Cyclosporine A induces endothelin-converting enzyme-1: Studies in vivo and in vitro.

AIM: Cyclosporine A (CsA) induces renal vasoconstriction and hypoxia and enhances the expression of endothelin-1 (ET-1) pro-hormone (pre-pro-ET-1), plausibly leading to a feed-forward loop of renal vasoconstriction, hypoxia and enhanced synthesis of the potent vasoconstrictor ET-1. Endothelin-converting enzyme (ECE)-1 cleaves big endothelin to generate endothelin (ET)-1 and is upregulated by hypoxia via hypoxia-inducible factor (HIF). We hypothesized that in addition to the direct induction of ET-1 synthesis, CsA might also intensify renal ECE-1 expression, thus contributing to enhanced ET-1 synthesis following CsA. METHODS: CsA was administered to Sprague Dawley rats (120 mg/kg/SC) for 4 days, and renal HIF and ECE-1 expression were assessed with Western blots and immunostaining. Human umbilical vein endothelial cells (HUVEC) and proximal tubular cell line (HK-2) were subjected to CsA, and ECE-1 induction was evaluated using real-time mRNA PCR and Western blots. RESULTS: Cyclosporine A intensified renal parenchymal ECE-1 expression in the rat kidney, particularly in distal nephron segments, along with renal hypoxia (detected by pimonidazole adducts) and HIF expression, in line with our recent observations showing episodic hypoxia in mice subjected to CsA. Furthermore, in cultured normoxic HUVEC and HK-2 cells, CsA dose-dependently induced both pre-pro-ET-1 and ECE-1 mRNA and protein expression, with enhanced ET-1 generation. CONCLUSION: CsA induces ECE-1 via both hypoxic and non-hypoxic pathways. ECE-1 may contribute to increased renal ET-1 generation following CsA, participating in a feed-forward loop of renal parenchymal hypoxia and ET synthesis.

Adaptive response to hypoxia and remote ischaemia pre-conditioning: a new hypoxia-inducible factors era in clinical medicine.

Transient ischaemia leads to tolerance to subsequent protracted ischaemia. This 'ischaemia pre-conditioning' results from the induction of numerous protective genes, involved in cell metabolism, proliferation and survival, in antioxidant capacity, angiogenesis, vascular tone and erythropoiesis. Hypoxia-inducible factors (HIF) play a pivotal role in this transcriptional adaptive response. HIF prolyl hydroxylases (PHDs), serving as oxygen sensors, control HIFα degradation. HIF-mediated ischaemic pre-conditioning can be achieved with the administration of PHD inhibitors, with the attenuation of organ injury under various hypoxic and toxic insults. Clinical trials are currently under way, evaluating PHD inhibitors as inducers of erythropoietin. Once their safety is established, their potential use might be further tested in clinical trials in various forms of acute ischaemic and toxic organ damage. Repeated transient limb ischaemia was also found to attenuate ischaemic injury in remote organs. This 'remote ischaemic pre-conditioning' phenomenon (RIP) has been extensively studied recently in small clinical trials, preceding, or in parallel with an abrupt insult, such as myocardial infarction, cardiac surgery or radiocontrast administration. Initial results are promising, suggesting organ protection. Large-scale multi-centre studies are currently under way, evaluating the protective potential of RIP in cardiac surgery, in the management of myocardial infarction and in organ transplantation. The mechanisms of organ protection provided by RIP are poorly understood, but HIF seemingly play a role as well. Thus, Inhibition of HIF degradation with PHD inhibitors, as well as RIP (in part through HIF), might develop into novel clinical interventions in organ protection in the near future.

Diabetes as a risk factor for medication-related osteonecrosis of the jaw.

Medication-related osteonecrosis of the jaw (MRONJ) is a severe devastating complication for which the exact pathogenesis is not completely understood. Multiple systemic and local factors may contribute to the development of MRONJ. A growing body of evidence supports diabetes mellitus (DM) as an important risk factor for this complication; however, the exact mechanism by which DM may promote MRONJ has yet to be determined. The current review elucidates the role of DM in the pathogenesis of MRONJ and the mechanisms by which DM may increase the risk for MRONJ. Factors related to DM pathogenesis and treatment may contribute to poor bone quality through multiple damaged pathways, including microvascular ischemia, endothelial cell dysfunction, reduced remodeling of bone, and increased apoptosis of osteoblasts and osteocytes. In addition, DM induces changes in immune cell function and promotes inflammation. This increases the risk for chronic infection in the settings of cancer and its treatment, as well as antiresorptive medication exposure, thus raising the risk of developing MRONJ. A genetic predisposition for MRONJ, coupled with CYP 450 gene alterations, has been suggested to affect the degradation of medications for DM such as thiazolidinediones and may further increase the risk for MRONJ.

