Claudio Taboga

Evidence for an Independent and Cumulative Effect of Postprandial Hypertriglyceridemia and Hyperglycemia on Endothelial Dysfunction and Oxidative Stress Generation Effects of Short- and Long-Term Simvastatin Treatment

Background—Postprandial hypertriglyceridemia and hyperglycemia are considered risk factors for cardiovascular disease. Evidence suggests that postprandial hypertriglyceridemia and hyperglycemia induce endothelial dysfunction through oxidative stress; however, the distinct role of these two factors is a matter of debate. Methods and Results—Thirty type 2 diabetic patients and 20 normal subjects ate 3 different meals: a high-fat meal; 75 g glucose alone; and high-fat meal plus glucose. Glycemia, triglyceridemia, nitrotyrosine, and endothelial function were assayed during the tests. Subsequently, diabetics took 40 mg/d simvastatin or placebo for 12 weeks. The 3 tests were performed again at baseline, between 3 to 6 days after the start, and at the end of each study. High-fat load and glucose alone produced a decrease of endothelial function and an increase of nitrotyrosine in normal and diabetic subjects. These effects were more pronounced when high fat and glucose were combined. Short-term simvastatin treatment had no effect on lipid parameters but reduced the effect on endothelial function and nitrotyrosine observed during each different test. Long-term simvastatin treatment was accompanied by a lower increase in postprandial triglycerides, which was followed by smaller variations of endothelial function and nitrotyrosine during the tests. Conclusions—This study shows an independent and cumulative effect of postprandial hypertriglyceridemia and hyperglycemia on endothelial function, suggesting oxidative stress as common mediator of such effect. Simvastatin shows a beneficial effect on oxidative stress and endothelial dysfunction, which may be ascribed to a direct effect as well as the lipid-lowering action of the drug.

Meal-Generated Oxidative Stress in Type 2 Diabetic Patients

OBJECTIVE Free radical production has been reported to be increased in diabetic patients and to be involved in the pathogenesis of diabetic complications. In this study, a standardized meal was administered to 10 type 2 diabetic patients and 10 healthy matched normal subjects to evaluate its effects on plasma oxidative stress generation. RESEARCH DESIGN AND METHODS In diabetic patients, at baseline and after the meal, plasma malondialdehyde (MDA), vitamin C, protein SH groups, uric acid, vitamin E, and total plasma radical-trapping parameter, which evaluates plasma antioxidant capacity due to known and unknown antioxidants present in the plasma as well as their mutual cooperation, were measured. RESULTS After the meal, plasma MDA and vitamin C increased, while protein SH groups, uric acid, vitamin E, and total plasma radical-trapping parameter decreased more significantly in the diabetic subjects than in control subjects. CONCLUSIONS This finding shows that in the absorptive phase, free radicals are produced in diabetic patients. Since plasma glucose, but not insulin, rose significantly more in diabetic subjects than in control subjects, hyperglycemia may play an important role in the generation of postprandial oxidative stress in diabetic patients.

Meal-induced oxidative stress and low-density lipoprotein oxidation in diabetes: The possible role of hyperglycemia

Oxidative stress and its contribution to low-density lipoprotein (LDL) oxidation have been implicated in the pathogenesis of vascular diabetic complications. However, the relationship between hyperglycemia, hyperinsulinemia, hyperlipidemia, and oxidative stress is still debated. If plasma glucose and/or insulin and/or lipid are some of the most important determinants of oxidative stress in diabetes, then their typical postprandial elevations in diabetes would be expected to favor oxidative stress and LDL oxidation. To test this hypothesis, in type 2 diabetic patients, we evaluated the effects of two different standard meals designed to produce different levels of postprandial hyperglycemia on the plasma oxidative status and LDL oxidation. The meals were administered in randomized order to each of 10 type 2 diabetic patients. Blood samples were collected at baseline and 60 and 120 minutes after the meals. In every sample, plasma levels of glucose, insulin, cholesterol, triglycerides, nonesterified fatty acids (NEFAs), malondialdehyde (MDA), and the total radical-trapping antioxidant parameter (TRAP) were measured. LDL susceptibility to oxidation was evaluated at baseline and after 120 minutes. Plasma glucose, insulin, triglycerides, and MDA increased and NEFAs and TRAP significantly decreased after either meal. The variations in plasma glucose, MDA, and TRAP were significantly greater and LDL was more susceptible to oxidation after the meal that produced a significantly higher degree of hyperglycemia. These results suggest that postprandial hyperglycemia may contribute to oxidative stress in diabetic patients, providing a mechanistic link between hyperglycemia and diabetic vascular disease.

