Plasma copeptin levels in patients with restless legs syndrome.

OBJECTIVES: Copeptin, the C-terminal fragment of antidiuretic hormone, is a new biomarker that has been found to be elevated in several disorders and could be related with prognosis. This study aimed to compare plasma copeptin levels in patients with restless legs syndrome (RLS) with healthy individuals and to investigate whether plasma copeptin levels were associated with the severity of disease. MATERIAL AND METHODS: 41 patients with primary RLS, who were followed in Bakirkoy Psychiatry and Neurology Research and Training Hospital and 41 age- and sex-matched healthy individuals were included into the study. RLS patients were divided into subgroups as mild-moderate, severe, and very severe according to the severity of symptoms. Sleep quality and excessive daytime sleepiness were determined according to Pittsburgh Sleep Quality Index and Epworth Sleepiness Scale, respectively, and sleep quality scores were analyzed statistically among the groups divided according to disease severity. Copeptin levels in all the patients were compared to the controls. RLS subgroups were compared with each other to evaluate association between copeptin levels and disease severity. RESULTS: Plasma copeptin levels in RLS patients were significantly higher than controls (P < .001). However, there was no association between copeptin levels and disease severity. Excessive daytime sleepiness was found as 14.63% and low sleep quality as 68.29% in patients. CONCLUSIONS: Hypothalamic-pituitary-adrenal axis activation and sympathetic hyperactivity in RLS might be responsible for increased Antidiuretic hormone (ADH) and copeptin release. We think that copeptin might have a potential role in the pathogenesis of RLS and be a biomarker for this disease.

Effects of bleomycin and antioxidants on the fatty acid profile of testicular cancer cell membranes.

Bleomycin is used in chemotherapy regimens for the treatment of patients having testicular germ-cell tumor (TGCT). There is no study in the literature investigating the effects of bleomycin on membrane lipid profile in testicular cancer cells. We investigated membrane fatty acid (FA) profiles isolated, derivatized and analyzed by gas chromatography of NTera-2 testicular cancer cells incubated with bleomycin (Bleo) for 24 h in the absence and presence of N-Acetyl-L-Cysteine (NAC) and curcumin (Cur) as commonly used antioxidant adjuvants. At the same time the MAPK pathway and EGFR levels were followed up. Bleomycin treatment increased significantly saturated fatty acids (SFA) of phospholipids at the expense of monounsaturated (MUFA) and polyunsaturated fatty acids (PUFA). Bleomycin also led to a significant increase in the trans lipid isomers of oleic and arachidonic acids due to its free radical producing effect. Incubation with bleomycin increased the p38 MAPK and JNK levels and downregulated EGFR pathway. Coincubation of bleomycin with NAC reversed effects caused by bleomycin. Our results highlight the important role of membrane fatty acid remodeling occurring during the use of bleomycin and its concurrent use with antioxidants which can adjuvate the cytotoxic effects of the chemotherapeutic agents.

Effect of haemodialysis on the oxidative stress and antioxidants in diabetes mellitus.

Oxidative stress has been defined as a loss of counterbalance between free radical or reactive oxygen species (ROS) production and antioxidant systems. It is involved in the pathogenesis of different chronic diseases. High levels of ROS production via different biochemical mechanisms accompany diseases like type 2 diabetes mellitus (DM) and end-stage renal disease (ESRD). Elevated oxidative status and reduced antioxidant defence systems in patients with DM and ESRD accelerate the prevalence of atherosclerosis and other chronic complications. Our aim was to reveal the effects of diabetes and haemodialysis (HD) separately and together on oxidative stress. In our study, we included 20 diabetic (DM) patients with no renal disease, 20 non-diabetic haemodialysis (HD), 20 diabetic haemodialysis (DHD) patients and 20 healthy volunteers. We have determined the levels of lipid peroxidation expressed as thiobarbituric acid-reactive substances (TBARS), oxidative protein damage as indicated by protein carbonyl (PCO) content and activities of antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSHPx) in all patient groups and healthy subjects. We found enhanced oxidative stress in all patient groups due to an increase in lipid peroxidation (TBARS) and increased oxidative protein damage in terms of PCO content and reduced activities of SOD, CAT and GSH-Px. Oxidative stress was more profound in diabetic patients undergoing haemodialysis. We conclude that both diabetes and dialysis increase oxidative stress and their combined effect on oxidative stress is the highest in magnitude as observed in diabetic patients undergoing haemodialysis.

