R. Lo Presti

Diabetes mellitus: oxidative stress and wine

SUMMARY This review focuses on the link between diabetes mellitus and oxidative stress and, in particular, on the role that moderate wine consumption may play in preventing diabetic complications and the onset of diabetes. With this aim, a search of PubMed was carried out for literature published up to March In diabetes mellitus, oxidative stress results both from exposure to hyperglycaemia through glycoxidation and sorbitol system activation, and from functional limitation of the hexose monophosphate shunt, leading to a decrease in glutathione synthesis. Oxidative stress alters the plasma lipoprotein profile (particularly low- density lipoproteins), the coagulative parameters (with an increased thrombotic risk), the endothelium (with a decrease in prostacyclin synthesis and an increase of thromboxane production) and the cell membranes (which undergo peroxidation). In diabetic patients, an altered oxidative pattern is present not only in the fasting state but also especially after food intake. In particular, food intake induces a decrease in the total radical-trapping antioxidant parameter (TRAP) and an elevation of hydroperoxides and thiobarbituric acid reactive substances (TBARS). Previously several clinical trials tried to improve the diabetic oxidative status using α-tocopherol, ascorbic acid and β-carotene supplementation. Some authors found, in normal subjects, a reduction of hydroperoxides postprandially when the meal included red wine. Other authors showed that the oxidative pattern present in type 2 diabetic patients was mitigated by red wine. These actions may reduce cardiovascular risk. Moreover, an inverse relationship was observed between alcohol consumption and the incidence of type 2 diabetes; this relationship was valid for a light to moderate intake and it seemed to depend on drinking regularly and to be independent of the type of alcoholic beverage. In conclusion, moderate and regular wine consumption could ameliorate the diabetic oxidative status. This lifestyle measure might contribute to preventing diabetic complications and the onset of diabetes.

Role of genetic polymorphisms in myocardial infarction at young age.

Acute myocardial infarction (AMI) in young adult presents a typical pattern of risk factors, clinical, angiographic and prognostic characteristics. In the last years we demonstrated that hemorheological profile is altered in these patients in a persistent way and independently of the number of risk factors and of the extent of coronary lesions. Thus, the hyperviscosity syndrome following AMI could be considered an intrinsic characteristic of these patients. Consequently it is possible to hypothesise the presence of a genetic background at the origin of this predisposition. If this background is able to influence the risk of ischemic heart disease, this should be particularly evident in young subjects. Since inflammatory mechanisms play a central role in mediating all phases of atherosclerosis, genes encoding for inflammatory or anti-inflammatory molecules are candidates for the risk of developing atherosclerosis. As atherosclerosis is the first cause of mortality in Western countries and if pro-inflammatory genotypes contribute to risk of coronary heart disease, alleles associated to disease susceptibility should not be included in the genetic background favouring longevity: People genetically predisposed to a weak inflammatory activity have fewer chances to develop cardiovascular disease and, therefore, have better chance for a long-life. According to this hypothesis, we studied in our population of young patients with AMI, the distribution of some polymorphisms influencing a inflammation and found an higher prevalence of pro-inflammatory polymorphisms (SNP A2080G of pyrin gene, SNP Gly670Arg of PECAM gene, C1019T of Cx 37 gene, SNP G1059C of PCR gene) and a lower prevalence of anti-inflammatory polymorphisms (Asp299Gly of TLR4 gene, SNP -1082 G/A of IL10 gene, CCR5Δ32). Results of these studies show that early myocardial infarction could be associated with a genetic predisposition to an intense inflammatory response, associated also to an hyperviscosity syndrome.

Techniques to evaluate erythrocyte deformability in diabetes mellitus

Abstract.Using several rheological techniques, we examined erythrocyte deformability in different groups of diabetic subjects. The macrorheological techniques used for this evaluation were respectively whole-blood filtration, filtration of erythrocyte suspensions, polyviscosimetry and diffractometry. Whole-blood filterability, at a negative pressure of 20 cm water, was decreased in type 2 diabetics; no difference was evident at a negative pressure of 10 cm water. The filtration of erythrocyte suspensions at low haematocrit (5%) did not show differences between normal and diabetic subjects. Polyviscosimetry, which explores the filterability of erythrocyte suspensions at high haematocrit (80%) through wide pores, demonstrated an impaired behaviour especially in type 2 diabetics. Diffractometry, which measures erythrocyte elongation induced by a defined shear stress through the diffraction pattern of a laser beam, showed an alteration in type 1 diabetic subjects. The microrheological methods employed for this evaluation were those based on fluorescence spectroscopy. Labeling intact red blood cells with fluorescent probes, we determined the membrane dynamic properties and using these techniques we found a reduction of erythrocyte membrane fluidity and a decrease of red cell membrane protein lateral mobility.

