Impaired insulin action in newly diagnosed type 1 (insulin-dependent) diabetes mellitus

Summary Hepatic and peripheral insulin sensitivity were investigated in five newly diagnosed Type 1 (insulin-dependent) diabetic subjects before and after 1 week of twice daily insulin therapy. Eight weight-matched control subjects were also studied. Hepatic glucose production and glucose utilization were measured basally and during two sequential 2-h insulin (25 and 40 mU· kg^–1· h^–1)/glucose infusion periods. In the untreated hyperglycaemic diabetic patients hepatic glucose production was 16.3±2.6, 8.1±1.1 and 3.6±2.8|mol· kg^–1· min^–1 respectively for each of the three periods (mean±SEM), and fell with treatment to 12.5±1.4, 0.5±0.5 and 0.5±0.5 mol· kg^–1· min^–1. Hepatic glucose production for normal subjects was 13.4±0.7, 2.3±0.8 and <0.1 mol-kg^–1· min^–1. Glucose utilization was 12.7±1.4,18.2±0.7 and 22.1±3.4mol· kg^–1· min^–1 before treatment in the diabetic subjects, and 11.8±1.7, 20.9±3.3 and 30.1±3.6 after treatment. These values compare with those in the euglycaemic control subjects (13.4±0.7, 18.7±1.6 and 36.3±2.7 mol · kg^–1· min^–1). The pre-treatment metabolic clearance rate of glucose in all diabetic studies with insulin levels >30mU/l was 2.6 ±0.4 and rose to 3.9 ±0.5 ml· kg^–1· min^–1 following insulin therapy. This was significantly lower than in the control subjects (6.7±0.8 ml· kg^–1 · min^–1; p<0.005). Basal nonesterified fatty acid levels were high in the untreated, but normal in the treated diabetic subjects, and fell in response to insulin infusion. Basal -hydroxybutyrate levels were high in both diabetic groups, but also fell in response to insulin infusion. Erythrocyte insulin receptor binding was normal in the untreated diabetic subjects, and was not changed by treatment. Therefore, treatment of newly diagnosed Type 1 diabetic subjects with insulin reverses the hepatic insensitivity to insulin. In contrast, treatment only partially improves peripheral glucose disposal. Since erythrocyte insulin receptor binding is normal, it is likely that a post-receptor defect in peripheral glucose metabolism exists in Type 1 diabetic patients despite insulin therapy and good diabetic control for a period of 1 week.     

Increased glucose carbon recycling in severely insulin deficient Type 1 (insulin-dependent) diabetic subjects

Summary Six Type 1 (insulin-dependent) diabetic subjects were studied in order to determine the contribution of recycling of glucose carbon to the overproduction of glucose which is characteristic of the fasting hyperglycaemia produced by insulin withdrawal. The subjects were studied on two occasions, once after an overnight insulin infusion and once following 24 h of insulin withdrawal. The difference in turnover rates of 1-¹⁴C-glucose and 3-³H-glucose was used as a measure of glucose recycling. Insulin withdrawal caused a marked metabolic derangement with a rise in non-esterified fatty acids from 0.69±0.23 to 1.11±0.21 mmol/l (mean±SEM, p<0.05), total ketones from 0.27±0.06 to 2.06±0.51 mmol/l (p<0.01), cortisol from 341±43 to 479±31 nmol/l (p<0.05) and growth hormone from 1.1±0.3 to 19+5-mu/l (p<0.05). Glucose turnover rose from 13.8±2.3 mol·kg^–1·min^–1 at a glucose of 6.9±0.7 mmol/l in the insulin infused study to 25.8±4.4 mol·kg^–1·min^–1 (p<0.05) at a glucose of 16.4±0.7 mmol/l in the insulin withdrawn study. Recycling also rose from 3.0±0.4 mol· kg^–1·min^–1 to 9.4±2.2 mol·kg^–1·min^–1 (p<0.05) when insulin withdrawn, accounting for 23±3% and 36±3% of glucose turnover, respectively. We conclude that in the severely insulin deficient Type 1 diabetic subject recycling of glucose carbon is a major contributor to the excess glucose production.     

Insulin resistance in Type 2 (non-insulin-dependent) diabetic patients with hypertriglyceridaemia

Summary Hypertriglyceridaemia, which is frequently seen in Type 2 (non-insulin-dependent) diabetes mellitus, is associated with insulin resistance. The connection between hypertriglyceridaemia and insulin resistance is not clear, but could be due to substrate competition between glucose and lipids. To address this question we measured glucose and lipid metabolism in 39 Type 2 diabetic patients with hypertriglyceridaemia, i. e. mean fasting serum triglyceride level equal to or above 2 mmol/l (age 59±1 years, BMI 27.4±0.5 kg/m², HbA_1c8.0±0.2%, serum triglycerides 3.2±0.2 mmol/l) and 41 Type 2 diabetic patients with normotriglyceridaemia, i. e. mean fasting serum triglyceride level below 2 mmol/l (age 58±1 years, BMI 27.0±0.7 kg/m², HbA_1c7.8±0.2 %, serum triglycerides 1.4±0.1 mmol/l). Insulin sensitivity was assessed using a 340 pmol·(m²)^–1· min^–1 euglycaemic insulin clamp. Substrate oxidation rates were measured with indirect calorimetry and hepatic glucose production was estimated using a primed (25 Ci)-constant (0.25 Ci/min) infusion of [3-³H]-glucose. Suppression of lipid oxidation by insulin was impaired in patients with hypertriglyceridaemia vs patients with normal triglyceride levels (3.5±0.2 vs 3.0±0.2mol·kg^–1· min^–1; p<0.05). Stimulation of glucose disposal by insulin was reduced in hypertriglyceridaemic vs normotriglyceridaemic patients (27.0±1.3 vs 31.9±1.6 mol·kg^–1·min^–1; p<0.05) primarily due to impaired glucose storage (9.8±1.0 vs 14.6±1.4mol·kg^–1·min^–1; p<0.01). In contrast, insulinstimulated glucose oxidation was similar in patients with hypertriglyceridaemia and in patients with normal triglyceride concentrations (16.9±0.8 vs 17.2±0.7mol·kg^–1·min^–1). Hepatic glucose production in the basal state and during the clamp did not differ between the two groups. We conclude therefore that oxidative substrate competition between glucose and lipids does not explain insulin resistance associated with hypertriglyceridaemia in Type 2 diabetes. The question remains whether the reduced nonoxidative glucose disposal observed in the patients with hypertriglyceridaemia is genetically determined or a consequence of increased lipid oxidation.     

