cyclic-gmp and Insulin-Resistance

Natriuretic peptides (NPs) are a group of peptide-hormones mainly secreted from the heart, signaling via c-GMP coupled receptors. NP are well known for their renal and cardiovascular actions, reducing arterial blood pressure as well as sodium reabsorption. Novel physiological functions have been discovered in recent years, including activation of lipolysis, lipid oxidation, and mitochondrial respiration. Together, these responses promote white adipose tissue browning, increase muscular oxidative capacity, particularly during physical exercise, and protect against diet-induced obesity and insulin resistance. Exaggerated NP release is a common finding in congestive heart failure. In contrast, NP deficiency is observed in obesity and in type-2 diabetes, pointing to an involvement of NP in the pathophysiology of metabolic disease. Based upon these findings, the NP system holds the potential to be amenable to therapeutical intervention against pandemic diseases such as obesity, insulin resistance, and arterial hypertension. Various therapeutic approaches are currently under development. This paper reviews the current knowledge on the metabolic effects of the NP system and discusses potential therapeutic applications.

Topics: Animals; Cyclic GMP; Drug Design; Exercise; Humans; Hypertension; Insulin Resistance; Metabolic Syndrome; Natriuretic Peptides; Obesity

Central obesity shows impaired platelet responses to the antiaggregating effects of nitric oxide (NO), prostacyclin, and their effectors--guanosine 3',5'-cyclic monophosphate (cGMP) and adenosine 3',5'-cyclic monophosphate (cAMP). The influence of weight loss on these alterations is not known. To evaluate whether a diet-induced body-weight reduction restores platelet sensitivity to the physiological antiaggregating agents and reduces platelet activation in subjects affected by central obesity, we studied 20 centrally obese subjects before and after a 6-month diet intervention aiming at reducing body weight by 10%, by measuring (i) insulin sensitivity (homeostasis model assessment of insulin resistance (HOMA(IR))); (ii) plasma lipids; (iii) circulating markers of inflammation of adipose tissue and endothelial dysfunction, and of platelet activation (i.e., soluble CD-40 ligand (sCD-40L) and soluble P-selectin (sP-selectin)); (iv) ability of the NO donor sodium nitroprusside (SNP), the prostacyclin analog Iloprost and the cyclic nucleotide analogs 8-bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP) and 8-bromoadenosine 3',5'-cyclic monophosphate (8-Br-cAMP) to reduce platelet aggregation in response to adenosine-5-diphosphate (ADP); and (v) ability of SNP and Iloprost to increase cGMP and cAMP. The 10 subjects who reached the body-weight target showed significant reductions of insulin resistance, adipose tissue, endothelial dysfunction, and platelet activation, and a significant increase of the ability of SNP, Iloprost, 8-Br-cGMP, and 8-Br-cAMP to reduce ADP-induced platelet aggregation and of the ability of SNP and Iloprost to increase cyclic nucleotide concentrations. No change was observed in the 10 subjects who did not reach the body-weight target. Changes of platelet function correlated with changes of HOMA(IR). Thus, in central obesity, diet-induced weight loss reduces platelet activation and restores the sensitivity to the physiological antiaggregating agents, with a correlation with improvements in insulin sensitivity.

Topics: Adenosine Diphosphate; Adipose Tissue; Adult; Blood Platelets; Cyclic AMP; Cyclic GMP; Diet, Reducing; Endothelium, Vascular; Epoprostenol; Female; Humans; Iloprost; Insulin Resistance; Male; Nitric Oxide; Nitroprusside; Obesity, Abdominal; Platelet Activation; Platelet Aggregation Inhibitors; Weight Loss

Preeclampsia (PE) is a common maternal disease that complicates 5 to 10% of pregnancies and remains as the major cause of maternal and neonatal mortality. Cost-effective interventions aimed at preventing the development of preeclampsia are urgently needed. However, the pathogenesis of PE is not well known. Multiple mechanisms such as oxidative stress, endothelial dysfunction and insulin resistance may contribute to its development. Regular aerobic exercise recovers endothelial function; improves insulin resistance and decreases oxidative stress. Therefore the purpose of this clinical trial is to determine the effect of regular aerobic exercise on endothelial function, on insulin resistance and on pregnancy outcome.. 64 pregnant women will be included in a blind, randomized clinical trial, and parallel assignment. The exercise group will do regular aerobic physical exercise: walking (10 minutes), aerobic exercise (30 minutes), stretching (10 minutes) and relaxation exercise (10 minutes) in three sessions per week. Control group will do the activities of daily living (bathing, dressing, eating, and walking) without counselling from a physical therapist.. NCT00741312.

Topics: Adolescent; Adult; Brachial Artery; Cyclic GMP; Endothelium, Vascular; Exercise; Female; Humans; Insulin Resistance; Nitric Oxide; Pregnancy; Single-Blind Method

The aim of the present study was to evaluate the effect of prolonged inhibition of beta-oxidation on glucose and lipid muscle forearm metabolism and cGMP and endothelin-1 forearm release in patients with type 2 diabetes mellitus and ischemic cardiomyopathy. Fifteen patients were randomly allocated in a double-blind cross-over parallel study with trimetazidine (20 mg tid) or placebo lasting 15 days. At the end of each period, all patients underwent euglycemic hyperinsulinemic clamps with forearm indirect calorimetry and endothelial balance of vasodilator and vasoconstricor factors. Compared with placebo, trimetazidine induced 1) an increase in insulin-induced forearm glucose uptake and glucose oxidation accompanied by a reduction in forearm lipid oxidation and citrate release and 2) a decrease of endothelin-1 release paralleled by a significant increase in forearm cGMP release. Forearm glucose oxidation significantly correlated with cGMP release (r=0.37, P<0.04), whereas forearm lipid oxidation positively correlated with endothelin-1 release (r=0.40, P<0.03). In conclusion, for the first time, we demonstrated that insulin-induced forearm glucose oxidation and forearm cGMP release were increased whereas forearm endothelin-1 release was decreased during trimetazidine treatment. Muscle's metabolic and vascular effects of trimetazidine add new interest in the use of trimetazidine in type 2 diabetic patients with cardiovascular disease.

Topics: 3-Hydroxybutyric Acid; Aged; Blood Glucose; Citric Acid; Cross-Over Studies; Cyclic GMP; Diabetes Mellitus, Type 2; Double-Blind Method; Endothelin-1; Endothelium, Vascular; Fatty Acids, Nonesterified; Forearm; Glucose; Glucose Clamp Technique; Humans; Insulin; Insulin Resistance; Lipid Metabolism; Male; Middle Aged; Muscle, Skeletal; Myocardial Ischemia; Oxidation-Reduction; Trimetazidine

Essential hypertension has a familial predisposition, but the phenotype of elevated blood pressure has delayed penetrance. Because the kidney is a crucial determinant of blood pressure homeostasis, we studied early glomerular alterations in still-normotensive young subjects at genetic risk of hypertension. Thirty-nine normotensive adults (mean age 29 to 31 years), stratified by genetic risk (parental family history [FH]) of hypertension (26 with positive FH [FH+], 13 with negative FH [FH-]), underwent intravenous infusion of mixed amino acids. Before and during amino acid administration, we measured glomerular filtration rate (GFR), putative second messengers of amino acids (nitric oxide [NO.] metabolites and cGMP), serum insulin and amino acid concentrations, and the FE(Li)+ as an index of renal proximal tubular reabsorption. The FH+ group had a blunted GFR rise in response to amino acids (2.43+/-8.16% versus 31.0+/-13.4% rise, P:=0.0126). The amino acid-induced change in GFR correlated (r=0.786, P:<0.01) with the change in urinary NO. metabolite excretion; a diminished rise in urinary NO. metabolite excretion in the FH+ group (P:=0.0105) suggested a biochemical mechanism for the different GFR responses between FH groups: a relative inability to convert arginine to NO. The FH+ group had a far lower initial cGMP excretion at baseline (261+/-21.1 versus 579+/-84.9 nmol. h(-1)/1.73 m(2), P:=0.001), although cGMP did not change during the amino acid infusion (P:=0.703). FH status, baseline GFR, and baseline serum insulin jointly predicted GFR response to amino acids (P:=0.0013), accounting for approximately 45% of the variance in GFR response. Decline in FE(Li)+, an inverse index of proximal tubular reabsorption, paralleled increase in GFR (r=-0.506, P:=0.01), suggesting differences in proximal tubular reabsorption during amino acids between the FH groups. GFR response to amino acid infusion was blunted in the FH+ group despite significantly higher serum concentrations of 6 amino acids (arginine, isoleucine, leucine, methionine, phenylalanine, and valine) in the FH+ group, suggesting a novel form of insulin resistance (to the amino acid-translocating action of insulin) in FH+ subjects. We conclude that blunted glomerular filtration reserve in response to amino acids is an early-penetrance phenotype seen even in still-normotensive subjects at genetic risk of hypertension and is linked to impaired formation of NO. in the kidney. Corresponding changes in GFR an

Topics: Adult; Amino Acids; Arginine; Cyclic GMP; Female; Genetic Markers; Glomerular Filtration Rate; Humans; Hypertension; Infusions, Intravenous; Insulin; Insulin Resistance; Kidney Glomerulus; Kidney Tubules, Proximal; Lithium Carbonate; Male; Middle Aged; Nitrates; Nitrites; Phenotype; Risk Factors; Second Messenger Systems

In this study, we have compared resistance to insulin-mediated glucose disposal and plasma concentrations of nitric oxide (NO) and cyclic-GMP in healthy volunteers with (n = 35) or without (n = 27) at least one sibling and one parent with type 2 diabetes. The 62 volunteers were further divided into groups of those with normal glucose tolerance or impaired glucose tolerance. Insulin-mediated glucose disposal was quantified by determining the insulin sensitivity index (ISI) in response to a low-dose, constant infusion of insulin (25 mU/kg x h) and glucose (4 mg/kg x min) for 150 min. The mean (+/-SEM) ISI [(mL kg(-1) min(-1)/pmol/L) x 10(3)] was significantly greater in those without a family history (30.3 +/- 2.3) as compared with nondiabetic volunteers with a family history of type 2 diabetes, whether they had normal glucose tolerance (17.0 +/- 7.2) or impaired glucose tolerance (9.5 +/- 1.4). In addition, basal NO levels, evaluated by the measurement of its stable end products [i.e. nitrite and nitrate levels (NO2-/ NO3-)], were significantly higher, and cyclic-GMP levels, its effector messenger, were significantly lower in those with a family history, irrespective of their degree of glucose tolerance, when compared with healthy volunteers without a family history of type 2 diabetes. Furthermore, when the 62 volunteers were analyzed as one group, there was a negative correlation between ISI and NO2-/NO3- levels (r = -0.35; P < 0.005) and a positive correlation between ISI and cyclic-GMP levels (r = 0.30; P < 0.02). These results have shown that alterations of the NO/cyclic-GMP pathway seem to be an early event in nondiabetic individuals with a family history of type 2 diabetes and these changes are correlated with the degree of insulin resistance.

Topics: Blood Glucose; Cyclic GMP; Diabetes Mellitus, Type 2; Diet; Female; Humans; Insulin; Insulin Resistance; Male; Middle Aged; Nitric Oxide

A decrease in intracellular levels of 3',5'-cyclic guanosine monophosphate (cGMP) has been implicated in the progression of diabetic nephropathy. Hyperglycemia significantly inhibits cGMP-dependent pathway activity in the kidney, leading to glomerular damage and proteinuria. The enhancement of activity of this pathway that is associated with an elevation of cGMP levels may be achieved by inhibition of the cGMP specific phosphodiesterase 5A (PDE5A) using selective inhibitors, such as tadalafil. Hyperglycemia decreased the insulin responsiveness of podocytes and impaired podocyte function. These effects were associated with lower protein amounts and activity of the protein deacetylase sirtuin 1 (SIRT1) and a decrease in the phosphorylation of adenosine monophosphate-dependent protein kinase (AMPK). We found that PDE5A protein levels increased in hyperglycemia, and PDE5A downregulation improved the insulin responsiveness of podocytes with reestablished SIRT1 expression and activity. PDE5A inhibitors potentiate nitric oxide (NO)/cGMP signaling, and NO modulates the activity and expression of SIRT1. Therefore, we investigated the effects of tadalafil on SIRT1 and AMPK in the context of improving the insulin sensitivity in podocytes and podocyte function in hyperglycemia. Our study revealed that tadalafil restored SIRT1 expression and activity and activated AMPK by increasing its phosphorylation. Tadalafil also restored stimulating effect of insulin on glucose transport in podocytes with high glucose-induced insulin resistance. Additionally, tadalafil improved the function of podocytes that were exposed to high glucose concentrations. Our results display novel mechanisms involved in the pathogenesis of glomerulopathies in diabetes, which may contribute to the development of more effective treatment strategies for diabetic nephropathy.

