A comparison of the effects of low- and conventional-dose thiazide diuretic on insulin action in hypertensive patients with NIDDM

Summary In conventional doses, thiazide diuretics impair glucose tolerance and decrease insulin sensitivity, making them an unpopular choice for treating diabetic patients with hypertension. However, use of low-dose thiazide diuretics may avoid the adverse metabolic effects seen with conventional doses. In a double-blind, randomised crossover study we assessed peripheral and hepatic insulin action in 13 hypertensive non-insulin-dependent diabetic patients after a 6-week placebo run-in and following two 12-week treatment periods with either low (1.25 mg) or conventional (5.0 mg) dose bendrofluazide. There were no differences between doses in their effects on systolic and diastolic blood pressure. Bendrofluazide 1.25 mg had significantly less effect on serum potassium, uric acid, fasting glucose and HbA_{1 c} concentrations than the 5.00 mg dose. Exogenous glucose infusion rates required to maintain euglycaemia were significantly different between doses (p < 0.05) with conventional-dose bendrofluazide worsening peripheral insulin resistance compared to baseline (23.8 ± 2.9 vs 27.3 ± 3.5 μmol · kg^{− 1}· min^{− 1}, p < 0.05) and low-dose bendrofluazide producing no change compared to baseline (26.8 ± 3.6 vs 27.3 ± 3.5 μmol · kg^{− 1}· min^{− 1}, p = NS). Postabsorptive endogenous glucose production was higher on treatment with bendrofluazide 5.0 mg compared to 1.25 mg (11.7 ± 0.5 vs 10.2 ± 0.3 μmol · kg^{− 1}· min^{− 1}, p < 0.05) and suppressed to a lesser extent following insulin (4.0 ± 0.7 vs 2.0 ± 0.4 μmol · kg^{− 1}· min^{− 1}, p < 0.05). Treatment with bendrofluazide 5.0 mg increased postabsorptive endogenous glucose production compared to baseline (11.7 ± 0.5 vs 10.6 ± 0.4 μmol · kg^{− 1}· min^{− 1}, p < 0.05) whereas bendrofluazide 1.25 mg did not (10.2 ± 0.3 vs 10.6 ± 0.4 μmol · kg^{− 1}· min^{− 1}, p = NS). At a dose of 1.25 mg bendrofluazide is as effective as conventional doses but has less adverse metabolic effects. In contrast to conventional doses which worsen both hepatic and peripheral insulin resistance, low-dose bendrofluazide has no effect on insulin action in non-insulin-dependent diabetic subjects. [Diabetologia (1995) 38: 853–859] Received: 6 September 1994 and in revised form: 29 December 1994     

Insulin resistance and coronary artery disease

Summary The purpose of the present study was to quantitate insulin-mediated glucose disposal in normal glucose tolerant patients with angiographically documented coronary artery disease (CAD) and to define the pathways responsible for the insulin resistance. We studied 13 healthy, normal weight, normotensive subjects with angiographically documented CAD and 10 age-, weight-matched control subjects with an oral glucose tolerance test and a 2-h euglycaemic insulin (40 mU · m⁻²· min⁻¹) clamp with tritiated glucose and indirect calorimetry. Lean body mass was measured with tritiated water. All CAD and control subjects had a normal oral glucose tolerance test. Fasting plasma insulin concentration (66 ± 6 vs 42 ± 6 pmol/l, p < 0.05) and area under the plasma insulin curve following glucose ingestion (498 ± 54 vs 348 ± 42 pmol · l⁻¹· min⁻¹, p < 0.001) were increased in CAD vs control subjects. Insulin-mediated whole body glucose disposal (27.8 ± 3.9 vs 38.3 ± 4.4 μmol · kg fat free mass (FFM)⁻¹· min⁻¹, p < 0.01) was significantly decreased in CAD subjects and this was entirely due to diminished non-oxidative glucose disposal (8.9 ± 2.8 vs 20.0 ± 3.3 μmol · kg FFM⁻¹· min⁻¹, p < 0.001). The magnitude of insulin resistance was positively correlated with the severity of CAD (r = 0.480, p < 0.05). In the CAD subjects basal and insulin-mediated rates of glucose and lipid oxidation were normal and insulin caused a normal suppression of hepatic glucose production. In conclusion, subjects with angiographically documented CAD are characterized by moderate-severe insulin resistance and hyperinsulinaemia and should be included in the metabolic and cardiovascular cluster of disorders that comprise the insulin resistance syndrome or ’syndrome X'. [Diabetologia (1996) 39: 1345–1350] Received: 6 February 1996 and in revised form: 29 May 1996     

Non-invasive tracing of liver intermediary metabolism in normal subjects and in moderately hyperglycaemic NIDDM subjects. Evidence against increased gluconeogenesis and hepatic fatty acid oxidation in NIDDM