Protein oxidation in Type 2 diabetic patients on hemodialysis.

BACKGROUND: Oxidative stress is considered to be a unifying link between diabetes mellitus (DM) and its complications, including nephropathy. There have been many reports on increased production of oxidants and decreased level of antioxidants in diabetic patients. The dialysis procedure contributes to oxidative stress. An increase in oxidative stress may contribute to the development of oxidative protein damage in diabetic patients. Our aim was to reveal the effects of diabetes and hemodialysis (HD) on oxidative modifications of plasma proteins. METHODS: We measured reactive carbonyl derivates (PCO), protein thiol (P-SH), and reduced glutathione (GSH) levels in Type 2 diabetic (DM) and diabetic hemodialysed patients (DHD) and in healthy control participants. RESULTS: Protein carbonyl (PCO) content increased significantly in all patient groups relative to the controls. The dialysis procedure caused an additional increase in PCO levels in DHD patients before and after dialysis compared with the level in DM patients. There was a significant decrease in P-SH levels in DHD patients compared with the level in healthy participants and DM patients. There was no significant difference in the whole blood GSH levels between the DM patients and control participants. It was significantly higher in DHD patients in comparison to the DM patients. CONCLUSIONS: We conclude that PCO level increases in DM patients, and this increase is more profound in DHD patients, indicating that both diabetes and dialysis contribute to increased protein oxidation. The low P-SH level in DHD patients, but not in DM patients, suggests that dialysis is responsible for this decrease. We propose plasma PCO derivate as a novel specific marker for oxidative protein damage.

N-acetylcysteine enhances multidrug resistance-associated protein 1 mediated doxorubicin resistance.

BACKGROUND: Resistance of cancer cells against anticancer agents is caused partly by multidrug resistance-associated protein 1 (MRP1). The exact mechanism of MRP1-involved multidrug resistance has not yet been clarified, although glutathione (GSH) is likely to have a role for the resistance to occur. N-acetylcysteine (NAC) is a pro-glutathione drug. DL-buthionine (S,R)-sulfoximine (BSO) inhibits GSH synthesis. The aim of our study was to investigate the effect of NAC and BSO on MRP1-mediated doxorubicin resistance in human embryonic kidney (HEK293) and its MRP1-transfected 293MRP cells. MATERIALS AND METHODS: Human embryonic kidney cells were transfected with a plasmid encoding the whole MRP1 gene. Both cells were incubated with doxorubicin in the presence or absence of NAC and/or BSO. The viability of both cells was determined under different incubation conditions. Glutathione, glutathione S-transferase (GST) and glutathione peroxidase (GPx) levels were measured in the cell extracts obtained from both cells incubated with different drugs. RESULTS: N-acetylcysteine increased the resistance of both cells against doxorubicin. DL-buthionine (S,R)-sulfoximine decreased NAC-enhanced MRP1-mediated doxorubicin resistance, indicating that induction of MRP1-mediated doxorubicin resistance depends on GSH synthesis. Doxorubicin decreased the cellular GSH concentration and increased GPx activity. Glutathione S-transferase activity was decreased by NAC. CONCLUSION: Our results demonstrate that NAC enhances MRP1-mediated doxorubicin resistance and this effect depends on GSH synthesis. DL-buthionine (S,R)-sulfoximine seems a promising chemotherapy improving agent in MRP1 overexpressing tumour cells.

Secretory phospholipase A2 and phospholipids in neural membranes in an experimental epilepsy model.

