F2-isoprostane Levels Research Articles

Oxidative stress and cellular stress response in diabetic nephropathy

Oxidative stress has been suggested to play a main role in the pathogenesis of type 2 diabetes mellitus and its complications. As a consequence of this increased oxidative status, a cellular-adaptive response occurs requiring functional chaperones, antioxidant production, and protein degradation. This study was designed to evaluate systemic oxidative stress and cellular stress response in patients suffering from type 2 diabetes-induced nephropathy and in age-matched healthy subjects. Systemic oxidative stress has been evaluated by measuring advanced glycation end-products (pentosidine), protein oxidation (protein carbonyls [DNPH]), and lipid oxidation (4-hydroxy-2-nonenal [HNE] and F2-isoprostanes) in plasma, lymphocytes, and urine, whereas the lymphocyte levels of the heat shock proteins (Hsps) heme oxygenase-1 (HO-1), Hsp70, and Hsp60 as well as thioredoxin reductase-1 (TrxR-1) have been measured to evaluate the systemic cellular stress response. We found increased levels of pentosidine (P < 0.01), DNPH (P < 0.05 and P < 0.01), HNE (P < 0.05 and P < 0.01), and F2-isoprostanes (P < 0.01) in all the samples from type 2 diabetic patients with nephropathy with respect to control group. This was paralleled by a significant induction of cellular HO-1, Hsp60, Hsp70, and TrxR-1 (P < 0.05 and P < 0.01). A significant upregulation of both HO-1 and Hsp70 has been detected also in lymphocytes from type 2 diabetic patients without uraemia. Significant positive correlations between DNPH and Hsp60, as well as between the degree of renal failure and HO-1 or Hsp70, also have been found in diabetic uremic subjects. In conclusion, patients affected by type 2 diabetes complicated with nephropathy are under condition of systemic oxidative stress, and the induction of Hsp and TrxR-1 is a maintained response in counteracting the intracellular pro-oxidant status.

Removing endocannabinoids and reducing oxidative stress with polymyxin-B-immobilized fibers in patients with septic shock

Arachidonylethanolamide (AEA) and 2-arachidonyl-glycerol (2-AG) are endocannabinoids involved in septic shock, and 8-epi prostaglandin F2α (F2-isoprostane) is a biomarker of oxidative stress. Because the antibiotic polymyxin-B binds to endotoxins and endocannabinoids, direct hemoperfusion therapy with polymyxin-B-immobilized fibers (PMX-DHP) decreases serum levels of endocannabinoids. To investigate the features of sepsis and to determine the proper usage of PMX-DHP, we compared perioperative changes in levels of endocannabinoids and F2-isoprostane in patients with septic shock

Polyunsaturated Fatty Acid Pattern in Liver and Erythrocyte Phospholipids from Obese Patients

Our aim was to study the fatty acid (FA) composition of liver phospholipids and its relation to that in erythrocyte membranes from patients with obese nonalcoholic fatty liver disease (NAFLD), as an indication of lipid metabolism alterations leading to steatosis. Eight control subjects who underwent antireflux surgery and 12 obese patients with NAFLD who underwent subtotal gastrectomy with a gastro-jejunal anastomosis in Roux-en-Y were studied. The oxidative stress status of patients was assessed by serum F2-isoprostanes levels (gas chromatography/negative ion chemical ionization tandem mass spectrometry). Analysis of FA composition of liver and erythrocyte phospholipids was carried out by gas-liquid chromatography. Patients with NAFLD showed serum F2-isoprostanes levels 84% higher than controls. Compared with controls, liver phospholipids from obese patients exhibited significantly 1) lower levels of 20:4n-6, 22:5n-3, 22:6n-3 [docosahexaenoic acid (DHA)], total long-chain polyunsaturated FA (LCPUFA), and total n-3 LCPUFA, 2) higher 22:5n-6 [docosapentaenoic acid (DPAn-6)] levels and n-6/n-3 LCPUFA ratios, and 3) comparable levels of n-6 LCPUFA. Levels of DHA and DPAn-6 in liver were positively correlated with those in erythrocytes (r = 0.77 and r = 0.90, respectively; p < 0.0001), whereas DHA and DPAn-6 showed a negative association in both tissues (r = -0.79, p < 0.0001 and r = -0.58, p < 0.01, respectively), associated with lower DHA/DPAn-6 ratios. Obese patients with NAFLD showed marked alterations in the polyunsaturated fatty acid pattern of the liver. These changes are significantly correlated with those found in erythrocytes, thus suggesting that erythrocyte FA composition could be a reliable indicator of derangements in liver lipid metabolism in obese patients.

