Introducing a simplified approach to insulin therapy in type 2 diabetes: a comparison of two single-dose regimens of insulin glulisine plus insulin glargine and oral antidiabetic drugs.

AIM: To investigate whether the addition of a single bolus of insulin glulisine (glulisine), administered at either breakfast or main mealtime, in combination with basal insulin glargine (glargine) and oral antidiabetic drugs (OADs), provides equivalent glycaemic control in patients with type 2 diabetes, irrespective of the time of glulisine injection. METHODS: A national, multicentre, randomized, open-label, parallel-group study of 393 patients with type 2 diabetes who were suboptimally controlled [haemoglobin A(1c) (HbA(1c)) > 6.5-9.0% and fasting blood glucose (BG) 7.0% at baseline and who reached HbA(1c)

Fatty acid-induced beta cell hypersensitivity to glucose. Increased phosphofructokinase activity and lowered glucose-6-phosphate content.

Diabetic states are characterized by a raised serum/islet level of long chain fatty acids and a lowered ED50 for glucose-induced insulin secretion. Prolonged culture (> 6 h) of islets with long chain fatty acids replicates the basal insulin hypersecretion. We examined this effect in rat islets cultured for 24 h with 0.25 mM oleate. Insulin secretion at 2.8 mM glucose was doubled in combination with a 60% lowered islet content of glucose-6-phosphate (G6P). Investigation of the lowered G6P showed: (a) increased glucose usage from 0.5 to 100 mM glucose with identical values measured by [2-3H]glucose and [5-3H]glucose, (c) indicating little glucose- 6-phosphatase activity, (b) unchanged low pentose phosphate shunt activity, (c) 50% increased phosphofructokinase (PFK) Vmax, (d) a normal ATP/ADP ratio, and (e) unchanged fructose 2,6 bisphosphate content. Triacsin C, an inhibitor of fatty acyl-CoA synthetase, prevented the increase in PFK activity and the lowered G6P content. These results suggest that long chain acyl-CoA mediates the rise in PFK activity, which in turn lowers the G6P level. We speculate that the inhibition of hexokinase by G6P is thus attenuated, thereby causing the basal insulin hypersecretion.

Minimal chronic hyperglycemia is a critical determinant of impaired insulin secretion after an incomplete pancreatectomy.

We now describe experiments that allow one to determine the consequences of B cell reduction alone vs. those that result from superimposed mild hyperglycemia. Male CD rats underwent a 60% pancreatectomy (Px); controls were sham operated. 1 wk later, either 10% sucrose (SUC) was substituted as fluid supply or tap water was continued (WAT). Plasma glucose and insulin values in Px-WAT remained equal to the sham groups; in Px-SUC the values were euglycemic for 25 d, but then nonfasting plasma glucose rose 15 mg/dl. After 6 wk, B cell mass in Px-WAT was reduced by 45% and non-B cell mass by 57%. In contrast, in Px-SUC both masses were comparable to the sham groups. The insulin response to 27.7 mM glucose was measured using the in vitro perfused pancreas. The responses were reduced in Px-WAT but in proportion to their reduced B cell mass; in contrast, it was 75% less than expected in Px-SUC. Also, the response to arginine given at 16.7 mM glucose was reduced only in Px-SUC. These results show that a lowering of B cell mass that does not result in hyperglycemia has no adverse effect on the remaining B cells. On the other hand, if even mild hyperglycemia develops, B cell function becomes impaired and results in inappropriately reduced insulin stores and insulin responses to marked stimuli.

Increased secretory demand rather than a defect in the proinsulin conversion mechanism causes hyperproinsulinemia in a glucose-infusion rat model of non-insulin-dependent diabetes mellitus.

