Kaya Emerk

Increased erythrocyte susceptibility to lipid peroxidation in human Parkinson's disease

Recent evidence suggests that among the factors that lead to neurodegenerative changes in Parkinsons disease are stimulation of lipid peroxidation and deficiency of glutathione and glutathione peroxidase in substantia nigra. We have investigated the effect of neurodegenerative changes on plasma and erythrocytes of patients with Parkinsons disease and compared the results with those of age-matched controls. Both plasma lipid peroxide levels and erythrocyte susceptibility to lipid peroxidation were significantly increased in Parkinsons disease. Erythrocyte fragility tests revealed that in 35% of the patients there was increased fragility. In addition, erythrocyte catalase activities were not changed whereas glutathione levels and glutathione peroxidase activities were decreased in Parkinsons disease. Our results suggest that erythrocyte membrane integrity may be impaired in Parkinsons disease.

The effect of vitamin E therapy on plasma and erythrocyte lipid peroxidation in chronic hemodialysis patients.

The effect of vitamin E therapy on plasma and erythrocyte (RBC) lipid peroxidation was investigated in patients undergoing chronic hemodialysis. Before vitamin E therapy, both plasma and RBC lipid peroxidation values of chronic hemodialysis patients were significantly higher than those of healthy controls. Treatment with vitamin E (300 mg/day) for 1 month resulted in a significant decrease of lipid peroxidation. Vitamin E therapy may be a promising approach to prevent peroxidation of membrane lipids in chronic renal failure.

Increased plasma and erythrocyte lipid peroxidation in hyperlipidemic individuals.

Recent work from several laboratories has suggested involvement of lipid peroxidation in the pathogenesis of atherosclerosis. In the present study, the relationship between lipid profile and lipid peroxidation was investigated in plasma and erythrocytes of hyperlipidemic individuals

Long-Term Effect of 17β-Estradiol and Thrombin on Tissue Factor Pathway Inhibitor Release from HUVEC☆

In spite of the increasing evidence that estrogens have protective effects on the vascular system, the evidence that estrogens may contribute to the risk of thrombosis is still being debated. We investigated the effect of 17beta-estradiol (E2) on tissue factor pathway inhibitor (TFPI) release from of cultured human umbilical vein endothelial cells (HUVEC). In this study HUVEC were harvested by collegenase treatment and cultured in multiwelled plates with medium 199 supplemented with 10% fetal calf serum and antibiotics. The cells were incubated in the presence or absence of E2 (1 and 100 nM) with/without thrombin (4 U/mL) for 6 or 24 hours. After the incubations TFPI level of media were measured by IMUBIND Total Eliza kit. Our results demonstrates that E2 at physiological concentrations decreases the release of TFPI from HUVEC significantly. Thrombin also decreases TFPI antigen levels detected in culture media. When combined with thrombin the effect of estrogen is not visible due the much higher effectivity of thrombin in diminishing TFPI levels. These results show that E2 shifts the hemostatic balance towards the procoagulant phase through lowering the TFPI levels secreted by the endothelium.

17β-Estradiol Increases Intracellular Free Calcium Concentrations of Human Vascular Endothelial Cells and Modulates its Responses to Acetylcholine

In this study, we have investigated the effect of 17 beta-estradiol (E2) on intracellular free calcium concentrations ([Ca2+]i) in human umbilical vein endothelial cells (HUVEC) using fura-2 fluorescence. E2 at concentrations of 1nM -1 microM was added subsequently to HUVEC cultures which were either deprived of estrogens or preincubated with E2 (100 nM) for 24 hours. In both groups of cultures, E2 stimulated significant increases in [Ca2+]i in a dose-dependent manner. The effects were more prominent in E2-deprived cells. Preincubation of cells with tamoxifen or the presence of it in the buffer during the experiments did not inhibit the response of the cells to E2. Experiments performed in Ca2+ free/EGTA buffer yielded transient increases in [Ca2+]i suggesting release of Ca2+ from intracellular stores was responsible for the initial peak, while sustained elevations were supported by Ca2+ influx from the extracellular space. Addition of La3+ abolished the sustained [Ca2+]i elevations. Carbachol (CCh) (1nM, 100 nM) did not induce changes in [Ca2+]i of estrogen-deprived cells but produced significant increases in [Ca2+]i of the same cells after incubation with E2 for 30 minutes. The cultures which were preincubated with E2 for 24 hours responded to carbachol directly. The results of our study indicate that E2 may modulate the functions of endothelial cells after only a brief exposure and also may be necessary for the response to acetylcholine especially at low concentrations.

Apolipoprotein E Genotyping in Turkish Myocardial Infarction Survivors and Healthy Controls.

