Lipids and insulin regulate mitochondrial-derived peptide (MOTS-c) in PCOS and healthy subjects.

OBJECTIVE: Polycystic ovarian syndrome (PCOS) is a heterogeneous endocrine disorder associated with mitochondrial dysfunction and insulin resistance (IR). MOTS-c, a mitochondrial peptide, promotes insulin sensitivity (IS) through activating AKT and AMPK-dependent pathways. The current study was designed to examine the response of MOTS-c to lipids (intralipid) followed by insulin in PCOS and healthy subjects. METHODS: All subjects underwent 5-hour intralipid/saline infusion with a hyperinsulinemic-euglycaemic clamp in the final 2 hours. Plasma samples were collected to measure circulating MOTS-c using a commercial ELISA kit. Subsequently, this was repeated following an eight-week exercise intervention. RESULTS: Intralipid significantly increased plasma MOTS-c both in controls and PCOS subjects, whilst the insulin infusion blunted the intralipid-induced response seen for both lipids and MOT-c. Intralipid elevated plasma MOTS-c to 232 ± 124% of basal in control (P < 0.01) and to 349 ± 206% of basal in PCOS (P < 0.001) subjects. Administration of insulin suppressed intralipid-induced MOTS-c from 232 ± 124% to 165 ± 97% (NS) in control and from 349 ± 206% to 183 ± 177% (P < 0.05) in PCOS subjects, respectively. Following exercise, intralipid elevated plasma MOTS-c to 305 ± 153% of basal in control (P < 0.01) and to 215 ± 103% of basal in PCOS (P < 0.01) subjects; insulin suppressed intralipid-induced MOTS-c only in controls. CONCLUSIONS: In conclusion, this is the first study to show increased lipid enhanced circulating MOTS-c whilst insulin attenuated the MOTS-c response in human. Further, eight weeks of moderate exercise training did not show any changes in circulating MOTS-c levels in healthy controls and in women with PCOS.

Effect of magnesium sulfate administration to improve insulin resistance in type 2 diabetes animal model: using the hyperinsulinemic-euglycemic clamp technique.

This study attempted to elucidate the possible mechanism of magnesium sulfate (MgSO4 ) administration on reducing insulin resistance in type 2 diabetic rats. Fifty Wistar rats were divided into five groups: NDC was fed the normal diet, CD received high-fat diet with 35 mg/kg of streptozotocin, CD-Mg animals received MgSO4 via drinking water, CD-Ins1, and CD-Ins2 animals treated with low or high dose of insulin. Body weight and blood glucose levels were measured weekly. Intraperitoneal glucose tolerance test (IPGTT), insulin tolerance test, and metabolic cage assessment were performed monthly. After 12 weeks, the hyperinsulinemic-euglycemic clamp was performed for all animals and blood sample was taken to measure glycated hemoglobin (HbA1c), plasma insulin, glucagon, calcium, and magnesium levels. Liver and gastrocnemius muscle were isolated to measure glucagon receptor (GR), Glucose 6 phosphatase (G6Pase), Phosphoenolpyruvate carboxykinase (Pepck) and Glucose transporter 4 (Glut4) genes expression and GLUT4 protein translocation into the cell membrane. Consuming of high-fat diet generated insulin-resistant rats. Magnesium or insulin therapy altered insulin resistance, blood glucose, IPGTT, gluconeogenesis pathway, GR, body weight, the percentage of body fat, and HbA1C in diabetic rats. Administrations of MgSO4 or insulin in Type 2 diabetes mellitus animals increase GLUT4 gene and protein expression. Mg could improve glucose tolerance via stimulation of Glut4 gene expression and translocation and also suppression of the gluconeogenesis pathway and GR gene expression. Mg also increased glucose infusion rate and displayed beneficial effects in the treatment of glucose metabolism and improved insulin resistance.

Insulin resistance and protein metabolism in chronic hemodialysis patients.