Ex vivo endothelin dependent contraction of the remodeled rat spiral artery.

INTRODUCTION: Similarities of the rat to the human placenta make rat pregnancy models relevant to the study of human gestational diseases. Understanding of species differences is necessary to extrapolate from animal models to humans. We observed alpha-smooth muscle actin (αSMA) expression in rat endovascular trophoblasts (EVasT) and investigated the spatial and temporal expression of smooth muscle (SM) proteins and their potential function in remodeled spiral artery. METHODS: Rat placentas were examined from gestational day 13 to term, and were immunostained for cytokeratin, αSMA, alpha heavy chain of SM myosin, non-muscle myosin, Rho proteins, regulators of SM gene expression, myocardin, an early marker of SM differentiation and endothelin receptors A and B (ETA, ETB). Transmission electron microscopy (TEM) was performed. Modified spiral artery rings were studied ex vivo for endothelin-1- induced contraction. RESULTS: EVasT expressed SM proteins co-localizing with cytokeratin confirming their trophoblastic origin from gestational day 13 to term. Thin fibers, consistent with actin fibers, were observed by TEM, in the cellular localization of αSMA in EVasT. Functional experiments revealed that addition of 10(-7) M endothelin-1 ex vivo reduced vascular lumen area by 11.1% ± 1.8% compared with control. This effect was reduced to only 1.0 ± 1.7% with ETA antagonist, and to 5.4 ± 1.7% contraction by ETB antagonist, p < 0.002, for all. DISCUSSION: The expression of SM proteins in EVasT along with the contractibility of the rat remodeled spiral artery ex vivo, suggest that some vascular tone is potentially maintained by endothelin-1, and may play a role in situations of dysregulation of the vasoactive systems.

Intrauterine growth restriction and shallower implantation site in rats with maternal hyperinsulinemia are associated with altered NOS expression.

Nitric oxide synthase (NOS) plays an important role in hypertensive disorders of pregnancy. In the context of the known association between hyperinsulinemia and hypertension, we studied the expression of the 3 isoforms of NOS (neuronal-nNOS, inducible-iNOS, and endothelial-eNOS) in the placenta and implantation site of our insulin-induced intrauterine growth restriction (IUGR) rat model in which the normal gestational blood pressure decline is abrogated. The fetuses of hyperinsulinemic dams were significantly smaller than those of normal pregnant dams (male fetal weight=4.8+/-0.5 g vs. 5.4+/-0.4 g, hyperinsulinemic vs. control, respectively; female fetal weight=4.5+/-0.5 g vs. 5.1+/-0.4 g, hyperinsulinemic vs control, respectively, p<0.0001). Their placentas weighed less than those of normal pregnant dams (0.44+/-0.08 g in hyperinsulinemic dams vs. 0.50+/-0.09 g, p<0.0001) and their implantation site, designated the mesometrial triangle, was also smaller. Endovascular trophoblasts were found more often and in greater depth in normal pregnant dams. Possibly as a compensatory mechanism, the endovascular trophoblasts formed cell groups rather than a monolayer and occupied a larger portion of the arterial perimeter in arteries of hyperinsulinemic dams. iNOS expression increased by 80% (p<0.0001) and 180% (p=0.045) in placenta and mesometrial triangle of hyperinsulinemic dams, respectively. The expression of eNOS was reduced by 17% (p=0.048) in the placenta and did not change significantly in the mesometrial triangle (p>0.05). nNOS expression was decreased by 37% (p=0.03) in the placenta and increased by 53% (p=0.035) in the mesometrial triangle of hyperinsulinemic dams. Immunohistochemistry revealed prominent expression of iNOS in the placental junctional zone and in interstitial and endovascular trophoblasts in the mesometrial triangle. Assuming a role in trophoblastic invasion, the increased expression of iNOS in hyperinsulinemic dams explains the "compensatory" pattern of trophoblastic invasion. Expression of eNOS was prominent in endothelial cells and weak in endovascular trophoblasts. In our model of gestational hyperinsulinemia-induced IUGR, we found not only differing expression of the 3 NOS isoforms in the cellular elements of the placenta and mesometrial triangle, but also divergent modes of altered NOS isoform expression. These findings suggest, in accordance with other publications, that each isoform may have a distinct function in the placenta and placental bed. The differing expression of the 3 NOS isoforms in the placenta and in the mesometrial triangle in rat IUGR seems to result from the hyperinsulinemia and the resulting IUGR phenotype.