Hyperglycemia-Induced Thrombin Formation in Diabetes: The Possible Role of Oxidative Stress

Diabetes is characterized by the existence of a thrombosis-prone condition, possibly related to hyperglycemia. However, the mechanism linking hyperglycemia to the activation of the coagulation cascade is still unclear. It has been recently suggested that diabetes is accompanied by increased oxidative stress. In this work, the possibility that oxidative stress may be involved in the hyperglycemia-induced coagulation activation has been evaluated. Prothrombin fragment 1 + 2 (F1+2), which represents a reliable marker of the amount of thrombin released in the circulation, has been chosen for studying thrombin formation in vivo. In nine type II diabetic patients and in seven healthy control subjects, matched for age and body mass index, three different experiments were performed: oral glucose tolerance test (OGTT), intravenous antioxidant glutathione (GSH) administration for 2 h, and OGTT plus intravenous GSH administration. Samples were drawn at −15 min and every 30 min from 0 to 180 min. During the OGTT, F1+2 significantly increased in both diabetic and healthy subjects. GSH administration during OGTT normalized this phenomenon. GSH administered alone significantly decreased F1+2 in diabetic patients, while no effect was observed in the normal subjects. These data suggest that hyperglycemia may induce thrombin activation, possibly inducing an oxidative stress, and that antioxidant GSH may counterbalance this effect.

Total Radical-Trapping Antioxidant Parameter in NIDDM Patients

OBJECTIVE The existence of an oxidative stress in diabetes is still debated. This is largely due to the lack of good tools to assay the level of oxidative stress. The use of total radical-trapping antioxidant parameter (TRAP) has recently been proposed to explore the antioxidant property of a plasma sample. TRAP may be either directly measured by a fluorescence-based method (TRAPm) or calculated (TRAPc) by a mathematical formula, taking into account the serum levels of four natural antioxidants: protein-bound SH (thiol) groups, uric acid, vitamin E, and vitamin C. The difference between TRAPm and TRAPc is due to antioxidants, which are still unidentified, and to the possible synergism among the antioxidants. RESEARCH DESIGN AND METHODS In this study, we evaluated malondialdehyde (MDA), TRAPm, TRAPc, protein-bound SH groups, uric acid, vitamin E, and vitamin C in 40 NIDDM patients and 40 matched normal control subjects. RESULTS TRAPm and TRAPc were significantly lower in diabetic patients. A good correlation between TRAPm and TRAPc was found in both NIDDM patients (r = 0.68, P < 0.0001) and control subjects (r = 0.74, P < 0.0001). Protein-bound SH groups and uric acid were significantly lower in diabetic subjects, while MDA and vitamin E level were significantly higher. After correction for serum triglycerides (MDA) and cholesterol (vitamin E), MDA lost significance, while vitamin E did not. Vitamin C was not different in the two groups. CONCLUSIONS These data show decreased TRAP levels in NIDDM patients, suggesting the existence of lower antioxidant defenses in diabetes. The decrease appears to be due to various antioxidants, some of them not yet clearly defined. TRAP may represent a more reliable estimation of serum antioxidant capacity than the measurement of each known antioxidants. The correlation found between TRAPm and TRAPc values suggests that TRAPc, easier to measure than TRAPm, might be adequately reliable for routine assessment of oxidative stress in diabetic patients.

Fibrinogen Plasma Levels as a Marker of Thrombin Activation in Diabetes

This study attempted to verify the existence of a correlation between fibrinogen, a major cardiovascular risk factor in diabetes, and indexes of thrombin generation and action, prothrombin fragment 1 + 2 (F1 + 2), and D-dimer (D-D), in a group of diabetic subjects compared with a matched control group. Forty insulin-dependent diabetes mellitus patients and 30 matched healthy control subjects participated in this study. The subjects were tested for the following parameters: fibrinogen, prothrombin F1 + 2, D-D, fasting glycemia, and HbA1c. In addition, 5 diabetic subjects who maintained stable fibrinogen plasma levels > 300 mg/dl for at least 6 months before the study were treated with 12,500 U/day subcutaneous heparin for 7 days. Diabetic subjects showed increased levels of fibrinogen, prothrombin F1 + 2, and D-D plasma levels. Simple linear regression analysis detected a positive correlation between fibrinogen and prothrombin F1 + 2, D-D, and glycosylated HbA1c. In the five diabetic subjects treated with heparin fibrinogen, prothrombin F1 + 2 and D-D levels decreased at the end of the treatment. All these parameters returned to baseline after 7 days of washout. These data indicate that fibrinogen plasma levels are correlated to parameters of thrombin activation in plasma in diabetic patients and suggest that high fibrinogen plasma levels might be a risk marker for cardiovascular disease in diabetes because it is an expression of an existing thrombophilia.