Summary To test whether gluconeogenesis is increased in non-insulin-dependent diabetic (NIDDM) patients we infused (post-absorptive state) healthy subjects and NIDDM patients with [6,6-²H₂]glucose (150 min) and [3-¹³C]lactate (6 h). Liver glutamine was sampled with phenylacetate and its labelling pattern determined (mass spectrometry) after purification of the glutamine moiety of urinary phenylacetylglutamine. After correction for ¹³CO₂ re-incorporation (control test with NaH¹³CO₃ infusion) this pattern was used to calculate the dilution factor (F) in the hepatic oxaloacetate pool and fluxes through liver Krebs cycle. NIDDM patients had increased lactate turnover rates (16.18 ± 0.92 vs 12.14 ± 0.60 μmol · kg⁻¹· min⁻¹, p < 0.01) and a moderate rise in glucose production (EGP) (15.39 ± 0.87 vs 12.52 ± 0.28 μmol · kg⁻¹· min⁻¹, p = 0.047). Uncorrected contributions of gluconeogenesis to EGP were 31 ± 3 % (control subjects) and 17 ± 2 % (NIDDM patients). F was comparable (1.34 ± 0.02 and 1.39 ± 0.09, respectively) and the corrected percent and absolute contributions of gluconeogenesis were not increased in NIDDM (25 ± 3 % and 3.8 ± 0.5 μmol · kg⁻¹· min⁻¹) compared to control subjects (41 ± 3 % and 5.1 ± 0.4 μmol · kg⁻¹· min⁻¹). The calculated pyruvate carboxylase over pyruvate dehydrogenase activity ratio was comparable (12.1 ± 2.6 vs 11.2 ± 1.4). Lastly hepatic fatty oxidation, as estimated by the model, was not increased in NIDDM (1.8 ± 0.4 vs 1.6 ± 0.1 μmol · kg⁻¹· min⁻¹). In conclusion, in the patients studied we found no evidence of increased hepatic fatty oxidation, or, despite the increased lactate turnover rate, an increased gluconeogenesis. [Diabetologia (1998) 41: 212–220] Received: 4 July 1997 and in revised form: 16 September 1997     

The effect of portal and peripheral insulin delivery on carbohydrate and lipid metabolism in a miniature pig model of human IDDM

Summary A pig model of insulin-dependent diabetes was used to examine the importance of the portal-systemic insulin gradient for whole-body metabolic control. Six pigs had jugular vein, portal vein, and carotid artery cannulae implanted before being made diabetic (150 mg kg^{− 1} streptozotocin). Each animal received 4 weeks of portal and 4 weeks of peripheral insulin delivery in random order. The blood glucose target range was 5–10 mmol · l^{− 1}, and serum fructosamine and fasting and postprandial blood glucose concentrations were not different between peripheral and portal insulin infusion. Insulin requirement was not different between the 4 week infusion periods, but fasting peripheral insulin levels after peripheral delivery (124 ± 16 (mean ± SEM) pmol · l^{− 1}) were significantly higher (p < 0.05) than in portally infused (73.8 ± 5.4 pmol · l^{− 1}) or pre-diabetic control animals (68.4 ± 3.6 pmol · l^{− 1}). Basal hepatic glucose output was also higher (p < 0.05) in peripherally (4.2 ± 0.4 mg · kg^{− 1}· min^{− 1}) than in portally infused animals (2.9 ± 0.4 mg · kg^{− 1}· min^{− 1}) or controls (3.0 ± 0.3 mg · kg^{− 1}· min^{− 1}). Clamp glucose metabolic clearance rate was, however, not different between the peripheral and portal insulin delivery routes (8.1 ± 1.0 vs 9.0 ± 0.7 ml · kg^{− 1}· min^{− 1}), although both were significantly lower (p < 0.05) than that measured in prediabetic control animals (11.7 ± 1.0 ml · kg^{− 1}· min^{− 1}). Lipid profiles and subfractions were similar in all three groups. It is concluded that the portal route of delivery is superior to the peripheral in maintaining more appropriate insulin concentrations and control of hepatic glucose output, although in the absence of euglycaemia it is still associated with significant metabolic abnormalities. [Diabetologia (1997) 40: 1125–1134] Received: 25 February 1997 and in revised form: 23 May 1997     

Renal function and insulin sensitivity during simvastatin treatment in Type 2 (non-insulin-dependent) diabetic patients with microalbuminuria

Summary The effect of simvastatin (10–20 mg/day) on kidney function, urinary albumin excretion rate and insulin sensitivity was evaluated in 18 Type 2 (non-insulin-dependent) diabetic patients with microalbuminuria and moderate hypercholesterolaemia (total cholesterol ≥5.5 mmol·l⁻¹). In a double-blind, randomized and placebo-controlled design treatment with simvastatin (n=8) for 36 weeks significantly reduced total cholesterol (6.7±0.3 vs 5.1 mmol·l⁻¹ (p<0.01)), LDL-cholesterol (4.4±0.3 vs 2.9±0.2 mmol·l⁻¹ (p<0.001)) and apolipoprotein B (1.05±0.04 vs 0.77±0.02 mmol·l⁻¹ (p<0.01)) levels as compared to placebo (n=10). Both glomerular filtration rate (mean±SEM) (simvastatin: 96.6±8.0 vs 96.0±5.7 ml·min⁻¹·1.73 m⁻², placebo: 97.1±6.7 vs 88.8±6.0 ml·min⁻¹·1.73 m⁻²) (NS) and urinary albumin excretion rate (geometric mean x/÷ antilog SEM) (simvastatin: 18.4x/÷1.3vs 16.2 x/÷1.2 μg·min⁻¹, placebo 33.1 x/÷ 1.3 vs 42.7 x/÷ 1.3 μg·min⁻¹)(NS) were unchanged during the study. A euglycaemic hyperinsulinaemic clamp was performed at baseline and after 18 weeks in seven simvastatin-and nine placebo-treated patients. Isotopically determined basal and insulin-stimulated glucose disposal was similarly reduced before and during therapy in both the simvastatin (2.0±0.1 vs 1.9±0.1 (NS) and 3.1±0.6 vs 3.1±0.7 mg·kg⁻¹·min⁻¹ (NS)) and the placebo group (1.9±0.1 vs 1.8±0.1 (NS) and 4.1±0.6 vs 3.8±0.2 mg·kg⁻¹·min⁻¹ (NS)). No different was observed in glucose storage or glucose and lipid oxidation before and after treatment. Further, the suppression of hepatic glucose production during hyperinsulinaemia was not influenced by simvastatin (−0.7±0.8 vs −0.7±0.5 mg·kg⁻¹·min⁻¹ (NS)). In conclusion, despite marked improvement in the dyslipidaemia simvastatin had no impact on kidney function or urinary albumin excretion rate and did not reduce insulin resistance in these microalbuminuric and moderately hypercholesterolaemic Type 2 diabetic patients.     

