Olfactory receptor 2 in vascular macrophages drives atherosclerosis by NLRP3-dependent IL-1 production.

Atherosclerosis is an inflammatory disease of the artery walls and involves immune cells such as macrophages. Olfactory receptors (OLFRs) are G protein­coupled chemoreceptors that have a central role in detecting odorants and the sense of smell. We found that mouse vascular macrophages express the olfactory receptor Olfr2 and all associated trafficking and signaling molecules. Olfr2 detects the compound octanal, which activates the NLR family pyrin domain containing 3 (NLRP3) inflammasome and induces interleukin-1ß secretion in human and mouse macrophages. We found that human and mouse blood plasma contains octanal, a product of lipid peroxidation, at concentrations sufficient to activate Olfr2 and the human ortholog olfactory receptor 6A2 (OR6A2). Boosting octanal levels exacerbated atherosclerosis, whereas genetic targeting of Olfr2 in mice significantly reduced atherosclerotic plaques. Our findings suggest that inhibiting OR6A2 may provide a promising strategy to prevent and treat atherosclerosis.

The Combination of Cigarette Smoking and Alcohol Consumption Synergistically Increases Reactive Carbonyl Species in Human Male Plasma.

Cigarette smoking and alcohol consumption are major risk factors for lifestyle-related diseases. Although it has been reported that the combination of these habits worsens risks, the underlying mechanism remains elusive. Reactive carbonyl species (RCS) cause chemical modifications of biological molecules, leading to alterations in cellular signaling pathways, and total RCS levels have been used as a lipid peroxidation marker linked to lifestyle-related diseases. In this study, at least 41 types of RCS were identified in the lipophilic fraction of plasma samples from 40 subjects using liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS). Higher levels of 10 alkanals, 5 trans-2-alkenals, 1 cis-4-alkenal, and 3 alkadienals were detected in the smoking/drinking group (N = 10) as compared to those with either habit (N = 10 each) or without both habits (N = 10) in the analysis of covariances adjusted for age and BMI. The levels of 3 alkanals, 1 trans-2-alkenal, 1 alkadienal, and 1 4-hydroxy-2-alkenal in the smoking/drinking group were significantly higher than those in the no-smoking/drinking and no-smoking/no-drinking groups. These results strongly indicate that the combination of cigarette smoking and alcohol drinking synergistically increases the level and variety of RCS in the circulating blood, and may further jeopardize cellular function.

Elevated serum 4HNE plus decreased serum thioredoxin: Unique feature and implications for acute exacerbation of chronic obstructive pulmonary disease.

Acute exacerbation of chronic obstructive pulmonary disease (AECOPD) is a global problem with high mortality. Its pathogenesis is not fully understood. To reveal new serum feature of AECOPD and their potential implications, we have analyzed 180 serum samples, and found that in the serum of AECOPD patients, 4-hydroxy-2-nonenal (4HNE)-protein adducts are dynamically increased as partial pressure of oxygen (PaO2) drops, which is accompanied by progressively decreasing thioredoxin reductase (TrxR1) and thioredoxin (Trx1), as compared with those of healthy people. This phenomenon is unique, because acute hypoxia patients have 1.1-fold or 1.7-fold higher serum TrxR1 or Trx1 activity, respectively, than healthy people, in keeping with low 4HNE level. Moreover, serum 4HNE-protein adducts may form disulfide-linked complexes with high-molecular-weight, the amount of which is significantly increased during AECOPD. Serum 4HNE-protein adducts include 4HNE-Trx1 adduct and 4HNE-TrxR1 adduct, but only the former is significantly increased during AECOPD. Through cell biology, biochemistry and proteomics methods, we have demonstrated that extracellular 4HNE and 4HNE-Trx1 adduct affect human bronchial epithelial cells via different mechanisms. 4HNE-Trx1 adduct may significantly alter the expression of proteins involved mainly in RNA metabolism, but it has no effect on TrxR1/Trx1 expression and cell viability. On the other hand, low levels of 4HNE promote TrxR1/Trx1 expression and cell viability, while high levels of 4HNE inhibit TrxR1/Trx1 expression and cell viability, during which Trx1, at least in part, mediate the 4HNE action. Our data suggest that increasing serum 4HNE and decreasing serum Trx1 in AECOPD patients are closely related to the pathological processes of the disease. This finding also provides a new basis for AECOPD patients to use antioxidant drugs.