OBJECTIVES: Previous studies have revealed an increase in several forms of phospholipase A2 activity associated with cell injury, but the secretory form of phospholipase A2 has not previously been studied in neurological disorders. We investigated the influence of seizures on secretory phospholipase A2 and phospholipid breakdown in synaptosome fractions prepared from rat hippocampus, cortex and cerebellum in pentylenetetrazol-induced epilepsy. MATERIAL AND METHODS: Secretory phospholipase A2 concentration was measured by a photometric enzyme immunoassay. The synaptosomes underwent extraction, and the phospholipids fractions for phosphatidylcholine, phosphatidylethanolamine and lysophosphatidylcholine were recovered from the thin layer chromatography plates. The amount of each phospholipid was quantified using the amount of recovered phosphate in each phospholipid spot. RESULTS: Secretory phospholipase A2 concentration was found to be significantly higher in the epileptic group when compared with the control group. The amounts of phospholipids were found to be highly variable in different brain regions. CONCLUSION: Our results suggest that epileptic seizures enhanced phospholipid breakdown and induced alterations in the distribution of phospholipids in different brain regions.

Maternal serum screening for Down's syndrome, open neural tube defects and trisomy 18.

Maternal serum screening identifies women at an increased risk of a pregnancy with Down's syndrome or trisomy 18 or an open neural tube defect. The triple test, consisting of maternal serum alpha-fetoprotein, unconjugated estriol and human chorionic gonadotropin was carried out by a chemiluminescence immunoassay method in our laboratory. The study consisted of 373 pregnant women. The gestational range for the study group was 14-22 weeks. The mean maternal age for the study group was 28.53 +/- 5.46 years (range 17.4 to 43.5 years); 9.1% of the women were considered at high risk for Down's syndrome based on the test results. In our study the detection rate for Down's syndrome by prenatal karyotyping was 66.6%. Maternal serum screening allows reduction of the number of women requiring amniocentesis without a significant decrease in the detection rate.

Alterations of antioxidant enzymes and oxidative stress markers in aging.

In accordance with the present state of scientific knowledge, the excessive production of free radicals in the organism, and the imbalance between the concentrations of these and the antioxidant defenses may be related to processes such as aging and several diseases. The aging process has been described by various theories. In particular, the free radical theory of aging has received widespread attention which proposes that deleterious actions of free radicals are responsible for the functional deterioration associated with aging. Although, the relationship between lipid peroxidation and aging have been investigated extensively, the studies have produced conflicting results. To investigate the correlation between the oxidative stress and aging, we have determined the levels of lipid peroxidation expressed as thiobarbituric acid reactive substances (TBARS; MDA) and conjugated dien; oxidative protein damage as indicated by carbonyl content and activities of antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in a sample of 100 healthy men and women ranging in age from 20 to 70years. In addition, vitamin E, C levels, reduced glutathione and sulphydryl content were determined. The oxidation end product of nitric oxide (nitrate) was also studied to investigate any role of nitrogen radicals in aging. Our data show that there is an age related increase in lipid peroxidation expressed as MDA and oxidative protein damage as indicated by carbonyl content. Aging is not linked to a decline in antioxidant enzymes except GPx. Our data suggests that the level of oxidative stress increase cannot entirely be attributed to a decrease in the activities of antioxidant defense system and probably various factors may contribute to this process.

Biochemical evidence of crossed cerebellar diaschisis in terms of nitric oxide indicators and lipid peroxidation products in rats during focal cerebral ischemia.

OBJECTIVES: Cerebral hypoperfusion in the contralateral cerebellar hemisphere after stroke is interpreted as a functional and metabolic depression, possibly caused by a loss of excitatory afferent inputs on the corticopontocerebellar pathway terminating in the cerebellar gray matter. This phenomenon is defined as crossed cerebellar diaschisis and can be diagnosed clinically by positron emission tomography, single-photon emission computed tomography, brain magnetic resonance imaging and electroencephalography in terms of regional cerebral blood flow or metabolic rate of oxygen measurements. MATERIALS AND METHODS: In the present study, nitric oxide indicators (nitrite and cyclic guanosine monophosphate) and lipid peroxidation products (malondialdehyde and conjugated dienes) were measured in rat cerebral cortices and cerebella after permanent right middle cerebral artery occlusion in order to assess the crossed cerebellar diaschisis. RESULTS: Nitrite values in ipsilateral cortex were significantly higher than those in contralateral cortex at 10 (P < 0.001) and 60 (P < 0.05) min of ischemia but no significant changes were observed in both cerebellum compared to the 0 min values. In both cerebral cortex and cerebellum cGMP levels at 10 and 60 min were significantly increased (P < 0.001). This increase was marked in ipsilateral cortex and contralateral cerebellum when compared with opposite cortex and cerebellum (P < 0.001). MDA values in ipsilateral cortex were significantly higher than those in contralateral cortex at 60 min of ischemia (P < 0.05). Contralateral cerebellar MDA values were found significantly higher than those in ipsilateral cerebellum at 0 (P<0.001) and 60 (P < 0.05) min of ischemia. In ipsilateral cortex, conjugated diene values at 0, 10, 60 min of ischemia were higher than those in contralateral cortex. On the other hand 0, 10, 60 min conjugated diene levels in contralateral cerebellum were significantly higher than those in ipsilateral cerebellum (P < 0.001). CONCLUSION: These findings support the interruption of the corticopontocerebellar tract as the mechanism of the crossed cerebellar diaschisis.