Evidence that insulin can directly inhibit hepatic glucose production

Summary In order to evaluate the role of portal insulin in the modulation of hepatic glucose production (HGP), measurements of plasma glucose and insulin concentrations and both HGP and peripheral glucose disappearance rates were made following an infusion of a dose of tolbutamide (0.74 mg · m⁻²· min⁻¹) in healthy volunteers that does not result in an increase in peripheral vein insulin concentrations or metabolic clearance rate of glucose. The results showed that the infusion of such a dose of tolbutamide was associated with a significant and rapid decline in both HGP (from 9.0 ± 0.5 to 7.7 ± 0.5 μmol · kg⁻¹· min⁻¹ or Δ = − 13.8 ± 4.5 %; p < 0.001 compared to saline) and plasma glucose concentration (from 5.1 ± 0.2 to 4.4 ± 0.1 mmol/l or Δ = − 13.0 ± 2.1 %; p < 0.01 compared to saline). Since neither HGP nor fasting glucose fell when tolbutamide-stimulated insulin secretion was inhibited by the concurrent administration of somatostatin, it indicated that tolbutamide by itself, does not directly inhibit HGP. Finally, HGP fell by 26.3 ± 6.0 % at 10 min after a dose of tolbutamide that elevated both peripheral and portal insulin concentrations, at a time when HGP had barely increased (Δ = + 6.9 ± 5.3 %). The difference in the magnitude of the two responses was statistically significant (p < 0.03), providing further support for the view that insulin can directly inhibit HGP, independent of any change in flow of substrates from periphery to liver. [Diabetologia (1997) 40: 1300–1306] Received: 8 April 1997 and in revised form: 20 June 1997     

Reduced intrahepatic fat content is associated with increased whole-body lipid oxidation in patients with type 1 diabetes

Aims/hypothesis Insulin resistance may be associated with ectopic fat accumulation potentially determined by reduced lipid oxidation. In patients with type 1 diabetes peripheral insulin resistance is associated with higher intramyocellular lipid content. We assessed whether these patients are also characterised by intrahepatic fat accumulation and abnormal fat oxidation. Methods Nineteen patients with type 1 diabetes (6 women, 13 men, age 35±7 years, BMI 23±3 kg/m², HbA₁c 8.7±1.4%) and 19 healthy matched individuals were studied by (1) euglycaemic–hyperinsulinaemic clamp combined with [6,6−²H₂]glucose infusion to assess whole–body glucose metabolism; (2) indirect calorimetry to assess glucose and lipid oxidation; and (3) localised ¹H−magnetic resonance spectroscopy of the liver to assess intrahepatic fat content. Results Patients with type 1 diabetes showed a reduced insulin−stimulated metabolic clearance rate of glucose (4.3±1.3 ml kg⁻¹ min⁻¹) in comparison with normal subjects (6.0±1.6 ml kg⁻¹ min⁻¹; p<0.001). Endogenous glucose production was higher in diabetic patients (p=0.001) and its suppression was impaired during insulin administration (66±30 vs 92±8%; p=0.047) in comparison with normal subjects. Plasma glucagon concentrations were not different between groups. The estimated hepatic insulin concentration was lower in diabetic patients than in normal subjects (p<0.05), as was the intrahepatic fat content (1.5±0.7% and 2.2±1.0% respectively; p<0.03), the latter in association with a reduced respiratory quotient (0.74±0.05 vs 0.84±0.06; p=0.01) and increased fasting lipid oxidation (1.5±0.5 vs 0.8±0.4 mg kg⁻¹ min⁻¹; p<0.01). Conclusions/interpretation In patients with type 1 diabetes, insulin resistance was not associated with increased intrahepatic fat accumulation. In fact, diabetic patients had reduced intrahepatic fat content, which was associated with increased fasting lipid oxidation. The unbalanced hepatic glucagon and insulin concentrations affecting patients with type 1 diabetes may be involved in this abnormality of intrahepatic lipid metabolism.     

Evidence for direct action of alloxan to induce insulin resistance at the cellular level

Summary To determine whether long-term insulin deficiency alters insulin movement across the endothelium, plasma and lymph dynamics were assessed in dogs after alloxan (50 mg/kg; n = 8) or saline injection (n = 6). Glucose tolerance (K_G) and acute insulin response were assessed by glucose injection before and 18 days after treatment. Two days later, hyperglycaemic (16.7 mmol/l) hyperinsulinaemic (60 pmol · min⁻¹· kg⁻¹) glucose clamps were carried out in a subset of dogs (n = 5 for each group), with simultaneous sampling of arterial blood and hindlimb lymph. Alloxan induced fasting hyperglycaemia (12.9 ± 2.3 vs 5.7 ± 0.2 mmol/l; p = 0.018 vs pre-treatment) and variable insulinopenia (62 ± 14 vs 107 ± 19 pmol/l; p = 0.079). The acute insulin response, however, was suppressed by alloxan (integrated insulin from 0–10 min: 155 ± 113 vs 2745 ± 541 pmol · l⁻¹· 10 min⁻¹; p = 0.0027), resulting in pronounced glucose intolerance (K_G: 0.99 ± 0.19 vs 3.14 ± 0.38 min⁻¹; p = 0.0002 vs dogs treated with saline). During clamps, steady state arterial insulin was higher in dogs treated with alloxan (688 ± 60 vs 502 ± 38 pmol/l; p = 0.023) due to a 25 % reduction in insulin clearance (p = 0.045). Lymph insulin concentrations were also raised (361 ± 15 vs 266 ± 27 pmol/l; p = 0.023), such that the lymph to arterial ratio was unchanged by alloxan (0.539 ± 0.022 vs 0.533 ± 0.033; p = 0.87). Despite higher lymph insulin, glucose uptake (R_d) was significantly diminished after injection of alloxan (45.4 ± 2.5 vs 64.3 ± 6.5 μmol · min⁻¹· kg⁻¹; p = 0.042). This was reflected in resistance of target tissues to the lymph insulin signal (ΔR_d/Δlymph insulin: 3.389 ± 1.093 vs 11.635 ± 2.057 · 10⁻⁶· l · min⁻¹· kg^–1· pmol⁻¹· l⁻¹; p = 0.012) which correlated strongly with the K_G (r = 0.86; p = 0.0001). In conclusion, alloxan induces insulinopenic diabetes, with glucose intolerance and insulin resistance at the target tissue level. Alloxan treatment, however, does not alter lymph insulin kinetics, indicating that insulin resistance of Type 1 (insulin-dependent) diabetes mellitus reflects direct impairment at the cellular level. [Diabetologia (1998) 41: 1327–1336] Received: 3 November 1997 and in final revised form: 2 June 1998     

