmodified state reduced GAPDH glycolytic activity and glycolytic flux and increased gluconeogenic GAPDH activity and glucose production. Hepatic expression of 4K-R GAPDH in mice increased GAPDH gluconeogenic activity and the contribution of gluconeogenesis to endogenous glucose production in the unfed state. Consistent with the increased reliance on the energy-consuming gluconeogenic pathway, plasma free fatty acids and ketones were elevated in mice expressing 4K-R GAPDH, suggesting enhanced lipolysis and hepatic fatty acid oxidation. In normal mice, food withholding and refeeding, as well as hormonal regulators of reciprocal glycolysis/gluconeogenesis, such as insulin, glucagon, and norepinephrine, had no effect on global GAPDH acetylation. However, GAPDH acetylation was reduced in obese and type 2 diabetic mice. These findings show that post-translational modification of GAPDH lysine residues regulates hepatic and systemic metabolism, revealing an unappreciated role for hepatic GAPDH in substrate selection and utilization.-Bond, S. T., Howlett, K. F., Kowalski, G. M., Mason, S., Connor, T., Cooper, A., Streltsov, V., Bruce, C. R., Walder, K. R., McGee, S. L. Lysine post-translational modification of glyceraldehyde-3-phosphate dehydrogenase regulates hepatic and systemic metabolism.