Topics: AMP-Activated Protein Kinases; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Diabetic Nephropathies; Glucose; Humans; Hyperglycemia; Insulin; Insulin Resistance; Podocytes; Sirtuin 1; Tadalafil

Robust associations between low plasma level of natriuretic peptides (NP) and increased risk of type 2 diabetes (T2D) have been recently reported in humans. Adipose tissue (AT) is a known target of NP. However it is unknown whether NP signalling in human AT relates to insulin sensitivity and modulates glucose metabolism. We here show in two European cohorts that the NP receptor guanylyl cyclase-A (GC-A) expression in subcutaneous AT was down-regulated as a function of obesity grade while adipose NP clearance receptor (NPRC) was up-regulated. Adipose GC-A mRNA level was down-regulated in prediabetes and T2D, and negatively correlated with HOMA-IR and fasting blood glucose. We show for the first time that NP promote glucose uptake in a dose-dependent manner. This effect is reduced in adipocytes of obese individuals. NP activate mammalian target of rapamycin complex 1/2 (mTORC1/2) and Akt signalling. These effects were totally abrogated by inhibition of cGMP-dependent protein kinase and mTORC1/2 by rapamycin. We further show that NP treatment favoured glucose oxidation and de novo lipogenesis independently of significant gene regulation. Collectively, our data support a role for NP in blood glucose control and insulin sensitivity by increasing glucose uptake in human adipocytes. This effect is partly blunted in obesity.

Topics: Adipocytes; Adipose Tissue; Biomarkers; Cyclic GMP; Diabetes Mellitus, Type 2; Gene Expression Regulation; Glucose; Humans; Insulin Resistance; Models, Biological; Natriuretic Peptides; Obesity; Proto-Oncogene Proteins c-akt; Receptors, Atrial Natriuretic Factor; Signal Transduction; TOR Serine-Threonine Kinases

Topics: Adipocytes; Animals; Blotting, Western; Bradykinin; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Dual-Specificity Phosphatases; Glucose; Guanylate Cyclase; Immunoprecipitation; Insulin Resistance; JNK Mitogen-Activated Protein Kinases; Male; Mitogen-Activated Protein Kinase Phosphatases; Nitric Oxide; Nitric Oxide Synthase Type III; Phosphodiesterase Inhibitors; Proto-Oncogene Proteins c-akt; Purinones; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; RNA, Messenger; Signal Transduction

We hypothesized that carbon monoxide (CO) might suppress chronic inflammation, which led to metabolic disturbances. Ovariectomy (OVX) was performed in mice to mimic chronic inflammation secondary to loss of ovarian function. OVX increased fat mass and the infiltration of highly inflammatory CD11c cells into adipose tissue (AT), resulting in a disturbance of glucose metabolism. Treatment of CO attenuated these; CO decreased recruitment of CD11c-expressing cells in AT and reduced expression of CD11c in bone marrow-derived macrophages, protecting them from M1 polarization. Upregulated cGMP and decreased reactive oxygen species were responsible for the inhibitory activity of CO on CD11c expression; knockdown of soluble guanylate cyclase or heme oxygenase-1 using small interfering RNAs reduced this inhibition substantially. Improved OVX-induced insulin resistance (IR) by CO was highly associated with its activity to attenuate AT inflammation. Our results suggest a therapeutic value of CO to treat postmenopausal IR by reducing AT inflammation.

Topics: Adipose Tissue, White; Adiposity; Aging; Animals; Antimetabolites; Carbon Monoxide; Cells, Cultured; Cyclic GMP; Female; Guanylate Cyclase; Heme Oxygenase-1; Injections, Intraperitoneal; Insulin Resistance; Macrophages; Membrane Proteins; Mice, Inbred C57BL; Organometallic Compounds; Ovariectomy; Panniculitis; Prodrugs; Reactive Oxygen Species; Receptors, Cytoplasmic and Nuclear; RNA Interference; Soluble Guanylyl Cyclase; Specific Pathogen-Free Organisms

Although recent studies have underscored the role of the heme-oxygenase (HO) inducer hemin, on insulin-signaling and glucose metabolism, the underlying mechanisms are not completely understood. In this study, two-dimensional-gel electrophoresis, massspectrometry and MSACOT-analyses were used to identify and characterize novel proteins modulated by hemin in spontaneoushypertensive rat (SHR), a model of essential hypertension with insulin resistance/impaired glucose metabolism. In addition, the effects of hemin on endothelin-1 (ET-1), protein-tyrosine-phosphatase-1B (PTP-1B), atrial-natriuretic-peptide (ANP) and its surrogate-marker urinary cGMP, and inflammatory cytokines including TNF-α, IL-6 and IL-1β were investigated. In hemin-treated SHR, several proteins related to oxidative-stress and metabolism were modulated. Particularly, hemin enhanced aldolase- B, fumarylacetoacetate hydrolase, purine-nucleoside phosphorylase, adenosine-kinase, argininosuccinate synthetase and carbonic anhydrase-3 all of which are enzymes involved in glucose/energy metabolism and pH homeostasis. Similarly, hemin potentiated antioxidant pathways including, NADP(+)-dependant isocitrate-dehydrogenase, catalase, glutathione-S-transferase-Yb1 and hsp70, a pleiotropic agent that regulates protein-folding, oxidative/pro-inflammatory events. Hemin also increased enzymes implicated in cell-growth such as the nitrilase-protein-family, but reduced betaine-homocysteine methyltransferase, an enzyme associated with insulin resistance and dysfunctional glucose metabolism. Furthermore, hemin increased ANP and its surrogate marker, urinary cGMP, but reduced ET-1, PTP-1B, TNF-α, IL-6, IL-1β, whereas the HO-inhibitor, chromium-mesoporphyrin abolished the effects. The potentiation of ANP, urinary-cGMP, aldolade-B, fumarylacetoacetate hydrolase, purine-nucleoside phosphorylase, adenosine-kinase, argininosuccinate synthetase, carbonic anhydrase-3, hsp70 and the corresponding reduction of betaine-homocysteine methyltransferase, PTP-1B, TNF-α, IL-6, IL-1β, and ET-1 may be responsible for the improved glucose metabolism in hemin-treated animals. Collectively, these findings underscore the pleiotropic effects of the HO-system in cellular homeostasis with important roles in metabolism and defence.

Topics: Animals; Cyclic GMP; Cytokines; Electrophoresis, Gel, Two-Dimensional; Endothelin-1; Essential Hypertension; Female; Glucose; Heme Oxygenase (Decyclizing); Hemin; Hypertension; Inflammation; Inflammation Mediators; Insulin Resistance; Male; Mass Spectrometry; Oxidative Stress; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Rats, Sprague-Dawley

Gal-geun-dang-gwi-tang (GGDGT), an herbal medicine, is used to treat hypertension, stroke, and other inflammatory disorders in the clinical setting. Recently, GGDGT was recognized by the Korea Institute of Oriental Medicine. This study aimed to evaluate the effects of GGDGT in a diabetic atherosclerosis model using apolipoprotein E knockout (ApoE-/-) mice fed a Western diet.. The mice were divided into four groups: control group, C57BL6J mice receiving a regular diet (RD); ApoE-/- group, ApoE-/- mice receiving a Western diet (WD); rosiglitazone group, ApoE-/- mice receiving rosiglitazone (WD + 10 mg · kg(-1) · day(-1)); GGDGT group, ApoE-/- mice receiving GGDGT (WD + 200 mg · kg(-1) · day(-1)).. Treatment with GGDGT significantly improved glucose tolerance and plasma lipid levels. In addition, GGDGT ameliorated acetylcholine-induced vascular relaxation of the aortic rings. Immunohistochemical staining showed that GGDGT suppressed intercellular adhesion molecule (ICAM)-1 expression; however, expression of endothelial nitric oxide synthase (eNOS) and insulin receptor substrate (IRS)-1 were restored in the thoracic aorta and skeletal muscle, respectively.. These findings suggest that GGDGT attenuates endothelial dysfunction via improvement of the nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) signalling pathway and improves insulin sensitivity in diabetic atherosclerosis.

Topics: Animals; Aorta, Thoracic; Apolipoproteins E; Atherosclerosis; Blood Glucose; Cyclic GMP; Diabetic Angiopathies; Diet, Western; Endothelium, Vascular; Hypertension; Insulin; Insulin Receptor Substrate Proteins; Insulin Resistance; Lipids; Male; Medicine, Korean Traditional; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase Type III; Phytotherapy; Plant Extracts; Vasodilation

Angiotensin II participates to the regulation of cardiovascular physiology and it is involved in molecular mechanisms of insulin resistance. Angiotensin (1-7), derived from angiotensin II metabolism, is able to counteract many of the haemodynamic and non-haemodynamic actions of angiotensin II. In this study, we investigated in human umbilical vein endothelial cells (HUVECs) the possible action of angiotensin (1-7) on the insulin signalling pathway.. We stimulated HUVECs with insulin, angiotensin II and angiotensin (1-7), testing the effects on endothelial nitric oxide synthase (eNOS) enzyme activation and on insulin receptor substrate-1 (IRS1) phosphorylation. Moreover, we analysed the involvement of angiotensin type1, type2, and Mas receptors in these actions. Finally, we measured the nitric oxide (NO) production, the intracellular cGMP and the PKG-related activity in HUVECs, and the subsequent functional vasoactive effect of angiotensin (1-7) in mesenteric arteries of mice. Angiotensin II inhibits the insulin-induced Akt and eNOS phosphorylation, reducing the NO production. On the other hand, angiotensin (1-7) counteracts the inhibitory effect of angiotensin II, being able to restore the insulin-induced Akt/eNOS activation and the NO production. This effect is mediated by the Mas receptor. The inhibitory effects of angiotensin II on insulin signalling are, at least in part, mediated by an increased serine phosphorylation of IRS₁. Angiotensin (1-7) inhibits the serine phosphorylation of IRS1 induced by angiotensin II.. In endothelial cells angiotensin (1-7) counteracts the negative effects of angiotensin II on insulin signalling and NO production. The balance between angiotensin II and angiotensin (1-7) could represent a key mechanism in the pathophysiological processes leading to endothelial dysfunction and insulin-resistance.

Topics: Angiotensin I; Angiotensin II; Animals; Cells, Cultured; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Enzyme Activation; Human Umbilical Vein Endothelial Cells; Humans; Insulin; Insulin Receptor Substrate Proteins; Insulin Resistance; Mesenteric Arteries; Mice; Nitric Oxide; Nitric Oxide Synthase Type III; Peptide Fragments; Phosphatidylinositol 3-Kinase; Phosphorylation; Proto-Oncogene Mas; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Receptors, G-Protein-Coupled; Signal Transduction; Vasodilation

To elucidate the synergistic effects of taurine and L-arginine on hypertension, 25% fructose were administered to male Wistar rats for 3 months to establish insulin resistance hypertensive models. Rats with the systolic blood pressure (SBP) higher than 150 mmHg were considered as model rats. Forty-two model rats were randomly divided into six groups and administered with 3% taurine, 2.7% taurine + 0.3% L-arginine, 2.1% taurine + 0.9% L-arginine, 1.5% taurine + 1.5% L-arginine and 3% L-arginine in drinking water respectively. The results showed that coadministration of taurine (1.5%) and L-arginine (1.5%) could bring the levels of SBP, blood glucose, and insulin down to normal levels after 4 weeks. The thickness of blood vessels increased significantly in model group, which could be reversed by taurine and L-arginine. Serum NO, cGMP, and ET levels could return to normal levels. These data indicated that both taurine and L-arginine could ameliorate vascular remodeling and showed obvious antihypertensive effects, and taurine (1.5%) and L-arginine (1.5%) in the drinking water showed a better result in the cure of hypertension.

