Effects of Antioxidants and Pro-oxidants on Oxidative Stress and DNA Damage using the Comet Assay : Studies on Blood Cells from Type 2 Diabetes Subjects and Mouse Lymphoma Cells

Diet and oral supplements comprise two distinct sources of antioxidants known to prevent oxidative stress. Beneficial effects from antioxidants have been seen for patients at risk for type 2 diabetes. The aim of this thesis was to evaluate the positive effects of antioxidants against oxidative stress and DNA damage in type 2 diabetes subjects. We also used antioxidants as tools to determine the mechanisms behind genotoxicity induced by mutagenic pro-oxidative agents in mouse lymphoma cells. Several techniques were used to measure oxidative stress and DNA damage, but the main technique used was alkaline comet assay. The results showed that the fruit and vegetable intake was inversely related to oxidative stress in type 2 diabetes subjects. However, oral supplementary intake of 20 antioxidants did not decrease oxidative stress biomarkers. In studies on mouse lymphoma cells, using the alkaline comet assay, DNA damage was induced by catechol and o-phenylenediamine (OPD), while 4-nitro-o-phenylenediamine (4-NOPD) induced only oxidative damage, showing different mechanisms of action behind the mutagenicity of the compounds. Also, oxidative stress was induced by catechol and 4-NOPD, whereas imbalances in the nucleotide pool were seen after exposure to OPD or 4-NOPD. Addition of antioxidants together with these pro-oxidants showed that β-carotene was able to reduce DNA damage at low concentrations of catechol, but increased DNA damage at high concentration. In comparison, addition of α-tocopherol slightly decreased catechol-induced DNA damage at all concentrations of catechol. However, no effect of α-tocopherol was seen on OPD-or 4-NOPD-induced DNA damage. In conclusion, antioxidants from fruits and vegetables, but not from oral supplements, reduced oxidative stress in type 2 diabetes patients, suggesting fruits and vegetables being a healthier source for antioxidant-intake, as compared to oral supplements. Different mechanisms of action for mutagenic pro-oxidants were shown in mouse lymphoma cells, introducing the nucleotide pool as an interesting target for oxidative stress. Reduction of catechol-induced DNA damage by β-carotene or α-tocopherol was shown, with a pro-oxidative action of β-carotene at high concentration of catechol, Interestingly, α-tocopherol was not able to decrease OPD- or 4-NOPD-induced DNA damage, supporting different mechanisms of action behind the genotoxicity from the three pro-oxidants

Oxidative DNA damage and other risk factors, in relation to lifestyle in diabetes type II and metabolic syndrome patients

Lifestyle factors are important in prevention and treatment of lifestyle related diseases as obesity, type II diabetes and cardiovascular diseases. Patients with these diseases or in the risk zone of obtaining them are classified as having the metabolic syndrome. Diet, physical activity and stress management are considered as important factors for preventing or treating their development. In the development and progress of underlying mechanisms in these diseases, oxidative stress and inflammation are considered as important factors since these patient groups have increased levels of oxidative stress and inflammation compared to healthy subjects. The aim of this thesis was to study the influence of dietary factors and physiological risk factors on oxidative stress and inflammation in patients with diabetes type II or metabolic syndrome. This has been investigated in two clinical studies, which have resulted in the two papers in this thesis. In Paper I, patients with metabolic syndrome were part of a residential intervention program for 17 days, with a 5-day follow up. Six months after the start of the program the patients had significantly decreased levels of oxidative stress on DNA. During the same period the patients also had significantly decreased levels of the physiological risk factors weight, BMI, waist- and hip circumferences and body fat, and increased levels of HDL cholesterol. Initial levels and quality of cholesterol were the only factors found to have a correlation to the decrease in oxidative stress. Since the changes probably were lifestyle related, a prolonged lifestyle modification for the patients might help them to maintain the positive results from the intervention. In Paper II patients with diabetes type II were studied and their dietary intake and relevant clinical risk variables were measured. Correlations between diet and plasma antioxidants with oxidative stress and inflammation were analysed. Plasma alpha-carotene and beta-carotene were found to be good biomarkers for fruit and vegetable intake. Further more, fruit and vegetable intake had a negative correlation with DNA oxidation and lipid peroxidation. Dietary vitamin C had a negative correlation with lipid peroxidation and plasma ascorbate with DNA oxidation. The plasma carotenoids were negatively correlated with inflammation. In conclusion, a residential intervention program with the purpose to change diet, physical activity level and to cope with stress can result in health benefits for metabolic syndrome patients. Furthermore, antioxidants in fruit and vegetables can be an important lifestyle factor, since it seems to have a positive impact on DNA oxidation, lipid peroxidation and inflammation in diabetes type II patients. Fruit and vegetable intake and ascorbate might affect oxidative stress and carotenoids might affect inflammation, this supporting the recommendations of a high fruit and vegetable intake in this patient group

High intake of fruit and vegetables is related to low oxidative stress and inflammation in a group of patients with type 2 diabetes

Background : Patients with type 2 diabetes have increased levels of oxidative stress and inflammation. A high fruit and vegetable intake may be beneficial. Objective : To study whether fruit and vegetable intake and levels of plasma antioxidants relate to markers of oxidative stress and inflammation in a group of patients with type 2 diabetes. Further, to investigate whether plasma antioxidants are good biomarkers for intake of fruit and vegetables. Design : Patients with type 2 diabetes were studied. Their dietary intake and levels of plasma antioxidants, and markers of oxidative stress and inflammation were analysed. Results : Fruit and vegetable intake was inversely related to oxidative stress. Plasma carotenoids were negatively correlated with inflammation. The plasma levels of -carotene and β-carotene showed strongly positive associations with fruit and vegetable intake. Conclusions : The results suggest that fruit and vegetable intake may decrease oxidative stress and inflammation in this group of patients. An increased intake of fruit and vegetables can therefore be beneficial for patients with type 2 diabetes, since these patients are documented to have raised oxidative stress and inflammation. The study support the usefulness of plasma -carotene and β-carotene as biomarkers for fruit and vegetable intake