[Evaluation of selected components in antioxidant systems of blood in patients with diabetes]. / Ocena wybranych ukladów antyoksydacyjnych we krwi chorych na cukrzyce.

Oxidative stress plays a major role in the development of chronic complications of diabetes. The aim of our study was to evaluate the selected components of the antioxidative system in well metabolically controlled diabetic patients. We also decided to assess the correlation between these parameters and duration of disease and the presence of it's late complications. The study was entered by 30 patients with type 1 diabetes (18 female and 12 male, aged 30.2 + 10.8 years with mean duration of disease 8.37 + 6.56 years, HbA1c 6.8 + 1.6%). 24 healthy, sex- and age-matched volunteers served as controls. We assessed the following parameters: reduced glutathione in erythrocyte lysate (colorimetric method by Bioxytech GSH-400), serum glutathione peroxidase (enzymatic immunological method by Bioxytech pl. GPx-EIA) and plasma superoxide dismutase activity (colorimetric method based on cytochrome c reduction). In comparison with controls, we found significantly higher reduced glutathione level (11.20 + 0.79 vs 3.92 + 0.62 mumol/l, p < 0.001) and markedly lower dismutase activity (27.49 + 1.32 vs 39.73 + 4.45 U/ml, p < 0.001). The levels of glutathione peroxidase did not differ significantly from values obtained in healthy subjects. We did not observe any correlation between the analysed parameters and duration of diabetes, HbA1c or presence of chronic complications of disease. The obtained results might indicate that antioxidative systems in the state of good metabolic control of diabetes have adaptive properties.

[Appearance of undiagnosed diabetes mellitus in the population of professionally active people in the urban areas]. / Wystepowanie cukrzycy nieznanej w populacji czynnych zawodowo osób w srodowisku miejskim.

Diabetes remains a great social and clinical problem. Therefore, there is a need to focus our efforts on prevention of the disease, especially of type 2 diabetes. Type 2 diabetes is characterized by accelerated development of atherosclerotic changes (macroangiopathy). Hyperglycaemia, hypertension, hyperlipidaemia and hyperfibrinogenaemia also play an important role in the development of macroangiopathy. Hyperinsulinemia, which accompanies the visceral type of obesity, is characteristic of type 2 diabetes. Considering all the above mentioned findings, prevention of type 2 diabetes should be based on the population level, concentrating especially on the groups with increased risk of obesity and/or diabetes (early primary prevention). However, in the present conditions, it seems that screening studies can be conducted only in the groups with high risk of type 2 diabetes (late primary prevention). They allow for relatively early detection of disturbances in carbohydrate metabolism. The aim of the study was to assess the prevalence of undiagnosed diabetes in the population of professionally active inhabitants in Pleszew. 2700 subjects, aged 35-65 years, entered the study. All patients claimed to be healthy. In the first phase of the study, the fasting capillary glycaemia was tested. Fasting blood glucose or oral glucose tolerance test was performed in all cases which fasting capillary glucose was higher then 5.5 mmol/l (100 mg/dl). The screening study revealed 91 cases with glycaemia higher than 6.8 mmol/l (3.4%). 387 subjects (14.3%) with glycaemia ranging from 5.5 to 6.8 mmol/l were qualified to perform the oral glucose tolerance test. Out of this group 138 persons did not come to the laboratory. Thus, the test was conducted in 249 causes (64.3%). The results obtained excluded another 197 subjects as no disturbances in the glucose metabolism were found. Based on the results of the oral glucose tolerance test 39 patients were diagnosed to have an impaired glucose tolerance (2 h glycaemia from 7.8 to 11.1 mmol/l) and in 13 cases diabetes was diagnosed (2 h glycaemia above 11.1 mmol/l). In conclusion, the screening study performed in professionally active adults aged > 35 years, who claimed to be healthy, clinically latent diabetes or impaired glucose tolerance was found in 5.3% cases. 92.8% patients with IGT or diabetes were obese or overweight (BMI > 25 kg/m2) and 32.4% had hypertension (RR > 140/90 mm Hg). In 64% of subjects the serum cholesterol concentration was higher than 5.2 mmol/l and in 18% subjects HDL cholesterol concentration was lower than 1.0 mmol/l and LDL cholesterol higher than 3.5 mmol/l. Elevated triglycerides concentration > 2.0 mmol/l was observed in 30%. In the group with newly diagnosed diabetes, mean age was 55.0 +/- 9.2 years. 27.9% had positive family history of diabetes, 26.5% were smokers, 44.1% were found to have disturbed lower limbs circulation and 30.9% had abnormal feeling of vibration, 7.8% patients with diabetes had symptoms of diabetic retinopathy and 20.1% had microalbuminuria. Body mass index (BMI) in newly diagnosed diabetic patients was 31.6 +/- 5.3 kg/m2 and waist to hip ratio (WHR) was 0.94 +/- 0.41 and indicated the visceral type of obesity. Mean fasting glycaemia was equal 7.26 +/- 1.93 mmol/l and mean HbA1c value was 6.2 +/- 0.7%. It exceeded the laboratory normal value in 17.6% of cases. In 91 patients with fasting glycaemia higher then 15.5 mmol/l insulinaemia was also assessed; its level was elevated in 10 patients. The project of study was prepared in 1996. However, in 1999 the new criteria for diagnosis and treatment of type 2 diabetes were established. The results of the performed study indicate that screening towards diabetes should be performed in subjects aged > 35 years with overweight or obesity and at least one additional risk factor of arteriosclerosis.

