To determine whether nitric oxide (NO) synthase activity exists in rat skeletal muscle, media from incubated rat extensor digitorum longus muscle preparations were assayed for NO with a chemiluminescent detection system. Although small amounts of NO were detected in media alone, the addition of muscle increased NO concentration in the media by 30-fold. The release of NO into the media diminished over time. Either arginine (10(-6) M), sodium nitroprusside (10(-6) M), or prior electrical stimulation in vivo caused 50-200% increases (P < 0.05) in NO concentration. NG-monomethyl-L-arginine monoacetate (10(-6) M), an NO synthase inhibitor, decreased both basal 2-deoxyglucose transport and NO efflux, indicating that NO may play a role in modulating skeletal muscle carbohydrate metabolism. These data indicate that NO is released from an incubated skeletal muscle preparation and presents the possibility that muscle-derived NO may play an important metabolic role.

Nitric oxide synthase (NOS) is expressed in skeletal muscle. However, the role of nitric oxide (NO) in glucose transport in this tissue remains unclear. To determine the role of NO in modulating glucose transport, 2-deoxyglucose (2-DG) transport was measured in rat extensor digitorum longus (EDL) muscles that were exposed to either a maximally stimulating concentration of insulin or to an electrical stimulation protocol, in the presence of NG-monomethyl-L-arginine, a NOS inhibitor. In addition, EDL preparations were exposed to sodium nitroprusside (SNP), an NO donor, in the presence of submaximal and maximally stimulating concentrations of insulin. NOS inhibition reduced both basal and exercise-enhanced 2-DG transport but had no effect on insulin-stimulated 2-DG transport. Furthermore, SNP increased 2-DG transport in a dose-responsive manner. The effects of SNP and insulin on 2-DG transport were additive when insulin was present in physiological but not in pharmacological concentrations. Chronic treadmill training increased protein expression of both type I and type III NOS in soleus muscle homogenates. Our results suggest that NO may be a potential mediator of exercise-induced glucose transport.

The 12/15-lipoxygenase enzymes react with fatty acids producing active lipid metabolites that are involved in a number of significant disease states. The latter include type 1 and type 2 diabetes (and associated complications), cardiovascular disease, hypertension, renal disease, and the neurological conditions Alzheimer’s disease and Parkinson’s disease. A number of elegant studies over the last thirty years have contributed to unraveling the role that lipoxygenases play in chronic inflammation. The development of animal models with targeted gene deletions has led to a better understanding of the role that lipoxygenases play in various conditions. Selective inhibitors of the different lipoxygenase isoforms are an active area of investigation, and will be both an important research tool and a promising therapeutic target for treating a wide spectrum of human diseases.

OBJECTIVE-Subjects with dietary obesity and pre-diabetes have an increased risk for developing both nerve conduction slowing and small sensory fiber neuropathy. Animal models of this type of neuropathy have not been described. This study evaluated neuropathic changes and their amenability to dietary and pharmacological interventions in mice fed a high-fat diet (HFD), a model of pre-diabetes and alimentary obesity.RESEARCH DESIGN AND METHODS-Female C57BL6/J mice were fed normal diets or HFDs for 16 weeks.RESULTS-HFD-fed mice developed obesity, increased plasma FFA and insulin concentrations, and impaired glucose tolerance. They also had motor and sensory nerve conduction deficits, tactile allodynia, and thermal hypoalgesia in the absence of intraepidermal nerve fiber loss or axonal atrophy. Despite the absence of overt hyperglycemia, the mice displayed augmented sorbitol pathway activity in the peripheral nerve, as well as 4-hydroxynonenal adduct nitrotyrosine and poly(ADP-ribose) accumulation and 12/15-lipoxygenase overexpression in peripheral nerve and dorsal root ganglion neurons. A 6-week feeding with normal chow after 16 weeks on HFD alleviated tactile allodynia and essentially corrected thermal hypoalgesia and sensory nerve conduction deficit without affecting motor nerve conduction slowing. Normal chow containing the aldose reductase inhibitor fidarestat (16 mg ⅐ kg Ϫ1 ⅐ day Ϫ1 ) corrected all functional changes of HFD-induced neuropathy.CONCLUSIONS-Similar to human subjects with pre-diabetes and obesity, HFD-fed mice develop peripheral nerve functional, but not structural, abnormalities and, therefore, are a suitable model for evaluating dietary and pharmacological approaches to halt progression and reverse diabetic neuropathy at the earliest stage of the disease. Diabetes 56: [2598][2599][2600][2601][2602][2603][2604][2605][2606][2607][2608] 2007 O ver the last decade, profound changes in the quality, quantity, and source of food consumed in many developed countries combined with a decrease in levels of physical activity have led to an increase in the prevalence of diabetes and its complications (1). Furthermore, some manifestations of peripheral diabetic neuropathy (PDN) and cardiovascular disease in overweight and obese subjects develop at the stage of impaired glucose tolerance (IGT), preceding overt diabetes (2-4). A high BMI is a well-recognized risk factor for median nerve sensory conduction slowing and carpal tunnel syndrome (5-7). Furthermore, nondiabetic obese subjects have been reported to display significantly decreased compound muscle action potential amplitude of tibial and peroneal nerves and decreased sensory action potential amplitude of median, ulnar, and sural nerves compared with nondiabetic individuals (8). In the same study, warm and cold sensations from the index and little fingers, warm sensation from the big toe, and thermal and pain thresholds from the little finger directly correlated with the insulin sensitivity index, which was reduced in obese subjects. A higher prevalence...