Hyperproinsulinemia in non-insulin-dependent diabetes mellitus (NIDDM) is due to an increased release of proinsulin from pancreatic beta cells. This could reside in increased secretory demand placed on the beta cell by hyperglycemia or in the proinsulin conversion mechanism. In this study, biosynthesis of the proinsulin conversion enzymes (PC2, PC3, and carboxypeptidase-H [CP-H]) and proinsulin, were examined in islets isolated from 48-h infused rats with 50% (wt/vol) glucose (hyperglycemic, hyperinsulinemic, and increased pancreatic proinsulin to insulin ratio), 20% (wt/vol) glucose (normoglycemic but hyperinsulinemic), and 0.45% (wt/vol) saline (controls). A decrease in the islet content of PC2, PC3, and CP-H from hyperglycemic rats was observed. This reduction did not correlate with any deficiency in mRNA levels or biosynthesis of PC2, PC3, CP-H, or proinsulin. Furthermore, proinsulin conversion rate was comparable in islets from hyperglycemic and control rats. However, in islets from hyperglycemic rats an abnormal increased proportion of proinsulin was secreted, that was accompanied by an augmented release of PC2, PC3 and CP-H. Stimulation of the beta cell's secretory pathway by hyperglycemia, resulted in proinsulin being prematurely secreted from islets before its conversion could be completed. Thus, hyperproinsulinemia induced by chronic hyperglycemia likely results from increased beta cell secretory demand, rather than a defect in the proinsulin processing enzymes per se.

Mechanism of impaired glucose-potentiated insulin secretion in diabetic 90% pancreatectomy rats. Study using glucagonlike peptide-1 (7-37).

Chronic hyperglycemia causes a near-total disappearance of glucose-induced insulin secretion. To determine if glucose potentiation of nonglucose secretagogues is impaired, insulin responses to 10(-9) M glucagonlike peptide-1 (GLP-1) (7-37) were measured at 2.8, 8.3, and 16.7 mM glucose with the in vitro perfused pancreas in rats 4-6 wk after 90% pancreatectomy (Px) and sham-operated controls. In the controls, insulin output to GLP-1 was > 100-fold greater at 16.7 mM glucose versus 2.8 mM glucose. In contrast, the increase was less than threefold in Px, reaching an insulin response at 16.7 mM glucose that was 10 +/- 2% of the controls, well below the predicted 35-40% fractional beta-cell mass in these rats. Px and control rats then underwent a 40-h fast followed by pancreas perfusion using a protocol of 20 min at 16.7 mM glucose followed by 15 min at 16.7 mM glucose/10(-9) M GLP-1. In control rats, fasting suppressed insulin release to high glucose (by 90%) and to GLP-1 (by 60%) without changing the pancreatic insulin content. In contrast, in Px the insulin response to GLP-1 tripled in association with a threefold increase of the insulin content, both now being twice normal when stratified for the fractional beta-cell mass. The mechanism of the increased pancreas insulin content was investigated by assessing islet glucose metabolism and proinsulin biosynthesis. In controls with fasting, both fell 30-50%. In Px, the degree of suppression with fasting was similar, but the attained levels both exceeded those of the controls because of higher baseline (nonfasted) values. In summary, chronic hyperglycemia is associated with a fasting-induced paradoxical increase in glucose-potentiated insulin secretion. In Px rats, the mechanism is an increase in the beta-cell insulin stores, which suggests a causative role for a lowered beta-cell insulin content in the impaired glucose-potentiation of insulin secretion.

Regulatory effects of glucose on the catalytic activity and cellular content of glucokinase in the pancreatic beta cell. Study using cultured rat islets.

Glucose regulates the cellular content of glucokinase in the pancreatic beta cell by altering the level of the enzyme. We investigated the existence of a second regulatory pathway, an alteration in the catalytic activity, by comparing Vmax and protein levels of glucokinase in rat islets cultured under high glucose conditions (16.7 mM) for 6, 14, and 24 h. The Vmax was increased by glucose at all time points. In contrast, glucokinase protein levels on Western blots were unchanged from the control value at 6 h but increased 40% at the later time points (P < 0.0002). Further evidence for a dual regulatory system was obtained with a reversal protocol. After a 6-h incubation at high glucose, an additional 3-h incubation at 5.5 mM glucose restored glucokinase Vmax to normal, but failed to change the Vmax after a 24-h incubation at high glucose. Finally, 10 microM cycloheximide partially prevented the increase in glucokinase Vmax induced by 24 h of high glucose, but had no effect at 6 h, suggesting the early increase in enzymatic activity did not require protein synthesis. In summary, glucose regulates both the catalytic activity and cellular content of glucokinase in the beta cell. Glucose-induced increases in glucokinase activity are an important element of the beta cell adaptive response to hyperglycemia.