Contribution of glucose/glucose 6-phosphate cycle activity to insulin resistance in Type 2 (non-insulin-dependent) diabetes mellitus

Summary It has been suggested that increased glucose/glucose 6-phosphate substrate cycling impairs net hepatic glucose uptake in Type 2 (non-insulin-dependent) diabetes mellitus and contributes to hyperglycaemia. To investigate glucose/glucose 6-phosphate cycle activity and insulin action in Type 2 diabetes we studied eight patients and eight healthy control subjects, using the euglycaemic glucose clamp and isotope dilution techniques with purified [2-³H]- and [6-³H] glucose tracers, in the post-absorptive state and eight patients and five healthy control subjects during consecutive insulin infusions at rates of 0.4 and 2.0 mU·kg^–1·min^–1. [2-³H]glucose and [6-³H]glucose radioactivity in plasma samples were determined using selective enzymatic detritiation, allowing calculation of glucose turnover rates for each isotope, the difference being glucose/glucose 6-phosphate cycling. Endogenous glucose production ([6-³H]glucose) was greater in diabetic than control subjects in the post-absorptive state (15.6±1.5 vs 11.3±0.4 mol·kg^–1·min^–1, p<0.05) and during the 0.4 mU insulin infusion (10.1±1.3 vs 5.2±0.3 mol·kg^–1·min^–1, p<0.01) indicating hepatic insulin resistance. Glucose/glucose 6-phosphate cycling was significantly greater in diabetic than in control subjects in the post-absorptive state (2.6±0.4 vs 1.6±0.2 mol·kg^–1·min^–1, p<0.05) but not during the 0.4 mU insulin infusion (2.0±0.4 vs 2.0±0.3 mol·kg^–1·min^–1). During the 2.0 mU insulin infusion endogenous glucose production was suppressed to a similar degree in both groups (2.6±0.5 vs 3.4±0.7 mol · kg^–1·min^–1) but glucose disappearance was lower in the diabetic subjects (30.8±2.0 vs 52.4±4.6 mol·kg^–1·min^–1, p<0.01). During the 2.0 mU insulin infusion glucose/glucose 6-phosphate cycling was greater in the diabetic subjects (3.8±0.7 vs 0.8±0.6 mol·kg^–1·min^–1, p<0.05). In conclusion, both hepatic and peripheral insulin action are impaired in Type 2 diabetes. Increased glucose/glucose 6-phosphate cycling is seen in the post-absorptive state and also during marked hyperinsulinaemia, when insulin resistance is predominantly due to reduced peripheral tissue glucose uptake.     

Differential effects of insulin therapy on hepatic and peripheral insulin sensitivity in Type 2 (non-insulin-dependent) diabetes

Summary Hepatic glucose production and metabolic clearance rate of glucose were measured using (3-³H) glucose at steady state, basally and during two sequential 2 h insulin (25 and 40mU · kg^–1 · h^–1)/glucose(2 and 3mg · kg^–1 · min^–1) infusion periods. Eight diabetic subjects were studied before and after 1 week of twice daily insulin therapy; six control subjects matched for age, weight and degree of obesity were also studied. In the diabetic patients, pre-treatment hepatic glucose production was 20.0 ± 2.2, 9.9 ± 2.9, and 1.4 ± 0.8 mol · kg^–1 · min^–1 respectively (± SEM) for each of the three periods, and fell significantly with treatment to 12.8 ± 1.7,4.0 ± 1.5 and 1.9 ± 1.0 mol · kg^–1 · min^–1. Hepatic glucose production in normal subjects was 13.2 ± 0.6, 2.2 ± 0.8 and < 1 mol · kg^–1 · min^–1. The pre-treatment metabolic clearance rate in all diabetic studies with insulin levels 30 mU/l was 1.10 ± 0.14 ml · kg^–1 · min^–1 and remained virtually unchanged following insulin therapy; this was significantly lower than in the control subjects (6.83 ± 1.02, p < 0.001). Basal non-esterified fatty acid levels were higher (p < 0.02) in the pre-treated diabetic patients compared to post-treated diabetic patients and control subjects. Non-esterified fatty acids in each group fell to similar levels during the insulin infusions, but the rate of fall was slower in the pre-treated diabetic patients. Insulin receptor binding to erythrocytes was normal in the diabetic subjects and unchanged by treatment. Therefore, following insulin treatment of uncontrolled Type 2 (non-insulin-dependent) diabetes, the initially increased basal hepatic glucose production, and decreased hepatic sensitivity, return towards normal. However, the glucose clearance remains low, despite good diabetic control, and appears to be a major factor in the continuing glucose intolerance. As insulin receptor binding is normal, the defect of glucose clearance in Type 2 diabetes appears compatible with a post-receptor defect of glucose metabolism.     