Hyperlipidaemia is associated with increased insulin-mediated glucose metabolism, reduced fatty acid metabolism and normal blood pressure in transgenic mice overexpressing human apolipoprotein C1

Aims/hypothesis. Insulin resistance for glucose metabolism is associated with hyperlipidaemia and high blood pressure. In this study we investigated the effect of primary hyperlipidaemia on basal and insulin-mediated glucose and on non-esterified fatty acid (NEFA) metabolism and mean arterial pressure in hyperlipidaemic transgenic mice overexpressing apolipoprotein C1 (APOC1). Previous studies have shown that APOC1 transgenic mice develop hyperlipidaemia primarily because of an impaired hepatic uptake of very low density lipoprotein (VLDL). Methods. Basal and hyperinsulinaemic (6 mU · kg^–1· min^–1), euglycaemic (7 mmol/l) clamps with 3-³H-glucose or 9,10-³H-palmitic acid infusions and in situ freeze clamped tissue collection were carried out. Results. The APOC1 mice showed increased basal plasma cholesterol, triglyceride, NEFA and decreased glucose concentrations compared with wild-type mice (7.0 ± 1.2 vs 1.6 ± 0.1, 9.1 ± 2.3 vs 0.6 ± 0.1, 1.9 ± 0.2 vs 0.9 ± 0.1 and 7.0 ± 1.0 vs 10.0 ± 1.1 mmol/l, respectively, p < 0.05). Basal whole body glucose clearance was increased twofold in APOC1 mice compared with wild-type mice (18 ± 2 vs 10 ± 1 ml · kg^–1· min^–1, p < 0.05). Insulin-mediated whole body glucose uptake, glycolysis (generation of ³H₂O) and glucose storage increased in APOC1 mice compared with wild-type mice (339 ± 28 vs 200 ± 11; 183 ± 39 vs 128 ± 17 and 156 ± 44 vs 72 ± 17 μmol · kg^–1· min^–1, p < 0.05, respectively), corresponding with a twofold to threefold increase in skeletal muscle glycogenesis and de novo lipogenesis from 3-³H-glucose in skeletal muscle and adipose tissue (p < 0.05). Basal whole body NEFA clearance was decreased threefold in APOC1 mice compared with wild-type mice (98 ± 21 vs 314 ± 88 ml · kg^–1· min^–1, p < 0.05). Insulin-mediated whole body NEFA uptake, NEFA oxidation (generation of ³H₂O) and NEFA storage were lower in APOC1 mice than in wild-type mice (15 ± 3 vs 33 ± 6; 3 ± 2 vs 11 ± 4 and 12 ± 2 vs 22 ± 4 μmol · kg^–1· min^–1, p < 0.05) in the face of higher plasma NEFA concentrations (1.3 ± 0.3 vs 0.5 ± 0.1 mmol/l, p < 0.05), respectively. Mean arterial pressure and heart rate were similar in APOC1 vs wild-type mice (82 ± 4 vs 85 ± 3 mm Hg and 459 ± 14 vs 484 ± 11 beats · min^–1). Conclusions/interpretation. 1) Hyperlipidaemic APOC1 mice show reduced NEFA and increased glucose metabolism under both basal and insulin-mediated conditions, suggesting an intrinsic defect in NEFA metabolism. Primary hyperlipidaemia alone in APOC1 mice does not lead to insulin resistance for glucose metabolism and high blood pressure. [Diabetologia (2001) 44: 437–443] Received: 14 September 2000 and in revised form: 23 November 2000     

Acute blockade by endothelin-1 of haemodynamic insulin action in rats

Aims/hypothesis Plasma levels of endothelin-1 are frequently elevated in patients with hypertension, obesity and type 2 diabetes. We hypothesise that this vasoconstrictor may prevent full perfusion of muscle, thereby limiting delivery of insulin and glucose and contributing to insulin resistance. Materials and methods The acute effects of endothelin-1 on insulin-mediated haemodynamic and metabolic effects were examined in rats in vivo. Endothelin-1 (50 pmol min⁻¹ kg⁻¹ for 2.5 h) was infused alone, or 30 min prior to a hyperinsulinaemic-euglycaemic insulin clamp (10 mU min⁻¹ kg⁻¹ for 2 h). Insulin clamps (10 or 15 mU min⁻¹ kg⁻¹) were performed after 30 min of saline infusion. Results Endothelin-1 infusion alone increased plasma endothelin-1 11-fold (p < 0.05) and blood pressure by 20% (p < 0.05). Endothelin-1 alone had no effect on femoral blood flow, capillary recruitment or glucose uptake, but endothelin-1 with 10 mU min⁻¹ kg⁻¹ insulin caused a decrease in insulin clearance from 0.35 ± 0.6 to 0.19 ± 0.02 ml/min (p = 0.02), resulting in significantly higher plasma insulin levels (10 mU min⁻¹ kg⁻¹ insulin: 2,120 ± 190 pmol/l; endothelin-1 + 10 mU min⁻¹ kg⁻¹ insulin: 4,740 ± 910 pmol/l), equivalent to 15 mU min⁻¹ kg⁻¹ insulin alone (4,920 ± 190 pmol/l). The stimulatory effects of equivalent doses of insulin on femoral blood flow, capillary recruitment and glucose uptake were blocked by endothelin-1. Conclusions/interpretation Endothelin-1 blocks insulin’s haemodynamic effects, particularly capillary recruitment, and is associated with decreased muscle glucose uptake and glucose infusion rate. These findings suggest that elevated endothelin-1 levels may contribute to insulin resistance of muscle by increasing vascular resistance and limiting insulin and glucose delivery.     