Elevated cerebrospinal fluid F2-isoprostane levels indicating oxidative stress in healthy siblings of multiple sclerosis patients

Lipid peroxidation has been implicated in the pathogenesis of multiple sclerosis (MS). Isoprostanes, isomers of prostaglandins, are produced by free radical-mediated peroxidation of fatty acids in vivo and can be quantified in biological fluids. This study examines the levels of cerebrospinal fluid (CSF) F2-isoprostanes (F2-iPs) in MS patients ( n = 46), their healthy siblings ( n = 46) and unrelated controls ( n = 50). The median CSF F2-iP concentration (range) was significantly higher in siblings of MS patients, as compared to healthy controls (40.0 [7.1–68.7] and 29.1 [6.4–60.3] pg/mL, respectively, p = 0.031). MS patients demonstrated F2-iP levels intermediate between siblings and controls. F2-iP levels in MS patients and siblings correlated significantly ( R = 0.360, p = 0.012). These results suggest that siblings of MS patients have an increased oxidative stress response to environmental and/or genetic factors that may be involved in MS pathogenesis.

The Effect of Voluntary Wheel Running on Muscle Mass, Mitochondrial Function and Oxidative Damage in Sod1???/???mice

An absence in a major antioxidant enzyme CuZnSOD (SOD1) leads to age-related muscle loss. Exercise training has been demonstrated to induce beneficial changes in both muscle mass and function. The aim of the present study was to examine the potential protective effect of long-term voluntary wheel running (VWR) on muscle mass, mitochondrial function (including ROS and ATP production, RCR), and, on oxidative damage markers (cytosolic protein carbonyls and tissue F2-isoprostanes) in old Sod1−/− mice (20 months of age). At three months of age, mice were randomly divided into four different groups: WT and Sod1−/− (sedentary), and, WT and Sod1−/− (VWR), for a period of 15 months. Sod1−/− mice ran approximately 30% less than WT mice on a daily average. VWR induced positive changes in muscle mass in all three muscles studied (soleus, tibialis anterior and gastrocnemius) for both Sod1−/− and WT. When normalized to body weight (muscle mass/body weight), VWR increased the ratio for all three muscles, in both WT and Sod1−/−mice. Mitochondrial ROS production was increased in untrained Sod1−/− when compared to WT, either during State 1 (no exogenously added substrate) or when using Complex I or II specific substrates (glutamate/malate or succinate+rotenone, respectively). However, in response to long term VWR mitochondrial ROS production was reduced in State1 and glutamate/malate supported respiration (but not with succinate+rotenone) for the Sod1−/− mice. These changes would suggest a beneficial effect on mitochondrial function, but ATP production rates and RCR (State 3/State4) were measured to be unchanged. VWR had no effect on oxidative damage to protein as measured by the levels of cytosolic protein carbonyls in WT animals, but resulted in an increase in this parameter in muscle from the Sod1−/− mice. Oxidative damage to lipids, as measured by the levels of F2-isoprostanes, was increased in muscle from Sod1−/− mice when compared to WT, regardless if trained or sedentary. VWR did not induce further increases in F2-isoprostane levels in any of the measured groups. Collectively, these data suggest that although VWR reduced ROS production by Sod1−/− mitochondria, elevated levels of oxidative damage may play a role in the discovered limitation in voluntary wheel running. Supported by NIH P01 AG20591 (H.V.R.), MDA Grant MDA3879 (H.V.R.), and ACSM(Texas) SRDA to YCJ