Mechanism of compensatory hyperinsulinemia in normoglycemic insulin-resistant spontaneously hypertensive rats. Augmented enzymatic activity of glucokinase in beta-cells.

The cause of compensatory hyperinsulinemia in normoglycemic insulin-resistant states is unknown. Using spontaneously hypertensive rats (SHR), we tested the hypothesis that a lowered beta-cell set-point for glucose causes a hypersecretion of insulin at a normal glucose level. Islets isolated from normoglycemic hyperinsulinemic SHR were compared to age-matched (12 wk old) Wistar-Kyoto (WK) rats. The ED50 for glucose-induced insulin secretion was 6.6 +/- 1.0 mM glucose in SHR versus 9.6 +/- 0.5 mM glucose in WK (P < 0.02). Glucokinase enzymatic activity was increased 40% in SHR islets (P < 0.02) without any change in the glucokinase protein level by Western blot. The level of the beta-cell glucose transporter (GLUT-2) was increased 75% in SHR islets (P < 0.036). In summary, the beta-cell sensitivity for glucose was increased in these normoglycemic insulin resistant rats by an enhanced catalytic activity of glucokinase. We have identified a regulatory system for glucokinase in the beta-cell which entails variable catalytic activity of the enzyme, is modulated in response to variations in whole-body insulin sensitivity, and is not dependent on sustained changes in the plasma glucose level.

Chronic hyperglycemia is associated with impaired glucose influence on insulin secretion. A study in normal rats using chronic in vivo glucose infusions.

We have proposed that chronic hyperglycemia alters the ability of glucose to modulate insulin secretion, and have now examined the effects of different levels of hyperglycemia on B cell function in normal rats using chronic glucose infusions. Rats weighing 220-300 g were infused with 0.45% NaCl or 20, 30, 35, or 50% glucose at 2 ml/h for 48 h, which raised the plasma glucose by 18 mg/dl in the 30% rats, 37 mg/dl in the 35% rats, and 224 mg/dl in the 50% group. Insulin secretion was then examined using the in vitro isolated perfused pancreas. Glucose-induced insulin secretion remained intact in the normoglycemic 20% glucose rats and it was potentiated in the mildly hyperglycemic 30% glucose rats. However, with even greater hyperglycemia in the 35% glucose group the insulin response to a high glucose perfusate was severely blunted, and it was totally lost in the most hyperglycemic 50% glucose rats. In a second protocol that examined glucose potentiation of arginine-stimulated insulin release, a similar impairment in the ability of glucose to modulate the insulin response to arginine was found with increasing levels of chronic hyperglycemia. On the other hand, the ability of a high glucose concentration to inhibit arginine-stimulated glucagon release was preserved in all glucose-infused rats, but the glucagon levels attained in response to the arginine at 2.8 mM glucose were much less in the 50% glucose rats than in all the other groups. These data clearly show that after 48 h of marked hyperglycemia, glucose influence upon insulin secretion in the rat is severely impaired. This model provides a relatively easy and reproducible method to study the effects of long-term hyperglycemia on B cell function.

The loss of GLUT2 expression by glucose-unresponsive beta cells of db/db mice is reversible and is induced by the diabetic environment.

Glucose-induced insulin secretion by beta cells of diabetic db/db mice was studied by a pancreas perfusion technique, and the levels of GLUT2 protein in pancreatic islets were assessed by immunofluorescence microscopy and protein blot analysis. Beta cells from diabetic mice had a high basal rate of insulin secretion; they did not respond to glucose stimulation but displayed a normal secretory response to arginine. At the same time, GLUT2 expression by db/db islets was lost whereas beta cells from nondiabetic db/+ mice expressed high levels of this transporter. GLUT2 levels in liver or kidney of diabetic mice were, however, mostly unaltered. Transplanting islets from db/db mice under the kidney capsule of db/+ mice restored normal GLUT2 levels. Conversely, transplantation of db/+ islets into db/db mice induced the disappearance of GLUT2 expression. When islets from db/+ mice were transplanted under the kidney capsule of streptozocin-diabetic mice, the immunodetection of GLUT2 also disappeared. We conclude that: (a) GLUT2 expression is decreased in glucose-unresponsive beta cells from db/db mice; (b) the decreased expression of GLUT2 is reversible; (c) the loss of GLUT2 expression is induced by the diabetic environment of db/db and streptozocin-induced diabetic mice. These observations together with previously published data suggest that a factor different from glucose or insulin regulates the beta cell expression of GLUT2.