Effects of dietary protein restriction on albumin and fibrinogen synthesis in macroalbuminuric type 2 diabetic patients

Aims/hypothesis Diabetic nephropathy is associated with hypoalbuminaemia and hyperfibrinogenaemia. A low-protein diet has been recommended in patients with diabetic nephropathy, but its effects on albumin and fibrinogen synthesis are unknown. Methods We compared the effects of a normal (NPD; 1.38 ± 0.08 g kg⁻¹ day⁻¹) or low (LPD; 0.81 ± 0.04 g kg⁻¹ day⁻¹) -protein diet on endogenous leucine flux (ELF), albumin and fibrinogen synthesis (l-[5,5,5,-²H₃]leucine infusion), and markers of inflammation in nine type 2 diabetic patients with macroalbuminuria. Six healthy participants on NPD served as control participants. Results In comparison with healthy participants, type 2 diabetic patients on an NPD had similar ELF, reduced serum albumin (38 ± 1.1 vs 42 ± 0.8 g/l; p < 0.05), similar fractional synthesis rates (FSR) and absolute synthesis rates (ASR) of albumin, and both increased plasma fibrinogen concentration [10.7 ± 0.6 vs 7.2 ± 0.5 μmol/l (3.64 ± 0.22 vs 2.45 ± 0.18 g/l); p < 0.05] and fibrinogen ASR [11.03 ± 1.17 vs 6.0 ± 1.8 μmol 1.73 m⁻² day⁻¹ (3.7 ± 0.4 vs 1.9 ± 0.3 g 1.73 m⁻² day⁻¹); p < 0.01]. After LPD, type 2 diabetic patients had the following changes in comparison with NPD: reduced proteinuria (2.74 ± 0.4 vs 4.51 ± 0.8 g/day; p < 0.05), ELF (1.93 ± 0.08 vs 2.11 ± 0.08 μmol kg⁻¹ min⁻¹; p < 0.05) and total fibrinogen pool; increased serum albumin (42 ± 1 vs 38 ± 1 g/l; p < 0.01) and albumin ASR (14.1 ± 1 vs 9.9 ± 1 g 1.73 m⁻² day⁻¹; p < 0.05); and reduced plasma IL-6 levels, which were correlated with albumin ASR (r = −0.749; p < 0.05). Conclusions/interpretation LPD in type 2 diabetic patients with diabetic nephropathy reduces low-grade inflammatory state, proteinuria, albuminuria, whole-body proteolysis and ASR of fibrinogen, while increasing albumin FSR, ASR and serum concentration. ISRCTN ID no: CCT-NAPN-16911     

Contrasting action of short- and long-term adrenaline infusion on dog skeletal muscle glucose metabolism

Summary There are important differences between the short- and long-term effects of adrenaline on determinants of glucose tolerance. To assess this metabolic adaptation at tissue level, the present study examined the effect of acute and prolonged in vivo elevation of adrenaline on glycogen metabolism and glycolysis in skeletal muscle. Adrenaline (50 ng · kg⁻¹ · min⁻¹) was infused for 2 h or 74 h and the results compared with 1 h 0.9% NaCl infusion in six trained dogs. Muscle glycogen content was reduced by long-term adrenaline (161 ± 17 vs NaCl 250 ± 24 μmol/g dry weight;p < 0.05) but not short-term adrenaline (233 ± 21) indicating a sustained effect of adrenaline on glycogen metabolism. Acutely, glycogen synthase I was reduced (short-term adrenaline 12 ± 6 vs NaC122 ± 7μmol glycosyl units · g⁻¹ · min⁻¹;p < 0.05) but returned to normal with prolonged adrenaline infusion (20 ± 5). In contrast, K_m for glycogen phosphorylasea was not changed acutely (short-term adrenaline 31 ± 6 vs NaCl 27 ± 7 mmol/1 inorganic phosphate) but was reduced during long-term infusion (19 ± 4;p < 0.05 vs short-term adrenaline). Thus, with short- and long-term adrenaline infusion, there were different enzyme changes, although likely to promote glycogenolysis in both cases. In the glycolytic pathway the substrates glucose 6-phosphate and fructose 6-phosphate did not change significantly and hexokinase was not inhibited. Acutely, phosphofructokinase had reduced V_max (short-term adrenaline 34 ± 6 vs NaCl 44 ± 5 U/g; p < 0.05) but was still above the maximal operating rate in vivo. With prolonged adrenaline infusion, the K_m for phosphofructokinase was reduced (long-term adrenaline 0.32 ± 0.03 vs NaCl 0.44 ± 0.07 mmol/l fructose 6-phosphate;p < 0.05). In this situation of relatively low glycolytic flux, the sustained glycogenolytic effect of prolonged adrenaline infusion mediated by increased glycogen phosphorylase a ctivity occurs without a significant accumulation of hexose monophosphates or impairment of glycolysis.     