Patients with diabetes are at higher risk for atherosclerotic disease than nondiabetic individuals with other comparable risk factors. Studies examining mechanisms underlying diabetes-accelerated atherosclerosis have been limited by the lack of suitable humanoid animal models. In this study, diabetes was superimposed on a well-characterized swine model of atherosclerosis by injection of the ␤-cell cytotoxin streptozotocin (STZ), resulting in a >80% reduction in ␤-cells and an increase in plasma glucose to diabetic levels. Animals were maintained without exogenous insulin for up to 48 weeks. Plasma glucose and cholesterol levels and lesion extent and severity were quantified in swine with diabetes and hyperlipemia alone and in combination compared with controls. Diabetes had no effect on plasma cholesterol levels, P atients with diabetes are at two-to sixfold greater risk of developing atherosclerosis than nondiabetic individuals (1,2), and the most common cause of death in adult diabetic patients is coronary heart disease (3). This excess risk occurs in both type 1 and type 2 diabetes (3). In contrast to nondiabetic subjects, heart disease in diabetic subjects appears earlier in life, affects women almost as often as men, and is more often fatal (3). As many as 80% of all type 2 diabetic patients will die of atherosclerotic macrovascular disease (4), and adult diabetic patients are more susceptible to developing other atherosclerotic risk factors, including hypertension and dyslipidemias (3). Furthermore, data on the efficacy of insulin control of diabetes in ameliorating atherosclerotic disease are inconsistent. Although improvement of glycemic control may reduce the risk of heart disease in type 1 diabetic patients (3), insulin-treated type 2 diabetic patients continue to have increased risk of cardiovascular events (3).The mechanisms underlying the increased risk and greater acceleration and severity of atherosclerotic disease in diabetes remain an enigma due, at least in part, to the lack of suitable humanoid animal models (5,6). In this study, we chose to induce diabetes in an atherosclerotic swine model, which we have extensively characterized over the last two decades (7-15). Swine has proven to be an excellent model for cardiovascular studies, in that the development, morphology, and function of the normal cardiovascular system in swine closely resembles that of humans (16). Swine are also, like humans, omnivores, develop spontaneous atherosclerosis with increased age (17), and have lipoprotein profiles and metabolism similar to humans (7,18 -20). When fed cholesterol-and lipidcontaining diets, they develop humanoid atherosclerosis in most arterial beds in time periods inversely proportional to cholesterol level (7)(8)(9)(10)(11)(12)(13)(14)21). Therefore, by use of streptozotocin (STZ) as a diabetes-inducing agent that we superimposed on the atherosclerotic swine model, we set out to determine the following: 1) whether diabetes could be reproducibly produced and maintained over prolonged time periods in...

Glycemic traits are used to diagnose and monitor type 2 diabetes, and cardiometabolic health. To date, most genetic studies of glycemic traits have focused on individuals of European ancestry. Here, we aggregated genome-wide association studies in up to 281,416 individuals without diabetes (30% non-European ancestry) with fasting glucose, 2h-glucose post-challenge, glycated hemoglobin, and fasting insulin data. Trans-ancestry and single-ancestry meta-analyses identified 242 loci (99 novel; P <5x10 -8 ), 80% with no significant evidence of between-ancestry heterogeneity. Analyses restricted to European ancestry individuals with equivalent sample size would have led to 24 fewer new loci. Compared to single-ancestry, equivalent sized trans-ancestry fine-mapping reduced the number of estimated variants in 99% credible sets by a median of 37.5%. Genomic feature, gene-expression and gene-set analyses revealed distinct biological signatures for each trait, highlighting different underlying biological pathways. Our results increase understanding of diabetes pathophysiology by use of trans-ancestry studies for improved power and resolution.

