Jason Y. Hokama

The antioxidant α-lipoic acid enhances insulin-stimulated glucose metabolism in insulin-resistant rat skeletal muscle

Insulin resistance of muscle glucose metabolism is a hallmark of NIDDM. The obese Zucker (fa/fa) rat—an animal model of muscle insulin resistance—was used to test whether acute (100 mg/kg body wt for 1 h) and chronic (5–100 mg/kg for 10 days) parenteral treatments with a racemic mixture of the antioxidant α-lipoic acid (ALA) could improve glucose metabolism in insulin-resistant skeletal muscle. Glucose transport activity (assessed by net 2-deoxyglucose [2-DG] uptake), net glycogen synthesis, and glucose oxidation were determined in the isolated epitrochlearis muscles in the absence or presence of insulin (13.3 nmol/1). Severe insulin resistance of 2-DG uptake, glycogen synthesis, and glucose oxidation was observed in muscle from the vehicle-treated obese rats compared with muscle from vehicle-treated lean (Fa/−) rats. Acute and chronic treatments (30 mgkg−1 · day−1, a maximally effective dose) with ALA significantly (P < 0.05) improved insulin-mediated 2-DG uptake in epitrochlearis muscles from the obese rats by 62 and 64%, respectively. Chronic ALA treatment increased both insulin-stimulated glucose oxidation (33%) and glycogen synthesis (38%) and was associated with a significantly greater (21%) in vivo muscle glycogen concentration. These adaptive responses after chronic ALA administration were also associated with significantly lower (15–17%) plasma levels of insulin and free fatty acids. No significant effects on glucose transporter (GLUT4) protein level or on the activities of hexokinase and citrate synthase were observed. Collectively, these findings indicate that parenteral administration of the antioxidant ALA significantly enhances the capacity of the insulinstimulatable glucose transport system and of both oxidative and nonoxidative pathways of glucose metabolism in insulin-resistant rat skeletal muscle.

Stimulation by α-lipoic acid of glucose transport activity in skeletal muscle of lean and obese Zucker rats

Abstract α-Lipoic acid (ALA), a potent biological antioxidant, improves insulin action of skeletal muscle glucose transport and metabolism in both human and animal models of insulin resistance. In order to obtain further insight into the potential intracellular mechanisms for the action of ALA on insulin-stimulated glucose transport in skeletal muscle, we investigated the effects of direct incubation with ALA (2 mM) on 2-deoxyglucose (2-DG) uptake by epitrochlearis muscle from either insulin-sensitive lean (Fa/-) or insulinresistant obese ( fa fa ) Zucker rats. ALA stimulated 2-DG uptake in muscle of lean animals by 76%, whereas ALA stimulated 2-DG uptake by only 48% in muscle from obese animals. The stimulation of 2-DG uptake due to ALA was enhanced 30–55% in the presence of insulin. In contrast, ALA action on 2-DG uptake was not additive with the effects of electrically-stimulated muscle contractions in either insulin-sensitive or insulin-resistant muscle. Wortmannin (1 μM), an inhibitor of phosphotidylinositol-3-kinase, completely inhibited insulin action on 2-DG uptake, but inhibited ALA action by only 25%. Collectively, these results indicate that although a portion of ALA action on glucose transport in mammalian skeletal muscle is mediated via the insulin signal transduction pathway, the majority of the direct effect of ALA on skeletal muscle glucose transport is insulin-independent.

Diabetes enhances leukocyte accumulation in the coronary microcirculation early in reperfusion following ischemia

BACKGROUND Diabetic hearts are particularly vulnerable to ischemia-reperfusion injury. For leukocytes to participate in ischemia-reperfusion injury, they must first sequester in the microcirculation. The aim of this study was to determine, by direct observation, if early leukocyte deposition was increased in the diabetic coronary microcirculation early in reperfusion following myocardial ischemia. METHODS Non-diabetic and streptozotocin (STZ)-induced diabetic rat hearts, subjected to 30 min of 37 degrees C, no-flow ischemia, were initially reperfused with blood containing labeled leukocytes. The deposition of fluorescent leukocytes in coronary capillaries and venules was directly visualized and recorded using intravital fluorescence microscopy. In addition, flow cytometry was used to measure CD11b adhesion molecule expression on polymorphonuclear (PMN) leukocytes from non-diabetic and STZ-diabetic rats. RESULTS In the non-diabetic, control hearts, early in reperfusion, leukocytes trapped in coronary capillaries and adhered to the walls of post-capillary venules. In the diabetic hearts, leukocyte trapping in capillaries and adhesion to venules were both significantly increased (P<0.05). PMN CD11b expression was also significantly increased in the diabetic blood compared to the non-diabetic blood (P<0.05). CONCLUSIONS Early in reperfusion following myocardial ischemia, leukocytes rapidly accumulate in greater numbers in the coronary microcirculation of the diabetic heart by both trapping in coronary capillaries and by adhering to venules. The enhanced retention of leukocytes in the diabetic coronary microcirculation increases the likelihood of inflammation-mediated reperfusion injury and may explain, in part, the poor recovery of diabetic hearts from an ischemic event.