Insulin-induced vasodilatation and endothelial function in obesity/insulin resistance. Effects of troglitazone

Summary Insulin resistance is associated with a decreased vasodilator response to insulin. Because insulin's vasodilator effect is nitric oxide dependent, this impairment may reflect endothelial dysfunction. Troglitazone, an insulin-sensitiser, might thus improve insulin-dependent and/or endothelium-dependent vascular function in insulin resistant obese subjects. For 8 weeks, fifteen obese subjects were treated with either 400 mg troglitazone once daily or placebo, in a randomised, double-blind, cross-over design. At the end of each treatment period, we measured forearm vasodilator responses (plethysmography) to intra-arterial administered acetylcholine and sodium nitroprusside; insulin sensitivity and insulin-induced vascular and neurohumoral responses (clamp); vasoconstrictor responses to N ^G-monomethyl-L-arginine (L-NMMA) during hyperinsulinaemia; and ambulatory 24-h blood pressure (ABPM). Baseline data (placebo) of obese subjects were compared with those obtained in lean control subjects. Obese subjects were insulin resistant compared with leans (whole-body glucose uptake: 26.8 ± 3.0 vs. 53.9 ± 4.3 μmol · kg^–1· min^–1, p < 0.001). Troglitazone improved whole-body glucose uptake (to 31.9 ± 3.3 μmol · kg^–1· min^–1, p = 0.028), and forearm glucose uptake (from 1.09 ± 0.54 to 2.31 ± 0.69 μmol · dL^–1· min^–1, p = 0.006). Insulin-induced vasodilatation was blunted in obese subjects (percent increase in forearm blood flow (FBF) in lean 66.5 ± 23.0 %, vs. 10.1 ± 11.3 % in obese, p = 0.04), but did not improve during troglitazone. Vascular responses to acetylcholine, sodium nitroprusside and L-NMMA did not differ between the obese and lean group, nor between both treatment periods in the obese individuals. In conclusion, in insulin resistant obese subjects, endothelial vascular function is normal despite impaired vasodilator responses to insulin. Troglitazone improved insulin sensitivity but it had no effects on endothelium-dependent and -independent vascular responses. These data do not support an association between insulin resistance and endothelial function. [Diabetologia (1998) 41: 569–576] Received: 19 September 1997 and in revised form: 22 December 1997     

Role of islet amyloid polypeptide secretion in insulin-resistant humans

Summary Although it is generally accepted that islet amyloid polypeptide is cosecreted with insulin, relatively few data on its kinetics are available. We therefore studied the dynamics of islet amyloid polypeptide release following oral and frequently sampled intravenous glucose tolerance tests in comparison to insulin and C-peptide using mathematical model techniques in 14 control subjects, 10 obese and 11 hypertensive patients. The fractional clearance rate of islet amyloid polypeptide (0.034±0.004 min^–1 in control subjects, 0.058±0.008 in the obese and 0.050±0.008 in the hypertensive patients) was significantly different (p<0.01) in each group compared with that of insulin (0.14±0.03 min^–1) and similar to that of C-peptide (0.061±0.007 min^–1), at least in the insulin-resistant subjects. Based on the insulin sensitivity index derived from the minimal model analysis of intravenous glucose tolerance test data, both the hypertensive (2.4±0.4 min^–1/(μU/ml); p<0.0005) and the obese (2.7±0.5; p<0.001) patients demonstrated severe insulin resistance compared to control subjects (8.1±1.3). Marked insulin hypersecretion was found in the hypertensive (57.6±5.2 nmol·l^–1 in 180 min; p<0.001) and obese (60.8±10.1; p<0.003) patients in comparison with control subjects (32.4±3.2). The release of islet amyloid polypeptide was significantly higher in the hypertensive (83.1±16.6 pmol/l in 180 min; p<0.02) and obese (78.6±13.1; p<0.005) patients than in control subjects (40.5±6.4). No correlation was found between islet amyloid polypeptide release and the insulin sensitivity index in any group. We conclude that, due to a significantly slower clearance of islet amyloid polypeptide in comparison to insulin, reliance on molar ratios between these two peptides might be misleading in the interpretation of islet amyloid polypeptide secretion especially under non-steady-state conditions. [Diabetologia (1994) 37: 188–194] Received: 10 June 1993 and in revised form: 20 August 1993     

Exercise under hyperinsulinaemic conditions increases whole-body glucose disposal without affecting muscle glycogen utilisation in type 1 diabetes

Aims/hypothesis We examined whole-body and muscle metabolism in patients with type 1 diabetes during moderate exercise at differing circulating insulin concentrations. Methods Eight men (mean ± SEM age 36.4 ± 1.5 years; diabetes duration 11.3 ± 1.4 years; BMI 24.6 ± 0.7 kg/m²; HbA_1c 7.9 ± 0.2% and VO₂ peak 44.5 ± 1.2 ml kg⁻¹ min⁻¹) with type 1 diabetes were studied on two occasions at rest (2 h) and during 45 min of cycling at 60% maximum VO₂ with insulin infused at the rate of either 15 (LO study) or 50 (HI) mU m⁻² min⁻¹ and blood glucose clamped at 8 mmol/l. Indirect calorimetry, insulin-glucose clamps and thigh muscle biopsies were employed to measure whole-body energy and muscle metabolism. Results Fat oxidation contributed 15 and 23% to total energy expenditure during exercise in the HI and LO studies, respectively. The respective carbohydrate (CHO) oxidation rates were 31.7 ± 2.7 and 27.8 ± 1.9 mg kg⁻¹ min⁻¹ (p < 0.05). Exogenous glucose utilisation rate during exercise was substantially greater (p < 0.001) in the HI study (18.4 ± 2.1 mg kg⁻¹ min⁻¹) than in the LO study (6.9 ± 1.2 mg kg⁻¹ min⁻¹). Muscle glycogen content fell by ∼40% during exercise in both trials. Muscle glycogen utilisation, muscle intermediary metabolism, and phosphorylation of protein kinase B/Akt, glycogen synthase kinase 3α/β and extracellular signal-regulated protein kinase 1 and 2 proteins were no different between interventions. Conclusions/interpretation In patients with type 1 diabetes, exercise under peak therapeutic insulin concentrations increases exogenous glucose utilisation but does not spare muscle glycogen utilisation. A disproportionate increase in exogenous glucose utilisation relative to the increase in CHO oxidation suggests an increase in glucose flux through non-oxidative pathways. Chokkalingam and Tsintzas are joint first authors.     