[Evaluation of expression of polymorphonuclear neutrophil surface receptors in patients with type 1 diabetes]. / Ocena ekspresji receptorów powierzchniowych granulocytów obojetnochlonnych u chorych na cukrzyce typu 1.

UNLABELLED: Polymorphonuclear neutrophils (PMN) play an important role in the pathogenesis of diabetic microvascular complications. Stimulation of these cells is associated with the appearance of specific receptors on their surface. The aim of the study was to evaluate the expression of the receptors specific for PMN: CD 11b, CD 18. The study was performed in a group of 23 type 1 diabetic patients, aged from 19 to 47 years (mean 30.7 +/- 8.6 years), including 15 females and 8 males (mean diabetes duration time 13.7 +/- 7.5 years; mean HbA1c 7.9 +/- 2.5%). The expression of PMN surface receptors was measured by flow cytometry using a ORTHO DIAGNOSTIC SYSTEM cytofluorimeter. The results were presented as a PMN percentage indicating expression of CD 11b and CD 18. In comparison to healthy controls, there was a significant increase in the number of PMN, both with CD 11b and CD 18 receptors present--(CD 11b: 93.2 +/- 3.4 vs 98.0 +/- 1.9% p < 0.05), (CD 18: 98.5 +/- 0.47 vs 99.4 +/- 0.7%, p < 0.05). CONCLUSION: In patients with type 1 diabetes, PMNs demonstrate a pronounced expression of surface receptors which may indicate an enhanced activity of these cells. The increase of expression of surface receptors is independent of diabetes duration time and HbA1c.

[Assessment of plasma fibronectin concentration in type 1 diabetic patients]. / Ocena stezenia fibronektyny w osoczu u chorych z typem 1 cukrzycy.

Fibronectin is a family of glycoproteins that are present on many cell surfaces, in extracellular matrix and in plasma. Fibronectin might play an important role in pathogenesis of chronic diabetic complications. The aim of this study was to estimate plasma concentration of fibronectin in patients with recently onset and with long duration of type 1 diabetes mellitus. Moreover, the correlation between fibronectin concentration and HbA1c, duration of diabetes and late diabetic complications was assessed. The study was performed in 18 patients with recently onset of diabetes (aged 23.8 +/- 5.3 years, HbA1c 10.63 +/- 0.83%) (group A) and 21 patients with long history of diabetes (aged 33.9 +/- 10.5 years, mean diabetes duration 9.8 +/- 5.8 years, HbA1c 9.24 +/- 2.48%) (group B). The plasma concentration of fibronectin was estimated with the use of the ELISA test. The plasma fibronectin concentration was significantly higher in type 1 diabetic patients in comparison with healthy subjects (381.00 +/- 27.42 and 297.50 +/- 25.32 micrograms/ml, respectively, p < 0.05). The values observed in the patients with recently onset and long duration of diabetes and in the subjects with and without diabetic complications did not differ significantly (p > 0.05, p > 0.05). We noticed positive correlation between fibronectin concentration and HbA1c (r = 0.35; p < 0.05) and negative correlation with patients age (r = -0.35; p < 0.05). The results of our study suggest, that plasma fibronectin concentration in diabetes is increased independently from microangiopathy but rather results from hyperglycaemia.