Hepatic and peripheral insulin resistance: A common feature of Type 2 (non-insulin-dependent) and Type 1 (insulin-dependent) diabetes mellitus

Summary Hepatic glucose production (³H-glucose technique) and insulin-mediated glucose uptake (insulin clamp technique) were measured in 38 Type 2 (non-insulin-dependent) and 11 Type 1 (insulin-dependent) diabetic patients. Fasting plasma glucose concentration was 8.3 ± 0.5 mmol/l in the former, and 9.6 ± 1.3 mmol/1 in the latter group; the respective fasting plasma insulin levels were 19 ± 2 mU/l (p < 0.005 versus 13 ± 1 mU/l in 33 age-matched control subjects), and 9 ± 1 mU/l (p < 0.01 versus 14 ± 1 mU/l in 36 younger control subjects). In the fasting state, hepatic glucose production was slightly increased (15%, 0.1 > p > 0.05) in the Type 2 diabetic patients and markedly elevated (65%, p < 0.001) in the Type 1 patients compared with their respective control groups. In both groups of diabetic subjects, the rates of hepatic glucose production were inappropriately high for the prevailing plasma glucose and insulin levels, indicating the presence of hepatic resistance to insulin. Basal plasma glucose clearance was also significantly reduced in both the Type 2 (34%) and the Type 1 (14%) diabetic subjects. The fasting plasma glucose concentration correlated directly with hepatic glucose production, and inversely with plasma glucose clearance. During the insulin clamp, plasma insulin was maintained at approximately 100 mU/l in all groups, while plasma glucose was maintained constant at the respective fasting levels. Total glucose uptake was reduced in both the Type 2 (4.57 ± 0.31 versus 6.39 ± 0.25 mg · min^–1 · kg^–1 in the control subjects, p < 0.01) and the Type 1 (4.77 ± 0.48 versus 7.03 ± 0.22 mg · min^–1 · kg^–1, p < 0.01) diabetic patients. Insulin-stimulated glucose clearance was reduced to a similar extent in Type 2 (54%) and Type 1 (61%) diabetic subjects, and correlated directly with fasting glucose clearance. These results show that insulin resistance is a common feature of both types of diabetes and can be demonstrated in the basal as well as the insulin-stimlated state. Both hepatic and peripheral resistance to the action of insulin contribute to diabetic hyperglycaemia.     

Enhanced hepatic insulin sensitivity,but peripheral insulin resistance in patients with Type 1 (insulin-dependent) diabetes

Summary Sensitivity to insulin in vivo was studied in 8 normal weight C-peptide negative Type 1 (insulin-dependent) diabetic patients (age 23±1 years, diabetes duration 6±2 years), and in 8 age, weight and sex matched healthy subjects, using the euglycaemic clamp and 3-³H-glucose tracer technique. Prior to the study diabetic patients were maintained normoglycaemic overnight by a glucose controlled insulin infusion. Sequential infusions of insulin in 3 periods of 2 h resulted in mean steady state insulin levels of 12±2 versus 11±1, 18±2 versus 18±2 and 28±3 versus 24±2 U/ml in diabetic patients and control subjects. Corresponding glucose utilization rates were 2.4±0.2 versus 2.4±0.1, 2.4±0.2 versus 3.0±0.3 and 2.9±0.3 versus 4.6±O.6 mg·kg^–1·min^–1, p<0.02. Portal insulin values in the three periods were calculated to 12±2 versus 25±3, 18±2 versus 32±3 and 28±3 versus 37±3 U/ml in the diabetic patients and control subjects using peripheral insulin and C-peptide concentrations and assuming a portal to peripheral insulin concentration gradient of 1 in diabetic patients and of 2.4 in control subjects. Corresponding glucose production rates were 2.5±0.2 versus 2.4±0.1, 1.6±0.1 versus 0.9±0.2 and 0.7±0.1 versus 0.4±0.2 mg·kg^–1·min^–1. Using this approach the insulin dose-response curve for the peripheral glucose utilization was right-ward shifted, while the dose-response curve for the hepatic glucose production as a function of portal insulin levels was left-ward shifted. We conclude that in vivo insulin action is increased in the liver but decreased in peripheral tissues in insulin treated Type 1 diabetic patients. Presumably these oppositely directed changes in insulin action are acquired defects, secondary to the present mode of peripheral insulin treatment.     