Topics: Animals; Antihypertensive Agents; Arginine; Blood Glucose; Blood Pressure; Cyclic GMP; Drug Synergism; Endothelins; Hypertension; Insulin; Insulin Resistance; Male; Nitric Oxide; Rats; Rats, Wistar; Systole; Taurine

Adiposity greatly increases the risk of atherothrombotic events, a pathological condition where a chronic state of oxidative stress is reported to play a major role. This study aimed to investigate the involvement of (NO)-soluble guanylyl cyclase (sGC) signaling pathway in the platelet dysfunction from high fat-fed (HFF) rats.. Male Wistar rats were fed for 10 weeks with standard chow (SCD) or high-fat diet (HFD). ADP (10 μM)- and thrombin (100 mU/ml)-induced washed platelet aggregation were evaluated. Measurement of intracellular levels of ROS levels was carried out using flow cytometry. Cyclic GMP levels were evaluated using ELISA kits.. High-fat fed rats exhibited significant increases in body weight, epididymal fat, fasting glucose levels and glucose intolerance compared with SCD group. Platelet aggregation induced by ADP (n = 8) and thrombin from HFD rats (n = 8) were significantly greater (P < 0.05) compared with SCD group. Platelet activation with ADP increased by 54% the intraplatelet ROS production in HFD group, as measured by flow cytometry (n = 6). N-acetylcysteine (NAC; 1 mM) and PEG-catalase (1000 U/ml) fully prevented the increased ROS production and platelet hyperaggregability in HFD group. The NO donors sodium nitroprusside (SNP; 10 μM) and SNAP (10 μM), as well as the NO-independent soluble guanylyl cyclase stimulator BAY 41-2272 (10 μM) inhibited the platelet aggregation in HFD group with lower efficacy (P < 0.05) compared with SCD group. The cGMP levels in response to these agents were also markedly lower in HFD group (P < 0.05). The prostacyclin analogue iloprost (1 μM) reduced platelet aggregation in HFD and SCD rats in a similar fashion (n = 4).. Metabolic abnormalities as consequence of HFD cause platelet hyperaggregability involving enhanced intraplatelet ROS production and decreased NO bioavailability that appear to be accompanied by potential defects in the prosthetic haem group of soluble guanylyl cyclase.

Topics: Adenosine Diphosphate; Animals; Antioxidants; Blood Platelets; Cyclic GMP; Diet, High-Fat; Enzyme Activation; Enzyme Activators; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Glucose Intolerance; Glucose Tolerance Test; Guanylate Cyclase; Insulin Resistance; Male; Nitric Oxide; Nitric Oxide Donors; Oxidative Stress; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Function Tests; Rats; Rats, Wistar; Reactive Oxygen Species; Receptors, Cytoplasmic and Nuclear; Signal Transduction; Soluble Guanylyl Cyclase; Thrombin; Time Factors; Weight Gain

Vascular smooth muscle cells (VSMCs) from the animal model of insulin resistance obese Zucker rats (OZR) show impaired ability of nitric oxide (NO) to increase cGMP and of cGMP to activate its specific kinase PKG, these defects being attributable to oxidative stress. We aimed to investigate the intracellular signalling downstream PKG in human and rat VSMC, and to clarify whether it is modified by insulin resistance and oxidative stress.. In aortic VSMC from humans, lean Zucker rats (LZR) and OZR, we measured by Western blots the activation induced by NO and cGMP of signalling molecules of PI3-K and MAPK pathways, with or without PKG inhibition, hydrogen peroxide and antioxidants. We explored the mechanism of the increased oxidative stress in VSMC from OZR by measuring superoxide anion concentrations (luminescence method) with or without inhibition of NADPH oxidase, xanthine oxidase, and mitochondrial electron transport chain complex and by measuring superoxide dismutase (SOD) expression (Western blot) and activity.. In VSMC from humans and LZR, the NO/cGMP/PKG pathway activates both PI3-K (Akt, mTOR) and MAPK (ERK-1/2, p38MAPK) signalling. This effect is attenuated in VSMC from OZR, in which the greater oxidative stress is mediated by NADPH oxidase and mitochondrial complex and by a reduced synthesis/activity of SOD. Impairment of the NO/cGMP/PKG signalling is reproduced in VSMC from LZR by hydrogen peroxide and reverted in VSMC from OZR by antioxidants.. In VSMC from an animal model of insulin resistance the NO/cGMP/PKG intracellular signalling is impaired due to an increased oxidative stress.

Topics: Analysis of Variance; Animals; Antioxidants; Blotting, Western; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Electron Transport Chain Complex Proteins; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Humans; Insulin Resistance; JNK Mitogen-Activated Protein Kinases; Male; Mitogen-Activated Protein Kinases; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; NADPH Oxidases; Nitric Oxide; Nitric Oxide Donors; Oxidants; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinase; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Rats; Rats, Zucker; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Superoxide Dismutase; Superoxides; Time Factors; Xanthine Oxidase

Hemodynamic nitrate tolerance has been shown to result in an insulin-resistant state. We studied whether nitrate tolerance induced by a 7-day continuous exposure to transdermal nitroglycerin influenced the meal-induced insulin sensitization phenomenon in rabbits.. Changes in insulin sensitivity in response to feeding in conscious rabbits were determined by rapid insulin sensitivity test, in both nitrate-tolerant and nitrate-intolerant animals. In a separate series of experiments with anesthetized rabbits with or without nitrate tolerance, the hyperinsulinemic euglycemic glucose clamping methods was used to study the effect of intraportal infusion of cholecystokinin (CCK) on whole-body insulin sensitivity.. Rabbits with normal feeding exhibited a 46 ± 6% increase in insulin sensitivity as compared with their matching fasting controls. A 7-day period of treatment with patches releasing 0.07 mg of nitroglycerin per hour yielded nitrate tolerance and a state of insulin resistance and no increase in insulin sensitivity in response to food. Intraportal infusion of CCK8 (0.3-3.0 μg/kg over 20 minutes) resulted in a dose-dependent increase in insulin sensitivity in normal but not in nitrate-tolerant, fasted anesthetized animals.. Nitrate tolerance blocks both the meal-induced insulin sensitization phenomenon and the insulin-sensitizing effect of intraportal CCK.

Topics: Animals; Blood Glucose; Blood Pressure; Cyclic GMP; Drug Tolerance; Eating; Fasting; Glucose Clamp Technique; Heart Rate; Insulin Resistance; Liver; Male; Nitroglycerin; Rabbits; Sincalide

Obesity is characterized by chronic inflammation of adipose tissue, which contributes to insulin resistance and diabetes. Although nitric oxide (NO) signaling has antiinflammatory effects in the vasculature, whether reduced NO contributes to adipose tissue inflammation is unknown. We sought to determine whether (1) obesity induced by high-fat (HF) diet reduces endothelial nitric oxide signaling in adipose tissue, (2) reduced endothelial nitric oxide synthase (eNOS) signaling is sufficient to induce adipose tissue inflammation independent of diet, and (3) increased cGMP signaling can block adipose tissue inflammation induced by HF feeding.. Relative to mice fed a low-fat diet, an HF diet markedly reduced phospho-eNOS and phospho-vasodilator-stimulated phosphoprotein (phospho-VASP), markers of vascular NO signaling. Expression of proinflammatory cytokines was increased in adipose tissue of eNOS-/- mice. Conversely, enhancement of signaling downstream of NO by phosphodiesterase-5 inhibition using sildenafil attenuated HF-induced proinflammatory cytokine expression and the recruitment of macrophages into adipose tissue. Finally, we implicate a role for VASP, a downstream mediator of NO-cGMP signaling in mediating eNOS-induced antiinflammatory effects because VASP-/- mice recapitulated the proinflammatory phenotype displayed by eNOS-/- mice.. These results imply a physiological role for endothelial NO to limit obesity-associated inflammation in adipose tissue and hence identify the NO-cGMP-VASP pathway as a potential therapeutic target in the treatment of diabetes.

Topics: Adipose Tissue; Animals; Cell Adhesion Molecules; Cyclic GMP; Dietary Fats; Disease Models, Animal; Endothelium, Vascular; Inflammation; Insulin Resistance; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Microfilament Proteins; Nitric Oxide; Nitric Oxide Synthase Type III; Obesity; Phosphodiesterase 5 Inhibitors; Phosphoproteins; Phosphorylation; Piperazines; Purines; Signal Transduction; Sildenafil Citrate; Sulfones

Proinflammatory activation of Kupffer cells is implicated in the effect of high-fat feeding to cause liver insulin resistance. We sought to determine whether reduced endothelial nitric oxide (NO) signaling contributes to the effect of high-fat feeding to increase hepatic inflammatory signaling and if so, whether this effect 1) involves activation of Kupffer cells and 2) is ameliorated by increased NO signaling.. Effect of NO/cGMP signaling on hepatic inflammation and on isolated Kupffer cells was examined in C57BL/6 mice, eNos(-/-) mice, and Vasp(-/-) mice fed a low-fat or high-fat diet.. We show that high-fat feeding induces proinflammatory activation of Kupffer cells in wild-type mice coincident with reduced liver endothelial nitric oxide synthase activity and NO content while, conversely, enhancement of signaling downstream of endogenous NO by phosphodiesterase-5 inhibition protects against high fat-induced inflammation in Kupffer cells. Furthermore, proinflammatory activation of Kupffer cells is evident in eNos(-/-) mice even on a low-fat diet. Targeted deletion of vasodilator-stimulated phosphoprotein (VASP), a key downstream target of endothelially derived NO, similarly predisposes to hepatic and Kupffer cell inflammation and abrogates the protective effect of NO signaling in both macrophages and hepatocytes studied in a cell culture model.. These results collectively imply a physiological role for endothelial NO to limit obesity-associated inflammation and insulin resistance in hepatocytes and support a model in which Kupffer cell activation during high-fat feeding is dependent on reduced NO signaling. Our findings also identify the NO/VASP pathway as a novel potential target for the treatment of obesity-associated liver insulin resistance.

Topics: Animals; Cell Adhesion Molecules; Cells, Cultured; Cyclic GMP; Cytokines; Dietary Fats; Endothelial Cells; Gene Expression Regulation; Hepatitis; Insulin Resistance; Kupffer Cells; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Microfilament Proteins; Molecular Targeted Therapy; Nitric Oxide; Nitric Oxide Synthase Type III; Obesity; Phosphodiesterase 5 Inhibitors; Phosphoproteins; Signal Transduction

Diet-induced obesity (DIO) in mice causes vascular inflammation and insulin resistance that are accompanied by decreased endothelial-derived NO production. We sought to determine whether reduced NO-cGMP signaling contributes to the deleterious effects of DIO on the vasculature and, if so, whether these effects can be blocked by increased vascular NO-cGMP signaling.. By using an established endothelial cell culture model of insulin resistance, exposure to palmitate, 100 micromol/L, for 3 hours induced both cellular inflammation (activation of IKK beta-nuclear factor-kappaB) and impaired insulin signaling via the insulin receptor substrate-phosphatidylinositol 3-kinase pathway. Sensitivity to palmitate-induced endothelial inflammation and insulin resistance was increased when NO signaling was reduced using an endothelial NO synthase inhibitor, whereas endothelial responses to palmitate were blocked by pretreatment with either an NO donor or a cGMP analogue. To investigate whether endogenous NO-cGMP signaling protects against vascular responses to nutrient excess in vivo, adult male mice lacking endothelial NO synthase were studied. As predicted, both vascular inflammation (phosphorylated I kappaB alpha and intercellular adhesion molecule levels) and insulin resistance (phosphorylated Akt [pAkt] and phosphorylated eNOS [peNOS] levels) were increased in endothelial NO synthase(-/-) (eNOS(-/-)) mice, reminiscent of the effect of DIO in wild-type controls. Next, we asked whether the vascular response to DIO in wild-type mice can be reversed by a pharmacological increase of cGMP signaling. C57BL6 mice were either fed a high-fat diet or remained on a low-fat diet for 8 weeks. During the final 2 weeks of the study, mice on each diet received either placebo or the phosphodiesterase-5 inhibitor sildenafil, 10 mg/kg per day orally. In high-fat diet-fed mice, vascular inflammation and insulin resistance were completely prevented by sildenafil administration at a dose that had no effect in mice fed the low-fat diet.. Reduced signaling via the NO-cGMP pathway is a mediator of vascular inflammation and insulin resistance during overnutrition induced by high-fat feeding. Therefore, phosphodiesterase-5, soluble guanylyl cyclase, and other molecules in the NO-cGMP pathway (eg, protein kinase G) constitute potential targets for the treatment of vascular dysfunction in the setting of obesity.