Nitric oxide metabolites and oxidative stress in mild essential hypertension

Many papers have showed non univocal data about oxidative stress status and nitric oxide metabolites in essential hypertension. Considering this preamble we examined the total antioxidant status (TAS), the lipid peroxidation (LP), expressed as thiobarbituric acid-reactive substances (TBARS), the stable end products of nitric oxide (NOx) and LP/NOx ratio in 25 subjects with untreated mild essential hypertension. The obtained data show a significant increase in TBARS (p < 0.001) and NOx (p < 0.001) in hypertensives and no variation in TAS and in TBARS/NOx ratio. None of these parameters was statistically related to the metabolic parameters or to the blood pressure values. The high level of lipid peroxidation observed in this group of hypertensives suggests the timely and specific employment of antihypertensive and antioxidant agents while the NOx increase seems to confirm the inflammatory status accompanying this clinical condition.

Screening and study enrolment in the Randomized Evaluation of Normal vs. Augmented Level (RENAL) Replacement Therapy Trial

Background and Objectives: Aspects of trial design, screening and study efficiency can affect recruitment and the findings of the trial itself. A clear understanding of the screening and study inclusion process will assist clinicians in interpreting trial results. Design: Prospective observational data collection on all patients screened for possible inclusion in a randomized controlled trial of normal vs. augmented renal replacement therapy in critically ill patients (the RENAL Trial). Setting: 35 hospitals in Australia and New Zealand. Participants: All patients screened for the RENAL Trial. Results: We screened 4,551 patients. Of these patients, 767 were ineligible because of lack of inclusion criteria and 2,085 because of exclusion criteria. Of the remaining 1,699, 1,508 (88.7%) were enrolled. The three most common exclusion criteria which prevented recruitment of potentially eligible patients were that the patient had end-stage kidney failure and was already on chronic dialysis (484; 23.2%), the patient’s body weight was either <60 or >120 kg (456; 21.8%), and the fact that the patient had already received renal replacement therapy during the index admission. Important modifiable impediments to recruitment were inability to obtain consent in 191 cases, unavailability of research staff in 124 cases, physician objection in 89 cases, and inability to deliver the trial protocol in 78 cases. Conclusion: The RENAL Trial’s enrolment efficiency was high and compared favourably with previous large intensive care units trials and with that of trials in patients with acute renal failure. The high rate of enrolment suggests that the results can be applied with confidence to most patients with de novo acute renal failure. The loss of close to 1.5% of patients due to consent issues highlights a common problem in critical care trials. The low rate of physician objection suggests clinical equipoise.

Nitric oxide metabolites, leukocyte activation markers and oxidative status in dialyzed subjects.

Aims: Our purpose was to evaluate, in a group of 42 end-stage renal disease patients who regularly undergo hemodialysis, some indexes of leukocyte activation, nitric oxide metabolites (NOx) and other parameters that reflect the oxidative stress before and after a standard hemodialysis session. Methods: Elastase and myeloperoxidase (MPO) were determined by means of ELISA. The NO production was evaluated by a micromethod which measures the concentration of NOx. The oxidation of polyunsaturated fatty acids was evaluated in plasma by detection of thiobarbituric acid-reactive substances (TBARS). Total antioxidant status (TAS) was obtained using spectrophotometry. Results: At baseline, we observed an increase of elastase, NOx, TBARS and TAS, without any variation of MPO. After the dialysis session, we found an increase in elastase and MPO, a decrease in NOx and TAS and no variation in TBARS. No modification occurred after subdividing the patients in two subgroups. Conclusions: Our data confirm the involvement of leukocytes and oxidative stress in hemodialysis patients.