OBJECTIVE: Loss of lean body mass (sarcopenia) is associated with increased morbidity and mortality in patients receiving chronic hemodialysis (CHD). Insulin resistance (IR), which is highly prevalent in patients receiving CHD, has been proposed to play a critical role in the development of sarcopenia. The aim of this study was to examine the effect of IR on amino acid metabolism in patients receiving CHD. DESIGN: This was a cross-sectional study. SUBJECTS: The study included 12 prevalent (i.e., patients that have been on dialysis for more than 90 days) African American patients receiving CHD. METHODS: IR was measured as glucose disposal rate (GDR) determined from hyperinsulinemic euglycemic clamp (HGEC) studies performed 3 consecutive times. Plasma amino acid (AA) concentrations were measured by real-time high-performance liquid chromatography (HPLC) throughout the clamp study. The primary outcome was percentage change in leucine concentrations during the clamp study. The main predictor was the GDR measured simultaneously during the HGEC studies. Mixed model analysis was used to account for repeated measures. RESULTS: All individual AA concentrations declined significantly in response to high-dose insulin administration (P < .001). There was a significant direct association between GDR by HECG studies and the percentage change in leucine concentration (P = .02). Although positive correlations were observed between GDR values and concentration changes from baseline for other AAs, these associations did not reach statistical significance. CONCLUSIONS: Our results suggest that the severity of IR of carbohydrate metabolism is associated with a lesser decline in plasma leucine concentrations, suggesting a similar resistance to protein anabolism. Insulin resistance represents a potential mechanism for sarcopenia commonly observed in patients receiving CHD.

Effects of n-3 PUFAs on postprandial variation of metalloproteinases, and inflammatory and insulin resistance parameters in dyslipidemic patients: evaluation with euglycemic clamp and oral fat load.

BACKGROUND: The oral fat load (OFL) is considered as one of the most accurate models of postprandial lipoprotein metabolism and it has been widely used to evaluate the postprandial fat load effect on single markers of inflammation. OBJECTIVE: To evaluate the effects of n-3 PUFAs, primarily eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), with a content of 400 mg of EPA and 450 mg of DHA in each capsule, on metalloproteinases and inflammatory biomarkers in patients affected by combined dyslipidemia both in a fasting state and after a standardized OFL in a randomized, placebo-controlled trial. METHODS: Placebo or n-3 PUFAs 3 g/day (1 g three times a day during the meals) was administered for 6 months. At the baseline, and after 2, 4, and 6 months we evaluated body mass index (BMI), body weight, fasting plasma glucose (FPG), fasting plasma insulin (FPI), homeostasis model assessment insulin resistance index (HOMA-IR), blood pressure, lipid profile, soluble intercellular adhesion molecule-1 (sICAM-1), interleukin 6 (IL-6), high-sensitivity C-reactive protein (hs-CRP), soluble vascular cell adhesion molecule-1 (sVCAM-1), sE-selectin, tumor necrosis factor-α (TNF-α), and metalloproteinases 2 and 9 (MMP-2 and 9). Furthermore, at the baseline and at the end of the study, all patients underwent an euglycemic hyperinsulinemic clamp and an oral fat load. RESULTS: Tg levels were lower (-54 mg/dL) and high-density lipoprotein cholesterol higher (+6 mg/dL) with n-3 PUFAs compared with placebo; n-3 PUFAs gave lower levels of FPG (-3 mg/dL), sICAM (-25 ng/mL), IL-6 (-0.3 pg/mL), hs-CRP (-0.6 mg/L), sVCAM-1 (-89 ng/mL), sE-selectin (-5.8 ng/mL), TNF-α (-0.3 ng/mL), MMP-2 (-185.1 ng/mL), and MMP-9 (-91.5 ng/mL), and a greater M value (+1.21 µmol/min/kg) compared with placebo. After the OFL, there was a decrease of Tg, MMPs, and all inflammatory parameters with n-3 PUFAs, but not with placebo. CONCLUSION: Supplementation with n-3 PUFA resulted in lower levels of FPG, plasma lipids, MMPs, and inflammatory parameters and in a better increase of M value compared to placebo, both in the fasting state and after an OFL.

Management of lercanidipine overdose with hyperinsulinaemic euglycaemia therapy: case report.

