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     

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     

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     

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     

Rat C peptide I and II stimulate glucose utilization in STZ-induced diabetic rats

Aims. To study the effects of physiological concentrations of rat proinsulin C peptide I and II, respectively, on whole body glucose utilization in streptozotocin diabetic and healthy rats. Methods. A sequential insulin clamp procedure was used (insulin infusion rates 3.0 and 30.0 mU · kg^–1· min^–1) in awake animals. C-peptide infusion rates were 0.05 and 0.5 nmol · kg^–1· min^–1. Blood glucose was clamped at 7.7 ± 0.3 mmol/l in the diabetic rats and at 3.9 ± 0.1 mmol/l in the healthy rats. Results. In diabetic rats infused at lower rates of C peptide and insulin, glucose utilization increased by 79–90 % (p < 0.001) compared with diabetic animals infused with saline and insulin. Increasing the rate of C-peptide infusion tenfold did not elicit a statistically significant further increase in glucose utilization. C peptide I and II exerted similar effects. The metabolic clearance rate for glucose in the diabetic animals infused with C peptide was not different from that of the healthy rats. During high-dose insulin infusion (30.0 mU · kg^–1· min^–1) glucose utilization increased considerably and no statistically significant C-peptide effects were observed. About 85 % of the increase in glucose utilization induced by C peptide could be blocked by treatment with N-monomethyl-l-arginine. Conclusions/interpretation. Physiological concentrations of homologous C peptide stimulate whole body glucose utilization in diabetic but not in healthy rats. C peptide I and II elicit similar effects. The influence of C peptide on glucose utilization may be mediated by nitric oxide. [Diabetologia (1999) 42: 958–964] Received: 8 January 1999 and in final revised form: 20 April 1999     

Glucoregulatory function of glucagon in hypo-, eu- and hyperthyroid miniature pigs

Summary The glucoregulatory function of glucagon was investigated in hypo-, eu- and hyperthyroid miniature pigs. Infusion glucagon, (3 ng x kg body weight⁻¹ · min⁻¹) transiently increased blood glucose (p<0.01) and hepatic glucose production (p<0.01) in euthyroidism, but was without effect in hyperthyroidism. Infusing glucagon plus somatostatin (2 ng x kg body weight⁻¹ · min⁻¹ and 0.2 μg x kg body weight⁻¹ · min⁻¹) transiently increased blood glucose (Δ 3.0 to 4.3 mmol/l) and hepatic glucose production (Δ 3.3 to 7.7 umol x kg body weight⁻¹ · min⁻¹) in all thyroid states, the effect was less pronounced in hyperthyroid pigs. By contrast, hypoglucagonaemia (74 to 107 pg/ml) at basal insulin (28 to 35 μU/ml) provoked hypoglycaemia (1.4 to 2.2 mmol/l) and a fall in glucose production (Δ 4.7 to 8.3 umol x kg body weight⁻¹ · min⁻¹), which was independent of the thyroid state; the effect was most pronounced in hyperthyroidism (p<0.01). Hepatic glycogen content, arterial gluconeogenic precursor concentrations as well as the glycaemic response (Δ 0.60 mmol/l) to alanine infusion (23 umol x kg body weight⁻¹ · min⁻¹) were all unaffected by hyperthyroidism. We conclude that moderate experimental hyperthyroidism reduces glucagon action due to reduced glycogen mobilisation. This may in part result from increased insulin sensitivity.     

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     

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.     

Insulin resistance is associated with high plasma ouabain-like immunoreactivity concentration in NIDDM

