Ilona Németh

Blood glutathione redox ratio as a parameter of oxidative stress in premature infants with IRDS

Oxygen toxicity is thought to play an important pathogenic role in several neonatal diseases, including idiopathic respiratory distress syndrome (IRDS). Therefore, the development of a reliable measure of the actual oxidative stress status of patients would be of great clinical significance. In order to obtain information about the oxidative stress during the first week of life in premature infants with IRDS, the blood concentrations of oxidized and reduced glutathione, as well as their molar ratios, were determined by a highly sensitive, specific enzymic assay. The fractional inspired oxygen concentrations needed to maintain adequate arterial oxygen tension and the arterio-alveolar oxygen ratios were chosen as parameters indicating the severity of illness in premature infants at a given time. There was a highly significant positive correlation between the glutathione redox ratios and the fractional inspired oxygen concentrations. A maturity-related difference was also found; the oxidized glutathione concentrations were the highest in the least mature infants, accompanied by a pronounced compensatory rise in the reduced glutathione concentrations as well. A significant negative correlation was found between the arterio-alveolar oxygen ratio and the glutathione redox ratio: i.e., an improvement in oxygenation was accompanied by a decrease in the glutathione redox ratio. The efficient recycling of reduced glutathione in erythrocytes providing antioxidant protection for premature infants, permits the use of the blood glutathione redox ratio as a noninvasive measure of in vivo oxidative stress.

Impaired early neurologic outcome in newborn piglets reoxygenated with 100% oxygen compared with room air after pneumothorax-induced asphyxia

Birth asphyxia is a serious problem worldwide, resulting in 1 million deaths and an equal number of neurologic sequelae annually. It is therefore important to develop new and better ways to treat asphyxia. In the present study we tested the effects of reoxygenation with room air or with 100% oxygen (O2) after experimental pneumothorax-induced asphyxia on the blood oxidative stress indicators, early neurologic outcome, and cerebral histopathology of newborn piglets. Twenty-six animals were studied in three experimental groups:1) sham-operated animals (SHAM, n = 6), 2) animals reoxygenated with room air after pneumothorax (R21, n = 10), and 3) animals reoxygenated with 100% O2 after pneumothorax (R100, n = 10). In groups R21 and R100, asphyxia was induced under anesthesia with bilateral intrapleural room air insufflation. Gasping, bradyarrhythmia, arterial hypotension, hypoxemia, hypercarbia, and combined acidosis occurred 62 ± 6 min (R21) or 65 ± 7 min (R100; mean ± SD) after the start of the experiments; then pneumothorax was relieved, and a 10-min reoxygenation period was started with mechanical ventilation with room air (R21) or with 100% O2 (R100). The newborn piglets then breathed room air spontaneously during the next 3 h. Blood oxidative stress indicators (oxidized and reduced glutathione, plasma Hb, and malondialdehyde concentrations) were measured at different stages of the experiments. Early neurologic outcome examinations (neurologic score of 20 indicates normal, 5 indicates brain-dead) were performed at the end of the study. The brains were next fixed, and various regions were stained for cerebral histopathology. In the SHAM group, the blood gas and acid-base status differed significantly from those measured in groups R21 and R100. In group R100, arterial Po2 was significantly higher after 5 (13.8 ± 5.6 kPa) and 10 min (13.2 ± 6.3 kPa) of reoxygenation than in group R21 (8.7 ± 2.8 kPa and 9.2 ± 3.1 kPa). The levels of all oxidative stress indicators remained unchanged in the study groups (SHAM, R21, and R100). The neurologic examination score in the SHAM group was 18 ± 0, in group R21 it was 13.5 ± 3.1, and in group R100 it was 9.5 ± 4.1 (significant differences between SHAM and R21 or R100, and between R21 and R100). Cerebral histopathology revealed marked damage of similar severity in both asphyxiated groups. We conclude that the blood oxidative stress indicators and cerebral histopathology did not differ significantly after a 10-min period of reoxygenation with room air or with 100% O2 after pneumothorax-induced asphyxia, but reoxygenation with 100% O2 might impair the early neurologic outcome of newborn piglets.