Topics: Animals; Aorta, Thoracic; Aortic Diseases; Cell Adhesion Molecules; Cells, Cultured; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Dietary Fats; Disease Models, Animal; Down-Regulation; Endothelial Cells; Enzyme Inhibitors; Humans; I-kappa B Kinase; Inflammation; Inflammation Mediators; Insulin; Insulin Receptor Substrate Proteins; Insulin Resistance; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; NF-kappa B; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase Type III; Palmitic Acid; Phosphatidylinositol 3-Kinases; Phosphodiesterase 5 Inhibitors; Phosphodiesterase Inhibitors; Phosphorylation; Piperazines; Proto-Oncogene Proteins c-akt; Purines; Signal Transduction; Sildenafil Citrate; Sulfones

Nitric oxide (NO) is a short-lived gaseous messenger with multiple physiological functions including regulation of blood flow, platelet adhesion and aggregation inhibition. NO synthases (NOS) catalyze the conversion of cationic amino acid L-arginine in L-citrulline and NO. Despite an increasing prevalence of obesity and metabolic syndrome (MetS) in the last decades, the exact mechanisms involved in the pathogenesis and cardiovascular complications are not fully understood. We have examined the effects of obesity and MetS on the L-arginine-NO-cGMP pathway in platelets from a population of adolescents.. A total of twenty six adolescent patients (13 with obesity and 13 with MetS) and healthy volunteers (n=14) participated in this study. Transport of L-arginine, NO synthase (NOS) activity and cGMP content in platelets were analyzed. Moreover, platelet function, plasma levels of L-arginine, metabolic and clinical markers were investigated in these patients and controls.. L-arginine transport (pmol/10(9) cells/min) in platelets via system y(+)L was diminished in obese subjects (20.8±4.7, n=10) and MetS patients (18.4±3.8, n=10) compared to controls (52.3±14.8, n=10). The y(+)L transport system correlated negatively to insulin levels and Homeostasis Model Assessment of Insulin Resistance (HOMA IR) index. No differences in NOS activity and cGMP content were found among the groups. Moreover, plasma levels of L-arginine were not affected by obesity or MetS.. Our study provides the first evidence that obesity and MetS lead to a dysfunction of L-arginine influx, which negatively correlates to insulin resistance. These findings could be a premature marker of future cardiovascular complications during adulthood.

Topics: Adolescent; Arginine; Biological Transport; Blood Platelets; Cardiovascular Diseases; Case-Control Studies; Cyclic GMP; Humans; Insulin Resistance; Metabolic Syndrome; Nitric Oxide; Obesity

Emerging evidence indicates that aldosterone causes oxidative stress by stimulating proinflammatory/oxidative mediators, including nuclear factor-kappaB, activating protein (AP-1), and c-Jun N-terminal kinase. Thus, in insulin-resistant type 2 diabetes (T2D), oxidative stress generated by hyperglycemia and aldosterone would potentiate the oxidative destruction of tissue and important regulators of glucose metabolism like adiponectin and insulin. Although heme oxygenase (HO)-1 is cytoprotective, its effects on T2D have not been fully characterized. Here we report an enduring antidiabetic effect of the HO inducer, hemin, on Zucker diabetic-fatty rat (ZDF), a model of insulin-resistant T2D. Chronically applied hemin to ZDF reduced and maintained significantly low fasting and postprandial hyperglycemia for 4 months after therapy. The antidiabetic effect was accompanied by enhanced HO activity, catalase, cyclic GMP, bilirubin, ferritin, total antioxidant capacity, and insulin. In contrast, reduced aldosterone alongside markers/mediators of oxidative stress, including 8-isoprostane, c-Jun N-terminal kinase, nuclear factor-kappaB, AP-1, and AP-2 were observed. Interestingly, in hemin-treated ZDF, inhibitory proteins of insulin-signaling, such as glycogen synthase kinase-3 and protein-tyrosine phosphatase-1B were reduced, whereas agents that promote insulin signaling including adiponectin, cAMP, AMP-activated protein kinase, aldolase-B, and glucose transporter-4 (GLUT4), were robustly increased. Correspondingly, hemin improved ip glucose tolerance, reduced insulin intolerance, and lowered insulin resistance (homeostasis model assessment of insulin resistance), and the inability of insulin to enhance GLUT4 was overturned. These results suggest that the suppression of hyperglycemia and aldosterone-induced oxidative stress alongside the potentiation of insulin-sensitizing pathways may account for the 4-month enduring antidiabetic effect. The synergistic interaction between the HO system, aldolase-B, adiponectin, AMP-activated protein kinase, and GLUT4 may be explored for novel strategies against postprandial/fasting hyperglycemia and insulin-resistant T2D.

Topics: Aldosterone; Animals; Cyclic GMP; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 2; Drug Evaluation, Preclinical; Fasting; Heme Oxygenase (Decyclizing); Hemin; Hyperglycemia; Insulin Resistance; Male; Muscle, Skeletal; Obesity; Oxidative Stress; Rats; Rats, Sprague-Dawley; Rats, Zucker; Signal Transduction

Hyperglycemia-induced oxidative stress is a common phenomenon in diabetes. Since oxidative stress depletes adiponectin and insulin levels, we investigated whether an upregulated heme oxygenase (HO) system would attenuate the oxidative destruction of adiponectin/insulin and improve insulin sensitivity and glucose metabolism in streptozotocin (STZ)-induced type 1 diabetes. HO was upregulated with hemin (15 mg/kg ip) or inhibited with chromium mesoporphyrin (CrMP, 4 micromol/kg ip). Administering hemin to STZ-diabetic rats reduced hyperglycemia and improved glucose metabolism, whereas the HO inhibitor CrMP annulled the antidiabetic effects and/or exacerbated fasting/postprandial hyperglycemia. Interestingly, the antidiabetic effects of hemin lasted for 2 mo after termination of therapy and were accompanied by enhanced HO-1 and HO activity of the soleus muscle, along with potentiation of plasma antioxidants like bilirubin, ferritin, and superoxide dismutase, with corresponding elevation of the total antioxidant capacity. Importantly, hemin abated c-Jun NH2-terminal kinase (JNK), a substance known to inhibit insulin biosynthesis, and suppressed markers/mediators of oxidative stress including 8-isoprostane, nuclear-factor (NF)-kappaB, activating protein (AP)-1, and AP-2 of the soleus muscle. Furthermore, hemin therapy significantly attenuated pancreatic histopathological lesions including acinar cell necrosis, interstitial edema, vacuolization, fibrosis, and mononuclear cell infiltration. Correspondingly, hemin increased plasma insulin and potentiated agents implicated in insulin sensitization and insulin signaling such as adiponectin, adenosine monophosphate-activated protein kinase (AMPK), cAMP, cGMP, and glucose transporter (GLUT)4, a protein required for glucose uptake. These were accompanied by improved glucose tolerance [intraperitoneal glucose tolerance text (IPGTT)], decreased insulin intolerance [intraperitoneal insulin tolerance test (IPITT)], and reduced insulin resistance [homeostasis model assessment of insulin resistance (HOMA-IR) index], whereas CrMP nullified the hemin-dependent antidiabetic and insulin-sensitizing effects. In conclusion, by concomitantly enhancing insulin and paradoxically potentiating insulin sensitivity, this study unveils a novel, unique, and long-lasting antidiabetic characteristic of upregulating HO with hemin that could be exploited against insulin-resistant and insulin-dependent diabetes.

Topics: Animals; Cyclic GMP; Diabetes Mellitus, Experimental; Drug Evaluation, Preclinical; Fasting; Glucose; Heme Oxygenase (Decyclizing); Hemin; Hyperglycemia; Hypoglycemic Agents; Insulin Resistance; JNK Mitogen-Activated Protein Kinases; Male; Muscle, Skeletal; NF-kappa B; Rats; Rats, Sprague-Dawley; Streptozocin; Transcription Factor AP-1

In type 2 diabetes (T2D), postprandial and fasting hyperglycemia are important predictors of cardiovascular diseases; however, few drugs are currently available to simultaneously suppress these conditions. Here, we report an enduring antidiabetic effect of the heme oxygenase (HO) inducer hemin on Goto-Kakizaki rats (GK), a nonobese insulin-resistant T2D model. HO breaks down the heme-moiety-generating antioxidants (biliverdin/bilirubin and ferritin) and carbon monoxide, which stimulate insulin secretion. Hemin induces HO-1 to potentiate HO activity and the HO-derived products. Chronically applied hemin (30 mg/kg ip) for a month reduced and maintained fasting glucose at physiological levels for 3 mo. Before therapy, glucose levels were 9.3 +/- 0.3 mmol/l (n = 14). At 1, 2, and 3 mo posttherapy, we recorded 6.7 +/- 0.13, 5.9 +/- 0.2, and 7.2 +/- 0.2 mmol/l, respectively. Hemin was also effective against postprandial hyperglycemia (14.6 +/- 1.1 vs. 7.5 +/- 0.4 mmol/l; n = 14; P < 0.01), and the effect remained sustained for 3 mo after therapy. The reduction of hyperglycemia was accompanied by enhanced HO-1, HO activity, and cGMP of the soleus muscle, alongside increased plasma bilirubin, ferritin, SOD, total antioxidant capacity, and insulin levels, whereas markers/mediators of oxidative stress like urinary-8-isoprostane and soleus muscle nitrotyrosine, NF-kappaB, and activator protein-1 and -2 were abated. Furthermore, inhibitors of insulin signaling including soleus muscle glycogen synthase kinase-3 and JNK were reduced, while the insulin-sensitizing adipokine, adiponectin, alongside AMPK were increased. Correspondingly, hemin improved glucose tolerance, suppressed insulin intolerance, reduced insulin resistance, and overturned the inability of insulin to enhance glucose transporter 4, a protein required for glucose uptake. Hemin also upregulated HO-1/HO activity and cGMP and lowered glucose in euglycemic Sprague-Dawley control rats albeit less intensely, suggesting greater selectivity of the HO system in diabetic conditions. In conclusion, reduced oxidative stress alongside the concomitant and paradoxical enhancement of insulin secretion and insulin-sensitizing pathways may account for the 3-mo-enduring antidiabetic effect. The synergistic interaction among HO, adiponectin, and GLUT4 may be explored against insulin-resistant diabetes.