Polymorphonuclear leukocyte integrin pattern, at baseline and after activation, in type 2 diabetic subjects with macrovascular complications

Abstract. In vascular atherosclerotic disease and in diabetes mellitus few studies have evaluated the polymorphonuclear leukocyte (PMN) adhesion molecule pattern. In this study we examined the PMN integrin expression at baseline and after activation in controls and type 2 diabetic subjects with macrovascular complications (MVC). We enrolled 21 subjects with type 2 diabetes mellitus and macrovascular complications, localized in peripheral, coronary and cerebral sites. The patients had peripheral occlusive arterial disease, chronic cerebrovascular disease or coronary heart disease. We evaluated the expression of some PMN integrins (CD11a, CD11b, CD11c, CD18), using flow cytofluorimetry, at baseline and after in vitro activation with 4-phorbol-12-myristate-13 acetate. Type 2 diabetic subjects with MVC showed, compared to normals, an increase of CD11a and CD18 and a decrease of CD11b and CD11c. After activation, in PMNs of normal subjects, we found an increase in the expression of all adhesion molecules, while in PMNS of type 2 diabetic subjects with MVC we observed an increase of CD11b and CD11c and a decrease of CD11a. In type 2 diabetic patients with MVC the basal upregulation of CD11a and CD18 may be related to the PMN spontaneous activation, while the behavior of CD11b may depend on its selfconsumption. After activation the CD11a modification may be due to its cleavage or to an altered integrin phosphorylation/dephosphorylation balance.

Polymorphonuclear leukocyte membrane fluidity before and after activation in subjects with insulin resistance.

Abstract The aim of this research was the evaluation of polymorphonuclear leukocyte (PMN) membrane fluidity in subjects with insulin resistance. Insulin sensitivity, in fact, may be influenced by plasma membrane fluidity. We enrolled 19 subjects with insulin resistance previously demonstrated during an euglycemic hyperinsulinemic clamp. PMN membrane fluidity was studied by labeling intact cells with the fluorescent probe 1-[4-(trimethyl-amino)phenyl]-6-phenyl-1,3,5-hexatriene and calculating the fluorescence polarization degree. The measurement was made before and after incubation of PMNs with two activating agents: 4-phorbol 12-myristate 13-acetate (PMN) and N-formyl-methionyl-leucyl-phenylalanine (fMLP). The baseline data showed a reduction of PMN memebrane fluidity in subjects wit insulin resistance. After PMN activation with PMA and fMLP, no significant variation in membrane fluidity was present in PMNs from normals, while in those from subjects with insulin resistance a slight decrease in PMN membrane fluidity was found only after activation with fMLP. The behavior of PMN membrane fluidity, before and after activation, distinguishes insulin-resistant subjects from normal controls, although the effect cannot be directly correlated with the degree of insulin resistance.

RHEOLOGICAL AND METABOLIC LEUCOCYTE DETERMINANTS IN DIABETES MELLITUS

In diabetics of type I and 2 we examined, in resting white blood cells (WBC), the filtration parameters (Initial Relative Flow Rate - lRFR, Clogging Rate - CR) employing the St. George Filtrometer, the polymorphonuclear cells (PMN) membrane fluidity, the PMN cytosolic Ca2+ content and the PMN membrane cholesterol/phospholipid ratio (C/PL). From the obtained data, it is evident that, while the lRFR of unfractionated WBC distinguishes normals from diabetics of type 1 and 2, the fIltration parameters of the PMN and mononuclear cells (MN) do not show any significant difference. PMN membrane fluidity, PMN cytosolic Ca2+ content and PMN C/PL do not discriminate normals from diabetics of type 1 and 2. No relationship is evident between WBC fIltration and metabolic parameters (fasting blood glucose level, serum cholesterol and triglycerides); no correlation is present between PMN fIltration parameters, PMN membrane fluidity and PMN metabolic determinants (cytosolic Ca2+, CIPL).

Polymorphonuclear Leukocyte Integrinsin Deep Venous Thrombosis

The polymorphonuclear leukocytes (PMN) have a role in the pathophysiology of deep venous thrombosis (DVT). We examined the phenotypical expression of PMN beta2-integrins (CD l l a, CDl l b, CD 11c) in a group of 19 subjects with leg DVT. PMN cells were incubated with fluorescent monoclonal antibodies against CD11a, CD11b, CD11c, and the evaluation was made by flow cytofluorimetry. The same integrins were determined after in vitro activation with 4-phorbol 12-myristate 13-acetate (PMA) and N-formylmethionyl-leucyl-phenylalanine (fMLP). In DVT subjects, at baseline, the phenotypical expression of CD11b was decreased and that of CD11c increased when compared with normal controls. In normal subjects PMN activation with PMA and fMLP led to a constant increase of all PMN adhesion molecules, while in DVT subjects the CDl l a did not show any change. These data might have therapeutical ap plications, especially with the aim of preventing post-thrombotic deterioration of vein function.