Two Aldehyde Clearance Systems Are Essential to Prevent Lethal Formaldehyde Accumulation in Mice and Humans.

Reactive aldehydes arise as by-products of metabolism and are normally cleared by multiple families of enzymes. We find that mice lacking two aldehyde detoxifying enzymes, mitochondrial ALDH2 and cytoplasmic ADH5, have greatly shortened lifespans and develop leukemia. Hematopoiesis is disrupted profoundly, with a reduction of hematopoietic stem cells and common lymphoid progenitors causing a severely depleted acquired immune system. We show that formaldehyde is a common substrate of ALDH2 and ADH5 and establish methods to quantify elevated blood formaldehyde and formaldehyde-DNA adducts in tissues. Bone-marrow-derived progenitors actively engage DNA repair but also imprint a formaldehyde-driven mutation signature similar to aging-associated human cancer mutation signatures. Furthermore, we identify analogous genetic defects in children causing a previously uncharacterized inherited bone marrow failure and pre-leukemic syndrome. Endogenous formaldehyde clearance alone is therefore critical for hematopoiesis and in limiting mutagenesis in somatic tissues.

Association of aldehyde exposure with cardiovascular disease.

The effect of aldehyde exposure on the cardiovascular system remains unclear. The objective of this study was to determine whether aldehyde exposure is associated with the prevalence of cardiovascular disease (CVD). We analyzed associations between aldehydes and CVD using data from 1962 adult participants in the National Health and Nutrition Examination Survey (NHANES) from 2013 to 2014. Multivariable logistic regression and restricted cubic spline models were used to examine the association between aldehydes and CVD. The prevalence of CVD was 10.3%. After adjusting for confounding factors, including age, sex, education level, race, diabetes mellitus, smoking, alcohol use, hypertension, body mass index, the poverty-income ratio, physical activity, energy intake, high-density cholesterol (HDL) and low-density cholesterol (LDL), compared with the lowest quartiles, the odds ratios (ORs) with 95% confidence intervals (CIs) for CVD across the quartiles were 0.52 (0.31, 0.87), 0.73 (0.43, 1.22), and 1.13 (0.68, 1.86) for benzaldehyde and 1.48 (0.87, 2.52), 1.70 (1.01, 2.92), and 2.13 (1.19, 3.86) for isopentanaldehyde. There was no significant association between other aldehydes and CVD. The restricted cubic spline plot showed a J-curve relationship between benzaldehyde and CVD. The inflection point for the curve was found at a benzaldehyde level of 0.98 ng/ml. The ORs (95% CIs) for CVD were 0.51 (0.31, 0.86) and 1.58 (1.15, 2.17) on the left and right sides of the inflection point, respectively. Our results demonstrate a J-curve relationship between benzaldehyde and CVD. Isopentanaldehyde is positively associated with CVD. Further study is warranted to verify this association and to elucidate its underlying mechanisms.

Association of aspirin resistance with 4-hydroxynonenal and its impact on recurrent cerebral infarction in patients with acute cerebral infarction.

OBJECTIVES: To investigate the association of aspirin resistance (AR) with the plasma 4-hydroxynonenal (4-HNE) level and its impact on recurrent cerebral infarction (CI) in patients with acute cerebral infarction (ACI) who were receiving aspirin therapy. METHODS: One hundred and fifty-four ACI patients who previously received aspirin therapy (100 mg/day) were enrolled. Whole urine (for measuring 11dhTXB2 and creatinine) along with blood (for measuring the plasma 4-HNE level) were collected at least 7 days after the patients received aspirin. A cutoff of 1500 pg/mg of 11dhTXB2/ creatinine was used to determine AR. A follow-up period to monitor recurrence CI events was 1 year. In addition, blood testing was performed when the patients were first admitted to hospital. RESULTS: Forty-six of the 154 enrolled patients (29.9%) were found to be AR. No statistical difference in age, sex, hypertension, diabetes mellitus, coronary disease, smoking status, NIHSS score, TOAST classification, platelet count, thrombocytocrit, LDL-C, HDL-C, TG, and TC was found between the AR and aspirin-sensitive (AS) patients, but the plasma 4-HNE level was found to be higher in the AR patients than AS patients (p < .05). Multiple logistic regression analysis showed that the 4-HNE level was associated with a higher risk of AR (OR = 1.034; 95% CI = 1.011-1.058; p < .05). Moreover, 1-year follow-up showed that AR was more prevalent in patients with recurrent CI (26 (56.6%)) than those without (20/(43.5%)) (p < .001). CONCLUSIONS: The plasma 4-HNE level is strongly associated with AR and thus may be a factor contributing to AR. Patients with AR have a greater risk of recurrence CI.