Summary The aim of the present study was to elucidate the pathophysiologic significance of circulating ouabain as a link between insulin resistance (IR) and hypertension (HT) in NIDDM. Euglycaemic (4.5 mmol/l) hyperinsulinaemic (360–580 pmol/l) clamping was performed using an artificial endocrine pancreas. Plasma ouabain-like immunoreactivity (OLI) was determined by radioimmunoassay using a highly specific antibody to ouabain. HT was defined as systolic blood pressure > 140 mm Hg and/or diastolic > 90 mm Hg or being treated with antihypertensive agents. The values (mean ± SEM) of glucose infusion rate (GIR) and plasma OLI were compared among the four groups classified using IR and HT as factors. Group I (IR−/HT−, n = 15):GIR 7.20 ± 0.36 mg · kg⁻¹· min⁻¹, OLI 130.8 ± 20.9 pmol/l, which was not different from that in eight normal control subjects (7.69 ± 0.40 mg · kg⁻¹· min⁻¹ and 142.6 ± 32.3 pmol/l, respectively); Group II (IR−/HT+, n = 13): 5.89 ± 0.36 mg · kg⁻¹· min⁻¹, 172.5 ± 35.0 pmol/l; Group III (IR+/HT−, n = 14) 1.91 ± 0.28 mg · kg⁻¹· min⁻¹, 576.6 ± 161.5 pmol/l (p < 0.01 vs Group I and II); Group IV (IR+/HT+, n = 15) 1.79 ± 0.22 mg · kg⁻¹· min⁻¹, 703.1 ± 170.1 pmol/l (p < 0.01 vs Group I and II), respectively. Six of 57 NIDDM patients studied exhibited very high (> 1500 pmol/l) plasma OLI concentrations, showed marked insulin resistance and were all hypertensive. When analysed as a whole, plasma OLI was negatively correlated with GIR (p < 0.001), but was not correlated with arterial blood pressure. These results demonstrate that plasma concentration of OLI is closely associated with the severity of IR but not with blood pressure elevation. It is, however, possible that in some fraction of NIDDM patients with insulin resistance, the elevation of blood pressure may be causally related to circulating OLI. [Diabetologia (1995) 38: 792–797] Received: 13 September 1994 and in revised form: 9 December 1994     

Near normalisation of blood glucose improves the potentiating effect of GLP-1 on glucose-induced insulin secretion in patients with type 2 diabetes

Aims/hypothesis The ability of glucagon-like peptide-1 (GLP-1) to enhance beta cell responsiveness to i.v. glucose is impaired in patients with type 2 diabetes mellitus compared with healthy individuals. We investigated whether 4 weeks of near normalisation of blood glucose (BG) improves the potentiation of glucose-stimulated insulin secretion by GLP-1. Methods Nine obese patients with type 2 diabetes and inadequate glycaemic control (HbA_1c 8.0 ± 0.4%) were investigated before and after 4 weeks of near normalisation of BG using insulin treatment (mean diurnal blood glucose 6.4 ± 0.3 mmol/l, HbA_1c 6.6 ± 0.3%). Nine matched healthy participants were also studied. Beta cell function was investigated before and after insulin treatment using stepwise glucose infusions and infusion of saline or GLP-1 (1.0 pmol kg⁻¹ min⁻¹), resulting in supraphysiological total GLP-1 concentrations of approximately 200 pmol/l. The responsiveness to glucose or glucose+GLP-1 was expressed as the slope of the linear regression line relating insulin secretion rate (ISR) and plasma glucose concentration (pmol kg⁻¹ min⁻¹ [mmol/l]⁻¹). Results In the diabetic participants, the slopes during glucose+saline infusion did not differ before and after insulin treatment (0.33 ± 0.07 and 0.39 ± 0.04, respectively; p = NS). In contrast, near normalisation of blood glucose improved beta cell sensitivity to glucose during glucose+GLP-1 infusion (1.27 ± 0.2 before vs 1.73 ± 0.31 after; p < 0.01). In the healthy participants, the slopes during the glucose+saline and glucose+GLP-1 infusions were 1.01 ± 0.14 and 4.79 ± 0.53, respectively. Conclusions/interpretation A supraphysiological dose of GLP-1 enhances beta cell responses to glucose in patients with type 2 diabetes, and 4 weeks of near normalisation of blood glucose further improves this effect. ClinicalTrials.gov ID no.: NCT00612625     

The effect of metabolic control on leucine metabolism in Type 1 (insulin-dependent) diabetic patients

Summary Leucine production rate, metabolic clearance rate and oxidation rate were measured in 10 Type 1 (insulin-dependent) diabetic patients after (1) 24 h insulin withdrawal, (2) conventional insulin therapy and (3) an overnight insulin infusion to maintain normoglycaemia, and in 10 control subjects. In the insulin-withdrawn patients, leucine concentration (259 ± 17 μmol/1), production rate (2.65 ± 0.29 p mol·min⁻¹ kg⁻¹) and oxidation rate (0.69 ± 0.10 μmol · min⁻¹ · kg⁻¹) were significantly greater (p < 0.001;p < 0.05;p < 0.005 respectively) than corresponding values in control subjects (127±6; 1.81 ± 0.12; 0.19 ± 0.02). Following conventional insulin therapy, leucine concentration (162 ± 12 μmol/1) and oxidation rate (0.43 ± 0.05 μmol · min⁻¹ · kg⁻¹) were lower than after insulin withdrawal but were still significantly greater than in control subjects (p<0.05;p<0.005). Although leucine concentration, production rate and metabolic clearance rate were normal after an overnight insulin infusion, leucine oxidation rate was still greater than normal (0.34 ± 0.06 μmol · min⁻¹ kg⁻¹;p<0.05). These results suggest that increased leucine concentration in insulin deficiency is due to elevated leucine production rate caused by increased proteolysis, and that leucine concentration is restored to normal by insulin treatment.     