Biochemical and Ultrastructural Lung Damage Induced by Rhabdomyolysis in the Rat

Rhabdomyolysis-induced oxidative stress is associated with morphological and functional damage to the kidney and other organs, but applications of this model in the lung are still lacking. The aim of the present study was to determine the relationship between oxidative stress and the morphological changes occurring in the lungs of rats subjected to rhabdomyolysis. Rhabdomyolysis was induced by intramuscular glycerol injection (50% v/v, 10 ml/kg), and the control group was injected with saline vehicle. Arterial blood samples were drawn at 0, 2, 4, and 6 hrs for measurement of arterial gases, creatine kinase activity, and plasma free F2-isoprostane levels. Six hours later, the lungs were removed to determine the wet-to-dry weight ratio, reduced glutathione (GSH) and GSH disulfide (GSSG) levels, and activity of antioxidant enzymes (catalase [CAT], superoxide dismutase [SOD], and GSH peroxidase [GSH-Px]). Protein carbonylation and lipid peroxidation were assessed in the lungs by measurement of carbonyl and malondialdehyde (MDA) production, respectively. Bronchoalveolar lavage, cell counts, and lung ultrastructural studies were also performed. Six hours after glycerol injection, arterial PO2 and PCO2 were 23% and 38% lower, respectively, and plasma free F2-isoprostane levels were 72% higher, compared with control values. In lungs, protein carbonyl and MDA production were 58% and 12% higher, respectively; the GSH:GSSG ratio and GSH-Px activity were 43% and 60% lower, respectively; and activities of CAT and SOD showed no significant differences compared with controls. Rhabdomyolysis-induced ultrastructural impairment of the lung showed Type II cell damage, extracytoplasmic lamellar bodies and lack of tubular myelin reorganization, endothelial cellular edema, and no disruption of the alveolar-capillary barrier. These results provide evidence that rhabdomyolysis could induce tissue injury associated with increased oxidative stress, suggesting the contribution of oxidative stress to the pathogenic mechanism of acute lung injury.

Brassica vegetable consumption reduces urinary F2-isoprostane levels independent of micronutrient intake

Isothiocyanates and indoles (e.g. indole-3-carbinol) from Brassica vegetables (e.g. broccoli) induce Phase I and Phase II enzymes responsible for the oxidation, reduction and metabolism of endogenous and exogenous carcinogens. Brassica vegetables also contain micronutrients that may provide additional DNA protection from reactive oxygen species. This randomized crossover trial (n = 20) compares the effects of a Brassica Vegetable (BV) intervention against a Micronutrient and Fiber Supplementation (M+F) intervention on urinary F2-isoprostane levels (F2-iP), a stable biomarker of systemic oxidative stress. Brassica intake was monitored by repeated 24 h recalls, urinary ITC levels and questionnaire. Urinary F2-iP levels were measured by mass spectrometry from first-morning urine samples collected at Baseline and after each intervention, and change in natural log transformed urinary F2-iP levels were analyzed using repeated measures regression. Brassica consumption increased from 2 grams/day (g/d) during the Baseline or M+F intervention periods to 218 g/d during the BV intervention, whereas exposure to most antioxidant vitamins and minerals was greatest during the M+F intervention. F2-iP levels significantly decreased by 22.0 or 21.8% during the BV intervention compared with Baseline or the M+F intervention (P = 0.05, P = 0.05, respectively). Urinary F2-iP levels did not significantly differ between Baseline and the M+F intervention (difference = 0.2%; P = 0.98). Brassica intake has been associated with reduced risk of colon, lung, bladder, breast, prostate and other cancers. Our results suggest that Brassica consumption reduces systemic oxidative stress independent of the vitamin and mineral content of these vegetables.