Effects of Lamotrigine on brain nitrite and cGMP levels during focal cerebral ischemia in rats.

Glutamate receptor antagonists are protective in animal models of focal cerebral ischemia. Lamotrigine (3,5-diamino-6-[2,3-dichlorophenyl]-1,2, 4-triazine) is an anticonvulsant drug that blocks voltage-gated sodium channels and inhibits the ischemia-induced release of glutamate. Experiments in primary neuronal cultures implicate nitric oxide (NO) as a mediator of glutamatergic neurotoxicity acting via N-Methyl-D-Aspartate (NMDA) receptors. The effect of glutamate release inhibitor, Lamotrigine upon NO and cGMP production has been examined in focal cerebral ischemia in rats. Focal cerebral ischemia was produced by the permanent occlusion of right middle cerebral artery (MCA) in urethane anesthetized rats. A number of indicators of brain NO production (nitrite, cGMP) were determined in ipsilateral and contralateral cerebral cortex and cerebellum after 0, 10, 60 min of focal cerebral ischemia. The same parameters were measured in rats treated with Lamotrigine (20 mg/kg, i.p.) 30 min before or just after the occlusion of the right MCA.

The effects of nitric oxide synthase inhibitor, L-NAME on NO production during focal cerebral ischemia in rats: could L-NAME be the future treatment of sudden deafness?

Recent evidence in primary neuronal cell culture implicates NO as a mediator of glutamatergic neurotoxicity acting via N-methyl-D-aspartate (NMDA) receptors. In this study, we investigated the effects of inhibition of NOsynthase activity in focal cerebral ischemia in rats. Focal cerebral ischemia was produced by permanent occlusion of right MCA in urethane anesthetized rats. A number of indicators of brain NO production, nitrite and cGMP were determined in ipsilateral and contralateral cerebral cortex and cerebellum after 0, 10 and 60 minutes of focal cerebral ischemia. The same parameters were measured in rats pre- and posttreated with the potent Nitric oxide synthase (NOS) inhibitor, NW-nitro-L-arginine methyl ester (L-NAME).

Inhibitory role of N omega-nitro-L-arginine methyl ester (L-NAME), a potent nitric oxide synthase inhibitor, on brain malondialdehyde and conjugated diene levels during focal cerebral ischemia in rats.

The effect of N omega-nitro-L-arginine methyl ester (L-NAME) on ischemic neuronal damage was studied in a rat model of permanent focal cerebral ischemia in terms of ipsilateral and contralateral cortical and cerebellar tissue lipid peroxides. Forty-five male Swiss Albino rats were assigned to one of four groups; sham operated as control, subjected to right middle cerebral artery occlusion or injection of L-NAME (10 mg/kg i.p.) either 30 min before or just after right middle cerebral artery occlusion. Changes in lipid peroxides were expressed as nanomoles of malondialdehyde and conjugated diene per milligram of protein. Malondialdehyde values following 60 min of ischemia relative to contralateral cortex and conjugated diene levels in 0, 10 and 60 min of ischemia were found to be higher in ipsilateral cortex than in contralateral cortex. On the other hand, contralateral cerebellar malondialdehyde levels after 0 and 60 min of ischemia and conjugated diene levels after 0, 10 and 60 min of ischemia were higher than those in ipsilateral cerebellum. Pharmacological inhibition of nitric oxide synthase by L-NAME before or just after permanent middle cerebral artery occlusion significantly decreased the malondialdehyde and conjugated diene levels in both the cortex and the cerebellum. No significant differences were found in malondialdehyde values between rats that had been pre- and post-treated with L-NAME, but conjugated diene levels in the post-treated group seemed to be significantly lower than those in the pretreated group. On the whole, these results suggest that malondialdehyde and conjugated diene represent early biochemical markers of lipid peroxidation in ischemic tissues, reflecting the radical-mediated tissue damage.