Involvement of Pazopanib and Sunitinib Aldehyde Reactive Metabolites in Toxicity and Drug-Drug Interactions in Vitro and in Patient Samples.

Tyrosine kinase inhibitors (TKI) are targeted anticancer drugs that have been successfully developed over the past 2 decades. To date, many of them (around 70%) require warnings for liver injury and five of them, including pazopanib and sunitinib, have Black Box Warning (BBW) labels. Although TKI-induced hepatotoxicity is the first cause of drug failures in clinical trials, BBW labels, and market withdrawals, the underlying mechanisms remain unclear. However, the recent discovery of new reactive metabolites (RM) with aldehyde structures during pazopanib and sunitinib metabolism offers new perspectives for investigating their involvement in the toxicity of these two TKI. These hard electrophiles have a high reactivity potential toward proteins and are thought to be responsible for cytochrome P450 inactivation, drug-drug interactions (DDI), and liver toxicity. We report here, for the first time, the presence of these aldehyde RM in human plasma samples obtained during drug monitoring. Docking experiments in the CYP3A4 active site were performed and showed that pazopanib and sunitinib fitting in the catalytic site are in accordance with their regioselective oxidation to aldehydes. They also suggested that aldehyde RM may react with lysine and arginine residues. Based on these results, we studied the reactivity of the aldehyde RM toward lysine and arginine residues as potential targets on the protein framework to better understand how these RM could be involved in liver toxicity and drug-drug interactions. Adduct formation with different hepatic and plasma proteins was investigated by LC-MS/MS, and adducts between pazopanib or sunitinib aldehyde derivatives and lysine residues on both CYP3A4 and plasma proteins were indeed shown for the first time.

Simultaneous determination of seven endogenous aldehydes in human blood by headspace gas chromatography-mass spectrometry.

Endogenous aldehydes (EAs) formed by the free-radical-mediated reaction are regarded as potential biological markers of several diseases. In this work, an automated and solvent-free analytical method was developed for quantitative analysis of seven EAs (C1-C7) in human blood by using gas chromatography-mass spectrometry coupled with a headspace generator sampler (HS-GC-MS). A 1:4 (blood:water) dilution and the 1,2­dibromopropane internal standard were introduced to reduce the influence of the matrix effect from complex biological fluids. The sample preparation steps were simplified. Various experimental parameters for derivatization and extraction conditions were studied such as HS extraction temperature and time, the amount of derivatization reagent, pH and the salt effect. Under these optimum conditions, seven low-mass aldehydes were separated and analyzed within 10 min. Additionally, this method achieved limits of detection in the range of 0.0692-0.864 µg/L, an excellent linearity with correlation coefficients higher than 0.9996 and appropriate repeatability and reproducibility values (RSD < 12% at low, middle and high levels). The HS-GC-MS method was applied to measure the concentrations of the seven aldehydes in blood from bladder cancer patients (n = 15) and control subjects (n = 15). Compared with the control subjects, the levels of butanal (p < 0.01), formaldehyde, acetaldehyde, propanal, pentanal, hexanal and heptanal (all p < 0.001) were increased significantly, indicating that EAs may be useful as biomarkers in the early diagnosis of bladder cancer.