Comparison of the effects of human recombinant insulin-like growth factor I and insulin on plasma amino acid concentrations and leucine kinetics in humans

Summary We examined the effects of recombinant human insulin-like growth factor I (IGF-I) and insulin on the plasma amino acid (AA) profile and leucine kinetics in eight normal subjects. IGF-I was infused at 52 pmol·kg^–1·min^–1, in combination with prime-continuous [1-¹⁴C] leucine infusion, to obtain steady-state plasma concentrations of total (54±3 nmol/l) and free (7.3±1 nmol/l) IGF-I (study 1). In response to IGF-I, plasma AA levels declined by 37±3% (1975±198 to 1368±120 mol/l) and total branched chain amino acids (BCAA) declined by 34±3% (390±21 to 256±13 mol/l). This hypoaminoacidaemic effect was associated with a decline in endogenous leucine flux of 17±2% (1.88±0.05 to 1.57±0.04 mol·kg^–1·min^–1) and leucine oxidation of 17±1% (0.31±0.02 vs 0.26±0.02 mol·kg^–1·min^–1) (both p<0.01 vs basal). The same subjects underwent a second study (study 2) in which insulin was infused at 6.22 pmol·kg^–1·min^–1 to obtain a steady-state plasma insulin concentration of 530±25 pmol/l while maintaining euglycaemia. The infusion rate was designed to match the declines in plasma BCAA levels and leucine turnover observed during IGF-I infusion. The rates of glucose infusion necessary to maintain euglycaemia during IGF-I and insulin infusion were 4.9±1.0 and 7.8±0.6 mg·kg^–1 ·min^–1, respectively. During insulin infusion total BCAA declined by 39% from 369±23 to 226±20 mol/l, leucine flux declined by 16±2% from 1.90±0.05 to 1.61±0.03 mol·kg^–1·min^–1, and leucine oxidation declined by 19±2% from 0.32±0.02 to 0.26±0.02 mol·kg^–1·min^–1. On a molar basis IGF-I was 7.3% as potent as insulin in inhibiting proteolysis. These results demonstrate that in humans: (i) the hypoaminoacidaemic response to IGF-I can be entirely ascribed to the inhibition of proteolysis; (ii) qualitatively, the effects of IGF-I and insulin on plasma AA profile and protein metabolism are similar; (iii) quantitatively, IGF-I is 14-fold less potent than insulin in suppressing protein degradation.Abbreviations IGF-I Insulin-like growth factor I - AA aminoacid - BCAA branched chain amino acids - KIC alphaketoisocaproate - ELF endogenous leucine flux - NOLD non-oxidative leucine disposal     

Determination and kinetic analysis of non-insulin mediated glucose uptake in Type 1 (insulin-dependent) diabetes mellitus

Summary In man, total glucose uptake is the sum of insulin mediated glucose uptake and non-insulin mediated glucose uptake. The latter pathway has not been examined in Type 1 (insulin-dependent) diabetes mellitus. In order to assess non-insulin mediated glucose uptake in Type 1 diabetes, we measured steady-state rates of glucose uptake during glucose clamps at 5.27, 9.71 and 12.5 mmol/l using low (0.25 mU· kg^–1·min^–1), intermediate (0.75 mU·kg^–1·min^–1) and high (1.50 mU·kg^–1·min^–1) insulin infusion rates in 10 subjects with Type 1 diabetes. For insulin infusion rates of 0.25, 0.75 and 1.50 mU·kg^–1·min^–1 as plasma glucose rose from 5.27 to 9.71 mmol/l, total glucose uptake increased by 35, 43 and 52 percent respectively (p<0.05 for each insulin infusion rate). For all three insulin infusion rates, there was no significant increase in total glucose uptake as plasma glucose increased from 9.71 to 12.5 mmol/l. At each glycaemic level, glucose uptake correlated significantly with plasma free insulin (r=0.81, p<0.01 at 5.71 mmol/l; r=0.84, p<0.01 at 9.71 mmol/l; r=0.73, p<0.02 at 12.5 mmol/l). Linear regression analysis to a point corresponding to plasma free insulin equalling zero, yielded values for non-insulin mediated glucose uptake (mmol·kg^–1·min^–1) of 0.11,0.14,0.18 at plasma glucose of 5.27, 9.7 and 12.5 mmol/l respectively. Thus, increasing plasma glucose concentrations were associated with increasing rates of non-insulin mediated glucose uptake. For each insulin infusion rate used, the percent of total glucose uptake accounted for by non-insulin mediated glucose uptake remained independent of plasma glucose concentration, but decreased as insulin infusion rate increased. During the insulin infusion at 0.25 mU·kg^–1·min^–1, this percentage ranged from 83.7 to 91.4%. Analysis of glucose uptake data derived for theoretical plasma insulin levels of 0, 40, 80 and 160 U/ml yielded linear Eadie-Hofstee plots (r=– 0.83 to – 0.99), suggesting that insulin increased Vmax but did not alter Km. Hence, in these subjects with Type 1 diabetes, glucose uptake, both insulin mediated and non-insulin mediated can be described by Michaelis-Menten kinetics. Comparison of values obtained for Vmax and Km in the present studies of Type 1 diabetes with those obtained from non-diabetic subjects indicates that non-insulin dependent glucose uptake in Type 1 diabetes is quantitatively similar to that of non-diabetic subjects.     

Hyperinsulinaemia and insulin insensitivity: studies in subjects with insulinoma

Summary Hepatic glucose turnover, peripheral insulin sensitivity and insulin receptor binding were measured in four subjects with insulinoma before and 3 months after surgical resection of the insulinoma. Basal hepatic glucose production, quantitated employing a primed constant infusion of tritiated glucose, was low pre-operatively (5.2±1·7mol·kg^–1· min^–1) but returned to normal post-operatively (14.9±2.8; normal subjects 13.9±0.8 mol·kg^–1·min^–1). Paired euglycaemic dose-response curves were developed for each subject. Insulin sensitivity, expressed as a right shift of the dose-response curve (ED₅₀), was low pre- and post-operatively. However, insulin responsiveness (V_max) remained normal (pre-operatively 13.9±2.2, post-operatively 13.8±0.8, normal subjects 16.7±0.8ml·kg^–1·min^–1). There was no consistent pattern in monocyte or erythrocyte receptor binding before or after surgery. These data suggest that the chronic hyperinsulinaemia causes suppression of hepatic glucose production, and a state of insulin insensitivity which appears to be due to a post-receptor defect.     

