07-681 Sigma-AldrichAnti-phospho-AMPK α (Thr172) Antibody

In coronary artery disease patients, enhanced external counterpulsation (EECP) improves peripheral arterial function and nitric oxide (NO) bioavailability, which have been implicated in the pathogenesis of abnormal glucose tolerance (AGT). We sought to evaluate the effects of EECP on outcomes of arterial function, glucose tolerance, and skeletal muscle morphology in subjects with AGT.18 subjects with AGT were randomly (2:1 ratio) assigned to receive either 7 wk (35 1-h sessions) of EECP (n = 12) or 7 wk of standard care (control; n = 6). Peripheral vascular function, biochemical assays, glucose tolerance, and skeletal muscle morphology were evaluated before and after EECP or control. EECP increased normalized brachial artery (27%) and popliteal artery (52%) flow-mediated dilation. Plasma nitrite/nitrate (NOx) increased (30%) and 8-isoprostane-PGF-F(2α), a marker of lipid peroxidation in the plasma, decreased (-23%). Fasting plasma glucose declined (-16.9 ± 5.4 mg/dl), and the homeostasis model assessment of insulin resistance (HOMA-IR) decreased (31%) following EECP. Plasma glucose 120 min after initiation of oral glucose tolerance testing decreased (-28.3 ± 7.3 mg/dl), and the whole body composite insulin sensitivity index (C-ISI) increased (21%). VEGF concentrations increased (75%), and vastus lateralis skeletal muscle biopsies demonstrated improvements in capillary density following EECP. No change was observed in cellular signaling pathways, but there was a significant increase GLUT-4 protein expression (47%) following EECP.Our findings provide novel evidence that EECP has a beneficial effect on peripheral arterial function and glucose tolerance in subjects with AGT.

The molecular signaling mechanisms by which insulin leads to increased glucose transport and metabolism and gene expression are not completely elucidated. We have characterized the nature of insulin signaling defects in skeletal muscle from Type 2 diabetic patients. Insulin receptor substrate (IRS-1) phosphorylation, phosphatidylinositol (PI) 3-kinase activity, and glucose transport activity are impaired as a consequence of functional defects, whereas insulin receptor tyrosine phosphorylation, mitogen-activated protein kinase (MAPK) phosphorylation, and glycogen synthase activity are normal. Using biotinylated photoaffinity labeling, we have shown that reduced cell surface GLUT4 levels can explain glucose transport defects in skeletal muscle from Type 2 diabetic patients under insulin-stimulated conditions. Current work is focused on mechanisms behind insulin-dependent and insulin-independent regulation of glucose uptake. We have recently determined the independent effects of insulin and hypoxia/AICAR exposure on glucose transport and cell surface GLUT4 content in skeletal muscle from nondiabetic and Type 2 diabetic subjects. Hypoxia and AICAR increase glucose transport via an insulin-independent mechanism involving activation of 5'-AMP-activated kinase (AMPK). AMPK signaling is intact, because 5-aminoimidazole-4-carboxamide 1-beta-D-ribonucleoside (AICAR) increased AMPK and acetyl-CoA carboxylase (ACC) phosphorylation to a similar extent in Type 2 diabetic and nondiabetic subjects. However, AICAR responses on glucose uptake were impaired. Our studies highlight important AMPK-dependent and independent pathways in the regulation of GLUT4 and glucose transport activity in insulin resistant skeletal muscle. Understanding signaling mechanisms to downstream metabolic responses may provide valuable clues to a future therapy for Type 2 diabetes.

Several adipocyte-secreted factors have been demonstrated to potentially link obesity, insulin resistance, and cardiovascular disease. Among those, adiponectin is an insulin-sensitizing and anti-inflammatory adipokine, concentrations of which are decreased in obesity-associated metabolic and vascular disorders. Recently, two adiponectin receptors (AdipoR) have been isolated and adenosine monophosphate kinase (AMPK), as well as acetyl coenzyme A carboxylase (ACC), appear to be critical downstream mediators for various effects of this adipokine. In addition to beneficial metabolic effects, adiponectin seems to be vasoprotective by interfering with various atherogenic processes. Of clinical interest, thiazolidinediones (TZDs) which are used in the treatment of type 2 diabetes stimulate adiponectin expression and secretion whereas several hormones dysregulated in insulin resistance and obesity downregulate this adipokine. The current knowledge on regulation and function of adiponectin in obesity, insulin resistance, and cardiovascular disease is summarized in this review and its clinical implications are discussed.

The AMP-activated protein kinase (AMPK) is a metabolic-stress-sensing protein kinase that regulates metabolism in response to energy demand and supply by directly phosphorylating rate-limiting enzymes in metabolic pathways as well as controlling gene expression.

AMPK (5'-AMP-activated protein kinase) is emerging as a metabolic master switch, by which cells in both mammals and lower organisms sense and decode changes in energy status. Changes in AMPK activity have been shown to regulate glucose transport in muscle and glucose production by the liver. Moreover, AMPK appears to be a key regulator of at least one transcription factor linked to a monogenic form of diabetes mellitus. As a result, considerable efforts are now under way to explore the usefulness of AMPK as a therapeutic target for other forms of this disease. Here we review this topic, and discuss new findings which suggest that AMPK may play roles in regulating insulin release and the survival of pancreatic islet beta-cells, and nutrient sensing by the brain.

An increasing body of evidence has revealed that activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK)-activated protein kinase increases fatty acid oxidation by lowering the concentration of malonyl coenzyme A (CoA), an inhibitor of carnitine palmitoyl transferase 1. Studies carried out primarily in skeletal muscle suggest that AMPK modulates the concentration of malonyl CoA by concurrently phosphorylating and inhibiting acetyl CoA carboxylase (ACC), the rate limiting enzyme in malonyl CoA synthesis, and phosphorylating and activating malonyl CoA decarboxylase (MCD), an enzyme involved in its degradation. We have recently observed that AMPK and MCD activities are increased and ACC activity diminished in skeletal muscle, liver and, surprisingly, in adipose tissue 30 min following exercise (treadmill run) in normal rats. In liver and adipose tissue these changes were associated with a decrease in the activity of glycerol-3-phosphate acyltransferase (GPAT), which catalyses the first committed reaction in glycerolipid synthesis and, which like ACC, is phosphorylated and inhibited by AMPK. Similar changes in ACC, MCD and GPAT were observed following the administration of 5-aminoimidazole 4-carboxamide-riboside (AICAR), further indicating that the exercise-induced alterations in these enzymes were AMPK-mediated. CONCLUSIONS: (1) AMPK plays a major role in regulating lipid metabolism in multiple tissues following exercise. (2) The net effect of its activation is to increase fatty acid oxidation and diminish glycerolipid synthesis. (3) The relevance of these findings to the regulation of muscle glycogen repletion in the post-exercise state and to the demonstrated ability of AMPK activation to decrease adiposity and increase insulin sensitivity in rodents remains to be determined.

The AMPK (5'AMP-activated protein kinase) is becoming recognized as a critical regulator of energy metabolism. However, many of these effects in muscle metabolism have been ascribed to AMPK based on the use of the unspecific activator AICAR (5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside). Using mouse models in which AMPK activity has been specifically blocked (kinase dead) or knocked out we and others have been able to conduct studies gaining more conclusive data on the role of AMPK in muscle metabolism. In this mini-review focus is on AMPK and its regulatory role for glucose transport and GS (glycogen synthase) activity in skeletal muscle, indicating that AMPK is a GS kinase in vivo which might influence GS activity during exercise and that AMPK is involved in AICAR/hypoxia-induced glucose transport but not or only partially in contraction-stimulated glucose transport.