Nonenzymatic antioxidants of blood in multiple sclerosis

Abstract Free radical action has been suggested as a causal factor in multiple sclerosis. We investigated the plasma level of lipid peroxides expressed in terms of malone dialdehyde and changes in blood nonenzymatic antioxidants (glutathione, α-tocopherol, retinol, plasma sulfhydryl groups, and uric acid) in multiple sclerosis patients with exacerbation or in remission, including a group treated with β-interferon. The malone dialdehyde level was increased by 38% (n.s.) during exacerbations. The blood concentration of oxidized glutathione was likewise elevated (P < 0.05), while the ratio of plasma α-tocopherol to cholesterol plus triglyceride was decreased (P < 0.001). These changes suggest increased free radical production and consumption of the scavenger molecules during the active phase of the disease. Blood reduced glutathione level was increased (P < 0.01) during exacerbation and remission as well. The rise in this thiol is likely to be a compensatory mechanism defending the cells from further oxidant injuries. β-Interferon increased plasma α-tocopherol levels (P < 0.001) but not the lipid corrected α-tocopherol value. Other parameters were not influenced by the drug.

Oxidative stress in juvenile essential hypertension.

Objective Oxidative stress, an antioxidant/pro-oxidant imbalance, in patients with juvenile essential hypertension was measured via several biochemical parameters. As the blood pressure is associated with the body mass index (BMI), results were compared with those on BMI-matched controls. Design and setting A prospective observational study at a university teaching hospital. Patients Children and adolescents with essential hypertension (mean ± standard deviation: age 14.4 ± 3.1 years, BMI 25.0 ± 6.9 kg/m2, n = 52) before any treatment, and controls with a similar BMI distribution (age 14.3 ± 4.3 years, BMI 24.4 ± 6.6 kg/m2, n = 48). Methods Measurements were made of the plasma levels of (1) nitrites + nitrates, an indirect measure of available nitric oxide; (2) lipid peroxidation end-products, as malondialdehydes and free thiols; and (3) the redox status of the red blood cell glutathione, as a new oxidative stress parameter. Results There were decreased plasma levels of nitrates and increased levels of lipid peroxidation end-products in the hypertensive patients, resulting in a consistent increase in the plasma lipid peroxidation/nitric oxide ratio as compared with the controls with the same BMI (P < 0.01). This ratio additionally correlated directly with both the systolic and diastolic blood pressures for the overall patient population (P < 0.001). A significant glutathione depletion in the red blood cells resulted in an elevated ratio of oxidized/reduced forms with a reduced in vitro antioxidant protective capacity in the hypertensive patients versus the BMI-matched controls (P < 0.001). Conclusions The presence of systemic oxidative stress was proven in hypertensive children and adolescents, irrespective of their BMI.

Nontoxic heat shock protein coinducer BRX-220 protects against acute pancreatitis in rats

BACKGROUND Nontoxic heat shock protein (HSP) inducer compounds open up promising therapeutic possibilities by activating one of the natural and highly conserved defense mechanisms of the organism. AIMS In the present experiments, we examined the effects of a HSP coinducer drug-candidate, BRX-220, on the cholecystokinin-octapeptide (CCK)-induced acute pancreatitis in rats. METHODS Male Wistar rats weighing 240 to 270 g were divided into two groups. In group B, 20 mg/kg BRX-220 was administered orally, followed by 75 microg/kg CCK subcutaneously three times, after 1, 3, and 5 h. This whole procedure was repeated for 5 d. The animals in group slashed circleB received physiological saline orally instead of BRX-220, but otherwise the protocol was the same as in group B. The rats were exsanguinated through the abdominal aorta 12 h after the last administration of CCK. We determined the serum amylase activity, the plasma trypsinogen activation peptide concentration, the pancreatic weight/body weight ratio, the DNA and total protein contents of the pancreas, the levels of pancreatic HSP60 and HSP72, the activities of pancreatic amylase, lipase, trypsinogen, and free radical scavenger enzymes (superoxide dismutase, catalase, and glutathione peroxidase), the degree of lipid peroxidation, protein oxidation, and the reduced glutathione level. Histopathological investigation of the pancreas was also performed in all cases. RESULTS Repeated CCK treatment resulted in the typical laboratory and morphological changes of experimentally induced pancreatitis. The pancreatic levels of HSP60 and HSP72 were significantly increased in the animals treated with BRX-220. In group B, the pancreatic total protein content and the amylase and trypsinogen activities were significantly higher vs. group slashed circleB. The plasma trypsinogen activation peptide concentration, and the pancreatic lipid peroxidation, protein oxidation, and the activity of Cu/Zn-superoxide dismutase were significantly decreased in group B vs. group slashed circleB, whereas the glutathione peroxidase activity was increased. The morphological damage in group B was significantly lower than that in group slashed circleB. CONCLUSION The HSP coinducer BRX-220, administered for 5 d, has a protective effect against CCK-induced acute pancreatitis.