ACE-inhibition increases hepatic and extrahepatic sensitivity to insulin in patients with Type 2 (non-insulin-dependent) diabetes mellitus and arterial hypertension

Summary To assess the effects of ACE-inhibition on insulin action in Type 2 (non-insulin-dependent) diabetes mellitus associated with essential hypertension, 12 patients with Type 2 diabetes (on diet and oral hypoglycaemic agents) and arterial hypertension were examined on two occasions, in a single blind, cross-over study, after two days of treatment with either captopril or a placebo. The study consisted of a euglycaemic-hyperinsulinaemic clamp (two sequential steps of insulin infusion at the rates of 0.25 mU·kg^–1·min^–1 and 1 mU·kg^–1·min^–1, 2 h each step), combined with an infusion of 3-³H-glucose to measure the rate of hepatic glucose production and that of peripheral glucose utilization. The results show that blood pressure was lower after captopril (sitting, systolic 148±5 mmHg, diastolic 89±2 mm Hg) compared to placebo (155±6 and 94±2 mm Hg) (p<0.05). Captopril treatment resulted in a more suppressed hepatic glucose production (2.7±0.4 vs 4.94±0.55 mol·kg^–1·min^–1), and a lower plasma non-esterified fatty acid concentration (0.143±0.05 vs 0.200±0.05 mmol/l) (captopril vs placebo, p<0.05) at the end of the first step of insulin infusion (estimated portal plasma insulin concentration 305±28 pmol/l); and in a greater glucose utilization (36.5±5.1 vs 28±3.6mol·kg^–1·min^–1, p<0.001) at the end of the second step of insulin infusion (arterial plasma insulin concentration of 604±33 pmol/l). We conclude that captopril improved insulin sensitivity in Type 2 diabetes associated with hypertension at the level of the liver and extrahepatic tissues, primarily muscle and adipose tissue. Thus, in contrast to other antihypertensive drugs such as diuretics and beta-blockers which may have a detrimental effect on insulin action, ACE-inhibitors appear to improve insulin action in Type 2 diabetes and essential hypertension, at least on a short-term basis.     

Decreased hepatic glucagon responses in Type 1 (insulin-dependent) diabetes mellitus

Summary The effect of glucagon infusion on hepatic glucose production during euglycaemia was evaluated in seven Type 1 (insulin-dependent) diabetic patients and in ten control subjects. In the diabetic subjects normoglycaemia was maintained during the night preceding the study by a variable intravenous insulin and glucose infusion. During the study endogenous insulin secretion was suppressed by somatostatin (450 g/h) and replaced by insulin infusion (0.15 mU·kg^–1·min^–1). ³H-glucose was infused for isotopic determination of glucose turnover. Plasma glucose was clamped at 5 mmol/1 for 2 h 30 min and glucagon (1.5 ng· kg^–1·min^–1) was then infused for the following 3 h. Hepatic glucose production and glucose utilisation were measured during the first, second and third hour of the glucagon infusion. Basal hepatic glucose production (just prior to glucagon infusion) was similar in diabetic (1.2±0.3 mg·kg^–1·min^–1) and control (1.6±0.1 mg·kg^–1·min^–1) subjects. In diabetic patients hepatic glucose production rose slowly to 2.1±0.5 mg·kg^–1·min^–1 during the first hours of glucagon infusion and stabilized at this level (2.4±0.5 mg·kg^–1·min^–1) in the third hour. In control subjects hepatic glucose production increased sharply to higher levels than in the diabetic subjects (3.4±0.3 mg·kg^–1·min^–1) during the first and second hour of glucagon infusion (p<0.05) and then gradually fell (2.9±0.4 mg·kg^–1·min^–1) during the third hour. In conclusion, when stimulated with glucagon at a physiologic plasma concentration diabetic patients had 1) an overall reduced hepatic glucose production response and 2) an abnormal sluggish response pattern. These abnormalities may imply inappropriate counter-regulation following a hypoglycaemic episode.     

Normal insulin sensitivity during the phase of glucose intolerance but insulin resistance at the onset of diabetes in the spontaneously diabetic BB rat

Summary In diabetes-prone BB rats, 30 to 50% of animals undergo autoimmune destruction of the pancreatic B-cells leading to a short period of glucose intolerance, followed by an abrupt onset of diabetes. We have examined whether the glucose intolerance period and the onset of diabetes are associated with changes in insulin sensitivity, using the euglycaemic hyperinsulinaemic clamp coupled with [3-³H] glucose infusion. Glucose intolerant rats were detected by a transient glycosuria one hour after an oral glucose load performed every four days. Insulin sensitivity studied in these rats the day following their detection was normal. Other diabetes-prone BB rats were tested daily and studied on the first day of glycosuria. In the basal state, glucose production was increased in diabetic rats (11.3±1.1 vs 7.1±0.8mg·min^–1·kg^–1, p<0.05). Tissue glucose utilization was similar in diabetic and control rats (8.3±0.5 vs 7.1±0.8mg·min^–1·kg^–1) despite a three fold higher glycaemia in the diabetic rats. During the hyperinsulinaemic clamps, glycaemia was clamped at 6.1–6.6 mmol/l in diabetic and control rats. A decreased insulin sensitivity was observed in diabetic rats at submaximal (200 U/ml) and maximal (1500 U/ml) insulin concentrations for both inhibition of hepatic glucose production and stimulation of glucose utilization. No autoantibodies against insulin could be detected in the plasma of diabetic rats. Plasma concentrations of glucagon, catecholamines, ketone bodies and fatty acids were similar in control and diabetic rats during the clamp studies. Our results suggest that the decrease of basal insulin concentration is responsible for the insulin resistance in the diabetic BB rat at onset of diabetes, either directly or through the increased glycaemia.     