The effects of detergents on t-butyl hydroperoxide-induced chemiluminescence.

We examined the effects of Triton X-100, digitonin, sodium dodecyl sulfate, taurocholic acid, and cetylpyridinium chloride on hemoglobin-catalyzed and t-butyl hydroperoxide-induced chemiluminescence. The experimental system contained hemoglobin, luminol, t-butyl hydroperoxide, and different concentrations of detergents (5-100 mg/dl) in TRIS-HCl buffer. Control assays were performed by excluding detergents. Chemiluminescence was detected using a liquid scintillation counter in single photon mode. All concentrations chosen for each detergent reduced the maximum chemiluminescence value and retarded the time that maximum chemiluminescence occurred. The most prominent reduction in maximum chemiluminescence was observed with 50 and 100 mg/dl digitonin. The smallest reduction was observed with 5 mg/dl sodium dodecyl sulfate, without retardation of the time that maximum chemiluminescence occurred. Our aim was to use detergents in membrane-containing experimental systems and hence to identify the detergent with the least effect on chemiluminescence. Our results suggest that sodium dodecyl sulfate is the most suitable detergent for chemiluminescence studies in membrane systems.

Serum and urine malondialdehyde levels in NIDDM patients with and without hyperlipidemia.

Malondialdehyde (MDA), a marker of lipid peroxidation, was measured as thiobarbituric acid reactive substance (TBARS) in 78 noninsulin-dependent diabetic patients, 38 hyperlipidemic patients, and 28 healthy subjects. Diabetic patients were divided into groups and subgroups according to the existence of hyperlipidemia and other complications. Serum and urine MDA concentrations were significantly higher in diabetic and nondiabetic patient groups than in the control group. By contrast to urine MDA levels, serum MDA level was significantly higher in hyperlipidemic diabetics than that of normolipidemic diabetics. Serum MDA levels in the hyperlipidemic diabetic group and urine MDA levels in both diabetic groups were significantly higher than those in hyperlipidemic nondiabetic group. In both diabetic groups, the existence of complications didn't affect serum and urine MDA levels. No correlation existed between serum and urine MDA levels in both patient groups and control subjects. This study confirmed the existence of lipid peroxidation disorders in diabetic patients.

ERP spectral analysis of cholesterol rich patients.

Visual event-related potentials (ERPs) were studied in twenty hypercholesterolemic (HC) patients and twenty age-matched healthy controls. ERPs were recorded in two different experimental conditions that the target stimulus (red light) was counted (Test 1) or uncounted (Test 2). Amplitude spectra of ERPs were computed by transient response-frequency characteristics (TRFC) method. Their maxima were found to occupy the frequency bands of 1-2, 3-4, 5-7, 8-12, 13-20, 20.5-32 Hz. The amplitude mean (decibel) of 1-2 and 3-4 frequency bands were decreased in Test 2 compared to Test 1 in the control group, but no significant amplitude differences were found between Test 1 and Test 2 in the HC group.

Event-related potentials in hypertriglyceridemia.

Event-related potentials (ERPs) of twenty-three hypertriglyceridemic (HTG) patients and twenty-three age-matched healthy controls were recorded in two different experimental conditions that the target stimulus was counted (Test 1) or uncounted (Test 2). Latencies of each wave were inside normal limits in all patients. No differences were found in the P3b amplitudes of Test 1 case among HTG and control subjects. Amplitude spectra of ERPs were computed by transient response-frequency characteristics (TRFC) method. Their maxima were found to occupy the frequency bands of 1-2, 3-4, 5-7, 8-12, 13-20, 20.5-32 Hz. When comparing Test 1 with Test 2, significant amplitude differences were found in 1-2 and 3-4 frequency bands of controls and 1-2 and 13-20 frequency bands of patient group.