Topics: Adiponectin; Animals; Bilirubin; Blood Glucose; Cyclic GMP; Diabetes Mellitus, Type 2; Fasting; Ferritins; Glucose Transporter Type 4; Glycogen Synthase Kinase 3; Heme Oxygenase (Decyclizing); Hemin; Hyperglycemia; Insulin; Insulin Resistance; Male; Muscle, Skeletal; Oxidative Stress; Postprandial Period; Rats; Rats, Sprague-Dawley; Rats, Wistar; Up-Regulation

Insulin-mediated signal transduction is positively correlated to adiponectin, adenosine monophosphate-activated protein kinase (AMPK), and glucose-transporter-4 (GLUT4) but negatively to oxidative/inflammatory mediators such as nuclear factor-kappaB, activating-protein (AP)-1, AP-2, and c-Jun-N-terminal-kinase. Although hemeoxygenase (HO) suppresses oxidative insults, its effects on insulin-sensitizing agents like AMPK and GLUT4 remains unclear and were investigated using Goto-Kakizaki rats (GK), a nonobese insulin-resistant type-2 diabetic model. HO was induced with hemin or inhibited with chromium mesoporphyrin (CrMP). The application of hemin to GK rats evoked a 3-month antidiabetic effect, whereas the HO-inhibitor, CrMP, exacerbated hyperglycemia and nullified insulin-signaling/glucose metabolism. Interestingly, the antidiabetic was accompanied by a paradoxical increase of insulin alongside the potentiation of insulin-sensitizing agents such as adiponectin, AMPK, and GLUT4 in the gastrocnemius muscle. Furthermore, hemin enhanced mediators/regulators of insulin signaling like cGMP and cAMP and suppressed oxidative insults by up-regulating HO-1, HO activity, superoxide dismutase, catalase, and the total antioxidant capacity in the gastrocnemius muscle. Accordingly, oxidative markers/mediators including nuclear factor-kappaB, AP-1, AP-2, c-Jun-N-terminal-kinase, and 8-isoprostane were abated, whereas CrMP annulled the cytoprotective and antidiabetic effects of hemin. Correspondingly, ip glucose tolerance, insulin tolerance, and homeostasis model assessment insulin resistance analyses revealed improved glucose tolerance, reduced insulin intolerance, enhanced insulin sensitivity, and reduced insulin resistance in hemin-treated GK rats. In contrast, CrMP, abolished the insulin-sensitizing effects and restored and/or exacerbated insulin resistance. Our study unveils a 3-month enduring antidiabetic effect of hemin and unmasks the synergistic interaction among the HO system, adiponectin, AMPK, and GLUT4 that could be explored to enhance insulin signaling and improve glucose metabolism in insulin-resistant diabetes.

Topics: Adiponectin; AMP-Activated Protein Kinase Kinases; Animals; Cyclic AMP; Cyclic GMP; Diabetes Mellitus, Type 2; Disease Models, Animal; Glucose; Glucose Transporter Type 4; Heme Oxygenase-1; Hemin; Insulin; Insulin Resistance; Male; Mesoporphyrins; Muscle, Skeletal; NF-kappa B; Protein Kinases; Rats; Rats, Inbred Strains; Rats, Sprague-Dawley; Rats, Wistar; Transcription Factor AP-1; Up-Regulation

Obesity and diabetes are major risk factors for the development of vascular disease in the lower limbs. Previous studies have demonstrated reduced nitric oxide (NO)-mediated vasodilation, increased adrenergic constriction, and inward, atrophic remodeling in the limb circulation of obese Zucker rats, but the component of the "metabolic syndrome" driving these changes is unclear. Because insulin resistance precedes the state of frank diabetes, the current study hypothesized that insulin resistance independent of obesity induced by fructose feeding would impair microvascular function in the skeletal muscle circulation in lean Zucker rats (LZR). A 66% fructose diet impaired glucose tolerance and induced moderate insulin resistance with no changes in whole-body hemodynamics of anesthetized rats (FF-LZR), compared to control LZR. NO-mediated vasodilation of isolated gracilis arteries, assessed in vitro with acetylcholine and sodium nitroprusside, was reduced approximately 20% in FF-LZR vs. LZR. NO-independent cGMP-mediated vasodilation was unimpaired. Pretreatment of isolated vessels with the superoxide scavenger, tempol, improved responses to both vasodilators. Reactivity to adrenergic stimulation was unaltered in FF-LZR vs. LZR, although constriction to endothelin was increased. Structural and passive mechanical characteristics of isolated gracilis arteries were similar in both LZR and FF-LZR. Taken together, these findings indicate that moderate insulin resistance is sufficient to impair endothelial function in an oxidant-dependent manner in the rat hindlimb circulation. Other aspects of skeletal muscle vascular function documented in obese models, specifically adrenergic tone and inward remodeling, must reflect either severe insulin resistance or other aspects of obesity. The factors accounting for nonendothelial vasculopathies remain unknown.

Topics: Acetylcholine; Animals; Arteries; Cyclic GMP; Diet; Endothelium, Vascular; Fructose; Hindlimb; Insulin Resistance; Male; Muscle, Skeletal; Nitric Oxide; Nitroprusside; Obesity; Oxidants; Rats; Rats, Zucker; Sweetening Agents; Vasoconstriction; Vasodilation; Vasodilator Agents

Natriuretic peptides (NPs) have been characterized as vascular hormones that regulate vascular tone via guanylyl cyclase (GC), cyclic GMP (cGMP), and cGMP-dependent protein kinase (cGK). Recent clinical studies have shown that plasma NP levels were lower in subjects with the metabolic syndrome. The present study was conducted to elucidate the roles for NP/cGK cascades in energy metabolism.. We used three types of genetically engineered mice: brain NP (BNP) transgenic (BNP-Tg), cGK-Tg, and guanylyl cyclase-A (GCA) heterozygous knockout (GCA(+/-)) mice and analyzed the metabolic consequences of chronic activation of NP/cGK cascades in vivo. We also examined the effect of NPs in cultured myocytes.. BNP-Tg mice fed on high-fat diet were protected against diet-induced obesity and insulin resistance, and cGK-Tg mice had reduced body weight even on standard diet; surprisingly, giant mitochondria were densely packed in the skeletal muscle. Both mice showed an increase in muscle mitochondrial content and fat oxidation through upregulation of peroxisome proliferator-activated receptor (PPAR)-gamma coactivator (PGC)-1alpha and PPARdelta. The functional NP receptors, GCA and guanylyl cyclase-B, were downregulated by feeding a high-fat diet, while GCA(+/-) mice showed increases in body weight and glucose intolerance when fed a high-fat diet. NPs directly increased the expression of PGC-1alpha and PPARdelta and mitochondrial content in cultured myocytes.. The findings together suggest that NP/cGK cascades can promote muscle mitochondrial biogenesis and fat oxidation, as to prevent obesity and glucose intolerance. The vascular hormone, NP, would contribute to coordinated regulation of oxygen supply and consumption.

Topics: Animals; Blood Glucose; Cells, Cultured; Cyclic GMP; Dietary Fats; Down-Regulation; Genetic Engineering; Glucose Intolerance; Insulin Resistance; Lipid Peroxidation; Mice; Mice, Knockout; Mitochondria; Molecular Sequence Data; Muscle Cells; Muscle, Skeletal; Natriuretic Peptide, Brain; Natriuretic Peptides; Obesity; Oxygen Consumption; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; PPAR delta; PPAR gamma; Protein Kinases; Receptors, Atrial Natriuretic Factor; Trans-Activators; Transcription Factors; Up-Regulation

Some in vivo and ex vivo studies demonstrated a resistance to the vasodilating effects of nitric oxide (NO) in insulin-resistant states and, in particular, obese Zucker rats (OZR). To evaluate the biochemical basis of this phenomenon, we aimed to identify defects of the NO/cGMP/cGMP-dependent protein kinase (PKG) pathway in cultured vascular smooth muscle cells (VSMCs) from OZR and lean Zucker rats (LZR) by measuring: 1) NO donor ability to increase cGMP in the absence and presence of inhibitors of soluble guanylate cyclase (sGC) and phosphodiesterases (PDEs); 2) NO and cGMP ability to induce, via PKG, vasodilator-stimulated phosphoprotein (VASP) phosphorylation at serine 239 and PDE5 activity; 3) protein expression of sGC, PKG, total VASP, and PDE5; 4) superoxide anion concentrations and ability of antioxidants (superoxide dismutase+catalase and amifostine) to influence the NO/cGMP/PKG pathway activation; and 5) hydrogen peroxide influence on PDE5 activity and VASP phosphorylation. VSMCs from OZR vs. LZR showed: 1) baseline cGMP concentrations higher, at least in part owing to reduced catabolism by PDEs; 2) impairment of NO donor ability to increase cGMP, even in the presence of PDE inhibitors, suggesting a defect in the NO-induced sGC activation; 3) reduction of NO and cGMP ability to activate PKG, indicated by the impaired ability to phosphorylate VASP at serine 239 and to increase PDE5 activity via PKG; 4) similar baseline protein expression of sGC, PKG, total VASP, and PDE5; and 5) higher levels of superoxide anion. Antioxidants partially prevented the defects of the NO/cGMP/PKG pathway observed in VSMCs from OZR, which were reproduced by hydrogen peroxide in VSMCs from LZR, suggesting the pivotal role of oxidative stress.

Topics: Animals; Cell Adhesion Molecules; Cells, Cultured; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Insulin Resistance; Male; Microfilament Proteins; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Nitric Oxide; Oxidative Stress; Phosphodiesterase Inhibitors; Phosphoproteins; Phosphorylation; Rats; Rats, Zucker; Signal Transduction

Mitochondrial dysfunction in the skeletal muscle has been implicated in a wide variety of pathological processes including insulin resistance in type 2 diabetes. A recent report indicates that calorie restriction can modulate mitochondrial function through the nitric oxide/cGMP-dependent pathway. Following up on these findings, we examined whether cGMP could rescue mitochondrial dysfunction in C2C12 myotubular cells induced by conditions of high-glucose and high-insulin. Treatment of the cells with cGMP promoted mitochondrial biogenesis and ATP synthesis without enhancing production of reactive oxygen species (ROS) in association with up-regulation of the genes involved in oxidative phosphorylation and ROS reduction. The increased mitochondria were revealed to have lower membrane potential, which is similar to the effect of calorie restriction, and reversed mitochondrial dysfunction caused by high-glucose and high-insulin. These results indicated that augmented cGMP-dependent cascades in the skeletal muscle may attenuate insulin resistance observed in patients with type 2 diabetes and metabolic syndrome.

Topics: Animals; Cell Line; Cells, Cultured; Cyclic GMP; Dose-Response Relationship, Drug; Glucose; Insulin; Insulin Resistance; Mice; Mitochondria, Muscle; Muscle Fibers, Skeletal; Reactive Oxygen Species

In the endothelium, insulin promotes nitric oxide (NO) production, through the insulin receptor/IRS-1/PI3-Kinase/Akt/eNOS signaling pathway. An inhibitor of insulin action, TRIB3, has recently been identified which affects insulin action by binding to and inhibiting Akt phosphorylation. We have recently described a Q84R gain-of-function polymorphism of TRIB3 with the R84 variant being associated with insulin resistance and an earlier age at myocardial infarction.. To investigate the TRIB3 R84 variant impact on endothelial insulin action, we cultured human umbilical vein endothelial cells (HUVECs) naturally carrying different TRIB3 genotypes (QQ-, QR-, or RR-HUVECs). TRIB3 inhibitory activity on insulin-stimulated Akt phosphorylation and the amount of protein which was coimmunoprecipitable with Akt were significantly greater in QR- and RR- as compared to QQ- HUVECs. After insulin stimulation, Akt and eNOS activation as well as NO production were markedly decreased in QR- and RR- as compared to QQ-HUVECs. TRIB3 molecular modeling analysis provided insights into the structural changes related to the polymorphisms potentially determining differences in protein-protein interaction with Akt.. Our data demonstrate that the TRIB3 R84 variant impairs insulin signaling and NO production in human endothelial cells. This finding provides a plausible biological background for the deleterious role of TRIB3 R84 on genetic susceptibility to coronary artery disease.