Total Plasma Antioxidant Capacity Predicts Thrombosis-Prone Status in NIDDM Patients

OBJECTIVE To explore the hypothesis that a relationship exists between free radical activity and abnormalities in hemostasis in NIDDM. RESEARCH DESIGN AND METHODS The use of the total radical-trapping antioxidant parameter (TRAP) has very recently been proposed to explore the antioxidant property of a plasma and their mutual cooperation. In the present study, TRAP, vitamin E, vitamin C, vitamin A, uric acid, protein-bound SH (thiol) groups, fibrinogen, prothrombin fragments F1 + 2, and D-dimer have been evaluated in 46 NIDDM patients and 47 healthy matched control subjects. RESULTS In NIDDM patients, TRAP, vitamin A, SH groups, and uric acid were significantly reduced, whereas the level of vitamin E was significantly increased. Vitamin C was similar in the two groups. Fibrinogen, prothrombin fragment 1 + 2, and D-dimer were increased in diabetic patients. TRAP, but no single other antioxidant, had a strong inverse association with fibrinogen, prothrombin fragment 1 + 2, and D-dimer. CONCLUSIONS These findings are consistent with the hypothesis that oxidative stress may condition coagulation activation in diabetics. However, the data suggest that it is the total antioxidant capacity rather than any single plasma antioxidant that is the most relevant parameter.

Increased prothrombin fragment 1 + 2 in type I diabetic patients.

Dr. Antonio Ceriello, Cattedra di Medicina Interna, Facoltà di Medicina, Università di Udine, P.le S. Maria della Misericordia, I-33100 Udine (Italy) The pathogenesis of vascular lesions in diabetic patients has been considered to be at least partly dependent on the alterations in the hemostatic system [1]. However, the existence and the relevance of a hypercoagulable state in diabetes mellitus have been the subject of much debate [1,2]. It has been demonstrated that the conversion of the coagulation zymogen prothrombin to thrombin is associated with the prominent production of a cleavage product namely prothrombin fragment 1+2 (Fl+2) [3]. It has been proposed recently that prothrombin Fl+2 plasma levels may be considered a very sensitive marker for hypercoagulable states in humans [4]. We evaluated prothrombin Fl+2‚(ELISA) levels in 19 insulin-dependent diabetic patients without signs of vascular complications (12 males and 7 females: age 23.4 ± 1.9 years, mean ± SE; body mass index 22.8 ± 1.4; duration of diabetes 6.4 ± 1.3 years; insulin regimen 25-55 U/day, mean 32.8 ± 2.5 U/ day; all subjects had 24-hour urinary albumin excretion rates less than 30 μg/min, and no microaneurysms were detected on full fundal photographs or íluorescein fundal angiogra-phy; they had systolic blood pressures < 120 mm Hg and diastolic blood pressures < 90 mm Hg; ischemic heart disease and peripheral vascular occlusion were excluded according to local criteria) compared to 10 matched healthy normal subjects (6 males and 4 females: age 24.2 ± 1.8 years, body mass index 23.2 ± 1.3). Prothrombin Fl+2 levels were significantly elevated in diabetic patients (0.69 ± 0.11 vs. 0.27 ± 0.03 nmol/l;p < 0.01), but no correlation was found between prothrombin Fl+2 plasma levels and both fasting glycemia (r = 0.17) and glycosylated Hb A1 c (r = 0.16). Increased FPA levels have been reported in diabetes [5]. These findings were considered more suggestive of a thrombin hyperac-tivity than of an increase of thrombin production [6]. This hypothesis was sustained by the evidence of normal [7] or depressed [6] levels of thrombinantithrombin complex in the presence of increased FPA [6] and fibrin monomers [7] in diabetes.