Serum glycated lipoproteins in type II diabetic patients with and without complications.

The extent of lipoprotein glycation was assessed in 82 patients diagnosed as diabetes mellitus type II. Glycated lipoprotein levels in serum were determined by agarose gel film electrophoresis in 48 non-diabetic control subjects, 56 diabetics with complications and 26 diabetics with no complications. All glycated lipoproteins were higher in the patients when the diabetics were compared to the control group, but glycated very low-density lipoprotein (VLDL) was the only glycated lipoprotein fraction that was observed to be higher in the diabetic patients with complications compared to the patients without complications (P < 0.01). Triglycerides and VLDL-cholesterol were also increased in the diabetics, but there was no significant difference in these variables when the patients without complications were compared to the control subjects. There was no difference in total cholesterol and low-density lipoprotein (LDL)-cholesterol levels of the diabetic patients compared to the controls.

Plasma and urine sialic acid in non-insulin dependent diabetes mellitus.

Urinary excretions of albumin, glycosaminoglycans (GAGS), total sialic acid (TSA), and lipid associated sialic acid (LASA) were measured in 78 non-insulin dependent diabetic patients (NIDDM) and 28 healthy subjects. TSA excretion was significantly higher in normoalbuminuric and microalbuminuric diabetic subjects than the control subjects and TSA excretion was correlated with urinary albumin excretion rate (AER). In normoalbuminuric diabetics, the duration of diabetes correlated significantly with both sialicaciduria and albuminuria. Although serum TSA levels were significantly higher in both diabetic groups than the control subjects, there was no correlation between serum and urinary TSA levels.

The effects of streptozocin-induced diabetes on renal plasma flow.

To evaluate the significance of collagen levels, nonenzymatic glycation, and effective renal plasma flow in diabetes, we studied 52 Streptozocin induced-diabetic rats. After 10 weeks of diabetes, rats were injected i.v. 0.5 microCi/g 99mTechnetium Mercaptoacetyl triglycine and effective renal plasma flow was calculated from the renograms obtained. The collagen content and hydroxymethyl furfural levels of kidney and tail tissue as well as renal plasma flow increased significantly in diabetic rats (p < 0.05). The increase in renal plasma flow indicates that hyperperfusion may play a role in diabetic nephropathy mechanism. The correlation between renal plasma flow and glycation was not as high as the correlations between blood glucose concentrations and hydroxymethyl furfural and collagen levels, suggesting that factors other than glycation are important in increasing the renal plasma flow.

Glycation of lipoproteins and accelerated atherosclerosis in non-insulin-dependent diabetes mellitus.

We used a new and remarkably simple method to examine the extent of in vivo lipoprotein glycation in type II diabetic patients with atherosclerosis and diabetic patients with no complications. Serum glycated lipoprotein levels were determined by agarose gel film electrophoresis in 48 non-diabetic control subjects and 39 diabetic patients, of whom 26 had no complications and 13 had atherosclerotic heart disease. Fasting serum glucose, glycohemoglobin and serum fructosamine concentrations (indicators of glycemia) and total cholesterol, triglyceride, low-density lipoprotein-, very low-density lipoprotein- and high-density lipoprotein-cholesterol concentrations and the low-density lipoprotein/high-density lipoprotein ratio (serum lipid profile) were also determined in the control and diabetic subjects. Glycated low-density lipoprotein and very low-density lipoprotein concentrations were significantly increased in diabetic patients compared with controls; but only glycated very low-density lipoprotein was significantly increased in atherosclerotic patients compared with diabetics without complications. The lipid profile parameters were not significantly increased in patients compared with controls. In diabetics, especially those with poorly controlled hyperglycemia and atherosclerosis, glycation of lipoprotein fractions might be more important than serum lipid and lipoprotein abnormalities. The significant correlation between atherosclerosis and glycated very low-density lipoprotein, suggests that very low-density lipoprotein glycation could be responsible for the development of atherosclerosis in diabetes.