Diabetes and radiocontrast media increase endothelin converting enzyme-1 in the kidney.

Plasma endothelin-1 levels rise in diabetes and after exposure to contrast media suggesting a role in progressive diabetic and acute radiocontrast nephropathies. Here we studied individual and combined effects of streptozotocin-induced diabetes and contrast media on renal endothelin converting enzyme-1 levels in the rat. In vivo, medullary (but not cortical) endothelin converting enzyme protein gradually increased 4 to 5-fold following the induction of diabetes or after the administration of contrast media but rose 15-fold when diabetic rats were given contrast media. Changes in mRNA expression paralleled those of the protein. Immunohistochemistry confirmed that increased tubular and endothelial cell endothelin converting enzyme-1 were most pronounced in the medulla. In vitro, endothelin-1 levels increased 3-fold following incubation of endothelial cells with media high in glucose or with contrast and 4-fold with their combination. Endothelin converting enzyme-1 protein and mRNA expression changed in a similar pattern while prepro endothelin-1 mRNA increased with each insult but not in an additive way. Our study shows that diabetes and contrast media up-regulate renal medullary endothelin converting enzyme-1 expression and synthesis.

Adaptation to hypoxia in the diabetic rat kidney.

Hypoxia of the kidney in diabetes could predispose it to develop acute and chronic renal failure. To examine the relationship between renal hypoxia and renal failure, we measured hypoxia (as a pimonidazole adducts), hypoxia-inducible factors (HIFs), and a hypoxia target gene heme oxygenase-1. The studies were performed in rats with streptozotocin (STZ)-induced diabetes, Cohen diabetes sensitive rats, and during short-term artificial hyperglycemia in rats induced by intravenous glucose and octreotide. STZ-treated rats received insulin, the superoxide dismutase mimetic tempol, or contrast medium. Radiocontrast media causes hypoxia and HIF induction. Hypoxia, HIFs, and heme oxygenase were undetectable in controls, but transiently activated in STZ-treated and the Cohen diabetes sensitive rats. Different patterns of HIFs and pimonidazole were observed between the three models. Insulin abolished pimonidazole and HIF induction, whereas tempol lead to increased HIFs and heme oxygenase induction at similar levels of pimonidazole. When compared with control rats, STZ-treated rats exhibited more intense and protracted renal pimonidazole, with augmented hypoxia inducible factor production and reduced GFR following contrast media. Our data suggest that both regional hypoxia and hypoxia adaptation transiently occur in early stages of experimental diabetes, largely dependent on hyperglycemia or after contrast media. Tempol may augment the HIF response in diabetes.

Guillain-Barré syndrome following hepatitis B vaccination.

A 52-year-old woman developed Guillain-Barré syndrome 10 weeks after immunization with recombinant hepatitis B vaccine. Common infectious causes of GBS were ruled out. The temporal relationship between GBS and hepatitis B virus (HBV) vaccination was suggestive of a vaccine-induced cause. The possible mechanisms of this very, rare complication are discussed.

Treatment of diabetes with vanadium salts: general overview and amelioration of nutritionally induced diabetes in the Psammomys obesus gerbil.