Aumento de estresse oxidativo no sangue de pacientes com ostomia / Increased oxidative stress in the blood of ostomy patients

ABSTRACT BACKGROUND: Ostomy is a surgical procedure that creates a stoma that aims to construct a new path for the output of feces or urine. The relationship of oxidative stress (OxS) markers in patients with ostomy is still poorly described. OBJECTIVE: The present study was aimed at investigating the changes in oxidative stress parameters in peripheral blood collected from ostomy patients when compared with a healthy control group. METHODS: It was evaluated 29 ostomy patients and 30 healthy control patients. The oxidative stress parameters evaluated were: lipid peroxidation [lipid hydroperoxide (LPO), 8-isoprostane (8-ISO) and 4-hydroxynonenal (4-HNE)], protein oxidation and nitration [carbonyl and 3-nitrotyrosine (3-NT)] and DNA oxidation [8-hydroxy-2'-deoxyguanosine (8-OHDG)] in serum from ostomy patients compared to health controls. RESULTS: The data showed an increase of LPO, 8-ISO, 4-HNE, 3-NT and 8-OHDG in serum collected from ostomy patients when compared to healthy controls. CONCLUSION: The findings support the hypothesis that ostomy triggers the oxidative stress observed in the blood collected from these patients.

RESUMO CONTEXTO: Ostomia é um procedimento cirúrgico que cria um estoma com objetivo de construir um novo caminho para a saída das fezes ou urina. A relação dos marcadores de estresse oxidativo em pacientes ostomizados ainda é pouco descrita. OBJETIVO: O presente estudo tem como objetivo investigar as alterações dos parâmetros de estresse oxidativo em sangue de pacientes ostomizados comparados a controles saudáveis. MÉTODOS: Foram avaliados 29 pacientes ostomizados e 30 controles saudáveis. Os parâmetros de estresse oxidativo avaliados foram: peroxidação lipídica [hidroperóxido de lipídio (LPO), 8-isoprostano (8-ISO) e 4-hidroxinonenal (4-HNE)], oxidação e nitração de proteínas [carbonila e 3-nitrotirosina (3-NT)] e oxidação do DNA [8-hidroxi-2'-desoxiguanosina (8-OHDG)] em soro de pacientes ostomizados comparados a controles saudáveis. RESULTADOS: Os dados mostraram um aumento de LPO, 8-ISO, 4-HNE, 3-NT e 8-OHDG em soro de pacientes ostomizados em comparação a controles saudáveis. CONCLUSÃO: Os achados sustentam a hipótese de que a ostomia desencadeia o estresse oxidativo observado no sangue coletado destes pacientes.

Role of activation of lipid peroxidation in the mechanisms of acute methanol poisoning.

CONTEXT: The role of activation of lipid peroxidation in the mechanisms of acute methanol poisoning has not been studied. OBJECTIVE: We measured the concentrations of lipid peroxidation markers in acutely intoxicated patients with known serum concentrations of methanol and leukotrienes. METHODS: Blood serum samples were collected from 28 patients hospitalized with acute intoxication and from 36 survivors 2 years after discharge. In these samples, concentrations of 4-hydroxy-trans-2-hexenal (HHE), 4-hydroxynonenal (HNE), and malondialdehyde (MDA) were measured using the method of liquid chromatography-electrospray ionization-tandem mass spectrometry. RESULTS: The maximum acute serum concentrations of all three lipid oxidative damage markers were higher than the follow-up serum concentrations: HNE 71.7 ± 8.0 ng/mL versus 35.4 ± 2.3 ng/mL; p < .001; HHE 40.1 ± 6.7 ng/mL versus 17.7 ± 4.1 ng/mL; p < .001; MDA 80.0 ± 7.2 ng/mL versus 40.9 ± 1.9 ng/mL; p < .001. The survivors without methanol poisoning sequelae demonstrated higher acute serum concentrations of the markers than the patients with sequelae. A correlation between measured markers and serum leukotrienes was present: HNE correlated with LTC4 (r = 0.663), LTD4 (r = 0.608), LTE4 (r = 0.771), LTB4 (r = 0.717), HHE correlated with LTC4 (r = 0.713), LTD4 (r = 0.676), LTE4 (r = 0.819), LTB4 (r = 0.746), MDA correlated with LTC4 (r = 0.785), LTD4 (r = 0.735), LTE4 (r = 0.814), LTB4 (r = 0.674); all p < .001. Lipid peroxidation markers correlated with anion gap (r= -0.428, -0.388, -0.334; p = .026, .045, .080 for HNE, HHE, MDA, respectively). The follow-up serum concentrations of lipid oxidation markers measured in survivors with and without visual/neurological sequelae 2 years after discharge did not differ. CONCLUSION: Our results demonstrate that lipid peroxidation plays a significant role in the mechanisms of acute methanol poisoning. The acute concentrations of three measured biomarkers were elevated in comparison with the follow-up concentrations. Neuronal membrane lipid peroxidation seems to activate leukotriene-mediated inflammation as a part of the neuroprotective mechanisms. No cases of persistent elevation were registered among the survivors 2 years after discharge.