Surface Tension, Glutathione Content and Redox Ratio of the Tracheal Aspirate Fluid of Premature Infants with IRDS

Objective: Determination of the surface tension (ST), the total glutathione (GL) content and the ratio of oxidized glutathione (GSSG) to reduced glutathione (GSH) in the tracheal aspirate (TA) of newborn infants with IRDS. Methods: The ST of the TA was determined by monitoring the fluid level pulsated in a capillary glass tube by means of a digitalized videocomputerized picture analysis program, a technique developed in our laboratory. The concentrations of GSSG and total GL in the TA were determined enzymatically with glutathione reductase. All results of laboratory tests were referred to the total phospholipid (PL) concentration. Patients, Experimental Material: TA samples were collected from 32 intubated premature and newborn infants admitted to the NICU with IRDS during the first 2 weeks of their lives. Control samples were obtained from 11 children prior to elective surgery. Results: The ST relative to the PL content (surface tension index, STI) was significantly lower in the newborns with IRDS than in the control group, and the concentration of GSH in the TA was also markedly decreased in all IRDS infants studied. The concentration of GSSG and the ratio of GSSG to GSH were significantly higher in the severe cases and in those with an unfavourable prognosis. Surfactant treatment had a protective effect against oxidative stress, it induced a decrease in both the GSSG concentration and in the GL redox ratio (GSSG/GSH) in the TA. There was a close correlation between the GSH content and the STI value of the samples studied. Conclusion: Oxidation and consequent depletion of GSH in the TA may be an aggravating factor in the development of the insufficient surface activity in intubated newborns with IRDS.

Platelet aggregation, blood viscosity and serum lipids in hypertensive and obese children

A group of 35 patients (median age 15.5 years, range 8–17 years) with juvenile essential hypertension, 15 with body mass index (BMI kg/m2) <25 and 20 with BMI >25, as well as 35 age and sex matched controls (BMI <25 n=20; BMI >25 n=15) were investigated to study the role of hypertension and obesity, separately and in combination, on in vitro platelet aggregation, blood and plasma viscosity, plasma lipid concentrations and lipid peroxidation as well as nitric oxide (NO) production. Obese children (hypertensive and controls) had significantly higher concentrations of total cholesterol and triglycerides. The levels of high density lipoprotein (HDL)-cholesterol were lower in obese hypertensive children than their non-obese counterparts. There was a significant increase in platelet aggregation and a decrease in NO levels in hypertensive patients (obese and non-obese) reflecting a significant negative correlation (r=−0.553 and −0.530, n=35; P<0.01, respectively). However, an increased tendency to aggregation was also evident in obese normotensive patients. A significant positive correlation was observed between the platelet aggregation and BMI (r=0.501, n=35; P<0.01). Plasma free thiols were decreased in hypertensive children independent of their BMI. An increased lipid peroxidation and higher blood and plasma viscosity were found only in obese patients with hypertension. Multivariate analysis revealed significant interactions in the effects of obesity and hypertension on platelet aggregation and thiol oxidation. Conclusion: in obese children an increased platelet aggregation and oxidative insult contribute to the development of hypertension and to the promotion of vascular damage.