BACKGROUND: Numerous investigations have demonstrated the beneficial effect of vanadium salts on diabetes in streptozotocin (STZ)-diabetic rats, in rodents with genetically determined diabetes and in human subjects. The amelioration of diabetes included the abolition of hyperglycemia, preservation of insulin secretion, reduction in hepatic glucose production, enhanced glycolysis and lipogenesis and improved muscle glucose uptake through GLUT4 elevation and translocation. The molecular basis of vanadium salt action is not yet fully elucidated. Although evidence has been provided that the insulin receptor is activated, the possibility exists that cytosolic non-receptor tyrosine kinase, direct phosphorylation of IRS-1 and activation of PI3-K, leading to GLUT4 translocation, are involved. The raised phosphorylation of proteins in the insulin signaling pathway appears to be related to the inhibition of protein tyrosine phosphatase (PTPase) activity by vanadium salts. NOVEL EXPERIMENTS: The model utilized in our study was Psammomys obesus (sand rat), a desert gerbil which becomes hyperglycemic and hyperinsulinemic on an ad libitum high energy (HE) diet. In contrast to the previously investigated insulin deficient models, vanadyl sulphate was used to correct insulin resistance and hyperinsulinemia, which led to beta-cell loss. Administration of 5 mg/kg vanadyl sulfate for 5 days resulted in prolonged restoration of normoglycemia and normoinsulinemia in most animals, return of glucose tolerance to normal, and a reduction of hepatic phosphoenolpyruvate carboxykinase activity. There was no change in food consumption and in regular growth during or after the vanadyl treatment. Pretreatment with vanadyl sulfate, followed by transfer to a HE diet, significantly delayed the onset of hyperglycemia. Hyperinsulinemic-euglycemic clamp of vanadyl sulfate treated Psammomys demonstrated an improvement in glucose utilization. However, vanadyl sulfate was ineffective when administered to animals which lost their insulin secretion capacity on protracted HE diet, but substantially reduced the hyperglycemia when given together with exogenous insulin. The in vitro insulin activation of liver and muscle insulin receptors isolated from vanadyl treated Psammomys was ineffective. The in vivo vanadyl treatment restored muscle GLUT4 total protein and mRNA contents in addition to membrane GLUT4 protein, in accordance with the increased glucose utilization during the clamp study. These results indicate that short-term vanadyl sulfate treatment corrects the nutritionally induced, insulin resistant diabetes. This action requires the presence of insulin for its beneficial effect. Thus, vanadyl action in P. obesus appears to be the result of insulin potentiation rather than mimicking, with activation of the signaling pathway proteins leading to GLUT4 translocation, probably distal to the insulin receptor.

Effect of inhibition of glutathione synthesis on insulin action: in vivo and in vitro studies using buthionine sulfoximine.

Decreased cellular GSH content is a common finding in experimental and human diabetes, in which increased oxidative stress appears to occur. Oxidative stress has been suggested to play a causative role in the development of impaired insulin action on adipose tissue and skeletal muscle. In this study we undertook to investigate the potential of GSH depletion to induce insulin resistance, by utilizing the GSH synthesis inhibitor, L-buthionine-[S,R]-sulfoximine (BSO). GSH depletion (20-80% in various tissues), was achieved in vivo by treating rats for 20 days with BSO, and in vitro (80%) by treating 3T3-L1 adipocytes with BSO for 18 h. No demonstrable change in the GSH/GSSG ratio was observed following BSO treatment. GSH depletion was progressively associated with abnormal glucose tolerance test, which could not be attributed to impaired insulin secretion. Skeletal muscle insulin responsiveness was unaffected by GSH depletion, based on normal glucose response to exogenous insulin, 2-deoxyglucose uptake measurements in isolated soleus muscle, and on normal skeletal muscle expression of GLUT4 protein. Adipocyte insulin responsiveness in vitro was assessed in 3T3-L1 adipocytes, which displayed decreased insulin-stimulated tyrosine phosphorylation of insulin-receptor-substrate proteins and of the insulin receptor, but exaggerated protein kinase B phosphorylation. However, insulin-stimulated glucose uptake was unaffected by GSH depletion. In accordance, normal adipose tissue insulin sensitivity was observed in BSO-treated rats in vivo, as demonstrated by normal inhibition of circulating non-esterified fatty acid levels by endogenous insulin secretion. In conclusion, GSH depletion by BSO results in impaired glucose tolerance, but preserved adipocyte and skeletal muscle insulin responsiveness. This suggests that alternative oxidation-borne factors mediate the induction of peripheral insulin resistance by oxidative stress.

Lipoic acid protects against oxidative stress induced impairment in insulin stimulation of protein kinase B and glucose transport in 3T3-L1 adipocytes.