Bromofatty aldehyde derived from bromine exposure and myeloperoxidase and eosinophil peroxidase modify GSH and protein.

α-Chlorofatty aldehydes (α-ClFALDs) and α-bromofatty aldehydes (α-BrFALDs) are produced in activated neutrophils and eosinophils. This study investigated the ability of α-BrFALD and α-ClFALD to react with the thiols of GSH and protein cysteinyl residues. Initial studies showed that 2-bromohexadecanal (2-BrHDA) and 2-chlorohexadecanal (2-ClHDA) react with GSH producing the same fatty aldehyde-GSH adduct (FALD-GSH). In both synthetic and cellular reactions, FALD-GSH production was more robust with 2-BrHDA compared with 2-ClHDA as precursor. NaBr-supplemented phorbol myristate acetate (PMA)-activated neutrophils formed more α-BrFALD and FALD-GSH compared with non-NaBr-supplemented neutrophils. Primary human eosinophils, which preferentially produce hypobromous acid and α-BrFALD, accumulated FALD-GSH following PMA stimulation. Mice exposed to Br2 gas had increased levels of both α-BrFALD and FALD-GSH in the lungs, as well as elevated systemic plasma levels of FALD-GSH in comparison to mice exposed to air. Similar relative reactivity of α-ClFALD and α-BrFALD with protein thiols was shown using click analogs of these aldehydes. Collectively, these data demonstrate that GSH and protein adduct formation are much greater as a result of nucleophilic attack of cysteinyl residues on α-BrFALD compared with α-ClFALD, which was observed in both primary leukocytes and in mice exposed to bromine gas.

Oxidative stress is increased in sarcopenia and associated with cardiovascular disease risk in sarcopenic obesity.

OBJECTIVES: To define whether circulating markers of oxidative stress correlate with sarcopenia in terms of glutathione balance and oxidative protein damage, and whether these biomarkers are associated with risk of cardiovascular disease (CVD). STUDY DESIGN: Population-based cross-sectional study. 115 out of 347 elderly subjects were classified as non-sarcopenic non-obese (NS-NO), sarcopenic non-obese (S-NO), non-sarcopenic obese (NS-O), and sarcopenic obese (S-O). MAIN OUTCOME MEASUREMENTS: Sarcopenia was defined as a relative skeletal muscle mass index (RASM) <7.25kg/m2 for men or <5.67kg/m2 for women, while obesity was diagnosed in those presenting with% fat >27 for men or >38 for women. The CVD risk was estimated by the carotid intima-media thickness (IMT) and the Framingham score. Blood reduced glutathione (GSH), oxidized glutathione (GSSG), plasma malondialdehyde-(MDA) and 4-hydroxy-2,3-nonenal-(HNE) protein adducts were analyzed. RESULTS: Significantly greater blood GSSG/GSH ratio and plasma MDA/HNE protein adducts were observed in sarcopenic than in non-sarcopenic patients. A logistic regression model showed a close relationship between serum HNE and MDA adducts and sarcopenia (OR=1.133, 95% CI 1.057-1.215, and OR=1.592, 95% CI 1.015-1.991, respectively). Linear and logistic regression analysis evidenced strong associations between the IMT or the Framingham CVD risk category and blood GSSG/GSH or serum HNE protein adducts in the S-O group. CONCLUSION: Circulating markers of oxidative stress are increased in sarcopenia and related to CVD risk in sarcopenic obesity, suggesting that redox balance analysis would be a useful part of a multidimensional evaluation in aging. Further research is encouraged to support interventional strategies to correct redox imbalance, which might contribute to the prevention or at least limitation of sarcopenia and its co-morbidities.

Autoantibodies against aldehyde-modified collagen type IV are associated with risk of development of myocardial infarction.