Topics: Adaptor Proteins, Signal Transducing; Binding Sites; Cardiovascular Diseases; Cell Cycle Proteins; Cells, Cultured; Cyclic GMP; Endothelial Cells; Enzyme Activation; Genotype; Glycogen Synthase Kinase 3; Humans; Insulin; Insulin Receptor Substrate Proteins; Insulin Resistance; Models, Molecular; Mutation; Nitric Oxide; Nitric Oxide Synthase Type III; Oncogene Protein v-akt; Phenotype; Phosphatidylinositol 3-Kinases; Phosphorylation; Protein Binding; Protein Conformation; Protein Serine-Threonine Kinases; Receptor, Insulin; Repressor Proteins; Signal Transduction

Angiotensin converting enzyme (ACE) inhibitors usually cause severe coughing and intolerance while antagonists for angiotensin AT(1) receptor do not stimulate the production of nitric oxide (NO). NO has been shown to regulate arterial hypertension and insulin resistance. Hence, new hybrids of antagonist for angiotensin AT(1) receptor and a NO donor may have potent anti-hypertensive effect and regulate glucose metabolism and insulin resistance. Herein, the effects of [6-(nitrooxymethyl)pyridin-2-yl] methyl 4'-[1-(1,7'-dimethyl-2'-propyl-1H,3'H-2,5'-bibenzo[d]imidazol-3'-yl)ethyl] biphenyl-2-carboxylate (WB1106), a novel NO-releasing derivative of telmisartan newly synthesized, on the vasocontraction, hypertension and diet-induced insulin resistance were examined in vitro using rat aortic strips and in normotensive and spontaneous hypertension rats (SHR rats). Apparently, WB1106 induced the vasorelaxation of contracted rat aortic strips in a dose- and time-dependent manner, which depended on the activity of guanylate cyclase, a characteristic of NO-related function. Furthermore, WB1106 reduced the contractile and blood pressure responses to angiotensin II, which relied on the release of telmisartan. Moreover, treatment with WB1106 significantly reduced the blood pressure with similar potency to telmitarsan and increased the contents of cGMP in SHR rats. Therefore, WB1106 possesses both the angiotensin AT(1) receptor antagonist activity of telmisartan and the NO-releasing property of a 'slow NO donor'. Importantly, in contrast to equimolar telmisartan, treatment with WB1106 significantly attenuated body weight gains and improved glucose tolerance in high-fat and carbohydrate-fed rats, reflecting a synergistic effect of NO and telmisartan. Potentially, WB1106 may be a potent anti-hypertensive drug for treatment of hypertension and diabetes-related cardiovascular diseases in the clinic.

Topics: Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Animals; Antihypertensive Agents; Aorta, Thoracic; Benzimidazoles; Benzoates; Blood Pressure; Cyclic GMP; Diet; Dietary Carbohydrates; Dietary Fats; Glucose; Insulin Resistance; Male; Muscle Contraction; Muscle Relaxation; Nitric Oxide; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Rats, Sprague-Dawley; Telmisartan; Vasoconstriction; Vasoconstrictor Agents

Although the role of nitric oxide (NO) in peripheral glucose uptake has been thoroughly described, little is known regarding the alterations in NO metabolism during the early onset of insulin resistance. During this study we investigated the alterations in NO synthesis and bioavailability in a model for dietary modulations of insulin sensitivity. For 6 weeks, rats were fed a standard diet (C), a high-sucrose diet inducing insulin resistance (HS), or high-sucrose diets supplemented with cysteine, which endowed protection against the high-sucrose-induced insulin resistance (Ti). Several markers of NO synthesis and bioavailability were assessed and confronted with markers of insulin sensitivity. After 5 weeks, although urinary cGMP excretion did not differ between the groups, insulin resistance in HS rats was associated with both a significant increase in NO oxidation, as determined by plasma nitrotyrosine concentrations, and in the inducible NO synthase (iNOS)/endothelial NO synthase (iNOS/eNOS) mRNA ratio in skeletal muscle compared with C rats. These alterations were prevented in rats fed the cysteine-rich diets. NO production, as assessed by urinary 15NO3* excretion following a [15N2-(guanido)]-arginine intra-venous bolus, independently and significantly correlated with insulin sensitivity but did not significantly differ between C, HS, and Ti rats; neither did the aortic eNOS protein expression or skeletal muscle insulin-induced eNOS activation. Our results indicate that in this model of dietary modulations of insulin sensitivity (i) NO production accounts for part of total inter-individual variation in insulin sensitivity, but (ii) early diet-related changes in insulin sensitivity are accompanied by changes in NO bioavailability.

Topics: Animals; Aorta; Biomarkers; Cyclic GMP; Cysteine; Diet; Diet Therapy; Disease Models, Animal; Enzyme Activation; Gene Expression Regulation, Enzymologic; Insulin Resistance; Male; Muscle, Skeletal; Muscle, Smooth, Vascular; Nitrates; Nitric Oxide; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Rats; Rats, Wistar; Sucrose; Sweetening Agents; Tyrosine

Activation of protein kinase C (PKC) in vascular tissue is associated with endothelial dysfunction and insulin resistance. However, the effect of vascular PKC activation on insulin-stimulated endothelial nitric oxide (NO) synthase (eNOS) regulation has not been characterized in obesity-associated insulin resistance. Diacylglycerol (DAG) concentration and PKC activity were increased in the aorta of Zucker fatty compared with Zucker lean rats. Insulin-stimulated increases in Akt phosphorylation and cGMP concentration (a measure of NO bioavailability) after euglycemic-hyperinsulinemic clamp were blunted in the aorta of fatty compared with lean rats but were partly normalized after 2 weeks of treatment with the PKCbeta inhibitor ruboxistaurin (LY333531). In endothelial cell culture, overexpression of PKCbeta1 and -beta2, but not PKCalpha, -delta, or -zeta, decreased insulin-stimulated Akt phosphorylation and eNOS expression. Overexpression of PKCbeta1 and -beta2, but not PKCalpha or -delta, also decreased Akt phosphorylation stimulated by vascular endothelial growth factor (VEGF). In microvessels isolated from transgenic mice overexpressing PKCbeta2 only in vascular cells, Akt phosphorylation stimulated by insulin was decreased compared with wild-type mice. Thus, activation of PKCbeta in endothelial cells and vascular tissue inhibits Akt activation by insulin and VEGF, inhibits Akt-dependent eNOS regulation by insulin, and causes endothelial dysfunction in obesity-associated insulin resistance.

Topics: Animals; Atherosclerosis; Blood Vessels; Cattle; Cells, Cultured; Cyclic GMP; Diglycerides; Enzyme Activation; Insulin Resistance; Male; Mice; Mice, Transgenic; Nitric Oxide Synthase Type III; Obesity; Phosphorylation; Protein Kinase C; Protein Kinase C beta; Proto-Oncogene Proteins c-akt; Rats; Rats, Zucker; Vascular Endothelial Growth Factor A

The endogenous insulin sensitizing machinery termed the hepatic insulin sensitizing substance (HISS) mechanism has been shown to be nitrergic and linked to sensory fibers in the anterior hepatic plexus. We studied whether this mechanism could pharmacologically be exploited by cicletanine, a cGMP-PDE inhibitor antihypertensive drug, in conscious rabbits. Whole body insulin sensitivity and peripheral glucose uptake were determined by hyperinsulinaemic euglycaemic glucose clamping, and cardiac radiolabelled deoxyglucose (DOG) uptake in neurogenic, achieved by perineurial capsaicin treatment of the anterior hepatic plexus through defunctionalization of hepatic sensory fibers, and metabolic, induced by dietary hypercholesterolemia, insulin resistance models after single oral doses of cicletanine (3, 10 and 30 mg kg(-1)) or rosiglitazone (3 mg kg(-1)). The effect of cicletanine on cardiac and vascular tissue NO, cGMP, cAMP was measured by means of spin trapping technique and radioimmunoassay, respectively. Insulin sensitivity and peripheral DOG uptake were significantly increased by 10 and 30 mg kg(-1) cicletanine in both healthy and hypercholesterolaemic rabbits, but not in those with neurogenic insulin resistance. Rosiglitazone had no effect in healthy and neurogenic insulin resistant rabbits although it improved insulin sensitivity in hypercholesterolemic animals. The 10 mg kg(-1) cicletanine dose induced no change in either cardiac or vascular tissue NO, cGMP or cAMP concentrations. Nevertheless, at a dose of 30 mg kg(-1) producing an insulin sensitizing effect of approximately the same amplitude as seen with 10 mg kg(-1), the drug significantly increased tissue NO and cGMP concentrations. Oral cicletanine attains its insulin sensitizing effect at doses lower than those necessary to activate the NO-cGMP pathway in the cardiovascular system. This metabolic effect requires functional integrity of hepatic sensory nerves.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Animals; Blood Pressure; Blood Vessels; Cyclic AMP; Cyclic GMP; Dose-Response Relationship, Drug; Enzyme Inhibitors; Glucose; Glucose Clamp Technique; Heart Rate; Hypercholesterolemia; Hyperinsulinism; Hypoglycemic Agents; Insulin Resistance; Lipids; Male; Myocardium; Nitric Oxide; Pyridines; Rabbits; Rosiglitazone; Thiazolidinediones

The hepatic parasympathetic nerves and hepatic nitric oxide synthase (NOS) are involved in the secretion of a hepatic insulin sensitizing substance (HISS), which mediates peripheral insulin sensitivity. We tested whether binding of ACh to hepatic muscarinic receptors is an upstream event to the synthesis of nitric oxide (NO), which, along with the activation of hepatic guanylate cyclase (GC), permits HISS release. Male Wistar rats (8-9 wk) were anesthetized with pentobarbital sodium (65 mg/kg). Insulin sensitivity was assessed using a euglycemic clamp [the rapid insulin sensitivity test (RIST)]. HISS inhibition was induced by antagonism of muscarinic receptors (atropine, 3 mg/kg i.v.) or by blockade of NOS [NG-nitro-L-arginine methyl ester (L-NAME), 1 mg/kg intraportally (i.p.v.)]. After the blockade, HISS action was tentatively restored using a NOdonor [3-morpholynosydnonimine (SIN-1), 5-10 mg/kg i.p.v.] or ACh (2.5-5 microg.kg(-1).min(-1) .i.p.v.). SIN-1 (10 mg/kg) reversed the inhibition caused by atropine (RIST postatropine 137.7 +/- 8.3 mg glucose/kg; reversed to 288.3 +/- 15.5 mg glucose/kg, n = 6) and by L-NAME (RIST post-L-NAME 152.2 +/- 21.3 mg glucose/kg; reversed to 321.7 +/- 44.7 mg glucose/kg, n = 5). ACh did not reverse HISS inhibition induced by L-NAME. The role of GC in HISS release was assessed using 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 5 nmol/kg i.p.v.), a GC inhibitor that decreased HISS action (control RIST 237.6 +/- 18.6 mg glucose/kg; RIST post-ODQ 111.7 +/- 6.2 mg glucose/kg, n = 5). We propose that hepatic parasympathetic nerves release ACh, leading to hepatic NO synthesis, which activates GC, triggering HISS action.

Topics: Acetylcholine; Animals; Cyclic GMP; Enzyme Inhibitors; Guanylate Cyclase; In Vitro Techniques; Injections, Intravenous; Insulin Resistance; Liver; Male; Molsidomine; Muscarinic Antagonists; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Oxadiazoles; Portal Vein; Quinoxalines; Rats; Rats, Wistar; Signal Transduction

We aimed to evaluate whether insulin influences vascular endothelial growth factor (VEGF) synthesis and secretion in cultured vascular smooth muscle cells (VSMCs) via nitric oxide (NO) and whether these putative effects are lost in insulin-resistant states.. In VSMC derived from human arterioles and from aortas of insulin-sensitive Zucker fa/+rats and insulin-resistant Zucker fa/fa rats incubated with different concentrations of human regular insulin with or without inhibitors of phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3-K), mitogen-activated protein kinase (MAPK), nitric oxide synthase (NOS) and guanosine 3',5'cyclic monophosphate(cGMP)-dependent protein kinase (PKG), we measured protein expression (Western blot) and secretion (ELISA) of VEGF.. We found that in VSMCs from humans and from insulin-sensitive Zucker fa/+rats, insulin increases VEGF protein expression and secretion, with mechanisms blunted by wortmannin and LY294002 (PI3-K inhibitors), PD98059 (MAPK inhibitor), L-NMMA (NOS inhibitor) and Rp-8pCT-cGMPs (PKG inhibitor). Also the NO donor sodium nitroprusside (SNP) and the cGMP analogue 8-Bromo-cGMP increase VEGF protein expression and secretion, with mechanisms inhibited by wortmannin and PD98059. The insulin effects on VEGF are impaired in VSMCs from Zucker fa/fa rats, which also present a reduced insulin ability to increase NO.. In VSMCs from humans and insulin-sensitive Zucker fa/+rats: (i) insulin increases VEGF protein expression and secretion via both PI3-K and MAPK; (ii) the insulin effects on VEGF are mediated by nitric oxide. The insulin action on both nitric oxide and VEGF is impaired in VSMCs from Zucker fa/fa rats, an animal model of metabolic and vascular insulin-resistance.