In vivo insulin action and muscle glycogen synthase activity in Type 2 (non-insulin-dependent) diabetes mellitus: effects of diet treatment

Summary Insulin resistant glucose metabolism is a key element in the pathogenesis of Type 2 (non-insulin-dependent) diabetes mellitus. Insulin resistance may be of both primary (genetic) and secondary (metabolic) origin. Before and after diet-induced improvement of glycaemic control seven obese patients with newly-diagnosed Type 2 diabetes were studied with the euglycaemic clamp technique in combination with indirect calorimetry and forearm glucose balance. Muscle biopsies were obtained in the basal state and again after 3 h of hyperinsulinaemia (200 mU/l) for studies of insulin receptor and glycogen synthase activities. Similar studies were performed in seven matched control subjects. Insulin-stimulated glucose utilization improved from 110±11 to 183±23 mg·m^–2·min^–1 (p<0.03); control subjects: 219+23 mg·m^–2·min^–1 (p=NS, vs post-diet Type 2 diabetes). Nonoxidative glucose disposal increased from 74±17 to 138+19 mg·m^–2·min^–1 (p<0.03), control subjects: 159±22 mg· m^–2·m^–1 (p=NS, vs post-diet Type 2 diabetic patients). Forearm blood glucose uptake during hyperinsulinaemia increased from 1.58±0.54 to 3.35±0.23 mol·l^–1·min^–1 (p<0.05), control subjects: 2.99±0.86 mol·l^–1·min^–1 (p=NS, vs post-diet Type 2 diabetes). After diet therapy the increase in insulin sensitivity correlated with reductions in fasting plasma glucose levels (r=0.97, p<0.001), reductions in serum fructosamine (r=0.77, p<0.05), and weight loss (r=0.78, p<0.05). Values of muscle glycogen synthase sensitivity to glucose 6-phosphate (A_0.5 for glucose 6-phosphate) were similar in the basal state. However, insulin stimulation of glycogen synthase was more pronounced after diet treatment (A_0.5: 0.43±0.06 (before) vs 0.30±0.04 mmol/l (after); p<0.03; control subjects: 0.22±0.03 mmol/l). Muscle insulin receptor binding and kinase activity were similar before and after diet treatment and comparable to values in the control group. The data suggest that impaired insulin stimulation of in vivo glucose turn-over and muscle glycogen synthase activity tend to be restored during successful diet treatment of patients with Type 2 diabetes.     

The effects of different plasma insulin concentrations on lipolytic and ketogenic responses to epinephrine in normal and Type 1 (insulin-dependent) diabetic humans

Summary This study was performed to verify: (1) the ability of different insulin concentrations to restrict the lipolytic and ketogenic responses to exogenous epinephrine administration; (2) whether the ability of insulin to suppress the lipolytic and ketogenic responses during epinephrine administration is impaired in Type 1 (insulin-dependent) diabetic patients. Each subject was infused on separate occasions with insulin at rates of 0.2, 0.4, and 0.8 mU·kg^–1·min^–1 while normoglycaemic. To avoid indirect adrenergic effects on endocrine pancreas secretions, the so-called islet clamp technique was used. Rates of appearance of palmitic acid, acetoacetate, and 3-hydroxybutyrate were simultaneously measured with an infusion of ¹³C-labelled homologous tracers. After a baseline observation period epinephrine was exogenously administered at a rate of 16 ng·kg^–1·min^–1. At low insulin levels (20 U/ml) the lipolytic response of comparable magnitude was detected in normal and Type 1 diabetic patients. However, the ketogenic response was significantly higher in Type 1 diabetic patients. During epinephrine administration, similar plasma glucose increments were observed in the two groups (from 4.74±0.53 to 7.16±0.77 mmol/l (p<0.05) in Type 1 diabetic patients and from 4.94±0.20 to 7.11±0.38 mmol/l (p<0.05) in normal subjects, respectively). At intermediate insulin levels (35 U/ml) no significant differences were found between Type 1 diabetic patients and normal subjects, whereas plasma glucose levels rose from 4.98±0.30 to 6.27±0.66 mmol/l (p<0.05) in Type 1 diabetic patients, and from 5.05±0.13 to 6.61±0.22 mmol/l (p<0.05) in normal subjects. At high insulin levels (70 U/ml) the lipolytic response was detectable only in Type 1 diabetic patients; the ketogenic response was reduced in both groups. During the third clamp, a significant rise in plasma glucose concentration during epinephrine infusion was observed in both groups. In conclusion this study shows that at low insulin levels Type 1 diabetic patients show an increased ketogenic response to epinephrine, despite their normal nonesterified fatty acid response. Instead, high insulin levels are able to restrict the ketogenic response to epinephrine in both normal and Type 1 diabetic subjects, although there is a still detectable lipolytic response in the latter. In the presence of plasma free insulin levels that completely restrict the ketogenic response in the same group, there is still a distinct glycaemic response. Plasma insulin levels in Type 1 diabetic patients are a major determinant of the metabolic response to epinephrine.     