XANTHINE OXIDASE ACTIVATION IN MILD GESTATIONAL HYPERTENSION

Objective: We hypothesized that activation of the xanthine oxidase (XO) enzyme system is a potential source of free radicals in pregnancy-induced hypertension (PIH). Methods: A prospective observational study was carried out on 16 pregnant women who met the criteria of gestational hypertension [rise in blood pressure (BP) of 30 mm Hg systolic or 15 mm Hg diastolic after 20 weeks gestation or BP>140/90 mm Hg if earlier pressure is unknown] without proteinuria or any signs of renal impairment. Fourteen women with a clinically normal pregnancy matched for maternal age, parity, and gestational age acted as pregnant controls. Nonpregnant control women were members of the laboratory staff (n=15). Main Outcome Measures: Concentrations of free sulfhydryl (SH) groups, purine catabolites, lipid peroxidation products in plasma, and blood carboxyhemoglobin levels were used to follow oxidative stress and potential hemolysis. A noninvasive measurement of functional XO activity was carried out (i.e., the urinary ratio of the two metabolites of caffeine was estimated). Results: A pronounced oxidative stress was demonstrated in plasma samples of patients with hypertension by the elevated concentrations of uric acid and lipid peroxidation products. A reduced level of free sulfhydryl groups and an increased concentration of hypoxanthine (HX) were shown in normotensive pregnant individuals. The XO activity index was substantially higher in overweight pregnant subjects with mild hypertension [0.849±0.096 (p<0.01)] than in normotensive pregnant women or in age-matched nonpregnant subjects [0.596±0.105, 0.542±0.049 (means±SD), respectively]. Conclusions: Our study of mildly hypertensive pregnant subjects provides additional evidence of the putative role of XO activation as a source of free radicals in the early stage of endothelial dysfunction.

Roles of Paraoxonase and Oxidative Stress in Adolescents with Uraemic, Essential or Obesity-Induced Hypertension

Background/Aims: Paraoxonase 1 (PON1) is associated with high-density lipoproteins in the plasma, and is capable of hydrolysing oxidized lipids and preventing the oxidation of low-density lipoproteins. Oxidative stress and the PON1 (activity and Q192R polymorphism) were analysed in adolescent patients with essential (n = 49) or obesity-induced hypertension (n = 79), uraemic patients (n = 20), and also in obese normotensive patients (n = 60) and age-matched controls (n = 57). Methods: The PON1 activity was measured via paraoxon hydrolysis. The PON1 genotype was determined by real-time PCR. The levels of oxidized and reduced glutathione, the end-products of nitric oxide, cysteine, homocysteine and lipid peroxidation in the plasma were measured and related to the PON1 status. Results: There were no significant differences between the patient groups and the control group in the genotype distributions and the allele frequencies of the Q192R polymorphism. The PON activity was significantly lower (p < 0.001) in the uraemic hypertensive group than in the controls. The MDA concentration was significantly higher in the uraemic hypertensive (p < 0.001) and obese hypertensive (p < 0.05) patients. The plasma NOx concentrations were significantly lower (p < 0.001) and the ratio MDA/NOx were significantly higher in all four patient groups. The GSH levels were significantly lower in the patients with hypertension (p < 0.001) and obesity-induced hypertension (p < 0.05) than in the controls, while the GSSG level (p < 0.01) and the ratio GSSG/GSH (p < 0.05) was significantly higher in the uraemic hypertensive group. The plasma homocysteine level was significantly higher (p < 0.001) in the uraemic hypertensive patients as compared with the controls. Conclusions: We found no significant correlation between the biochemical parameters and neither genotypes nor enzyme activities. The PON1 status and the levels of certain biochemical parameters are independently associated with the hypertension in hypertensive and obese hypertensive patients, and the elevated levels of lipid peroxides and plasma homocysteine may contribute to the increased risk of cardiovascular complications in patients on haemodialysis.

Effect of allopurinol treatment in premature infants with idiopathic respiratory distress syndrome.

A randomized prospective study of the effectiveness of allopurinol (Ap), a potent and specific inhibitor of the enzyme xanthine oxidase, was performed in premature infants endangered by hypoxia. The drug was given at a dose of 20 mg/kg/day orally for 3 days. In the Ap-treated group the expected decrease in the serum concentration and urinary excretion of uric acid was accompanied by a decrease in the mortality rate of infants with idiopathic respiratory distress syndrome. In these patients a concomitant improvements in renal function, as indicated by an increased urinary flow rate and creatinine output, was also obvious. It is suggested that the observed beneficial effect is due to the specific inhibition of xanthine oxidase associated with Ap therapy leading to reduced generation of superoxide radicals and decreased urinary loss of purine.