The importance of dietary antioxidants on plasma antioxidant capacity and lipid peroxidationin vivoin middle-aged men

Background: High intake of foods rich in antioxidants is associated with a reduced risk of several diseases, including coronary heart disease and some cancers. Objective: To study associations between dietary antioxidants and antioxidant-rich food items and antioxidant capacity (AOC), as well as lipid peroxidation in vivo. Design: A total of 86 men, with a mean age of 60 years, who were part of a larger population-based study in Sweden, participated. Fourteen 24 h recalls were collected by telephone, evenly distributed during 1 year. AOC was measured in plasma using an enhanced chemiluminescence assay and biomarkers of non-enzymic in vivo lipid peroxidation were determined by measuring F2-isoprostanes (8-iso-prostaglandin F2a) in plasma. Results: Higher intakes of ascorbic acid, tocopherols and b-carotene, and of the combined intake of different antioxidant-rich foods, were related to a higher plasma AOC. The levels of F2-isoprostanes were related neither to dieta ry intake of antioxidants nor to antioxidant-rich foods. Conclusions: Individuals with a higher intake of several different antioxidant-rich foods and of dietary antioxidants had a higher plasma AOC than those with the lowest intake, supporting the importance of a balanced diet rich in various antioxidants. Moreover, the results indicate that non-enzymic lipid peroxidation in vivo, measured as F2-isoprostanes in plasma, is not easily affected by the diet. Keywords: antioxidant status; diet; food habits; isoprostanes; oxidative stress

Cimetidine Does Not Prevent Lung Injury in Newborn Premature Infants

Animal studies have shown that induction of cytochrome P450 (CYP) in the lung by oxygen exposure may result in the release of free radical oxidants and arachidonic acid metabolites, which can cause lung injury that is reduced by treatment with cimetidine, a CYP inhibitor. To determine whether cimetidine would reduce lung injury in human infants at risk for chronic lung disease, we conducted a randomized clinical trial in which we administered either cimetidine or a placebo for 10 d beginning < 24 h after birth to 84 newborn infants weighing < or = 1250 g who were receiving O2 and mechanical ventilation. Cimetidine had no significant effect on severity of respiratory insufficiency assessed at 10 d postnatal age. F2-isoprostane levels (a marker of oxidant injury) in tracheal aspirates were significantly higher in the cimetidine group at 4 d and at 10 d. There were no significant differences between the groups in tracheal aspirate levels of inflammatory markers (leukotriene B4, IL-8, and nucleated cell count) or arachidonic acid metabolites. We conclude that cimetidine does not reduce lung injury in newborn premature infants receiving O2 and mechanical ventilation. It is possible that cimetidine was not an adequate CYP inhibitor in this context.

Inhibitory effect of melatonin on diquat‐induced lipid peroxidation in vivo as assessed by the measurement of F2‐isoprostanes

Melatonin is a powerful antioxidant and free radical scavenger. A large body of in vivo and in vitro evidence shows that melatonin effectively inhibits membrane lipid peroxidation; this damage was based on the measurement of malondialdehyde and/or 4-hydroxynonenal levels. In the current study, for the first time using a more sensitive and specific biomarker, i.e. F2-isoprostanes, we investigate the effect of melatonin on diquat-induced lipid peroxidation in Fischer 344 rats. When diquat (40 mg/kg body weight) was intraperitoneally injected into rats, the levels of liver F2-isoprostanes were significantly increased at 1, 3, and 6 hr while plasma free F2-isoprostanes concentrations were augmented at 3, 6, and 12 hr after administration of the toxin. In addition, the plasma alanine aminotransferase activity level was measured as a parameter of hepatoxicity; the activity of this enzyme was augmented at 3, 6, and 12 hr after diquat administration when compared with levels of this constituent in untreated control rats. Pretreatment with melatonin (20 mg/kg) 30 min before diquat administration resulted in a significant reduction in both tissue and plasma F2-isoprostanes levels, and plasma alanine aminotransferase activity. These findings, using a sensitive and specific index of lipid peroxidation, show that the hepatoxicity of diquat, at least partially, is a consequence of reactive oxygen species-induced lipid damage. Melatonin's protective effects likely relate to its direct free radical scavenging ability and/or due to other antioxidative processes induced by the indole.