Liver and muscle insulin sensitivity,glycogen concentration and glycogen synthase activity in a rat model of non-insulin-dependent diabetes

Summary Mild diabetes was induced in adult rats with streptozotocin (45 mg/kg body weight), and insulin sensitivity, glycogen deposition and glycogen synthase activity assessed in liver and muscle 5 weeks later. Diabetic rats had significantly elevated fasting blood glucose concentrations (5.6±0.1 versus 3.6±0.1 mmol/l, p<0.001), and blood glucose concentrations 2 h after a 1 g/kg glucose load (12.0±0.6 versus 3.7±0.2 mmol/l, p<0.001). After a 20-h fast hepatic glucose output was significantly elevated (58±3 versus 47±3 mol·min^–1·kg^–1, p<0.05), and failed to suppress at high insulin concentrations during a euglycaemic clamp (hepatic glucose output 21±4 versus 2±4 mol·min^–1·kg^–1, p<0.02). Liver glycogen was lower in the diabetic rats by the end of the clamp (16±3 versus 38±6 mol/g wet wt, p<0.05). At the end of the clamp total glucose turnover was lower in the diabetic rats (107±4 versus 161±17 mol·min^–1· kg^–1, p<0.01), as was skeletal muscle glycogen synthase activity (0.46±0.04 versus 0.67±0.05 U/g wet wt, p<0.01) and glycogen concentration (22±2 versus 33±3 mol/g wet wt, p<0.05). Blood lactate and pyruvate responses suggested that glycolytic pathways were similarly affected. Thus, insulin insensitivity develops in both liver and skeletal muscle after 5 weeks of mild streptozotocin-induced diabetes.     

Both acute and chronic near-normoglycaemia are required to improve insulin resistance in Type 1 (insulin-dependent) diabetes mellitus

Summary To determine the impact of both short- and longterm near-normoglycaemia on insulin resistance in Type 1 (insulin-dependent) diabetes hepatic glucose production (mg · kg^–1 · min^–1) and peripheral glucose utilisation (M-value, mg · kg^–1 · min^–1) were estimated during an euglycaemic hyperinsulinaemic clamp (10 mU · kg · min) in patients with either good (HbA_1c<5.8%, groups A and B) or poor (HbA_1c>7.5%, groups C and D) long-term metabolic control (time > 12 months) and in healthy subjects (HbA_1c: 5.08±0.20%; n=8). To this end blood glucose was stabilized at 6.7 mmol/l by overnight (t=12 h) i.v. regular insulin in groups (n=8 each) A (HbA_1c: 5.49±0.46%) and C (HbA_1c: 8.83±1.20%),while groups B (HbA_1c:5.55±0.19%) andD (HbA_1c: 8.51±1.09%) were kept overnight on long-acting insulin without feed-back control of blood glucose before euglycaemic clamping. Thereby, pre-equilibration of blood glucose at 6.7 mmol/l was shown to normalize basal hepatic glucose production (A: 2.27±0.48; C 2.50±0.57 mg · kg^–1 · min^–1) despite different HbA_1c values, whereas basal hepatic glucose production stayed elevated in groups B (3.09±0.38 mg · kg^–1 · min^–1) and D (3.21±0.58 mg · kg^–1 · min^–1) with poor actual glycaemia (B: 10.9±4.6; D: 12.1±4.6 mmol/l). To restitute peripheral glucose utilisation close to normal (healthy subjects: 13.99±2.13; A: 12.12±2.67; B: 8.72±3.0; C: 10.27±1.69; D: 7.10±2.31 mg · kg^–1 · min^–1; healthy subjects vs A: NS; healthy subjects vs B, C, D: p<0.05) both long-term (HbA_1c<5.8%) and acute nearnormoglycaemia by 12-h i. v. insulin pre-treatment were required (group A). We conclude that good long-term glucose control per se is unable to normalize hepatic and peripheral glucose metabolism in Type 1 diabetic patients unless actual near-normoglycaemia is provided consistently, e.g. by i.v. overnight infusion of regular insulin.     

The effect of prehepatic insulin administration on alanine flux rates in diabetic dogs

Summary The in vivo flux rates of glucose (6-³H-glucose) and of alanine (U-¹⁴C-alanine) were measured in insulin-dependent chronically diabetic dogs which were infused with insulin employing a bedside-type artificial B cell and either the peripheral or the portal venous route. In comparison with non-diabetic control animals the diabetic dogs had near-normal patterns of glucose metabolism and pancreatic glucagon regardless of the route of insulin administration. They also showed reduced basal portal but moderately elevated peripheral insulin levels on peripheral and near-normal peripheral values on portal insulin infusion. Both concentration and production rates of alanine were reduced on peripheral (0.142±0.016mmol/l, 4.73±0.49 mol·kg^–1·min^–1, p< 0.05) but normal on portal insulin (0.206±0.030 mmol/l, 6.33±0.63 mol·kg^–1 ·min^–1). The alanine clearance was slightly elevated or normal in the diabetic dogs, and the glucose production from alanine showed a strongly delayed response to an exogenous glucose load on either route of insulin administration. It is concluded that the peripheral hyperinsulinism during posthepatic insulin administration stimulates glucose utilisation to a normal extent, but inhibits the provision of amino groups in resting muscle. Alanine synthesis is thereby reduced, and the carbon moieties are shunted from glucose into circulating lactate. Long-term studies are needed to elucidate the role of the liver under these conditions.     