The apolipoprotein (Apo) E gene is known to be polymorphic. Three common alleles determine six phenotypes which can easily be detected by restriction fragment length polymorphism. We performed apo E genotyping in myocardial infarction survivors and healthy controls for the first time in the Turkish population. DNA was amplified by polymerase chain reaction (PCR) and the PCR product was digested with restriction enzymes HhaI to detect apo E2, E3, E4 and with TaqI to detect apo E1. Relative allele frequency for the patient group was found to be 0.91 for E3, 0.07 for E2, 0.02 for E4 and for the control group 0.875 for E3, 0.067 for E2, 0.058 for E4. Copyright 1995 S. Karger AG, Basel

Cumene hydroperoxide-induced chemiluminescence in human erythrocytes: Effect of antioxidants and sulfhydryl compounds

1. The time-course of cumene hydroperoxide-induced changes in lipid peroxidation, protein sulfhydryl groups and chemiluminescence intensity was determined in human erythrocytes. 2. Increase in lipid peroxidation was maximal within 60 min of incubation and was paralleled by a decrease in protein sulfhydryl groups and an increase in chemiluminescence formation. 3. A standard assay system was established to investigate the protective effects of antioxidants and scavenger compounds on cumene hydroperoxide-induced chemiluminescence formation. 4. Chain-breaking antioxidants (i.e. butylated hydroxytoluene) and sulfhydryl compounds (i.e. dithiothreitol) were able to suppress chemiluminescence formation. 5. Our results suggested that secondary free radicals, as well as sulfhydryl groups of proteins are involved in cumene hydroperoxide-induced chemiluminescence formation.

Protective Effect of Defibrotide on Perfusion Induced Endothelial Damage

In the present study, in vitro effects of Defibrotide (D) on perfusion-induced changes in the morphology of endothelium were investigated by scanning (SEM) and transmission (TEM) electron microscope. Human umbilical cord veins were incubated or perfused with platelet-rich plasma alone (PRP) or platelet-rich plasma with Defibrotide (PRP+D) at 3ml/min or 14ml/min and the changes observed were compared. SEM examination of luminal surfaces demonstrated that perfusion with high flow rates may damage endothelial cells and lead to morphological changes which may be prevented by the presence of Defibrotide in the perfusate. Also, the marked reduction in the number of adhered platelets on luminal surface of veins incubated or perfused with Defibrotide compared to veins treated with platelet-rich plasma only revealed that Defibrotide has anti-thrombotic effects. TEM examination of ruthenium red (RR) stained thin sections of veins demonstrated that perfusion disrupts the glycosaminoglcan (GAG) coat on endothelial cells. But the presence of D in the perfusate preserves the integrity of GAG, indicating further cytoprotective effects of the drug on endothelium.

Effects of Aminoguanidine on Lipid and Protein Oxidation in Diabetic Rat Kidneys

Nonenzymatic glycation of tissue and plasma proteins may stimulate the production of oxidant and carbonyl stress in diabetes. The aim of this study was to evaluate the effects of aminoguanidine (AG) on lipid peroxidation, protein oxidation and nitric oxide (NO) release in diabetic rat kidneys. After induction of diabetes with streptozotocin, female Wistar rats were divided into 2 groups. Group DAG (n=9) rats were given AG hydrogen carbonate (1 g/L) in drinking water and group D (n=8) was diabetic control rats given only tap water. Group H (n=8) was followed as healthy controls. At the end of an 8 week period, NO release, lipid and protein oxidation were determined in kidney tissues. NO release was significantly lower in diabetic rats compared with healthy controls (p<0.05). Lipid peroxidation was significantly high in group D (3.9 ± 0.3 nmol MDA/g tissue) compared with the group DAG (2.6 ± 0.1 nmol MDA/g tissue, p<0.01) and group H (2.4 ± 0.2 nmol MDA/g tissue). Protein oxidation was significantly higher in diabetics than healthy controls (563.8 ± 23.9, 655.8 ± 7.2 , 431.5 ± 8.8 mmol carbonyl / g tissue for group DAG, D and H, respectively, p< 0.05). A positive correlation between albuminuria and thiobarbituric acid reactive substance (TBARS) levels (r= 0.54,p<0.005) and carbonyl content (r=0.70, p<0.0005) in kidney homogenate were observed. Although AG treatment had no effect on NO release, it significantly decreased lipid peroxidation in diabetic rat cortices. Consequently increased lipid peroxidation -as well as- protein oxidation could be involved in the pathogenesis of diabetic albuminuria.

INTERACTION OF 3H-DEFIBROTIDE WITH CULTURED HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS

Defibrotide is a profibrinolytic and antithrombotic drug which seems to modulate endothelial cell function. In this study, a method for radioactive labeling of the drug and its interaction with cultured endothelial cells is proposed. 3H-Acetic anhydride was used to label defibrotide. Endothelial cells obtained by collagenase treatment of human umbilical cord veins were cultured in 24-welled plastic culture dishes. Binding experiments were carried out by incubating cell cultures with media containing various concentrations of labeled defibrotide. Our results showed that labeled defibrotide has a KL value of 4.2 micrograms/ml for endothelial cells. Although the presence of a specific transporter is possible, the high molecular weight of the fraction used suggests that the interaction is binding to a specific receptor.