Assessment of insulin sensitivity and secretion with the labelled intravenous glucose tolerance test: improved modelling analysis

Summary A new modelling analysis was developed to assess insulin sensitivity with a tracer-modified intravenous glucose tolerance test (IVGTT). IVGTTs were performed in 5 normal (NGT) and 7 non-insulin-dependent diabetic (NIDDM) subjects. A 300 mg/kg glucose bolus containing [6,6-²H₂]glucose was given at time 0. After 20 min, insulin was infused for 5 min (NGT, 0.03; NIDDM, 0.05 U/kg). Concentrations of tracer, glucose, insulin and C-peptide were measured for 240 min. A circulatory model for glucose kinetics was used. Glucose clearance was assumed to depend linearly on plasma insulin concentration delayed. Model parameters were: basal glucose clearance (Cl_b), glucose clearance at 600 pmol/l insulin concentration (Cl₆₀₀), basal glucose production (P_b), basal insulin sensitivity index (BSI = Cl_b/basal insulin concentration); incremental insulin sensitivity index (ISI = slope of the relationship between insulin concentration and glucose clearance). Insulin secretion was calculated by deconvolution of C-peptide data. Indices of basal pancreatic sensitivity (PSI_b) and first (PSI₁) and second-phase (PSI₂) sensitivity were calculated by normalizing insulin secretion to the prevailing glucose levels. Diabetic subjects were found to be insulin resistant (BSI: 2.3 ± 0.6 vs 0.76 ± 0.18 ml · min^–1· m^–2· pmol/l^–1, p < 0.02; ISI: 0.40 ± 0.06 vs 0.13 ± 0.05 ml · min^–1· m^–2· pmol/l^–1, p < 0.02; Cl₆₀₀: 333 ± 47 vs 137 ± 26 ml · min^–1· m^–2, p < 0.01; NGT vs NIDDM). P_b was not elevated in NIDDM (588 ± 169 vs 606 ± 123 μmol · min^–1· m^–2, NGT vs NIDDM). Hepatic insulin resistance was however present as basal glucose and insulin were higher. PSI₁ was impaired in NIDDM (67 ± 15 vs 12 ± 7 pmol · min^–1· m^–2· mmol/l^–1, p < 0.02; NGT vs NIDDM). In NGT and in a subset of NIDDM subjects (n = 4), PSI_b was inversely correlated with BSI (r = 0.95, p < 0.0001, log transformation). This suggests the existence of a compensatory mechanism that increases pancreatic sensitivity in the presence of insulin resistance, which is normal in some NIDDM subjects and impaired in others. In conclusion, using a simple test the present analysis provides a rich set of parameters characterizing glucose metabolism and insulin secretion, agrees with the literature, and provides some new information on the relationship between insulin sensitivity and secretion. [Diabetologia (1998) 41: 1029–1039] Received: 17 September 1997 and in final revised form: 28 April 1998     

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]     

The set point for maternal glucose homeostasis is lowered during late pregnancy in the rat: the role of the islet beta-cell and liver