[Evaluation of nitric oxide metabolites concentration in patients with type I diabetes]. / Ocena stezenia metabolitów tlenku azotu u chorych z typem 1 cukrzycy.

Nitric oxide (NO) mainly known as a relaxing factor, serves a wide variety of other functions in different tissues. It is quickly metabolized to NO2- and NO3- in humans. The aim of the study was to estimate plasma nitrite anion (NO2-) concentration in type 1 diabetic patients. The study was performed in 30 well metabolically controlled patients (18 female and 12 male, aged 30.2 +/- 10.6 years, duration of diabetes 8.4 +/- 6.8 years. HbA1c 6.5 +/- 1.2%) (group A) and 20 poorly metabolically controlled patients (12 female and 8 male, aged 29.8 +/- 9.8 years, duration of diabetes 8.0 +/- 4.8 years, HbA1c 11.2 +/- 1.6%) (group B). The concentration of NO2- was measured with the use of a calorimetric micromethod, where nitrate reductase catalyses the conversion of NO3- to NO2-. The NO2- plasma concentration was significantly higher in diabetic patients in comparison to controls. The highest NO2- concentration was noticed in the group of well metabolically controlled diabetic patients (group A, group B, healthy subjects: 41.99 +/- 4.02, 33.33 +/- 2.44, 16.68 +/- 2.16 mumol/l, respectively, p < 0.05, p < 0.0001). The nitrite concentrations did not correlate with HbA1c (r = -0.03, p > 0.05). The results support the concept that metabolism of nitrite oxides in diabetes is disturbed independently from the degree of metabolic control.

The influence of insulin-dependent diabetes mellitus (IDDM) duration on superoxide anion and hydrogen peroxide production by polymorphonuclear neutrophils.

We address the question whether oxygen metabolism of polymorphonuclear neutrophils (PMN) is influenced by disease duration in patients with insulin-dependent diabetes mellitus (IDDM). PMN were isolated from patients with IDDM of various durations and from healthy controls. We measured PMN production of superoxide anions (O2-) by cytochrome c reduction (see Babior, B.M. et al. (1973) J. Clin. Invest. 52, 741-746) and PMN production of hydrogen peroxide (H2O2) by phenol red oxygenation (see Pick, E. (1980) J. Immunol. Methods 38, 161-169) in three groups of IDDM patients subdivided according to disease duration (group A: IDDM less that 10 years; group B: IDDM of 10-15 years; group C: IDDM of more than 15 years) and in control healthy subjects (group H). Unstimulated O2- production in all IDDM patients was not statistically different from control values (A: 4.3 +/- 0.4 nmol/10(6) PMN per 30 min, nmol/10(6) PMN per 30 min; C: 4.9 +/- 0.9 nmol/10(6) PMN per 30 min; and H: 3.5 +/- 0.2 nmol/10(6) PMN per 30 min, respectively). In contrast, stimulated O2- production was significantly lower in both patients with 10-15 years, and patients with more than 15 years, duration of IDDM than in controls (B: 25.7 +/- 2.5 nmol/10(6) PMN per 30 min; C: 21.1 +/- 3.4 nmol/10(6) PMN per 30 min and H: 42.2 +/- 1.1 nmol/10(6) PMN per 30 min, respectively) correlating with disease duration (r = -0.44, P < 0.033). The stimulated O2- production in patients with less than 10 years duration of IDDM (A: 35.7 +/- 1.9 nmol/10(6) PMN per 30 min) was slightly lower than in controls. H2O2 production of unstimulated PMN (A: 4.0 +/- 0.5 nmol/10(6) PMN per 30 min; B: 4.4 +/- 0.8 nmol/10(6) PMN per 30 min and C: 4.4 +/-1.0 nmol/10(6) PMN per 30 min, respectively) was much higher than those in controls. In contrast, stimulated H2O2 production did not differ statistically from the value noticed in healthy subjects. The results obtained might indicate that production of H2O2 by unstimulated cells is increased in diabetic patients while generation of O2- by stimulated neutrophils is markedly impaired, suggesting that toxic oxygen species production might be influenced by disease duration.

Metabolic control quality and free radical activity in diabetic patients.