The Blood Contribution to Early Myocardial Reperfusion Injury Is Amplified in Diabetes

Cardiovascular disease is excessive in diabetes, and blood cell function is altered. It is not clear, however, if alterations in the blood contribute to the excessive cardiovascular complications of this disease. In this study, we compared the contribution of nondiabetic and diabetic blood to myocardial reperfusion injury. The recovery of cardiac contractile function following no-flow ischemia was studied in isolated diabetic and nondiabetic rat hearts perfused with diabetic or nondiabetic diluted whole blood. Hearts were isolated from 10- to 12-week-old diabetic (streptozotocin, 65 mg/kg, i.v.) and nondiabetic rats and perfused with a Krebs-albumin-red cell solution (K2RBC, Hct 20%). After a 30-min pre-ischemic control period, during which cardiac pump function was evaluated, diabetic and nondiabetic hearts were perfused for 5 min with diluted whole blood (DWB; Hct 20%) collected from either diabetic or nondiabetic donor animals. Coronary flow was then stopped and the hearts subjected to 30 min of no-flow ischemia. Following ischemia, the hearts were reperfused with the K2RBC perfusate. Cardiac contractile function was evaluated throughout the 60-min reperfusion period. Six groups were studied: diabetic and nondiabetic hearts perfused before ischemia with either K2RBC, nondiabetic DWB (NDDWB), or diabetic DWB (DDWB). Perfusion with DWB prior to ischemia impaired the recovery of contractile function in all cases. The impairment to recovery was greater with DDWB than with NDDWB. Although diabetic hearts perfused with K2RBC throughout recovered quite well, the effect of DDWB perfusion in the diabetic hearts was dramatic. In an effort to determine why diabetic blood impaired functional recovery, measures of blood filterability and the generation of reactive oxygen species (ROS) were made. We found that diabetic blood was less filterable than nondiabetic blood; that is, the diabetic blood cells tended to plug the 5-μm filter pores more readily than the nondiabetic blood cells. Also, we found that the diabetic blood was capable of generating significantly greater ROS (oxygen free radicals) than nondiabetic blood (P < 0.05). These findings suggest that the blood contribution to myocardial reperfusion injury is amplified in diabetes. A tendency for diabetic blood cells to plug capillary-sized pores and show enhanced oxygen free radical production may account for the excessive contribution of diabetic blood to reperfusion injury in the heart.

Chronic expression of platelet adhesion proteins is associated with severe ischemic heart disease in type 2 diabetic patients: Chronic platelet activation in diabetic heart patients

Cardiac ischemia is a serious complication of type 2 diabetes. However, the pathophysiology underlying the increased severity of myocardial ischemia in diabetes is not clear. This study tested the hypothesis that platelet adhesion protein expression is chronically increased in older type 2 diabetic patients with established ischemic heart disease (IHD) compared to age-matched, nondiabetic patients with IHD. We compared the chronic expression of two platelet adhesion proteins, P-selectin and GPIIb/IIIa, in whole blood and the platelet reactivity to an acute stimulus. We found that the expression of platelet P-selectin was chronically increased in the nondiabetic patients with IHD compared to normal subjects. P-selectin expression was further increased in the diabetic patients with IHD compared to the nondiabetic IHD patients (P<.05). The results were stratified to examine the potential effect of aspirin usage on adhesion protein expression. We found that the expression of the activated GPIIb/IIIa complex was significantly reduced in those diabetic cardiac patients who were taking aspirin (P<.05). These findings indicate that, in patients with IHD, platelet adhesion proteins are chronically expressed and that the level of expression is increased more in IHD patients with type 2 diabetes. This complication of diabetes may exacerbate thrombus formation during a recurrent event, increasing the severity of ischemic injury. The results give further support to the use of aspirin in type 2 diabetics with established cardiac disease.

Pentoxifylline reduces coronary leukocyte accumulation early in reperfusion after cold ischemia

BACKGROUND Ischemia/reperfusion injury can complicate recovery in cardiac operations. Ischemia induces endothelial dysfunction, which may contribute to leukocyte accumulation during reperfusion. Leukocyte-mediated injury may then occur. Using intravital microscopy we previously reported increased leukocyte retention in coronary capillaries and venules during early reperfusion during warm ischemia/reperfusion. In this study we investigated whether cold cardioplegic protection would limit leukocyte sequestration in coronary microvessels early in reperfusion. Pentoxifylline (PTX) has antiinflammatory effects and may limit endothelial dysfunction during ischemia/reperfusion. The effect of cardioplegia modification with PTX was also examined. METHODS Isolated rat hearts were subjected to 90 minutes of 4 degrees C ischemia after arrest with cardioplegia. Hearts were reperfused with diluted whole blood containing fluorescent-labeled leukocytes. Leukocyte retention in coronary microvessels was observed with intravital microscopy. Three groups were studied, nonischemic control, cold ischemia, and PTX-modified cold ischemia. RESULTS In coronary capillaries, leukocyte trapping was nearly doubled in unmodified cold ischemia versus control. PTX modification significantly reduced leukocyte accumulation. In coronary venules, greater leukocyte adhesion was observed in unmodified cold ischemia compared to nonischemic controls. PTX modification significantly reduced leukocyte adhesion. CONCLUSIONS Cold cardioplegia did not prevent leukocyte retention in the coronary microcirculation early in reperfusion. PTX modification of cardioplegia significantly reduced leukocyte sequestration in coronary capillaries and venules. Preserving endothelial function during ischemia may limit leukocyte accumulation and ischemia/reperfusion injury after cardiac operation.

Cold cardioplegic preservation aggravates alloantibody mediated cardiac dysfunction due to an increase in vascular resistance

IN HEART TRANSPLANTATION, hyperacute rejection due to preformed antibodies is characterized by a rapid, irreversible, and complete graft dysfunction. However, when antibodies appear later in the posttransplantation course, the dysfunction they mediate can be reversible (acute humoral rejection). Hori et al recently reported that ischemia-reperfusion injury played a major role in the etiology of hyperacute rejection of cardiac xenografts. However, whether ischemia-reperfusion injury enhances the severity of hyperacute allograft rejection has not yet been investigated. The aim of this study was to investigate the role of cold ischemiareperfusion injury in alloantibody-mediated cardiac dysfunction in an isolated heart model.