Copper addition prevents the inhibitory effects of interleukin 1-β on rat pancreatic islets

Summary Since copper [Cu(II)] is a necessary cofactor for both intra-mitochondrial enzymes involved in energy production and hydroxyl scavenger enzymes, two hypothesised mechanisms for action of interleukin-Iβ (IL-1β), we studied whether CU(II) addition could prevent the inhibitory effect of IL-1β on insulin release and glucose oxidation in rat pancreatic islets. Islets were incubated with or without 50 U/ml IL-1β, in the presence or absence of various concentrations of Cu(II)-GHL (Cu(II) complexed with glycyl-l-histidyl-l-lysine, a tripeptide known to enhance copper uptake into cultured cells). CuSO₄ (1–1000 ng/ml) was used as a control for Cu(II) effect when present as an inorganic salt. At the end of the incubation period, insulin secretion was evaluated in the presence of either 2.8 mmol/l (basal insulin secretion) or 16.7 mmol/l glucose (glucose-induced release). In control islets basal insulin secretion was 92.0±11.4 pg · islet⁻¹ h⁻¹ (mean ± SEM,n=7) and glucose-induced release was 2824.0±249.0 pg · islet⁻¹ h⁻¹. In islets pre-exposed to 50 U/ml IL-1β, basal insulin release was not significantly affected but glucose-induced insulin release was greatly reduced (841.2±76.9,n=7,p<0.005). In islets incubated with IL-1β and Cu-GHL (0.4 μmol/l, maximal effect) basal secretion was 119.0±13.1 pg · islet⁻¹ h⁻¹ and glucose-induced release was 2797.2±242.2, (n=7,p<0.01 in respect to islets exposed to IL-1β alone). In contrast to data obtained with Cu(II)-GHL, increasing concentrations of CuSO4 (up to 10 μmol/l) did not influence the inhibitory effect of IL-1β on glucose-stimulated insulin release. Glucose oxidation (in the presence of 16.7 mmol/l glucose) was 31.5±2.4 pmol · islet⁻¹·90min⁻¹ in control islets and 7.0±0.9 (p<0.01) in IL-1β-exposed islets. In islets exposed to IL-1β and Cu-GHL glucose oxidation was similar to control islets (31.9±1.9). In contrast, Cu-GHL did not prevent the IL-1β-induced increase in nitric oxide production. Nitrite levels were 5±1.7, 26±5 and to 29±4 pmol · islet⁻¹·48 h⁻¹ (mean ± SEM,n=5) in the culture medium from control IL-1β and IL-1β+Cu-GHL exposed islets, respectively. These data indicate that the Cu(II) complexed to GHL is able to prevent the inhibitory effects of IL-1β on insulin secretion and glucose oxidation, but not on NO production. The mechanism of action of Cu-GHL is still unclear, but it might restore the activity of the enzymatic systems inhibited by IL-1β. [Diabetologia (1995) 38∶39–45]     

Differences in protein and energy metabolism following portal versus systemic administration of insulin in diabetic dogs

Aims/hypothesis In non-diabetic people, insulin levels in the liver are two-fold higher than those in the systemic circulation. In contrast, patients with type 1 diabetes have similar hepatic and systemic insulin levels because insulin is administered peripherally. The aim of this study was to compare the effects of systemic (SI) and pre-portal (PI) insulin administration on energy, glucose and protein metabolism in chronic insulin-dependent ketosis-prone diabetic dogs. Materials and methods We applied glucose-controlled insulin infusion, indirect calorimetry and stable isotope and radioisotope techniques to measure energy, protein and glucose metabolism. We maintained near-normoglycaemia at identical levels under both study conditions for 20 h. Results SI was associated with lower oxygen consumption (130±13 vs 161±8 ml/min), CO₂ production (99±10 vs 130±8 ml/min), respiratory quotient (0.76±0.02 vs 0.81±0.01) and energy expenditure (870±90 vs 1089±60 kcal/24 h) (p<0.05 for all differences). PI increased the respiratory quotient from the insulin-deprived state, whereas SI did not. Glucose kinetics were similar for SI and PI, whereas leucine oxidation (36±4 vs 54±5 μmol kg⁻¹ min⁻¹) and the fractional synthesis rates of liver tissue protein (0.68±0.6 vs 0.83±0.07%/h), albumin (0.55±0.06 vs 0.68±0.4%/h), and fibrinogen (1.73±0.23 vs 2.59±0.25%/h) were all lower during SI than PI (p<0.05). Conclusions/interpretation The route of insulin administration did not alter glucose metabolism but did affect protein synthesis in the liver. The potential impact of this altered liver protein metabolism on chronic complications needs careful evaluation. A similar decrease in energy expenditure resulting from systemic insulin administration during tight glycaemic control is a potential cause of weight gain.     