Effect of High Dose Alpha Tocopherol Therapy on Carotid Atherosclerosis, Biomarkers of Oxidative Stress and Inflammation in Patients with Coronary Artery Disease

Oxidative stress and inflammation are crucial in atherogenesis. High dose Alpha tocopherol (RRR-AT) supplementation has antioxidant and anti-inflammatory effects. Prospective studies with lower doses of AT on atherosclerosis have been equivocal. Thus, we tested the effect of RRR-AT supplementation(1200 IU/day for 2 years)on carotid atherosclerosis in a placebo-controlled, double blind study of 90 stable coronary artery disease patients. Intimal medial thickness (IMT) of both carotids, was determined by B-mode sonography at 0, 1, 1.5 and 2 yrs. At 6-month intervals blood was obtained for AT levels, C-reactive protein (CRP) and 24-hour urine for F2 -isoprostanes. AT levels significantly increased in the AT group (2.4-fold) but not in placebo. There was no significant difference in total or left carotid IMT between placebo and AT groups. There was a significant increase in IMT progression in Placebo with time for total and right CCA-IMT (p<0.05). Also, there was a significant decrease in IMT progression in the right CCA between placebo and AT groups over 2 years (p=0.017). HsCRP levels and F2-isoprostane levels significantly decreased with AT therapy compared to placebo (32% and 26%,p<0.01). Thus, high dose RRR-AT supplementation in CAD patients significantly reduced biomarkers of oxidative stress and inflammation, there was only a marginal benefit with regards to carotid atherosclerosis progression.

Lipid peroxidation in mice fed a choline-deficient diet as evaluated by total hydroxyoctadecadienoic acid

Objective The relevance of oxidative stress in mice fed a choline-deficient diet (CDD) was investigated in relation to the oxidative stress marker, hydroxyoctadecadienoic acid (HODE) in comparison with F2-isoprostanes. Further, the protective effects of antioxidants against oxidative damage were assessed by using HODE. Methods We recently proposed total HODE as a biomarker for oxidative stress in vivo. Biological samples such as plasma, urine, and tissues were first reduced and then saponified to convert various oxidation products of linoleates to HODE. In the present study, this method was applied to measure oxidative damage in mice induced by CDD for 1 mo. Results CDD, when compared with choline-controlled diet (CCD), increased liver weight and fatty acid accumulation but the increase in body weight was less significant. Remarkable increases in HODE and 8-iso-prostaglandin F 2α in liver and plasma were observed when mice were fed with the CDD for 1 mo compared with the CCD. The HODE level was about two to three orders higher than the F2-isoprostane level. This increase was decreased to the level of the CCD when α-tocopherol or 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran, a potent synthetic antioxidant, was mixed with the CDD. The stereoisomer ratio of HODE (9-and-13 ( Z, E)-HODE/9-and-13 ( E, E)-HODE) was decreased by CDD compared with CCD, which was spared by the addition of α-tocopherol and 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran. However, the increase in plasma glutamic-pyruvic transaminase and fatty acids in liver induced by the CDD was not recovered by any antioxidant. Conclusions This study clearly demonstrated that oxidative stress was involved in fatty liver formation induced by the CDD and that HODE was a good biomarker for an oxidative stress in vivo.

Release of Free F2-isoprostanes from Esterified Phospholipids Is Catalyzed by Intracellular and Plasma Platelet-activating Factor Acetylhydrolases

F2-isoprostanes are produced in vivo by nonenzymatic peroxidation of arachidonic acid esterified in phospholipids. Increased urinary and plasma F2-isoprostane levels are associated with a number of human diseases. These metabolites are regarded as excellent markers of oxidant stress in vivo. Isoprostanes are initially generated in situ, i.e. when the arachidonate precursor is esterified in phospholipids, and they are subsequently released in free form. Although the mechanism(s) responsible for the release of free isoprostanes after in situ generation in membrane phospholipids is, for the most part, unknown, this process is likely mediated by phospholipase A2 activity(ies). Here we reported that human plasma contains an enzymatic activity that catalyzes this reaction. The activity associates with high density and low density lipoprotein and comigrates with platelet-activating factor (PAF) acetylhydrolase on KBr density gradients. Plasma samples from subjects deficient in PAF acetylhydrolase do not release F2-isoprostanes from esterified precursors. The intracellular PAF acetylhydrolase II, which shares homology to the plasma enzyme, also catalyzes this reaction. We found that both the intracellular and plasma PAF acetylhydrolases have high affinity for esterified F2-isoprostanes. However, the rate of esterified F2-isoprostane hydrolysis is much slower compared with the rate of hydrolysis of other substrates utilized by these enzymes. Studies using PAF acetylhydrolase transgenic mice indicated that these animals have a higher capacity to release F2-isoprostanes compared with nontransgenic littermates. Our results suggested that PAF acetylhydrolases play key roles in the hydrolysis of F2-isoprostanes esterified on phospholipids in vivo.