Increased proinsulin/insulin ratio in pancreas extracts of hyperglycemic rats.

The plasma ratio of proinsulin/insulin is raised in people with NIDDM. A relative hypersecretion of proinsulin is thought to be the cause, because pancreas extracts from diabetic rats have a raised proinsulin/insulin ratio. We tested the hypothesis that the pancreatic proinsulin/insulin mismatch results from hyperglycemia-induced beta-cell degranulation. Normal rats made hyperglycemic with 48-h glucose infusions had a raised pancreatic percentage of proinsulin. In contrast, rats infused with enough glucose to induce compensatory hyperinsulinemia without changing the plasma glucose level had a normal percentage of proinsulin. The raised percentage of proinsulin in the hyperglycemic rats reflected a reduction in pancreatic insulin content. Administering an inhibitor of insulin release, diazoxide, to hyperglycemic rats blocked the fall in pancreatic insulin content and prevented the rise in the percentage of proinsulin. Normal rats infused with tolbutamide for 3 days and enough glucose to maintain euglycemia had a 50% reduction in pancreatic insulin content. The beta-cell degranulation from this nonhyperglycemic mechanism resulted in a raised pancreatic percentage of proinsulin. In summary, chronic hyperglycemia causes beta-cell degranulation primarily because of hyperstimulated insulin release. The net result is a rise in the ratio of immature (proinsulin-rich) to mature (insulin-rich) granules, which is reflected as an increased relative proportion of proinsulin. Mobilization of these proinsulin-enriched granules may explain the relative hypersecretion of proinsulin that occurs with diabetes.

Parallel reduction of pancreas insulin content and insulin secretion in 48-h tolbutamide-infused normoglycemic rats.

The overworked-beta-cell hypothesis proposes that lowered glucose-potentiated insulin secretory responses in diabetes are secondary to hyperstimulated insulin secretion and depletion of the beta-cell insulin stores. We tested this hypothesis in normal rats using a 48-h infusion of 200 mg x kg(-1) x day(-1) tolbutamide in 20% glucose. Insulin secretion was measured by in vitro pancreas perfusion. Twice daily blood glucose values were equal in the tolbutamide-infused and control rats. Pancreas insulin content was 47 +/- 7% that of the controls (P < 0.004). Insulin responses to 16.7 mmol/l glucose, 16.7 mmol/l glucose/10 mmol/l arginine, and 5.5 mmol/l glucose/10 mmol/l arginine were reduced in parallel, except for the first phase response to 16.7 mmol/l glucose/arginine. Pancreas amylin content was unchanged in the tolbutamide-infused rats as was amylin secretion, resulting in higher than normal stored and secreted amylin-to-insulin molar ratios. Importantly, a raised amylin-to-insulin ratio and a relatively unimpaired first versus second phase insulin response for high glucose/arginine both occur in diabetic rats. Thus, our results support the overworked-beta-cell hypothesis by showing chronic beta-cell stimulation without hyperglycemia replicates part of the beta-cell dysfunction found with diabetes.

Unresponsiveness to glucose in a streptozocin model of diabetes. Inappropriate insulin and glucagon responses to a reduction of glucose concentration.

The neonatal streptozocin (STZ) rat model of NIDDM has been previously found to have a markedly reduced insulin response to an acute increase in glucose concentration. We studied the effect of an acute reduction in glucose concentration on insulin and glucagon secretion in this model and contrasted the results with the effects of epinephrine and somatostatin using the in vitro isolated, perfused pancreas. The reduction in perfusate glucose concentration from 11.1 to 2.8 mM caused a rapid suppression of insulin release in the control rats, but had no inhibitory effect in the STZ group. Epinephrine (55 nM) and somatostatin (110 nM) caused similar decreases in insulin secretion in both groups. The glucose reduction also caused an increase in glucagon release in the controls, but had no effect in the STZ rats. Epinephrine, however, stimulated glucagon secretion in both groups in a similar fashion, and inhibition by somatostatin was also comparable. The baseline insulin and glucagon concentrations were enhanced in a separate series of experiments by the addition of arginine (5 mM) to the perfusate, and while the insulin and glucagon responses to the glucose reduction remained lost, appropriate inhibition of insulin secretion was demonstrated in the STZ rats with epinephrine. These data indicate that A- and B-cells in this rat model of NIDDM are selectively unresponsive to both increases and decreases in glucose concentration, while the responsiveness to nonglucose agents remains intact.