BACKGROUND: Oxidation of LDL particles entrapped in the extracellular matrix of the arterial wall is a key factor in the development of atherosclerosis. Lipid oxidation products, such as malondialdehyde (MDA), react with surrounding extracellular matrix proteins and cause modifications that are recognized by the immune system. MDA modification of collagen type IV is increased in carotid lesions from symptomatic patients and correlates with autoantibodies against MDA-modified collagen type IV in plasma. OBJECTIVE: The aim of this study was to determine whether autoantibodies against MDA-modified collagen type IV predict risk of development of myocardial infarction (MI). METHODS: Plasma levels of MDA-modified collagen type IV IgM and IgG antibodies were analysed by enzyme-linked immunosorbent assay in 385 subjects with incident MI during 13 years of follow-up and 410 age- and sex-matched controls in the Malmö Diet and Cancer study. RESULTS: MDA-modified collagen type IV IgG levels were higher in cases with incident MI than in controls. Subjects in the highest tertile of MDA-modified collagen type IV IgG had an increased risk of MI (hazard ratio 1.56, 95% confidence interval 1.22-2.00, P for trend 0.0004). This association remained significant after adjusting for factors included in the Framingham risk score and diabetes. High levels of MDA-collagen type IV IgG were associated with increased carotid intima-media thickness and elevated plasma levels of matrix metalloproteinase 10 and 12. CONCLUSIONS: Immune responses against MDA-modified collagen type IV are associated with more severe carotid disease and increased risk of MI. These immune responses may reflect LDL oxidation in the artery wall, but could also affect the atherosclerotic disease process.

Variant Aldehyde Dehydrogenase 2 (ALDH2*2) in East Asians Interactively Exacerbates Tobacco Smoking Risk for Coronary Spasm　- Possible Role of Reactive Aldehydes.

BACKGROUND: Coronary spastic angina (CSA) is common among East Asians and tobacco smoking (TS) is an established risk factor for CSA. Aldehyde dehydrogenase 2 (ALDH2) plays a key role in removing reactive toxic aldehydes and a deficient variant ALDH2 genotype (ALDH2*2) is prevalent among East Asians. We examined the interaction between TS andALDH2*2as a risk factor for CSA to better understand the disease pathogenesis.Methods and Results:The study subjects comprised 410 patients (258 men, 152 women; mean age, 66.3±11.5) in whom intracoronary injection of acetylcholine was performed on suspicion of CSA.ALDH2genotyping was performed by direct application of the Taqman polymerase chain reaction system. Of the study subjects, 244 had CSA proven and 166 were non-CSA. The frequencies of male sex,ALDH2*2, alcohol flushing syndrome, TS, coronary organic stenosis, and plasma levels of uric acid were higher (P<0.001, P<0.001, P<0.001, P<0.001, P<0.001, and P=0.015, respectively) and that of high-density lipoprotein cholesterol lower (P=0.002) in the CSA than non-CSA group. Multivariable logistic regression analysis revealed thatALDH2*2and TS were significant risk factors for CSA (P<0.001 and P=0.002, respectively).ALDH2*2exacerbated TS risk for CSA more than the multiplicative effects of each. CONCLUSIONS: ALDH2*2synergistically exacerbates TS risk for CSA, probably through aldehydes.

Oxidative stress leads to reduction of plasmalogen serving as a novel biomarker for systemic lupus erythematosus.

Oxidative stress is elevated in systemic lupus erythematosus (SLE) patients, and associated extensively with SLE pathogenesis. However, no common indicators of oxidative stress are yet in routine clinical use because of their instability, nonspecificity, and non-representation of all SLE symptoms. Moreover, the method for reproducible analysis of reactive oxygen species is still lacking. Lipids and their metabolites are essential components of biological systems, many of which serve as molecular targets of oxidative stress and play crucial roles in signaling, inflammation, and immune responses. Thus, determining the changed levels of lipids and their metabolites may serve the needs for SLE research. In the pilot study, shotgun lipidomics of sera from 30 SLE patients and 30 controls was performed and revealed a marked reduction of ethanolamine plasmalogen (pPE) species from 85.03±3.06 to 62.39±4.34 nmol/mL serum in controls and patients, respectively, accompanying significant increases in lysoPE (LPE) content (~46mol%) and 4-hydroxynonenal (an indictor of oxidative stress) in patients. Representative proinflammatory cytokines were also determined, revealing significant elevation of IL-6, IL-10, and TNF-α in SLE patients. Multivariate and multiple regression analyses showed for the first time that significant correlation among the SLE disease activity index, IL-10 levels, and pPE content exists, providing insights into SLE pathogenesis. The study also indicates that the changes of pPE (molecular targets of oxidative stress) and their peroxidation products may serve as novel biomarkers for diagnosis of SLE.