Increased F2-Isoprostane Levels in the Rat Brain and Plasma Caused by Oxidative Stress and Aging, and Inhibitory Effect of Vitamin E

Summary To determine whether lipid peroxidation induced by reactive oxygen species (ROS) is a causal factor of neurodegeneration during brain aging, we investigated whether F2isoprostanes, non-cyclooxygenase-derived prostanoids, are formed in the rat brain and plasma by hyperoxia as oxidative stress, and whether their formation is associated with vitamin E status in vivo in association with changes that occur during brain aging. Young rats subjected to hyperoxia for 48 h revealed a marked increase in the levels of F2-isoprostanes in the brain, but not in plasma. A similar increase in F2-isoprostane level was observed in aged rats kept in normal atmosphere. Vitamin E supplementation to young rats markedly inhibited F2-isoprostane formation even after hyperoxia. In contrast, vitamin E-deficient young rats kept in normal atmosphere showed a significant increase in F2-isoprostane level in the brain. These findings indicate that F2-isoprostane formation in the brain has important implications in the etiology of neurodegenerative diseases including Alzheimer’s disease during aging, and that the analysis of F2-isoprostane level in plasma does not always reflect neuronal damage cause by oxidative stress. Vitamin E may protect neuronal damage in the brain caused by oxidative stress experienced for a long period during aging.

High levels of F2-Isoprostanes in Jamaican adults with diabetes mellitus

Inadequate glycaemic control in Jamaican adults with type 2 diabetes mellitus (DM) prompted assessment of glycaemiccontrol, oxidative stress, cardiovascular (CV) and renal risk in adults attending a hospital clinic. A random sample of 133patients men (n=35) and women (n=98), with diabetes mellitus was selected from a population of 510 patients. Fasting bloodsamples (n=122) were evaluated for metabolic control and dyslipidaemia. Oxidative stress was evaluated by measurement ofurinary F2 isoprostane (n=124). The data were analysed using SPSS. The mean age of the participants was 56.7 ± 14.3 years,with mean duration of diabetes of 12.2 years. Mean fasting blood sugar was 8.6 ± 4.3 mmol/L. 77% of patients had HbA1c >6.5%. 69% of patients were being treated with insulin with no difference in HbA1c levels in these patients compared to thosereceiving other hypoglycaemic agents. 90% of men (median 1004pg/mg creatinine) and 99% of women (median1501.3pg/mg) had isoprostane levels above the median for subjects with CV risk. 54% of patients had total cholesterol levels≥ 5.2 mmol/L, 16% triglyceride levels ≥ 1.5 mmol/L, 25% HDL levels ≤ 1.0mmol/L and 86% LDL ≥ 2.5mmol/L. 63 % ofpatients had BP >130/85 mmHg. 81% were overweight or obese, with 80% of the men having waist circumferences >88 cm,whereas 87% of the women had waist circumferences of 84.5cm. Microalbuminuria was increased in 37% of the subjects.The high prevalence of overweight, central obesity, elevated LDL and hypertension in these patients indicated high CV andrenal risk. The risk was significantly higher in persons with inadequate glycaemic control. Isoprostane levels were high in themajority of subjects but did not correlate with HbA1c or any other variable. (Int J Diabetes 14: 51-54, 2006 )