This case report describes the first reported overdose of the dihydropyridine calcium channel blocker (CCB) lercanidipine. A 49 yr old male presented to the Emergency Department 3 hrs after the ingestion of 560 mg of lercanidipine. In the department he had a witnessed seizure within 15 minutes of arrival attributed to the overdose. Following immediate recovery of consciousness after the seizure, he had refractory hypotension and bradycardia which failed to respond to fluid resuscitation, glucagon therapy, and intravenous calcium. He went on to require vasopressor support with noradrenaline and was treated with high dose insulin therapy which was successful in achieving cardiovascular stability. Vasopressor therapy was no longer required within one half life of lercanidipine, and the total stay on intensive care was one day before transfer to a ward.Calcium channel blocker overdose is an uncommon but life-threatening overdose. Treatment for severe toxicity is similar to b-blocker overdose. Hypotension is treated with intravenous fluid therapy, intravenous calcium and possibly glucagon with vasopressor or inotropic support as required. Atropine is used to attempt reversal of bradycardia. High doses of intravenous insulin with intravenous dextrose as required (hyperinsulinaemic euglycaemia or HIET), has also been successfully reported. Experimental animal data suggests that HIET is of benefit and potentially superior to fluid therapy, calcium, glucagon and potentially vasopressor therapy. HIET effectively and sustainably reverses hypotension, bradycardia and improves myocardial contractility and metabolism. Current advice in calcium channel blocker overdose is to begin therapy early in toxicity, starting with a 1.0 IU/kg insulin bolus followed by an infusion of 0.5 IU/kg/hr of insulin and dextrose as required titrated to clinical response.

Chinese herbal extracts (SK0506) as a potential candidate for the therapy of the metabolic syndrome.

The metabolic syndrome has reached epidemic proportions worldwide, but currently there is a lack of effective therapies for this multifactorial endocrine disease. TCM (traditional Chinese medicine) has been utilized to treat a wide variety of diseases for centuries in the People's Republic of China, subsequently becoming a promising source for the development of new therapeutic agents. Chinese medicinal herbs Gynostemma pentaphyllum, Coptis chinensis and Salvia miltiorrhiza have been shown to have anti-atherosclerotic and antidiabetic properties. In this study, we have investigated the metabolic effects of a mixture of these three herbal extracts (SK0506) in a rodent model of the metabolic syndrome induced by an HFD (high-fat diet). SD (Sprague-Dawley) rats that were fed on an HFD for 4 weeks gained 33% more weight compared with chow-fed rats (P<0.05). Four weeks treatment with SK0506 prevented weight gain with decreased visceral fat (P<0.01 compared with vehicle treatment). SK0506 also significantly reduced plasma triacylglycerols (triglycerides), NEFAs (non-esterified fatty acids) and cholesterol. SK0506 exerted similar effects to RSG (rosiglitazone) on impaired glucose intolerance. SK0506 also significantly enhanced glucose uptake and glycogen synthesis in adipose tissue during hyperinsulinaemic-euglycaemic clamp. Western blotting analysis revealed that SK0506 enhanced GLUT4 (glucose transporter 4) expression in adipose tissue, and RSG markedly up-regulated GLUT4 translocation in skeletal muscle. Overall, the present study has discovered that SK0506 can reverse several components of the metabolic syndrome primarily through acting on hyperlipidaemia and visceral obesity. The results from the present study suggest that it is worthwhile to conduct a randomized clinical trial to confirm the potential that SK0506 may be a new oral agent for treating the metabolic syndrome and preventing Type 2 diabetes.

Hypothalamic nutrient sensing activates a forebrain-hindbrain neuronal circuit to regulate glucose production in vivo.