Evolution of abnormal insulin secretory responses during 48-h in vivo hyperglycemia.

Recent in vitro studies have shown that insulin release caused by continuous exposure to high glucose concentration markedly falls within a few hours. We wanted to determine if a similar effect occurs in vivo with chronic intravenous infusions in normal rats. Male CD rats (200-250 g) were infused with 50% glucose at 2 ml/h for 6, 14, 24, or 48 h, whereas controls received 0.45% NaCl, and insulin responses were tested with the in vitro isolated perfused pancreas. Plasma glucose averaged 352 +/- 20 mg/dl after 4 h and 396 +/- 11 mg/dl after 24 h versus 137 +/- 5 mg/dl in controls; plasma insulin at the same times was 8.94 +/- 1.44 and 12.1 +/- 2.62 ng/ml versus 1.69 +/- 0.19 ng/ml in controls. The incremental insulin response caused by an increase in perfusate glucose from 2.8 to 16.7 mM was not significantly reduced after 24 h of glucose infusion; in contrast, paradoxical suppression was seen after 48 h. A second protocol examined glucose potentiation by giving 10 mM arginine at 2.8 and 16.7 mM glucose; a hyperresponse to arginine at the lower glucose level was present after just 14 h of infusion. Therefore, these results do not support the hypothesis that beta-cells lose their sensitivity to glucose within hours of being exposed to higher than normal glucose concentrations.

Upregulated hexokinase activity in isolated islets from diabetic 90% pancreatectomized rats.

Glucokinase is the beta-cell glucose sensor, i.e., the site in glucose metabolism that determines the glucose set-point (sensitivity) for insulin secretion. Hexokinase is also present, but it normally contributes little to glucose metabolism because of end-product inhibition by glucose 6-phosphate. There is a lowered glucose set-point for insulin secretion in 90% pancreatectomized (Px) diabetic rats. We investigated the mechanism by measuring hexokinase and glucokinase activity in islet extracts. Glucokinase activity was minimally raised in Px islets (Vmax 125% of sham-operated control rats). In contrast, hexokinase Vmax was 250% of the control value, suggesting that the increased hexokinase activity caused the beta-cell glucose hypersensitivity. Additional evidence was obtained with a 40-h fast that was performed because of a previous observation that the inhibitory effect of fasting on insulin secretion was impaired in Px rats. Glucokinase activity fell normally in the Px rats (32 +/- 4% reduction in sham vs. 37 +/- 4% in Px rats) as opposed to hexokinase activity, which was unaffected in either group. In summary, a feature of hyperglycemia is upregulated islet hexokinase activity. The result is that hexokinase assumes partial control over the glucose set-point for insulin secretion. As such, regulatory effects on insulin secretion, such as fasting, that are mediated through glucokinase activity may be altered.

Increased catalytic activity of glucokinase in isolated islets from hyperinsulinemic rats.

The high Km glucose phosphorylation enzyme glucokinase is believed to be the beta-cell glucose sensor, i.e., the site in glucose metabolism that determines the sensitivity and specificity of glucose-induced insulin secretion. We investigated the regulation of this enzyme by measuring glucokinase Vmax and protein levels in isolated islets from hyperinsulinemic rats. Rats were infused for 48 h with 2 ml/h of 20% glucose, 50% glucose, or 0.45% NaCl (control rats). At the end of the infusion, 20% glucose-infused rats were normoglycemic and hyperinsulinemic (2.3-fold rise in basal plasma insulin level). Their islets had a 2.3-fold increase in insulin secretion at 8.3 mM glucose (51 +/- 10% of capacity vs. 22 +/- 5% in NaCl rats, P < 0.03), a 75% increase in glucokinase Vmax and little if any increase in glucokinase protein level (111 +/- 3% of control). The rats infused with 50% glucose had marked hyperglycemia and higher basal plasma insulin levels. Their islets were maximally stimulated by 8.3 mM glucose in combination with a 270% increase in glucokinase Vmax and a 69 +/- 11% increase in glucokinase protein level. Hexokinase Vmax was also doubled. Thus, compensatory increases in beta-cell glucose phosphorylation are a key mechanism for adaptive hyperinsulinemia. Our results show two types of regulation for the beta-cell high Km phosphorylation enzyme, glucokinase. The content of glucokinase protein is controlled by the plasma glucose level. Variable catalytic activity of this protein was also observed in this study.