AIMS/HYPOTHESIS: Oxidative stress has been shown to impair insulin-stimulated glucose transporter 4 translocation in 3T3-L1 adipocytes. This study explores the potential of the antioxidant lipoic acid to protect the cells against the induction of insulin resistance when given before exposure to oxidative stress. METHODS: 3T3-L1 were exposed for 16 h to lipoic acid after which cells were exposed for 2 h to continuous production of H2O2 by adding glucose oxidase to the culture medium. RESULTS: These conditions resulted in a 50-70% reduction in insulin-stimulated glucose transport activity associated with a decrease in reduced glutathione content from 37.4 +/- 3.1 to 26.4 +/- 4.9 nmol/mg protein, (p < 0.005). Lipoic acid pretreatment increased insulin-stimulated glucose transport following oxidative stress, reaching 84.8 +/- 4.4% of the control, associated with an increase in reduced glutathione content. Oxidation impaired the 4.89 +/- 0.36-fold insulin-stimulated increase in glucose transporter 4 content in plasma membrane lawns of control cells. Lipoic acid pretreatment was, however, associated with preserved insulin-induced glucose transporter 4 translocation in cells exposed to oxidation, yielding 80% of its content in controls. Although tyrosine phosphorylation patterns were not affected by lipoic acid pretreatment, insulin-stimulated protein kinase B/Akt serine 473 phosphorylation and activity were considerably impaired by oxidation but protected by lipoic acid pretreatment. A protective effect was not observed with either troglitazone, its isolated vitamin E moiety, or with vitamin C. CONCLUSION/INTERPRETATION: This study shows the ability of lipoic acid to provide partial protection against the impaired insulin-stimulated glucose transporter 4 translocation and protein kinase B/Akt activation induced by oxidative stress, potentially by its capacity to maintain intracellular redox state.

Lipoic acid acutely induces hypoglycemia in fasting nondiabetic and diabetic rats.

Lipoic acid (LA) is a unique antioxidant that increases peripheral glucose utilization in diabetic patients. This study was conducted to investigate whether the inhibition of glucose production could be an additional mechanism for the action of LA. Intravenous (i.v.) LA injection (100 or 60 mg/kg body weight) to fasting nondiabetic or streptozotocin (STZ)-induced diabetic rats caused a rapid reduction in blood glucose with no effect on circulating insulin levels. In vivo conversion of fructose to glucose was not inhibited by LA, whereas the gluconeogenesis flux from alanine was completely prevented. Reduced liver pyruvate carboxylase (PC) activity in vivo is suggested by the finding that LA induced a decrease in liver coenzyme A (CoA) content (44% and 28% reduction in nondiabetic and diabetic rats, respectively, compared with vehicle-treated animals) and liver acetyl CoA content (80% and 67% reduction in nondiabetic and diabetic rats, respectively). A reduction in plasma free carnitine (42% and 22% in nondiabetic and diabetic rats, respectively) was observed in LA-treated animals, and acylcarnitine levels were increased twofold. This could be attributed to elevated levels of C16 and C18 acylcarnitine, without a detectable accumulation of lipoylcarnitine. Under such conditions, a significant increase in the plasma free fatty acid (FFA) concentration (204% in nondiabetic and 151% in diabetic animals) with no elevation in beta-hydroxybutyrate levels was noted. In conclusion, this study suggests that short-term administration of LA at high dosage to normal and diabetic rats causes an inhibition of gluconeogenesis secondary to an interference with hepatic fatty acid oxidation. This may render LA an antihyperglycemic agent for the treatment of diabetic subjects, who display glucose overproduction as a major metabolic abnormality.

Lipoic acid prevention of neural tube defects in offspring of rats with streptozocin-induced diabetes.

OBJECTIVE: Increased oxidant stress has been suggested to play a role in the pathogenesis of disturbed embryogenesis in diabetic pregnancies. The present study was conducted to determine whether administration of lipoic acid, a naturally occurring antioxidant, would reduce the incidence of diabetic embryopathy in the streptozocin-induced diabetic rat model. STUDY DESIGN: After conception, rats were randomly distributed to 5 groups. From day 1, rats were daily injected intraperitoneally with either lipoic acid, 30 mg/kg, or vehicle. At day 6, rats from groups 3, 4, and 5 were made diabetic by a single intraperitoneal injection of streptozocin. Group 4 rats were injected with lipoic acid from day 1 to day 6, after vehicle treatment until day 17. At day 17 of gestation, rats were killed. The fetuses were released from the yolk sacs and surrounding decidua and were examined for size, resorption rate, and neural tube defects. RESULTS: Pregnant diabetic rats treated with vehicle lost weight during pregnancy (-3.2 +/- 1.9 g/d), as opposed to normal pregnancy-related weight gain (3.5 +/- 0.5 g/d). Treatment with lipoic acid protected against diabetes-induced weight loss, without a measurable effect on fed-state glucose concentrations. Daily treatment with lipoic acid (pregnancy days 1 to 17) was efficient in reducing the resorption rate from 24.0% +/- 9.5% in vehicle-treated diabetic rats to 10.2% +/- 4.8% in lipoic acid-treated diabetic rats (P <.05). The rate of neural tube defects in diabetic rats treated with lipoic acid throughout the pregnancy was reduced from 26.0% +/- 7.0% to 10.2% +/- 3.2% (P <.05). In rats treated only during pregnancy days 1 to 5 (before diabetes induction), lipoic acid failed to exert its protective effects against neural tube defects, which emphasizes the importance of the presence of lipoic acid during the organogenesis period. The atherosis of placental vasculature demonstrated in the vehicle-treated diabetic rats was absent from placentas obtained from lipoic acid-treated diabetic animals. CONCLUSIONS: Our data demonstrate a protective effect of lipoic acid against diabetic embryopathy, fetal losses, and ultrastructural alteration of diabetic placentas.