OBJECTIVE: Hypothalamic nutrient sensing regulates glucose production, but the neuronal circuits involved remain largely unknown. Recent studies underscore the importance of N-methyl-d-aspartate (NMDA) receptors in the dorsal vagal complex in glucose regulation. These studies raise the possibility that hypothalamic nutrient sensing activates a forebrain-hindbrain NMDA-dependent circuit to regulate glucose production. RESEARCH DESIGN AND METHODS: We implanted bilateral catheters targeting the mediobasal hypothalamus (MBH) (forebrain) and dorsal vagal complex (DVC) (hindbrain) and performed intravenous catheterizations to the same rat for infusion and sampling purposes. This model enabled concurrent selective activation of MBH nutrient sensing by either MBH delivery of lactate or an adenovirus expressing the dominant negative form of AMPK (Ad-DN AMPK α2 [D¹57A]) and inhibition of DVC NMDA receptors by either DVC delivery of NMDA receptor blocker MK-801 or an adenovirus expressing the shRNA of NR1 subunit of NMDA receptors (Ad-shRNA NR1). Tracer-dilution methodology and the pancreatic euglycemic clamp technique were performed to assess changes in glucose kinetics in the same conscious, unrestrained rat in vivo. RESULTS: MBH lactate or Ad-DN AMPK with DVC saline increased glucose infusion required to maintain euglycemia due to an inhibition of glucose production during the clamps. However, DVC MK-801 negated the ability of MBH lactate or Ad-DN AMPK to increase glucose infusion or lower glucose production. Molecular knockdown of DVC NR1 of NMDA receptor via Ad-shRNA NR1 injection also negated MBH Ad-DN AMPK to lower glucose production. CONCLUSIONS: Molecular and pharmacological inhibition of DVC NMDA receptors negated hypothalamic nutrient sensing mechanisms activated by lactate metabolism or AMPK inhibition to lower glucose production. Thus, DVC NMDA receptor is required for hypothalamic nutrient sensing to lower glucose production and that hypothalamic nutrient sensing activates a forebrain-hindbrain circuit to lower glucose production.

An abbreviated hyperinsulinemic-euglycemic clamp results in similar myocardial glucose utilization in both diabetic and non-diabetic patients with ischemic cardiomyopathy.

BACKGROUND: Positron emission tomography (PET) with insulin-stimulated (18)F-2-deoxyglucose (FDG) uptake is the gold standard for myocardial viability. However, insulin stimulation is infrequently performed due to time and inconvenience. We therefore assessed the clinical applicability of an abbreviated hyperinsulinemic-euglycemic clamp. METHODS AND RESULTS: Dynamic FDG PET was performed in 50 patients with ischemic cardiomyopathy (ejection fraction: .30 +/- .10) using an abbreviated hyperinsulinemic-euglycemic clamp with separate Non-Diabetic (n = 26) and Diabetic (n = 24) protocols (American Society of Nuclear Cardiology guidelines), and supplemental potassium. In regions with normal resting perfusion ((13)N-ammonia uptake >or=80% maximal segment), there were no differences in either maximal (Non-Diabetic: .60 +/- .20 vs Diabetic: .60 +/- .17 micromol/min/g, P = .93) or mean rates of myocardial glucose uptake (MGU) (Non-Diabetic: .52 +/- .18 vs Diabetic: .52 +/- .14 micromol/min/g, P = .63) between the protocols. Multivariate analysis showed that diastolic blood pressure alone (maximal MGU, r (2) = .20, P = .001) or with NYHA Heart Failure Class (mean MGU, r (2) = .25, P = .003) could account for some of the variability in normal-region MGU. Potassium supplementation safely attenuated the decline in plasma levels. CONCLUSIONS: This abbreviated hyperinsulinemic-euglycemic clamp produced similar MGU values in normal resting myocardium in non-diabetic and diabetic subjects, which are no different than published rates with a standard insulin clamp. Thus, this abbreviated approach is sufficient to overcome myocardial insulin resistance.

Review of the mechanism of action and clinical efficacy of recombinant human hyaluronidase coadministration with current prandial insulin formulations.

For patients with type 1 or type 2 diabetes, achieving good glycemic control is critical for successful treatment outcomes. As many patients remain unable to reach glycemic goals with currently available rapid-acting analog insulins, ultrafast insulin products are being developed that provide an even faster pharmacokinetic profile compared with current rapid prandial insulin products. The overall strategy of these ultrafast insulin products is to better mimic the normal physiologic response to insulin that occurs in healthy individuals to further improve glycemic control. Recombinant human hyaluronidase (rHuPH20) is a genetically engineered soluble hyaluronidase approved by the U.S. Food and Drug Administration as an adjuvant to increase the absorption and dispersion of other injected drugs; mammalian hyaluronidases as a class have over 6 decades of clinical use supporting the safety and/or efficacy of hyaluronidase coadministration. Clinical findings have demonstrated that coadministration of rHuPH20 with insulin or an insulin analog achieved faster systemic absorption, reduced inter- and intrapatient variability of insulin absorption, and achieved faster metabolic effects compared with injection of either insulin formulation alone. The magnitude of this acceleration is similar to the incrementally faster absorption of prandial insulin analogs as compared with regular insulin. In addition, coadministration of rHuPH20 with regular insulin or insulin analog also improved the achievement of prandial glycemic targets. Thus, rHuPH20 coadministration shows promise as a method of establishing a more rapid insulin profile to prandial insulin in patients with diabetes and has the potential to yield substantial improvements in postprandial glycemic excursion.