Diazoxide causes recovery of beta-cell glucose responsiveness in 90% pancreatectomized diabetic rats.

Chronic hyperglycemia causes near-total disappearance of glucose-induced insulin secretion. The etiology has been suggested to be a nonsustainable stimulation of insulin release that causes beta-cells to become unresponsive to glucose through an undefined mechanism. We used an inhibitor of insulin secretion, diazoxide, to test this hypothesis in 90% pancreatectomized (Px) rats. Px rats were given 5 days of diazoxide (30 mg/kg orally twice a day) or tap water starting on postoperative day 8, 15, or 22. In vitro pancreas perfusions were conducted 36 h posttreatment (2, 3, or 4 weeks after surgery) using a protocol of 15 min of 16.7 mM glucose followed by 15 min of 16.7 mM glucose plus 10 mM arginine. In 2-week Px rats, insulin responses to 16.7 mM glucose and to glucose/arginine were both appropriate for the reduced beta-cell mass, i.e., no defect in beta-cell glucose responsiveness had yet occurred. Diazoxide had no affect on insulin release at this time. Between 2 and 3 weeks after pancreatectomy, insulin output to 16.7 mM glucose fell 75%, and that to glucose/arginine fell 50%. Diazoxide given at this time partially blocked the fall in glucose-induced insulin secretion and totally prevented that with arginine. The increased insulin secretion caused by diazoxide was accompanied by 1) lower nonfasting plasma glucose values, 2) improved glucose tolerance after oral glucose load, and 3) a 50% increase in pancreatic insulin content. Our results support the concept that excessive insulin secretion is a major cause of the hyperglycemia-induced loss of beta-cell glucose responsiveness. A leading candidate for the mechanism of this effect is depleted pancreatic insulin stores. Overstimulation of insulin secretion provides a new target for pharmacological therapy aimed at reducing glucose intolerance in non-insulin-dependent diabetes mellitus.

Shared biochemical properties of glucotoxicity and lipotoxicity in islets decrease citrate synthase activity and increase phosphofructokinase activity.

Diabetic states are characterized by a raised serum/islet level of triglycerides and a lowered EC50 (concentration at half-maximal stimulation) for glucose-induced insulin secretion. Culturing islets with long-chain fatty acids (FAs) replicates the basal insulin hypersecretion. In a previous study, we showed that the mechanism involved deinhibition of hexokinase by a 60% decrease in glucose-6-phosphate (G-6-P). The key event was proposed to be an increased phosphofructokinase (PFK) Vmax secondary to an upregulatory effect of the FA metabolite, long-chain acyl-coenzyme A (LC-CoA). We now show another contributory factor, a lowered content of the PFK inhibitor citrate. Citrate synthase Vmax and citrate levels were lowered 45% in rat islets cultured with 250 micromol/l oleate for 24 h. Both effects were reversed by triacsin C, an inhibitor of fatty acyl-CoA synthetase, the enzyme that generates LC-CoA. Culturing islets with high doses of glucose (16.7 mmol/l) for 48 h should also raise cytosolic LC-CoA. As predicted, citrate synthase Vmax was lowered and PFK Vmax was increased, both in a triacsin C-reversible fashion. These results show shared selected functional and biochemical properties in beta-cells of so-called glucotoxicity and lipotoxicity.

Glucose-fatty acid cycle to inhibit glucose utilization and oxidation is not operative in fatty acid-cultured islets.