Summary The aim of this study was to determine the effects of late pregnancy on the ability of insulin to suppress maternal hepatic glucose production in the rat. Unlike in most previous studies, suppression of hepatic glucose production was measured at levels of glycaemia above the relatively hypoglycaemic basal pregnant level. Glucose kinetics were measured using steady-state tracer methodology in chronically catheterised, conscious virgin control and pregnant rats, firstly, during basal and low-dose hyperinsulinaemic euglycaemic clamp conditions and secondly, during a three-step glucose infusion protocol (glucose infusion rates of 0, 60 and 150 μmol · kg⁻¹· min⁻¹). During the clamps, plasma glucose levels were not different (6.1 ± 0.4 vs 6.5 ± 0.3 mmol/l, pregnant vs virgin; N. S.), but plasma insulin levels were higher in the pregnant rats (242 ± 30 vs 154 ± 18 pmol/l, pregnant vs virgin; p < 0.05) most probably due to stimulated endogenous insulin release in this group. Hepatic glucose production was suppressed from basal levels by 41 % in virgin and 90 % in pregnant rats. During the glucose infusion studies, at matched insulin levels (147 ± 10 vs 152 ± 14 pmol/l), but at plasma glucose levels which were much lower in the pregnant rats (5.5 ± 0.2 vs 8.4 ± 0.6 mmol/l, pregnant vs virgin; p < 0.0001), hepatic glucose production was shown to be suppressed by a similar degree in both groups (41 ± 5 vs 51 ± 5 % from basal, pregnant vs virgin; N. S.). Both the plasma insulin and percentage suppression of hepatic glucose production dose responses to plasma glucose were markedly shifted to the left indicating that the plasma glucose set point is lowered in pregnancy. In conclusion, suppression of hepatic glucose production by insulin is not impaired and the set point for plasma glucose homeostasis is lowered during late pregnancy in the rat. [Diabetologia (1996) 39: 785–792] Received: 2 October 1995 and in final revised form: 1 February 1996     

Involvement of non-esterified fatty acid oxidation in glucocorticoid-induced peripheral insulin resistance in vivo in rats

Summary The mechanism by which glucocorticoids induce insulin resistance was studied in normal rats administered for 2 days with corticosterone then tested by euglycaemic hyperinsulinaemic clamps. Corticosterone administration induced a slight hyperglycaemia, hyperinsulinaemia and increased non-esterified fatty acid levels. It impaired insulin-stimulated total glucose utilization (corticosterone 15.7±0.7; controls 24.6±0.8 mg·kg⁻¹·min⁻¹), as well as residual hepatic glucose production (corticosterone 4.9±1.0; controls 2.0±0.7 mg·kg⁻¹·min⁻¹). During the clamps, insulin did not decrease the elevated non-esterified fatty acid levels in corticosterone-administered rats (corticosterone 1.38±0.15, controls 0.22±0.04 mmol/l). Corticosterone administration decreased the in vivo insulin-stimulated glucose utilization index by individual muscles by 62±6%, and the de novo glycogen synthesis by 78±2% (n=8–9 muscles). GLUT4 protein and mRNA levels were either unchanged or slightly increased by corticosterone administration. Inhibition of lipid oxidation by etomoxir prevented corticosterone-induced muscle but not hepatic insulin resistance. In conclusion, glucocorticoid-induced muscle insulin resistance is due to excessive nonesterified fatty acid oxidation, possibly via increased glucose fatty-acid cycle ultimately inhibiting glucose transport, or via decreased glycogen synthesis, or by a direct effect on glucose transporter translocation or activity or both.     

Overexpression of mitochondrial FAD-linked glycerol-3-phosphate dehydrogenase does not correct glucose-stimulated insulin secretion from diabetic GK rat pancreatic islets

Summary Glucose-stimulated insulin secretion is impaired in GK (Goto-Kakizaki) rats, perhaps because of abnormalities in glucose metabolism in pancreatic islet beta cells. The glycerol phosphate shuttle plays a major role in glucose metabolism by reoxidizing cytosolic NADH generated by glycolysis. In the pancreatic islets of GK rats, the activity of mitochondrial FAD-linked glycerol-3-phosphate dehydrogenase (mGPDH), the key enzyme of the glycerol phosphate shuttle, is decreased and this abnormality may be responsible, at least in part, for impaired glucose-stimulated insulin secretion. To investigate this possibility, we overexpressed mGPDH in islets isolated from GK rats via recombinant adenovirus-mediated gene transduction, and examined glucose-stimulated insulin secretion. In islets isolated from diabetic GK rats at 8 to 10 weeks of age, glucose-stimulated insulin secretion was severely impaired, and mGPDH activity was decreased to 79 % of that in non-diabetic Wistar rats. When mGPDH was overexpressed in islets from GK rats, enzyme activity and protein content increased 2- and 6-fold, respectively. Basal (3 mmol/l glucose) and glucose-stimulated (20 mmol/l) insulin secretion from the Adex1CAlacZ-infected GK rat islets were, respectively, 4.4 ± 0.7 and 8.1 ± 0.7 ng · islet⁻¹· 30 min⁻¹, and those from mGPDH-overexpressed GK rat islets 4.7 ± 0.3 and 9.1 ± 0.8 ng · islet⁻¹· 30 min⁻¹, in contrast to those from the Adex1CAlacZ-infected non-diabetic Wistar rat islets (4.7 ± 1.6 and 47.6 ± 11.9 ng · islet⁻¹· 30 min⁻¹). Thus, glucose-stimulated insulin secretion is severely impaired in GK rats even in the stage when mGPDH activity is modestly decreased, and at this stage, overexpression of mGPDH cannot restore glucose-stimulated insulin secretion. We conclude that decreased mGPDH activity in GK rat islets is not the defect primarily responsible for impaired glucose-stimulated insulin secretion. [Diabetologia (1998) 41: 649–653] Received: 20 October 1997 and in revised form: 22 December 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.     