Hydrogen Peroxide Reduces Lower Esophageal Sphincter Tone in Human Esophagitis

We have previously used the normal lower esophageal sphincter (N-LES) of human organ donors to examine the physiologic signal transduction of lower esophageal sphincter (LES) circular muscle. Now, for the first time, we have obtained a human LES specimen with esophagitis (E-LES) and characterized its pathophysiologic mechanical and inflammatory profiles. E-LES was examined histologically, and its in vitro circular muscle contraction and production of inflammatory mediators were compared with those of N-LES. E-LES exhibited scattered erosions and displayed inflammatory cells in the epithelial layer, basal zone hyperplasia, and elongation of lamina propria papillae, characteristic of chronic reflux esophagitis. E-LES muscle strips developed lower in vitro tone (0.78 g) than N-LES (3.3 +/- 0.2 g). E-LES tone was essentially restored to normal by the H2O2 scavenger catalase, suggesting that H2O2 was responsible for reduction of tone. NOX5 cDNA was higher and H2O2 levels were 4 times higher in E-LES circular muscle (0.85 nmol/mg protein) than in N-LES (0.19 +/- 0.05 nmol/mg protein). When N-LES smooth muscle was incubated in H2O2 (70 micromol/L, 2 hours), platelet activating factor (PAF), prostaglandin E2 (PGE2), and F2-isoprostane increased 2.5, 5.2, and 36 times, respectively. In E-LES, levels of PAF, PGE2, and F2-isoprostane were 4, 6, and 40 times, respectively, higher than in N-LES. PAF, PGE2, and F2 isoprostane produced dose-dependent reductions in tone of N-LES muscle strips. We conclude that an excessive production of H2O2 triggers an increased production of PAF, PGE2, and F2-isoprostane, which are responsible for reducing LES tone in human esophagitis.

Levels of F2-isoprostanes in systemic sclerosis: correlation with clinical features

Oxidative stress may be one of the important complex pathogenetic mechanisms that lead to damage in scleroderma; free radicals may provoke endothelial injury, fibroblast proliferation and fragmentation of autoantigens favouring induction of autoantibodies. The present study investigates the oxidant status in scleroderma patients by measuring the urinary concentration of 8-isoprostaglandin-F2alpha, an F2-isoprostane, and a product of free radical-mediated peroxidation of arachidonic acid. Forty-three scleroderma patients (42 women and 1 man, mean age 54.1 yr, mean disease duration 9.0 yr) underwent clinical evaluation and instrumental investigations in order to assess skin, vascular, lung and heart involvement. Von Willebrand factor was evaluated as marker of vascular dysfunction in 36 out of the 43 cases. The urinary level of 8-isoprostaglandin-F2alpha was measured in all scleroderma patients and in the 43 age- and sex-matched healthy controls. Urinary levels of 8-isoprostaglandin-F2alpha were higher in scleroderma patients than in the healthy control group (341.7 vs 147.6 pg/mg creatinine; P < 0.001). Values of 8-isoprostaglandin-F2alpha were strongly correlated with the nailfold videocapillaroscopy pattern and lung involvement (P = 0.002 and 0.003, respectively), showing increasing levels with the progression of pulmonary severity. Correlation between 8-isoprostaglandin-F2alpha level and von Willebrand factor narrowly failed to reach statistical significance (P = 0.05). There was no correlation between 8-isoprostaglandin-F2alpha concentration and disease activity, vascular, skin and heart involvement, disease pattern or autoantibody profile. Our study further supports the role of oxidant stress in the pathogenesis of scleroderma, showing a strong correlation between a marker of free radical damage with both the severity of lung involvement and the videocapillaroscopic patterns.