Surplus dietary tryptophan inhibits stress hormone kinetics and induces insulin resistance in pigs.

Recently we have shown that surplus dietary tryptophan (TRP) reduced the plasma concentrations of cortisol and noradrenaline in pigs. Stress hormones are known to affect insulin sensitivity and metabolism. We now investigated the long-term effects of surplus dietary TRP on 1) plasma and urinary stress hormone kinetics, 2) insulin sensitivity for glucose and amino acid clearance, and 3) whole body nitrogen balance. Pigs were fed for 3weeks a high (13.2%) vs normal (3.4%) TRP to large neutral amino acids (LNAA) diet, leading to reduced fasting (14 h) plasma cortisol (17.1+/-3.0 vs 28.9+/-4.3 ng/mL, p<0.05) and noradrenaline (138+/-14 vs 225+/-21 pg/mL, p<0.005) concentrations, lower daily urinary noradrenaline (313+/-32 vs 674+/-102 ng/kg day, p<0.001) and adrenaline (124+/-13 vs 297+/-42 ng/kg day, p<0.001) but higher dopamine (5.8+/-0.5 vs 1.5+/-0.2 microg/kg day, p<0.001) excretions, respectively. Insulin sensitivities for both glucose and amino acid clearance, (as measured by the intraportal hyperinsulinaemic (1 mU/kg min) euglycaemic euaminoacidaemic clamp technique), were lower by 22% in pigs on the high vs normal TRP/LNAA diet (14.8+/-1.4 vs 18.9+/-0.9, p<0.05 and 69.7+/-4.3 vs 89.7+/-6.8 mL/kg min, p<0.05, respectively) without affecting urinary nitrogen excretion (35.5+/-1.0 vs 36.6+/-1.0% of dietary nitrogen intake, p=ns). In conclusion, long-term feeding of surplus dietary TRP inhibits both baseline adrenocortical and sympathetic nervous system activity, it induces insulin resistance for both glucose and amino acid clearance but it does not affect whole body protein catabolism. This indicates that the bioactive amino acid TRP contributes to homeostasis in neuroendocrinology and insulin action and that low baseline adrenocortical and sympatho-adrenal axis activity are associated with insulin resistance.

Bench-to-bedside review: hyperinsulinaemia/euglycaemia therapy in the management of overdose of calcium-channel blockers.

Hyperinsulinaemia/euglycaemia therapy (HIET) consists of the infusion of high-dose regular insulin (usually 0.5 to 1 IU/kg per hour) combined with glucose to maintain euglycaemia. HIET has been proposed as an adjunctive approach in the management of overdose of calcium-channel blockers (CCBs). Indeed, experimental data and clinical experience, although limited, suggest that it could be superior to conventional pharmacological treatments including calcium salts, adrenaline (epinephrine) or glucagon. This paper reviews the patho-physiological principles underlying HIET. Insulin administration seems to allow the switch of the cell metabolism from fatty acids to carbohydrates that is required in stress conditions, especially in the myocardium and vascular smooth muscle, resulting in an improvement in cardiac contractility and restored peripheral resistances. Studies in experimental verapamil poisoning in dogs have shown that HIET significantly improves metabolism, haemodynamics and survival in comparison with conventional therapies. Clinical experience currently consists only of a few isolated cases or short series in which the administration of HIET substantially improved cardiovascular conditions in life-threatening CCB poisonings, allowing the progressive discontinuation of vasoactive agents. While we await further well-designed clinical trials, some rational recommendations are made about the use of HIET in severe CBB overdose. Although the mechanism of action is less well understood in this condition, some experimental data suggesting a potential benefit of HIET in beta-adrenergic blocker toxicity are discussed; clinical data are currently lacking.