Insulin like activity in (−) epicatechin

Summary Water extract of the bark of plant ofPterocarpus marsupium Roxb is used as an antidiabetic drug in indigenous medicine in India. (−) Epicatechin, its active principle, has been found to be insulinogenic. The presentin vitro study reports some insulin like activities of (−) epicatechin. Like insulin, (−) epicatechin stimulates oxygen uptake in fat cells and tissue slices of various organs, increases glycogen content of rat diaphragm in dose-dependent manner with corresponding increase in U¹⁴-C glucose uptake, and inhibits theophylline induced lipolysis in isolated fat pads in dose-dependent manner. Experiments on competitive binding of¹²⁵I-insulin and (−) epicatechin to liver cell plasma membrane indicate that insulin does not share binding site with (−) epicatechin. (−) Epicatechin at a concentration of up to 1 mM does not effect the release of glucagon from the isletsin vitro. Thus, (−) epicatechin has insulinogenic as well as insulin like properties. C.D.R.I. Communication no 4499.     

Reversibility of insulin resistance in obese diabetic patients: role of plasma lipids

Summary The aim of the present study was to measure whole body glucose uptake (M) and oxidation rate by euglycaemic hyperinsulinaemic clamp and indirect calorimetry in 7 morbidly obese subjects (BMI > 40 kg/m²) at three time points: before bilio-pancreatic diversion (BPD) surgery (Ob); 3 months after surgery (PO_I); and after reaching stable body weight, at least 2 years after surgery (PO_II). A group of 7 control subjects (C), matched groupwise for sex, age and BMI with PO_II patients, was also studied. The M value at PO_I was significantly higher than at Ob (49.12 ± 8.57 vs 18.14 ± 8.57 μmol · kg⁻¹· min⁻¹). No statistical difference was observed between the PO_II and C groups. Similarly, glucose oxidation rate was significantly increased at PO_I with respect to Ob (24.2 ± 7.23 vs 9.42 ± 3.91 μmol · kg⁻¹· min⁻¹) and was not significantly different between PO_II and C. Basal levels of non-esterified fatty acids (NEFA) decreased significantly both from Ob to PO_I and from PO_I to PO_II (1517.1 ± 223.9 vs 1039.6 ± 283.4 vs 616.0 ± 77.6 μmol · l⁻¹). The same applied to basal plasma triglycerides (2.07 ± 0.77 vs 1.36 ± 0.49 vs 0.80 ± 0.19 g · l⁻¹). Weight decreased mainly in the late postoperative period (PO_I to PO_II 124.28 ± 11.22 to 69.71 ± 11.78, 83 % of total decrement), rather than in the early postoperative period (Ob to PO_I 135.25 ± 14.99 to 124.28 ± 11.22 kg, 17 % of total decrement). We also report the clinical case of a young woman of normal weight, who underwent BPD for chylomicronaemia (secondary to familial lipoprotein lipase deficiency), whose M value, plasma insulin and blood glucose levels were normalized upon normalization of serum NEFA and triglyceride levels as determined by the therapeutic lipid malabsorption. In conclusion, in obese diabetic patients lipid malabsorption induced by BPD causes a definite enhancement of insulin sensitivity and glucose tolerance. This improvement in metabolism is noticeable before the surgery has major effects on body weight. These observations suggest that lowered plasma lipids, rather than weight loss per se, are the cause of the reversibility of insulin resistance. [Diabetologia (1997) 40: 599–605] Received: 31 July 1996 and in revised form: 30 December 1996     

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.