Topics: Animals; Blotting, Western; Cells, Cultured; Cyclic GMP; Enzyme-Linked Immunosorbent Assay; Humans; Insulin; Insulin Resistance; MAP Kinase Kinase Kinases; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Nitric Oxide; Nitric Oxide Donors; Nitroprusside; Rats; Rats, Zucker; Vascular Endothelial Growth Factor A

We investigated whether the platelets from obese subjects are sensitive as those from controls to the antiaggregating effects of N-acetyl-L-cysteine (NAC)-an antioxidant thiol that increases availability of endogenous nitric oxide (NO)-and of superoxide dismutase (SOD) and amifostine which act as scavengers of superoxide anion.. In platelets from obese subjects (n=20, body mass index [BMI]=34.2+/-1.9 kg/m(2), homeostasis model assessment [HOMA] index=5.5+/-1.1) and controls (n=20, BMI=21.4+/-0.6 kg/m(2), HOMA index=1.4+/-0.2), we investigated the effects of NAC on aggregation and on 3',5'-cyclic guanosine monophosphate (cGMP) synthesis and the interplay between NAC and the organic nitrates glyceryl trinitrate (GTN) and sodium nitroprusside (SNP). Similar experiments were carried out with SOD and amifostine.. We found that a 3-min platelet exposure to NAC decreased aggregation and increased cGMP in controls, but not in obese subjects. Only more prolonged incubations exerted a small effect also in obese subjects. GTN and SNP increased platelet cGMP in both groups, but their effect was much lower in obese subjects. NAC (3 mmol/l), SOD (150 U/ml), and amifostine (50 micromol/l) enhanced the increase of cGMP elicited by NO donors, but again, the effect was much lower in obese subjects.. Since antioxidants do not restore the effects of NO in platelets from obese subjects, we hypothesize that oxidative stress is not the unique cause of platelet resistance to NO in obesity and suggest that a resistance to the NO action at the guanylate cyclase level could play a role in this phenomenon, potentially involved in the increased atherothrombotic risk linked to obesity.

Topics: Acetylcysteine; Adenosine Diphosphate; Adult; Amifostine; Antioxidants; Collagen; Cyclic GMP; Drug Resistance; Female; Free Radical Scavengers; Humans; Insulin Resistance; Male; Nitric Oxide; Nitric Oxide Donors; Nitroglycerin; Nitroprusside; Obesity; Platelet Aggregation; Platelet Aggregation Inhibitors; Superoxide Dismutase

Insulin resistance is associated with an increased risk of atherothrombotic vascular disease, but the mechanisms are poorly understood. We determined how insulin in vivo regulates platelet activation in nonobese and obese subjects by using methods mimicking thrombus formation. Twelve nonobese (aged 42+/-2 years, body mass index 24.0+/-0.4 kg/m(2)) and 14 obese (aged 43+/-1 years, body mass index 37.2+/-1.5 kg/m(2)) subjects were studied under euglycemic hyperinsulinemic (3-hour insulin infusion of 1 mU. kg(-1). min(-1)) conditions. Before and at the end of hyperinsulinemia, the following were determined: (1) platelet-related early hemostasis (shear rate of approximately 4000 s(-1)) by platelet function analysis; (2) platelet deposition to collagen during whole-blood perfusion (shear rate of 1600 s(-1)); (3) aggregation responses to collagen, thrombin receptor-activating peptide, ADP, and epinephrine; and (4) platelet cGMP concentrations. Insulin action on glucose metabolism was 69% lower in the obese subjects (1.6+/-0.2 mg. kg(-1). min(-1)) than in the nonobese subjects (5.4+/-0.4 mg. kg(-1). min(-1), P<0.0001). The in vivo insulin infusion inhibited platelet deposition to collagen from 4.3+/-0.6x10(6) to 3.5+/-0.4x10(6) per square centimeter in the nonobese subjects (P<0.05) but failed to do so in the obese subjects (5.2+/-0.8x10(6) versus 5.5+/-0.7x10(6) per square centimeter, P=NS; P<0.01 versus nonobese subjects). Epinephrine- and ADP-primed closure times by platelet function analysis were prolonged by insulin in the nonobese but not the obese subjects (P<0.05 for between-group difference). In the nonobese subjects, insulin decreased aggregation to all agonists and significantly increased platelet cGMP concentrations (2.5+/-0.3 versus 3.2+/-0.5 pmol/10(9) for before versus after insulin, respectively; P<0.01). In the obese subjects, insulin did not alter collagen-induced aggregation or cGMP concentrations (1.9+/-0.2 versus 1.8+/-0.1 pmol/10(9) for before versus the end of in vivo hyperinsulinemia, respectively; P=NS). These data demonstrate that normal in vivo insulin action inhibits platelet interaction with collagen under conditions mimicking thrombus formation and reduces aggregation to several agonists. These platelet-inhibitory actions of insulin are blunted or absent in obese subjects and could provide 1 mechanism linking insulin resistance to atherothrombotic disease.

Topics: Adenosine Diphosphate; Adult; Analysis of Variance; Blood Platelets; Body Mass Index; Collagen; Cyclic GMP; Enzyme-Linked Immunosorbent Assay; Epinephrine; Glucose; Glucose Clamp Technique; Hemostasis; Humans; Insulin; Insulin Resistance; Obesity; Peptide Fragments; Platelet Aggregation

Pioneer studies have proposed that multiple metabolic abnormalities, such as insulin resistance, increased Na(+)-H(+) exchanger activity and abnormal intracellular calcium homeostasis, are frequently associated with a subset of essential hypertensive patients with low plasma renin activity (PRA). However, it is unclear whether insulin resistance is related to the low renin status in the very early phase of genetical hypertension. Besides, there is controversy on the subject of the in vivo effect of acute hyperinsulinaemia on sodium-related factors. We investigated the relationship between sodium-related parameters and insulin sensitivity, and the effects of euglycaemic hyperinsulinaemia on cyclic guanosine monophosphate (cGMP) and atrial natriuretic peptide (ANP) levels in 17 young, lean, normotensive male subjects, who displayed extreme predispositions for the development of hypertension. PRA was significantly lower in the positive than in the negative family history group (P < 0.05). Insulin sensitivity (M-value) was correlated with PRA before euglycaemic hyperinsulinaemic clamping (r = 0.577, P < 0.05), and was also inversely correlated with fractional excretion of sodium (FE(Na)) before clamping (r = -0.51, P < 0.05). Euglycaemic hyperinsulinaemia significantly decreased PRA (P < 0.0001) and increased cGMP (P < 0.05) and ANP levels (P < 0.01). In conclusion, insulin sensitivity may be partially determined by PRA levels and FE(Na) before clamping in young, lean, normotensive male subjects. Acute euglycaemic hyperinsulinaemia decreases PRA, and increases cGMP and ANP levels from the fasting condition.

Topics: Adult; Atrial Natriuretic Factor; Cyclic GMP; Glucose Clamp Technique; Homeostasis; Humans; Hyperinsulinism; Hypertension; Insulin Resistance; Male; Predictive Value of Tests; Renin; Sensitivity and Specificity; Sodium; Thinness

We have recently made the novel observation that a pro-oxidant challenge with hydroquinone in combination with buthionine sulfoximine (each at 50 mg/kg i.p. daily for 7 days) provokes the onset of type II diabetes mellitus in a model of insulin resistance, the obese Zucker rat. Since endothelial dysfunction in oxidant stress may aggravate in vivo insulin resistance, we have now investigated endothelium-dependent and nitric oxide (NO)-mediated vascular responses in the obese Zucker rat in vivo following this pro-oxidant insult. Pro-oxidant-treated animals exhibited defective vasodepression to the endothelium-dependent agent acetylcholine and to a lesser extent, the NO donor glyceryl trinitrate, together with a reduction in circulating levels of cGMP. Our data therefore suggest that the progression to type II diabetes mellitus in the obese Zucker rat mediated by a pro-oxidant insult is associated with impairments in agonist-stimulated, endothelium-dependent vasodilation and vascular NO signalling.

Topics: Acetylcholine; Animals; Antioxidants; Area Under Curve; Body Weight; Buthionine Sulfoximine; Cyclic GMP; Diabetes Mellitus, Type 2; Endothelium, Vascular; Enzyme Inhibitors; Hemodynamics; Hydroquinones; Insulin Resistance; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Obesity; Oxidants; Rats; Rats, Zucker

Nitric oxide activates guanylate cyclase to form cGMP, comprising a signalling system that is believed to be a distinct mechanism for increasing glucose transport and metabolism in skeletal muscle. The effects of a selective cGMP phosphodiesterase inhibitor, zaprinast, on basal glucose utilization was investigated in incubated rat soleus muscle preparations isolated from both insulin-sensitive (lean Zucker; Fa/?) and insulin-resistant (obese Zucker; fa/fa) rats. Zaprinast at 27 microM significantly increased cGMP levels in incubated soleus muscle isolated from lean, but not obese, Zucker rats. Muscles were incubated with 14C-labelled glucose and various concentrations of zaprinast (3, 27 and 243 microM). Zaprinast (at 27 and 243 microM) significantly increased rates of net and 14C-labelled lactate release and of glycogen synthesis in lean Zucker rat soleus muscle; glucose oxidation was also increased by 27 microM zaprinast. In addition, regardless of concentration, the phosphodiesterase inhibitor failed to increase any aspect of 14C-labelled glucose utilization in soleus muscles isolated from obese Zucker rats. The maximal activity of nitric oxide synthase (NOS) was significantly decreased in insulin-resistant obese Zucker muscles. Thus the lack of effect of zaprinast in insulin-resistant skeletal muscle is consistent with decreased NOS activity. To test whether there is a defect in insulin-resistant skeletal muscle for endogenous activation of guanylate cyclase, soleus muscles were isolated from both insulin-sensitive and insulin-resistant Zucker rats and incubated with various concentrations of the NO donor sodium nitroprusside (SNP; 0.1, 1, 5 and 15 mM). SNP significantly increased rates of net and 14C-labelled lactate release, as well as glucose oxidation in muscles isolated from both insulin-sensitive and insulin-resistant rats. A decreased response to SNP was observed in the dose-dependent generation of cGMP within isolated soleus muscles from insulin-resistant rats. A possible link between impaired NO/cGMP signalling and abnormal glucose utilization by skeletal muscle is discussed.

Topics: Animals; Cell Fractionation; Cyclic GMP; Female; Glucose; Glycogen; Insulin; Insulin Resistance; Lactic Acid; Muscle, Skeletal; Nitric Oxide; Nitric Oxide Synthase; Nitroprusside; Obesity; Phosphodiesterase Inhibitors; Purinones; Rats; Rats, Zucker; Signal Transduction

1. The effects of human insulin and elevated D-glucose on L-arginine transport and synthesis of nitric oxide (NO) and prostacyclin (PGI2) have been investigated in human umbilical vein endothelial cells isolated from gestational diabetic pregnancies. 2. The increase in the Vmax for L-arginine transport (9.0 +/- 1.1) pmol (micrograms protein)-1 min-1) in diabetic endothelial cells cultured in 5 mM D-glucose was unaffected following 24 h exposure to 25 mM D-glucose. 3. Gestational diabetes-induced increases in basal intracellular cGMP and L-citrulline levels (inhibitable by L-NAME) and [Ca2+], were unaffected by elevated D-glucose. In contrast, PGI2 release was inhibited in diabetic cells exposed to either 5 or 25 mM D-glucose. 4. Elevated D-glucose attenuated histamine (10 microM, 5 min)-stimulated accumulation of cGMP and L-citrulline in endothelial cells isolated from gestational diabetic pregnancies. 5. The membrane hyperpolarization (-79 +/- 0.9 mV) sustained in diabetic endothelial cells in culture was insensitive to elevated D-glucose. 6. Elevated D-glucose abolished the stimulatory effect of human insulin (1 nM, 8 h) on L-[3H]leucine incorporation in diabetic endothelial cells cultured in 5 mM D-glucose. 7. Human insulin reduced the elevated rates of L-arginine transport and cGMP accumulation in diabetic cells cultured in 5 mM D-glucose but failed to reduce increased rates of transport or NO production in cells exposed to 25 mM D-glucose or cycloheximide. 8. Our findings demonstrate that hyperglycaemia impairs the actions of human insulin on umbilical vein endothelial cells isolated from gestational diabetic pregnancies. Changes in insulin sensitivity and/or its signalling cascade may be affected by hyperglycaemia associated with gestational diabetes, resulting in insulin resistance in endothelial cells derived from the fetal vasculature.