Increased oxidative stress has been reported in vivo in the diabetic state via the production of reactive oxygen species (ROS). Such stress is bound to play a key role on activation of circulating monocytes, leading to the accelerated atherosclerosis observed in diabetics. However the exact molecular mechanisms of monocyte activation by high glucose is currently unclear. Here, we demonstrate that chronic high glucose (CHG) causes a dramatic increase in the release of the inflammatory cytokine tumor necrosis factor ␣ (TNF␣), at least in part through enhanced TNF␣ mRNA transcription, mediated by ROS via activation of transcription factors nuclear factor B (NF-B) and activating protein-1 (AP-1). TNF␣ accumulation in the conditioned media was increased 10-fold and mRNA levels were increased 11.5-fold by CHG. The following observations supported that both NF-B and AP-1 mediated enhanced TNF␣ transcription by CHG: 1) A 295-base pair fragment of the proximal TNF␣ promoter containing NF-B and AP-1 sites reproduced the effects of CHG on TNF␣ transcription in a luciferase reporter assay, 2) mutational analyses of both NF-B and the AP-1 sites abrogated 90% of the luciferase activity, 3) gel-shift analysis using the binding sites showed activation of NF-B and AP-1 in CHG nuclear extracts, and 4) Western blot analyses demonstrated elevated nuclear levels of p65 and p50 and decreased cytosolic levels of IB␣ in CHG-treated monocytes. That ROS acted as a key intermediate in the CHG pathway was supported by the following evidence: 1) increased superoxide levels similar to those observed with PMA or TNF␣, 2) increased phosphorylation of stress-responsive mitogen-activated protein kinases p38 and JNK-1, 3) counteraction of the effects of CHG on TNF␣ production, the 295TNFluc reporter activity, activation of NF-B, and repression of IB␣ by antioxidants and p38 mitogen-activated protein kinase inhibitors. The study suggests that ROS function as key components in the regulatory pathway progressing from elevated glucose to monocyte activation.

Abstract-Type 2 diabetes is associated with significantly accelerated rates of macrovascular complications such as atherosclerosis. Emerging evidence now indicates that atherosclerosis is an inflammatory disease and that certain inflammatory markers may be key predictors of diabetic atherosclerosis. Proinflammatory cytokines and cellular adhesion molecules expressed by vascular and blood cells during stimulation by growth factors and cytokines seem to play major roles in the pathophysiology of atherosclerosis and diabetic vascular complications. However, more recently, data suggest that inflammatory responses can also be elicited by smaller oxidized lipids that are components of atherogenic oxidized low-density lipoprotein or products of phospholipase activation and arachidonic acid metabolism. These include oxidized lipids of the lipoxygenase and cyclooxygenase pathways of arachidonic acid and linoleic acid metabolism. These lipids have potent growth, vasoactive, chemotactic, oxidative, and proinflammatory properties in vascular smooth muscle cells, endothelial cells, and monocytes. Cellular and animal models indicate that these enzymes are induced under diabetic conditions, have proatherogenic effects, and also mediate the actions of growth factors and cytokines. This review highlights the roles of the inflammatory cyclooxygenase and 12/15-lipoxygenase pathways in the pathogenesis of diabetic vascular disease. Key Words: lipoxygenase Ⅲ diabetes Ⅲ diabetes complications Ⅲ inflammation Ⅲ lipids D iabetes is associated with significantly accelerated rates of cardiovascular complications such as atherosclerosis and hypertension. In particular, type 2 diabetes is associated with 2-to 4-fold increase in coronary artery disease. 1 This has been attributed to the clustering of several risk factors, including insulin resistance, hypertension, obesity, and dyslipidemia. 2,3 Multiple mechanisms contribute to vascular and arterial disease in the diabetic population. 2 Basic biochemical mechanisms have been described by which hyperglycemiainduced oxidant stress activates several downstream signals that mediate diabetic complications. 4 -8 Furthermore, advanced glycation end products formed by glucose-induced modification of proteins can act via their receptors such as RAGE and induce cellular oxidant stress, inflammation, and vascular dysfunction in diabetes. 9 -12 Recent evidence from laboratory and clinical studies demonstrates that diabetic atherosclerosis is not simply a disease of hyperlipidemia but also an inflammatory disorder involving multiple mediators such as C-reactive protein, cytokines such as tumor necrosis factor alpha, and interluekin-6 (IL-6). 2,13,14 A recent gene profiling study showed that high glucose treatment of monocytes leads to increased expression of multiple inflammatory cytokines, chemokines, and related factors, many of which are regulated by the proinflammatory transcription factor, nuclear factor-kappa B (NF-B). 15 The recognition now that highly effective antidiabetic agents, such as thi...