Lipid peroxidation biomarkers in adolescents with or at high-risk for bipolar disorder.

BACKGROUND: Prior work suggests that adult bipolar disorder (BD) is associated with increased oxidative stress and inflammation. This exploratory study examined markers of lipid and protein oxidation and inflammation in adolescents with and at varying risk for BD type I (BD-I). METHODS: Blood was obtained from four groups of adolescents (9-20 years of age): (1) healthy comparison subjects with no personal or family history of psychiatric disorders (n=13), (2) subjects with no psychiatric diagnosis and at least one parent with BD-I ('high-risk', n=15), (3) subjects with at least one parent with BD-I and a diagnosis of depressive disorder not-otherwise-specified ('ultra-high-risk', n=20), and (4) first-episode patients exhibiting mixed or manic symptoms that received a diagnosis of BD-I (n=16). Plasma levels of lipid peroxidation (LPH, 4-HNE, 8-ISO), protein carbonyl, and inflammation (IL-1α-ß, IL-6, IL-10, IFNγ, TNFα) were assessed using analysis of variance and covariance models. RESULTS: LPH was lower in adolescents with fully syndromal BD than controls, while LPH levels in the at-risk groups were between healthy controls and fully syndromal BD. Post-hoc analysis showed a non-significant increase in the (4-HNE+8-ISO)/LPH ratio suggesting a potential conversion of LPH into late-stage markers of lipid peroxidation. There were no significant differences among protein carbonyl content and inflammatory markers. CONCLUSIONS: In adolescents, fully syndromal BD is associated with significant reductions in LPH levels, and LPH levels decrease along the spectrum of risk for BD-I. Quantifying lipid peroxidation in longitudinal studies may help clarify the role of LPH in BD risk progression.

Different doxorubicin formulations affect plasma 4-hydroxy-2-nonenal and gene expression of aldehyde dehydrogenase 3A1 and thioredoxin reductase 2 in rat.

Increased oxidative stress is indisputably an important mechanism of doxorubicin side effects, especially its cardiotoxicity. To prevent impairment of non-tumorous tissue and to improve the specificity in targeting the tumor tissue, new drug nanotransporters are developed. In many cases preclinical therapeutic advantage has been shown when compared with the administration of conventional drug solution. Three forms of doxorubicin--conventional (DOX), encapsulated in liposomes (lipoDOX) and in apoferritin (apoDOX) were applied to Wistar rats. After 24 h exposition, the plasma level of 4-hydroxy-2-nonenal (4-HNE) as a marker of lipoperoxidation and tissue gene expression of thioredoxin reductase 2 (TXNRD2) and aldehyde dehydrogenase 3A1 (ALDH3A1) as an important part of antioxidative system were determined. Only conventional DOX significantly increases the level of 4-HNE; encapsulated forms on the other hand show significant decrease in plasma levels of 4-HNE in comparison with DOX. They also cause significant decrease in gene expression of ALDH3A1 and TXNRD2 in liver as a main detoxification organ, and a mild influence on the expression of these enzymes in left heart ventricle as a potential target of toxicity. Thus, 4-HNE seems to be a good potential biomarker of oxidative stress induced by various forms of doxorubicin.

4-Hydroxynonenal: A Superior Oxidative Biomarker Compared to Malondialdehyde and Carbonyl Content Induced by Carbon Tetrachloride in Rats.