Insulin regulation of glucose and lipid metabolism in massive obesity

Summary Eight obese patients and 12 normal individuals underwent a euglycaemic insulin clamp (20 and 40 mU · m^2–1 · min^–1) along with continuous infusion of 3-³H-glucose and 1-¹⁴C-palmitate and indirect calorimetry. Basal plasma glucose concentration (4.7±0.3 vs 4.4±0.2 mmol/l) was similar in the two groups, whereas hepatic glucose production was slightly higher in obese individuals (1.11±0.06 vs 0.84±0.05 mmol/min) in spite of higher plasma insulin levels (17±2 vs 6±1 mU/l; p<0.01). Insulin inhibition of hepatic glucose production was impaired in obese subjects. Glucose disposal by lean body mass was markedly reduced both at baseline (11.7±1.1 vs 15.6±0.6 mol · kg^–1 · min^–1; p<0.05) and during clamp (15.0±1.1 vs 34.4±2.8 and 26.7±3.9 vs 62.2±2.8 mol · kg^–1 · min^–1; p<0.01) Oxidative (12.2±1.1 vs 17.8±1 and 16.1±1.1 vs 51.1±1.7 mol · kg^–1 · min^–1; p<0.05–0.002) and non-oxidative glucose metabolism (3.9±1.1 vs 15.0±2.8 and 12.8±3.3 vs 38.3±2.2 mol · kg^–1 · min^–1; p<0.01–0.001) were impaired. Basal plasma concentrations of non-esterified fatty acids (635±75 vs 510±71 mol/l) and blood glycerol (129±17 vs 56±5 mol/l; p<0.01) were increased in obese patients. Following hyperinsulinaemia, plasma non-esterified fatty acids (244±79 vs 69±16 and 140±2 vs 36±10 mol/l; p<0.01) and blood glycerol levels (79±20 vs 34±6 and 73±22 vs 29±5 mol/l; p<0.01) remained higher in obese subjects. Baseline non-esterified fatty acid production rate per kg of fat body mass was significantly larger in normal weight subjects (37.7±6.7 vs 14.0±1.8 mol/l; p<0.01) and insulin inhibition was reduced in obese patients (–41±9 vs –74±3 and –53±11 vs –82±3%; p<0.05). Basal plasma non-esterified fatty acid utilization by lean body mass was similar in the two groups (9.8±0.9 vs 8.8±2.0 mol · kg^–1 · min^–1), whereas during clamp it remained higher in obese patients (6.0±1.2 vs 2.8±2.5 and 4.9±1.3 vs 1.5±0.6 mol · kg^–1 · min^–1; p<0.1–0.05). Lipid oxidation was higher in obese individuals in spite of hyperinsulinaemia (3.7±0.3 vs 2.4±0.4 and 2.3±0.4 vs 0.9±0.3 mol · kg^–1 · min^–1; p<0.05– 0.02). An inverse correlation was found between lipid oxidation and glucose oxidation (r=0.82 and 0.93; p<0.001) and glucose utilization (r=0.54 and 0.83; p<0.05–0.001) both in obese and control subjects. A correlation between lipid oxidation and non-oxidative glucose metabolism was present only in normal weight individuals (r=0.75; p<0.01). We conclude that in obesity all tissues (muscles, liver, and adipose tissue) are resistant to insulin action. Insulin resistance involves glucose as well as lipid metabolism.     

Effect of storage temperature of insulin on pharmacokinetics and pharmacodynamics of insulin mixtures injected subcutaneously in subjects with Type 1 (insulin-dependent) diabetes mellitus

Summary These studies were undertaken to assess the influence of storage temperature of insulin vials on pharmacokinetics and pharmacodynamics of a mixture of lente insulin (Monotard HM) and regular insulin (Actrapid HM) injected subcutaneously. Seven subjects with Type 1 (insulin-dependent) diabetes mellitus were studied twice after overnight normalization of plasma glucose. A mixture of lente insulin (0.22 U/kg) and regular insulin (0.11 U/kg) was prepared from insulin vials kept either refrigerated (4 °C) or at room temperature (18 °C) and injected subcutaneoulsy (abdomen). Euglycaemia was maintained for the following 16 h by glucose infusion at variable rate. With refrigerated insulin, the plasma free insulin peak was greater (53±5 versus 45±6 mU/l) and occurred earlier (2.5±0.2 versus 6±0.3 h), and the glucose infusion rate showed a greater (16.5±1.2 versus 14.5±0.9 mol·kg^–1·min^–1) and earlier peak (3.2±0.2 versus 6±0.4 h) as compared to that occurring with the non-refrigerated insulin (p<0.05). However, 6 h after insulin injection, both plasma free insulin and glucose infusion rate were 30% lower with the mixture of refrigerated as compared to that of non-refrigerated insulin (p<0.05). In contrast, when NPH-insulin (Protaphane HM) was mixed with regular insulin and injected in 4 out of the 7 diabetic patients, the storage temperature of insulin vials had no effect on the pharmacokinetics and pharmacodynamics of the mixture. Thus, the storage temperature of insulin vials profoundly influences the effects of the mixture lente/regular insulin, but does not affect the pharmacokinetics and pharmacodynamics of the mixture NPH/regular insulin.