Topics: Amino Acids; Antimetabolites; Arginine; Calcium; Citrulline; Cyclic GMP; Deoxyglucose; Endothelium, Vascular; Epoprostenol; Female; Glucose; Humans; Insulin Resistance; Membrane Potentials; Onium Compounds; Organophosphorus Compounds; Pregnancy; Pregnancy in Diabetics; Umbilical Veins

Previous studies in our laboratory showed that the platelet anti-aggregating effect exerted by insulin, mediated by a nitric oxide (NO)-induced increase of guanosine-3',5'-cyclic monophosphate (cGMP), is lost in the insulin-resistant of obesity and obese NIDDM. It is not clear 1) whether the alterations observed in obese NIDDM patients are attributable to the obesity-related insulin resistance or to diabetes per se and 2) whether insulin-resistant states present a normal or a blunted response to NO. This study has been conducted to investigate 1) the platelet sensitivity to insulin in lean NIDDM and 2) the platelet sensitivity to an NO donor, glyceryl trinitrate (GTN), in obesity and in both lean and obese NIDDM.. We determined 1) ADP-induced platelet aggregation and platelet cGMP content in platelet-rich plasma (PRP) obtained from 11 lean NIDDM patients, after a 3-min incubation with insulin (0, 240, 480, 960, 1,920 pmol/l) and 2) ADP-induced platelet aggregation and platelet cGMP content in PRP obtained from 9 obese subjects, 11 lean and 8 obese NIDDM patients, and 18 control subjects, after a 3-min incubation with 0, 20, 40, and 100 mumol/l GTN.. Insulin dose-dependently decreased platelet aggregation in lean NIDDM patients (P = 0.0001): with 1,920 pmol/l of insulin, ADP ED50 was 141.5 +/- 6.4% of basal values (P = 0.0001). Furthermore, insulin increased platelet cGMP (P = 0.0001) from 7.5 +/- 0.2 to 21.1 +/- 3.7 pmol/10(9) platelets. These results were similar to those previously described in healthy subjects. GTN reduced platelet aggregation in all the groups (P = 0.0001) at all the concentrations tested (P = 0.0001), but GTN IC50 values were much higher in insulin-resistant patients: 36.3 +/- 5.0 mumol/l in healthy control subjects, 26.0 +/- 6.0 mumol/l in lean NIDDM patients (NS vs. control subjects), 123.6 +/- 24.0 mumol/l in obese subjects (P = 0.0001 vs. control subjects), and 110.1 +/- 19.2 mumol/l in obese NIDDM patients (P = 0.0001 vs. control subjects). GTN dose-dependently increased platelet cGMP in all the groups (P = 0.0001 in control subjects, lean NIDDM patients, and obese subjects; P = 0.04 in obese NIDDM patients). Values reached by obese subjects and obese NIDDM patients, however, were lower than those reached by control subjects (with 100 mumol/l of GTN, P = 0.001 and P = 0.0001, respectively). In healthy control subjects and in obese subjects, the insulin:glucose ratio, used as an indirect measure of insulin sensitivity, was positively correlated to GTN IC50 (r = 0.530, P = 0.008), further suggesting that the sensitivity to NO is reduced in the presence of insulin resistance.. The insulin anti-aggregating effect is preserved in lean NIDDM; platelet sensitivity to GTN in preserved in lean NIDDM but is reduced in the insulin-resistant states of obesity and obese NIDDM. Resistance to nitrates, therefore, could be considered another feature of the insulin-resistance syndrome.

Topics: Adenosine Diphosphate; Adult; Analysis of Variance; Blood Glucose; Blood Platelets; Blood Pressure; Body Mass Index; Cholesterol; Cholesterol, HDL; Cyclic GMP; Diabetes Mellitus; Diabetes Mellitus, Type 2; Female; Glycated Hemoglobin; Humans; Insulin; Insulin Resistance; Male; Nitroglycerin; Obesity; Platelet Aggregation; Platelet Aggregation Inhibitors; Reference Values; Thinness; Triglycerides

1. The effects of nitric oxide (NO) on vascular reactivity and platelet function in the obese (cp/cp) and lean (+/?) JCR:LA-cp rats were investigated. 2. Phenylephrine (PE; 0.1 nM-10 microM) induced contraction of isolated aortic rings in both genotypes (cp/cp and +/?) of JCR:LA-cp rats. The sensitivity to contraction with PE was enhanced in cp/cp compared with +/? rings. Rings from both genotypes showed an increased contraction upon removal of the endothelium. 3. Acetylcholine (ACh; 0.1 nM-10 microM)-induced endothelium-dependent relaxation of rings was not significantly different in the two genotypes. Both were inhibited to a similar extent by NG-nitro-L-arginine methyl ester (L-NAME; 0.01-1 mM) when administered in vitro. 4. The nitric oxide synthase (NOS) inhibitor (L-NAME; 0.3, 1 or 3 mg ml(-1), p.o.) when administered in vivo increased blood pressure in cp/cp rats but not in +/? rats. 5. L-NAME resulted in greater inhibition of ACh-induced relaxation in cp/cp rings compared with +/? rings. 6. L-NAME treatment in vivo caused a decrease in cyclic GMP and NOS activity in rings from cp/cp but not +/? rats. 7. The NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP; 0.1 nM-10 microM)-induced relaxation of rings from +/? rats, an effect enhanced by the treatment with L-NAME in vivo. 8. Oral administration of L-NAME did not enhance the vasorelaxant effect of SNAP on rings of aorta from cp/cp animals. 9. Platelet aggregation and NOS activity were similar in both genotypes and were not modified by oral administration of L-NAME. 10. These results show that unimpaired generation of NO is crucial for maintenance of vascular tone particularly under conditions of vascular insult exemplified by insulin resistance, obesity and dyslipidemia detected in cp/cp rats.

Topics: Acetylcholine; Animals; Aorta, Thoracic; Blood Platelets; Blood Pressure; Cyclic GMP; Diabetes Mellitus; Enzyme Inhibitors; Genotype; Insulin Resistance; Muscle Contraction; Muscle, Smooth, Vascular; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Obesity; Penicillamine; Phenylephrine; Rats; S-Nitroso-N-Acetylpenicillamine; Vasoconstrictor Agents; Vasodilator Agents

The infusion of L-arginine induces the production of nitric oxide and stimulates the immediate secretion of insulin. To examine the relationship between insulin resistance and endothelium-dependent vascular relaxation in patients with essential hypertension, we evaluated the renal and insulin responses to L-arginine, 500 mg/kg infused intravenously over 30 minutes, in 23 patients with mild essential hypertension who were neither obese nor diabetic and in 20 normotensive control subjects. We found no difference between the two groups in blood glucose or insulin in the fasting condition. The renovascular relaxation induced by L-arginine was significantly less in patients with essential hypertension than in normotensive control subjects. The increase in plasma cyclic GMP in response to L-arginine was lower in hypertensive patients than in normotensive subjects. Although the serum concentrations of glucose in response to L-arginine were similar in the two groups, the serum insulin response of the essential hypertensives was significantly higher than that of the normotensive subjects. In all subjects, the peak cyclic GMP response to L-arginine was significantly correlated with the peak delta glucose/ delta insulin ratio response to L-arginine (r = .69, P < .001). Findings suggested that an impairment of endothelium-dependent renal vascular relaxation and a reduced sensitivity to insulin are present in patients with essential hypertension. A link may be present between the abnormality of the L-arginine/nitric oxide/cyclic GMP pathway and insulin resistance in patients with essential hypertension.

Topics: Arginine; Blood Glucose; Blood Pressure; Cyclic GMP; Female; Heart Rate; Humans; Hypertension; Insulin; Insulin Resistance; Insulin Secretion; Male; Middle Aged; Renal Circulation; Vasodilation

Non-insulin-dependent diabetes mellitus (NIDDM) is characterized by insulin resistance hyperinsulinaemia and a high frequency of hypertension. It has recently been shown that insulin exerts a sodium-retaining effect, which is preserved in NIDDM: We sought to determine whether insulin affected renal sodium handling differently in hypertensive and normotensive NIDDM patients.. After a baseline period of 2 h, eight normotensive (N-) NIDDM patients and eight NIDDM patients with hypertension (H-) underwent a euglycaemic clamp with infusion of two sequential doses of insulin (50 and 500 mU/kg/h) or vehicle (time control) during 2-h periods each. Fractional clearances of sodium and lithium were determined according to standard methods. Fractional lithium clearance was used to assess segmental tubular sodium handling.. Insulin induced similar decrements in fractional sodium excretion (N-NIDDM: 43+/-5.9 and 57+/-9.1%,H-N IDDM: 48+/-16.4 and 62+/-12.5%, low and high insulin dose respectively). Distal tubular sodium absorption increased simultaneously. A fall in fractional proximal sodium reabsorption was observed in N-NIDDM (4.4+/-2.7 and 29.8+/-5.1%, low and high insulin dose respectively), which was attenuated in H-NIDDM (-5.0+/-7.3 and -2.1+/-13.9% respectively). The latter appeared to be related to a defective atrial natriuretic factor (ANF) and renal cyclic GMP response. A modest decrease in blood pressure occurred during insulin infusion that was not related to changes in ANF or FeLi.. The findings suggest that insulin-induced sodium retention may contribute to hypertension in NIDDM if the homeostatic response to offset this effect fails.

Topics: Atrial Natriuretic Factor; Blood Pressure; Cardiovascular System; Case-Control Studies; Cyclic GMP; Diabetes Mellitus, Type 2; Female; Humans; Hypertension; Insulin; Insulin Resistance; Kidney; Male; Middle Aged; Natriuresis; Renin

Patients with leprechaunism have hyperinsulinemia and extreme insulin resistance. The mechanism of the insulin resistance has not been delineated. To examine postreceptor events in this unusual syndrome we have assayed the enzyme guanylate cyclase [E.C.4.6.12], which is modulated by insulin, and the concentration of the intracellular messenger cyclic GMP in liver from two children with leprechaunism and extreme insulin resistance. Both patients exhibited down regulation of the red blood cell insulin receptors, but normal insulin receptor binding to Ebstein-Barr transformed IM-9 lymphocytes and monocytes. There was no evidence of antireceptor or antiinsulin antibodies. Activity of liver guanylate cyclase expressed as pmol/mg protein/10 min incubation in the soluble and particulate fractions were, respectively, Ark-1 133 +/- 18, 25 +/- 6; Ark-2 129 +/- 17, 23 +/- 8; control children (six average) 287 +/- 16, 55 +/- 9. The concentration of cyclic GMP was also 50% lower (0.08 +/- 0.03 in Ark-1 and 0.07 +/- 0.04 in Ark-2), compared to 0.19 +/- 0.07 pmol/mg protein/min in the control livers. There was no change in adenylate cyclase activity in children with leprechaunism versus the control children. These data suggest an abnormality of a postreceptor event in this rare genetic disease. These data, however, do not rule out that in some cases of leprechaunism a receptor binding abnormality may be the primary defect. We speculate that a defect in insulin action distal to plasma membrane receptor binding may be etiological in this unusual syndrome.

Topics: Abnormalities, Multiple; Cyclic GMP; Face; Female; Guanylate Cyclase; Hirsutism; Humans; Infant, Newborn; Insulin Resistance; Liver; Receptor, Insulin

Topics: Adrenergic alpha-Agonists; Adrenergic beta-Agonists; Amino Acids; Animals; Binding Sites; Blood Glucose; Calcium; Carbohydrate Metabolism; Cyclic AMP; Cyclic GMP; Diabetes Mellitus; Gastrointestinal Hormones; Gluconeogenesis; Growth Hormone; Hormones; Humans; Hypoglycemia; Hypoglycemic Agents; Insulin; Insulin Resistance; Insulin Secretion; Islets of Langerhans; Lipid Metabolism; Obesity; Proteins; Receptors, Cell Surface; Sulfonylurea Compounds