The role of oxidative/reductive balance derangement in the pathogenesis of diabetic microangiopathy has often been discussed in the last few years. Therefore, we decided to evaluate the influence of intensive insulin therapy on selected indicators of free radical production. The levels of plasma hydrogen peroxide (H2O2) and serum malonyldialdehyde (MDA) were estimated in 15 patients with Type 1 and 15 with Type 2 diabetes before and after 2 weeks of intensive treatment. The initial H2O2 and MDA levels in all cases were significantly higher than in controls. After 2 weeks of treatment, the values for both estimated parameters were significantly lower; however, they were still higher than in the control group. Our results seem to confirm the previous suggestions concerning the relation between metabolic disturbances and oxidative stress in diabetic patients.

Evaluation of the effect of oral enalapril on neutrophil functions: comparison with the in vitro effect of enalapril and enalaprilat.

Polymorphonuclear neutrophils (PMN) participate in the development of myocardial injury by releasing free oxygen radicals and by involvement in the no-reflow phenomenon. Neutrophil-mediated myocardial injury, therefore contributes to the pathogenesis of heart failure. We investigated the effect of oral treatment with enalapril on neutrophil free oxygen radical production, aggregation and adherence in patients with moderate heart failure (New York Heart Association-NYHA II and III degrees). Samples were taken before and 48 h after a single 10 mg oral dose. Oral enalapril inhibited hydrogen peroxide released by unstimulated PMN, but did not affect stimulated H2O2 release, superoxide anion production, adhesion or aggregation of PMN. Enalaprilat in vitro stimulated PMN to release H2O2 and superoxide anions. Furthermore, in the in vitro conditions both enalaprilat and enalapril inhibited hydrogen peroxide release by stimulated cells. We conclude that, despite certain modifications of neutrophil function in vitro, oral administration of enalapril seems to exert a limited biological effect on circulating neutrophils.

Intravenous isosorbide dinitrate inhibits neutrophil aggregation and plasma-mediated stimulation of superoxide anion production.

Polymorphonuclear neutrophils are known to be activated during myocardial ischaemia causing release of free oxygen radicals and capillary plugging by cell aggregates and therefore to exacerbate ischaemic myocardial injury. Nitric oxide has been shown to modulate neutrophil activation within the ischaemic myocardium and therefore reduce myocardial injury during ischaemia. Drugs that act as nitric oxide donors may therefore modify neutrophil activation. We evaluated the effect of intravenous treatment with isosorbide dinitrate on neutrophil aggregation and plasma-mediated stimulation of neutrophil superoxide anion production in patients with ischaemic heart disease. Samples were obtained from patients before treatment and 15 and 30 min after receiving intravenous isosorbide dinitrate. Isosorbide dinitrate decreased neutrophil aggregation visualized in whole blood (25.3 +/- 3.6, 19.0 +/- 2.6 and 18.5 +/- 2.6 per 300 cells, respectively, P < 0.01). When patient's plasma was incubated with neutrophils obtained from healthy donors, superoxide anion release was 18.99 +/- 6.23, 11.38 +/- 2.79 and 11.49 +/- 3.15 nmol O2-/10(6) cells, respectively (P < 0.01). Therefore, intravenous isosorbide dinitrate inhibited both plasma-mediated stimulation of neutrophil superoxide anion production and neutrophil aggregation.

Evidence for plasma-mediated neutrophil superoxide anion production during myocardial infarction.

The participation of polymorphonuclear neutrophils (PMN) in the development of free-oxygen-mediated myocardial injury is well documented, but direct evidence that PMN-oriented stimuli released to the peripheral blood are able to stimulate PMN free oxygen radical production is missing. We have previously reported that peripheral blood plasma obtained from patients with acute myocardial infarction has chemotactic activity for neutrophils and augments PMN adherence. To investigate whether neutrophilic stimuli released to peripheral blood may induce PMN superoxide anion (O2-) production, we incubated PMN from healthy donors with plasma from patients with acute transmural infarction. PMN O2- production was measured by cytochrome c reduction. PMN O2- was higher under the influence of plasma obtained on the day of admission (24.66 +/- 12.41) and 1 day after onset of acute ischemia (22.91 +/- 10.37) as compared with those observed after incubation with saline (4.18 +/- 1.37; negative control) or zymosan-activated plasma (11.07 +/- 3.4; activated complement cascade positive control). In the following days, plasma-mediated PMN O2- decreased: 13.27 +/- 1.96; 12.37 +/- 3.54 and 8.18 +/- 1.56 after incubation with plasma obtained 2, 3 and 7 days after onset of symptoms, respectively. The results indicate an additional possibility of monitoring the inflammatory response to myocardial necrosis.