Metabolic effects of gamma-linolenic acid-alpha-lipoic acid conjugate in streptozotocin diabetic rats.

Data suggesting the involvement of increased oxidative stress in the pathophysiology of diabetes has raised interest in the potential therapeutic benefit of antioxidants. Although beneficial metabolic effects of antioxidant supplementation have been suggested, an antioxidant mode of action, particularly in skeletal muscle, has not been documented. In the present study, we evaluate the metabolic effects of a gamma-linolenic acid-alpha-lipoic acid conjugate (GLA-LA) in streptozotocin-induced diabetic rats, and assess its potential mode of action by comparing its effects with equimolar administration of LA and GLA alone. Ten days of oral supplementation of 20 mg/kg body weight GLA-LA, but not LA or GLA alone, caused a mild reduction in fasting blood glucose concentration as compared with vehicle-treated diabetic rats (375 +/- 11 vs. 416 +/- 16 mg/dl, p = 0.03), with no change in fasting plasma insulin levels. A peripheral insulin-sensitizing effect could be observed with GLA-LA, LA, and GLA treatments, as demonstrated by a significant (p < 0.04) 23%, 13%, and 10% reduction, respectively, in the area under the glucose curve following an intravenous insulin tolerance test. This effect was associated with a 67% and 50% increase in GLUT4 protein content in the membranes of gastrocnemius muscle of GLA-LA and LA-treated animals, respectively; however, no change was observed with GLA treatment alone. Interestingly, both GLA-LA and LA treatments corrected a diabetes-related decrease in the gastrocnemius muscle low-molecular-weight reduced thiols content. These data demonstrate insulin-sensitizing properties of the GLA-LA conjugate by distinct mechanisms attributable to each of its components, which are associated with antioxidant effects.

Lipoic acid reduces glycemia and increases muscle GLUT4 content in streptozotocin-diabetic rats.

Alpha lipoic acid (lipoate [LA]), a cofactor of alpha-ketodehydrogenase, exhibits unique antioxidant properties. Recent studies suggest a direct effect of LA on glucose metabolism in both human and experimental diabetes. This study examines the possibility that LA positively affects glucose homeostasis in streptozotocin (STZ)-induced diabetic rats by altering skeletal muscle glucose utilization. Blood glucose concentration in STZ-diabetic rats following 10 days of intraperitoneal (i.p.) injection of LA 30 mg/kg was reduced compared with that in vehicle-treated diabetic rats (495 +/- 131 v 641 +/- 125 mg/dL in fed state, P = .003, and 189 +/- 48 v 341 +/- 36 mg/dL after 12-hour fast, P = .001). No effect of LA on plasma insulin was observed. Gastrocnemius muscle crude membrane GLUT4 protein was elevated both in control and in diabetic rats treated with LA by 1.5- and 2.8-fold, respectively, without significant changes in GLUT4 mRNA levels. Gastrocnemius lactic acid was increased in diabetic rats (19.9 +/- 5.5 v 10.4 +/- 2.8 mumol/g muscle, P < .05 v nondiabetic rats), and was normal in LA-treated diabetic rats (9.1 +/- 5.0 mumol/g muscle). Insulin-stimulated 2-deoxyglucose (2 DG) uptake into isolated soleus muscle was reduced in diabetic rats compared with the control group (474 +/- 15 v 568 +/- 52 pmol/mg muscle 30 min, respectively, P = .05). LA treatment prevented this reduction, resulting in insulin-stimulated glucose uptake comparable to that of nondiabetic animals. These results suggest that daily LA treatment may reduce blood glucose concentrations in STZ-diabetic rats by enhancing muscle GLUT4 protein content and by increasing muscle glucose utilization.