Hyperinsulinemia Provokes Synchronous Increases in Central Inflammation and β-Amyloid in Normal Adults

Inflammation has been implicated as a pathogenetic factor in Alzheimer disease, possibly via effects on beta-amyloid (Abeta). Hyperinsulinemia induces inflammation and is a risk factor for Alzheimer disease. Thus, insulin abnormalities may contribute to Alzheimer disease pathophysiology through effects on the inflammatory network. To determine the effects of induced hyperinsulinemia with euglycemia on Abeta, transthyretin, and inflammatory markers and modulators in plasma and cerebrospinal fluid (CSF). Randomized crossover trial. Veterans Affairs hospital clinical research unit. Sixteen healthy adults ranging from 55 to 81 years of age (mean age, 68.2 years). On separate mornings, fasting participants received randomized infusions of saline or insulin (1.0 mU.kg(-1).min(-1)) with variable dextrose levels to maintain euglycemia, achieving plasma insulin levels typical of insulin resistance. Plasma and CSF were collected after an approximately 105-minute infusion. Plasma and CSF levels of interleukin 1alpha, interleukin 1beta, interleukin 6, tumor necrosis factor alpha, F2-isoprostane (CSF only), Abeta, norepinephrine, transthyretin, and apolipoprotein E. Insulin increased CSF levels of F2-isoprostane and cytokines (both P<.01), as well as plasma and CSF levels of Abeta42 (both P<.05). The changes in CSF levels of Abeta42 were predicted by increased F2-isoprostane and cytokine levels (both P<.01) and reduced transthyretin levels (P = .02). Increased inflammation was modulated by insulin-induced changes in CSF levels of norepinephrine and apolipoprotein E (both P<.05). Moderate hyperinsulinemia can elevate inflammatory markers and Abeta42 in the periphery and the brain, thereby potentially increasing the risk of Alzheimer disease.

Comparative Proteomic Analysis of Intra- and Interindividual Variation in Human Cerebrospinal Fluid

Cerebrospinal fluid (CSF) is a potential source of biomarkers for many disorders of the central nervous system, including Alzheimer disease (AD). Prior to comparing CSF samples between individuals to identify patterns of disease-associated proteins, it is important to examine variation within individuals over a short period of time so that one can better interpret potential changes in CSF between individuals as well as changes within a given individual over a longer time span. In this study, we analyzed 12 CSF samples, composed of pairs of samples from six individuals, obtained 2 weeks apart. Multiaffinity depletion, two-dimensional DIGE, and tandem mass spectrometry were used. A number of proteins whose abundance varied between the two time points was identified for each individual. Some of these proteins were commonly identified in multiple individuals. More importantly, despite the intraindividual variations, hierarchical clustering and multidimensional scaling analysis of the proteomic profiles revealed that two CSF samples from the same individual cluster the closest together and that the between-subject variability is much larger than the within-subject variability. Among the six subjects, comparison between the four cognitively normal and the two very mildly demented subjects also yielded some proteins that have been identified in previous AD biomarker studies. These results validate our method of identifying differences in proteomic profiles of CSF samples and have important implications for the design of CSF biomarker studies for AD and other central nervous system disorders.

Oxidant injury occurs rapidly after cardiac arrest, cardiopulmonary resuscitation, and reperfusion*

Investigations conducted in cellular models show that reperfusion of ischemic tissue is associated with a burst of reactive oxidant species within minutes after reperfusion. Oxidant injury may play a role in the poor outcome typical of people resuscitated from cardiac arrest. The objective of the present study was to determine the presence and timing of oxidant injury in an in vivo model of cardiac arrest. Randomized controlled. University medical center laboratory. Domestic swine. We evaluated oxidant injury during and after 8 mins of cardiac arrest using a gas chromatography/mass spectrometry F2-isoprostane assay and compared these results with a matched control group. Baseline mean arterial, venous, and brain tissue F2-isoprostane levels were not significantly different when the cardiac arrest group was compared with the control group. However, in the group subjected to cardiac arrest and cardiopulmonary resuscitation we found significant (p < .0006) two- to three-fold increases in venous and arterial F2-isoprostane levels, which peaked between 15 and 30 mins after reperfusion and returned to baseline within 90 mins (p < .0006). Overall mean (+/- SE) brain tissue F2-isoprostane levels increased significantly to 370 +/- 60 vs. 140 +/- 60 ng/g tissue in the cardiac arrest group compared with the control group (p = .026). This study shows that F2-isoprostane measurement could be used to assess oxidant injury in an animal model of cardiac arrest and that oxidant injury occurs rapidly after cardiac arrest and reperfusion.