Effect of short-term ACE inhibitor treatment on peripheral insulin sensitivity in obese insulin-resistant subjects

Aims/hypothesis This study was designed to investigate the effect of short-term ACE inhibitor treatment on insulin sensitivity and to examine possible underlying metabolic and haemodynamic effects in obese insulin-resistant subjects.Methods A randomised, double-blind placebo-controlled trial was performed in 18 obese insulin-resistant men (age, 53 ± 2 years; BMI, 32.6 ± 0.8 kg/m²; homeostasis model assessment of insulin resistance, 5.6 ± 0.5; systolic blood pressure [SBP], 140.8 ± 3.2; diastolic blood pressure [DBP], 88.8 ± 1.6 mmHg), who were free of any medication. The aim was to examine the effects of 2 weeks of ACE inhibitor treatment (ramipril, 5 mg/day) on insulin sensitivity, forearm blood flow, substrate fluxes across the forearm, whole-body substrate oxidation and intramuscular triacylglycerol (IMTG) content.Results Ramipril treatment decreased ACE activity compared with placebo (−22.0 ± 1.7 vs 0.2 ± 1.1 U/l, respectively, p < 0.001), resulting in a significantly reduced blood pressure (SBP, −10.8 ± 2.1 vs −2.7 ± 2.0 mmHg, respectively, p = 0.01; DBP, −10.1 ± 1.3 vs −4.2 ± 2.1 mmHg, respectively, p = 0.03). Ramipril treatment had no effect on whole-body insulin-mediated glucose disposal (before: 17.9 ± 2.0, after: 19.1 ± 2.4 μmol kg body weight⁻¹ min⁻¹, p = 0.44), insulin-mediated glucose uptake across the forearm (before: 1.82 ± 0.39, after: 1.92 ± 0.29 μmol 100 ml forearm tissue⁻¹ min⁻¹, p = 0.81) and IMTG content (before: 45.4 ± 18.8, after: 48.8 ± 27.5 μmol/mg dry muscle, p = 0.92). Furthermore, the increase in carbohydrate oxidation (p < 0.001) and forearm blood flow (p < 0.01), and the decrease in fat oxidation (p < 0.001) during insulin stimulation were not significantly different between treatments.Conclusions/interpretation Short-term ramipril treatment adequately reduced ACE activity and blood pressure, but had no significant effects on insulin sensitivity, forearm blood flow, substrate fluxes across the forearm, whole-body substrate oxidation and IMTG content in obese insulin-resistant subjects.     

Dissociation between insulin sensitivity of glucose uptake and endothelial function in normal subjects

Summary Insulin increases limb blood flow in a time- and dose-dependent manner. This effect can be blocked by inhibiting nitric oxide synthesis. These data raise the possibility that insulin resistance is associated with endothelial dysfunction. To examine whether endothelial function and insulin sensitivity are interrelated we quantitated in vivo insulin-stimulated rates of whole body and forearm glucose uptake at a physiological insulin concentration (euglycaemic hyperinsulinaemic clamp, 1 mU · kg^–1· min^–1 insulin infusion for 2 h) and on another occasion, in vivo endothelial function (blood flow response to intrabrachial infusions of sodium nitroprusside, acetylcholine, and N-monomethyl-l-arginine) in 30 normal male subjects. Subjects were divided into an insulin-resistant (IR) and an insulin-sensitive (IS) group based on the median rate of whole body glucose uptake (31 ± 2 vs 48 ± 1 μmol · kg^–1· min^–1, p < 0.001). The IR and IS groups were matched for age, but the IR group had a slightly higher body mass index, percentage of body fat and blood pressure compared to the IS group. The IR group also had diminished insulin-stimulated glucose extraction (p < 0.05) compared to the IS group, while basal and insulin-stimulated forearm blood flow rates were identical. There was no difference between the IR and IS groups in the forearm blood flow response to endothelium-dependent (acetylcholine and N-monomethyl-l-arginine) or -independent (sodium nitroprusside) vasoactive drugs. In conclusion, the ability of insulin to stimulate glucose uptake at physiological insulin concentrations and endothelium-dependent vasodilatation are distinct phenomena and do not necessarily coexist. [Diabetologia (1996) 39: 1477–1482] Received: 31 May 1996 and in revised form: 10 July 1996     

Type I diabetes mellitus does not alter initial splanchnic glucose extraction or hepatic UDP-glucose flux during enteral glucose administration

Aims/hypothesis. Our aim was to determine whether an alteration in splanchnic glucose metabolism could contribute to postprandial hyperglycaemia in people with Type I (insulin-dependent) diabetes mellitus. Methods. Splanchnic glucose extraction, hepatic glycogen synthesis and endogenous glucose production were compared in 8 Type I diabetic patients and in 11 control subjects. Endogenous hormone secretion was inhibited with somatostatin while insulin ( ∼ 550 pmol/l) and glucagon ( ∼ 130 ng/l) concentrations were matched with exogenous hormone infusions. Glucose containing [3-³H] glucose was infused into the duodenum at a rate of 20 μmol · kg^-1· min^-1. Plasma glucose concentrations were maintained at about 8.5 mmol/l in both groups by means of a separate variable intravenous glucose infusion. Results. Initial splanchnic glucose uptake, calculated by subtracting the systemic rate of appearance of [3-³H] glucose from the rate of infusion of [3-³H] glucose into the duodenum, did not differ in the diabetic and non-diabetic patients (4.1 ± 0.8 vs 3.0 ± 1.0 μmol/kg/min). In addition, hepatic glycogen synthesis, measured using the acetaminophen glucuronide method did not differ (10.7 ± 2.4 vs 10.1 ± 2.7 μmol · kg^-1· min^-1). On the other hand, suppression of endogenous glucose production, measured by an intravenous infusion of [6,6-²H₂] glucose, was greater (p < 0.05) in the diabetic than in the non-diabetic subjects (1.7 ± 1.6 vs 5.8 ± 1.9 μmol · kg^-1· min^-1). Conclusion/interpretation. When glucose, insulin and glucagon concentrations are matched in individuals with relatively good chronic glycaemic control, Type I diabetes does not alter initial splanchnic glucose uptake of enterally delivered glucose or hepatic glycogen synthesis. Alterations in splanchnic glucose metabolism are not likely to contribute to postprandial hyperglycaemia in people with well controlled Type I diabetes. [Diabetologia (2001) 44: 729–737] Received: 10 November 2000 and in revised form: 22 January 2001