Comparison of the effects on glucose and lipid metabolism of equipotent doses of insulin detemir and NPH insulin with a 16-h euglycaemic clamp.

AIMS/HYPOTHESIS: The association of insulin detemir with non-esterified fatty acid binding sites on albumin may limit its transfer from the circulation into the extravascular extracellular space in adipose tissue and muscle, due to the capillary endothelial cell barrier. In the liver, the open sinusoids may expose hepatocytes to insulin detemir, enabling it to have a greater effect in the liver than in peripheral tissues. METHODS: We investigated the effects of equipotent doses of insulin detemir and NPH insulin on hepatic glucose rate of appearance (Ra), peripheral glucose rate of disposal (Rd) and glycerol Ra (a measure of lipolysis) using stable isotope techniques. We also investigated the effects of these insulins on NEFA concentrations in seven healthy volunteers during a 16-h euglycaemic clamp. A higher dose of insulin detemir was also studied. RESULTS: There was no difference in the glucose infusion profile between insulin detemir and NPH. Insulin detemir had a greater effect on mean suppression of glucose Ra (mean difference 0.24 mg kg(-1) min(-1); CI 0.09-0.39; p<0.01), and minimum glucose Ra, with minimum low dose detemir -0.10+/-0.15 mg.kg(-1).min(-1) and minimum NPH 0.17+/-0.10 mg.kg(-1).min(-1) (p<0.02). However, it had a lesser effect on mean suppression of NEFA concentrations (mean difference -0.10 mmol/l; CI -0.03 to -0.17; ANOVA, p<0.02) than NPH. The effect of insulin detemir on glucose Rd and glycerol Ra was not different from NPH. Following high-dose detemir, total glucose infused and maximum glucose Rd were higher (p<0.02, p<0.03) and plasma NEFA concentrations lower (p<0.01) than with low-dose determir. CONCLUSIONS/INTERPRETATION: This study suggests that insulin detemir, when compared to NPH insulin, has a greater effect on the liver than on peripheral tissues and thus has the potential to restore the physiological insulin gradient.

Pre-clinical methods for the determination of insulin sensitivity.

We compared the hyperinsulinaemic euglycaemic glucose clamping (HEGC) procedure and the rapid insulin sensitivity test (RIST) to characterize insulin sensitivity in anaesthetized rats. The changes in insulin sensitivity were then supplemented with the direct measurement of insulin-stimulated glucose uptake using tissue accumulation of radioactive 2-deoxyglucose in skeletal muscle samples obtained from animals undergone either procedure. Studies of the recently described endogenous insulin sensitizer mechanism termed hepatic insulin sensitizing (HISS) mechanism, by the two methods yielded data for evaluation. The HISS mechanism is defined as an increase in tissue insulin sensitivity in response to post-prandial hepatic release of an undefined substance through a nitrergic pathway. For the HEGC method, insulin was infused to attain a stable plasma insulin immunoreactivity of 100 microU/ml determined by radioimmunoassay, whereas with the RIST method the HISS mechanism was activated by a 50 mg/kg i.v. insulin bolus. Euglycaemia was kept constant by means of glucose infusion. With the HEGC and the RIST methods, insulin sensitivity was defined as the average rate of glucose infusion and the amount of glucose/kg body weight/40 min (RIST index) infused to maintain euglycaemia and preinvestigation blood glucose level, respectively. During HEGC 16+/-4.2 mg/kg/min glucose was able to maintain euglycaemia, which decreased to 8+/-2.9 (p<0.05) after administration of 10 mg/kg NG-nitro-L-arginine methyl ester (L-NAME) (i.p.), a NO synthase inhibitor. Conversely, the RIST index decreased by 55+/-6.9% (p<0.05) after L-NAME. Similarly, 2-deoxyglucose uptake by the gastrocnemius muscle was decreased by 49.9+/-5.8 (p<0.05) and 52.3+/-7.4% (p<0.05) with the HEGC and the RIST methods, respectively. The results show that both the HEGC and the RIST methods supplemented with tissue radioactive 2-deoxyglucose uptake determinations are appropriate methods to characterize the alteration of insulin sensitivity in context of the HISS mechanism.