The glucose-fatty acid cycle of Randle entails two elements: decreased pyruvate dehydrogenase (PDH) activity, which inhibits glucose oxidation, and inhibition of phosphofructokinase (PFK) by a rise in citrate so that glucose-6-phosphate (G-6-P) levels increase, thereby inhibiting hexokinase activity and hence glucose utilization. Chronic exposure of islets to long-chain fatty acids (FA) is reported to lower PDH activity, but the effect on glucose oxidation and glucose-induced insulin secretion is uncertain. We investigated rat islets that were cultured for 4 days with 0.25 mmol/l oleate/5.5 mmol/l glucose. Glucose oxidation was doubled at 2.8 mmol/l glucose and unchanged at 27.7 mmol/l glucose in the FA-cultured islets despite a 35% decrease in assayed PDH activity. Pyruvate content was increased 60%, which may well compensate for the decreased PDH activity and maintain flux through the citric acid cycle. However, a greater diversion of pyruvate metabolism through the pyruvate-malate shuttle is suggested by unchanged pyruvate carboxylase Vmax and a fourfold higher release of malate from isolated mitochondria. The FA-cultured islets also showed increased basal glucose usage and insulin secretion together with a lowered level of G-6-P and 50% reductions in citrate synthase Vmax and the citrate content. Thus, the effects of chronic FA exposure on islet glucose metabolism differ from the glucose-fatty acid interactions reported in some other tissues.

Relative hypersecretion of proinsulin in rat model of NIDDM.

The plasma ratio of proinsulin to insulin is raised in individuals with non-insulin-dependent diabetes mellitus (NIDDM). Increased secretion of proinsulin relative to insulin is thought to be the cause, although differential changes in clearance have not been ruled out. This study was conducted in a rat model of NIDDM, 90% pancreatectomized (Px) rats, to investigate the pathophysiology of this observation. Proinsulin storage and secretion were assessed with high-performance liquid chromatography separation of the insulins and the proinsulins, followed by quantification of the peaks by insulin radioimmunoassay. In Px rats, the relative proportion of proinsulin in pancreas extracts was twice that of control (sham-operated) rats (15.6 +/- 1.4 vs. 8.3 +/- 1.4%, P less than 0.01). Samples obtained from the portal vein during in vitro pancreas perfusion also had an elevated proinsulin fraction (Px, 10.3 +/- 3.0; sham, 3.0 +/- 0.6%; P less than 0.006). In summary, 90% Px rats share many pathophysiological features with NIDDM, including loss of normal proinsulin homeostasis. Our results suggest that chronic hyperglycemia causes an intrinsic change in beta-cells that is characterized by the increased storage and secretion of proinsulin.

Abnormal insulin secretion in a streptozocin model of diabetes. Effects of insulin treatment.

We have proposed that chronic hyperglycemia causes the abnormal glucose influence on arginine-stimulated insulin secretion in the neonatal streptozocin (STZ) rat model of NIDDM and therefore studied the effect of 24 h of mild insulin-induced hypoglycemia on this defect. Ultralente insulin, 20 U/kg, was given at 9 a.m. and 10 U/kg at 5 p.m., and insulin and glucagon secretion were then studied the next morning using the in vitro isolated, perfused pancreas. The fed plasma glucose concentrations decreased in the STZ rats from 191 +/- 13 to 101 +/- 9 mg/dl and from 133 +/- 4 to 99 +/- 8 mg/dl in the controls. As expected, 10 mM arginine caused a trivial insulin response at 2.8 mM glucose in the treated and untreated control groups compared with the marked one at 16.7 mM. The response to arginine at 2.8 mM glucose in the untreated STZ rats, however, was strikingly elevated (7.65 +/- 2.29 versus 0.41 +/- 0.16 ng/ml in the untreated controls) and it was not potentiated by the high glucose background, but the result at 2.8 mM glucose in the treated STZ rats was similar to that of the treated controls (0.46 +/- 0.12 versus 0.16 +/- 0.03 ng/ml). A return of glucose influence on IBMX-stimulated insulin secretion was also noted. Glucose-induced insulin release, however, was not restored in the treated STZ rats, but it was markedly suppressed in the controls by the insulin treatment. Glucose influence on the glucagon response to arginine was maintained in the STZ model even though the glucagon release to a lowered glucose concentration was lost.(ABSTRACT TRUNCATED AT 250 WORDS)