Carbon tetrachloride (CCl4), a halogenated substance that generates free radical species during metabolism in vivo, induces hepatotoxicity, produces oxidative DNA damage, and increased levels of protein carbonyl, malondialdehyde (MDA), and 4-hydroxynonenal (4-HNE). In this study, Sprague-Dawley rats received single or repeated ip injections of carbon tetrachloride (CCl4), and formation and persistence of carbonyls, MDA, and 4-HNE in plasma were measured using gas chromatography-mass spectrometry. After a single injection of 500 mg/kg CCl4 the in vivo half-lives of MDA and carbonyl content were 1.5 d and 2 d, respectively, while that of 4-HNE was approximately 10 d. Treatment with CCl4 (50, 100, 500, or 1000 mg/kg) dose-dependently increased these oxidative biomarkers in blood. However, formation of protein carbonyls and MDA was less sensitive than 4-HNE to CCl4. Levels of serum glutamic oxaloacetic transaminase (SGOT) and glutamic pyruvic transaminase (SGPT) (hepatotoxicity markers) rose with CCl4 doses. After a single injection (500 mg/kg), the peak level of SGOT was observed after 8 h but SGPT after 24 h. Overall, 4-HNE was more dose-sensitive and showed greater formation subchronically than other biomarkers. Multiple ip treatments with 300 mg CCl4 /kg (d 1, 3, 6, 10, 14, and 21) demonstrated that 4-HNE formation was highest (18-fold, peak/control) and subchronic up to d 21 (last treatment day), unlike other biomarkers. Data suggest that 4-HNE, MDA, and carbonyl content may be useful oxidative biomarkers for exposure to free radical generating halogenated compounds. However, 4-HNE appears to be a more sensitive and sustainable biomarker for toxicological and risk assessments.

Superoxide induces protein oxidation in plasma and TNF-α elevation in macrophage culture: Insights into mechanisms of neurotoxicity following doxorubicin chemotherapy.

Chemotherapy-induced cognitive impairment (CICI) is a quality of life-altering consequence of chemotherapy experienced by a large percentage of cancer survivors. Approximately half of FDA-approved anti-cancer drugs are known to produce ROS. Doxorubicin (Dox), a prototypical ROS-generating chemotherapeutic agent, generates superoxide (O2(-)•) via redox cycling. Our group previously demonstrated that Dox, which does not cross the BBB, induced oxidative damage to plasma proteins leading to TNF-α elevation in the periphery and, subsequently, in brain following cancer chemotherapy. We hypothesize that such processes play a central role in CICI. The current study tested the notion that O2(-)• is involved and likely responsible for Dox-induced plasma protein oxidation and TNF-α release. Addition of O2(-)• as the potassium salt (KO2) to plasma resulted in significantly increased oxidative damage to proteins, indexed by protein carbonyl (PC) and protein-bound HNE levels. We then adapted this protocol for use in cell culture. Incubation of J774A.1 macrophage culture using this KO2-18crown6 protocol with 1 and 10 µM KO2 resulted in dramatically increased levels of TNF-α produced. These findings, together with our prior results, provide strong evidence that O2(-)• and its resulting reactive species are critically involved in Dox-induced plasma protein oxidation and TNF-α release.

Oxidative stress: Determination of 4-hydroxy-2-nonenal by gas chromatography/mass spectrometry in human and rat plasma.

The lipid peroxidation product 4-hydroxynonenal (HNE) is a biomarker of oxidative stress which is essentially involved in the pathophysiology of many diseases. The analysis of HNE is challenging because this aldehyde is extremely reactive and thus unstable. Hence, we adopted a gas chromatography-mass spectrometry (GC-MS) method based on derivatization of HNE with pentafluorobenzyl-hydroxylamine-HCl followed by trimethylsilylation to trimethylsilyl ethers. Ions representative for a negative ion chemical ionization mode were recorded at m/z = 152 for HNE and at m/z = 162 for the deuterated analogon (HNE-d11) as internal standard. This excellent stable and precise GC-MS method was carefully validated for HNE, and showed good linearity (r(2) = 0.998), and high specificity and sensitivity. Within-day precisions were 4.4-6.1% and between-day precisions were 5.2-10.2%. Accuracies were between 99% and 104% for the whole calibration range (2.5-250 nmol/L) of HNE. To examine the versatility of this modified GC-MS method, we analyzed HNE in ethylenediaminetetraacetic acid (EDTA) plasma in a well-defined collective of migraine patients; recently published. The results underline our former observations that women with migraine are afflicted with increased levels of HNE. Patients with thyroidal dysfunction showed no significant HNE alterations. This was confirmed by normal HNE EDTA plasma levels in hyper- und hypothyroid Sprague-Dawley rats. Taken together, the GC-MS method presented herein is of excellent quality to record oxidative stress-related bioactive HNE levels. This is important for a reorientation of oxidative stress analytics in other human diseases first of atherosclerosis and cancer.