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Background Myocardial iron overload in patients with thalassemia major (TM) is one of the most important complications. The purpose of the study was to identify advanced echocardiography parameters for early identification of myocardial dysfunction during follow-up of patients with TM. Methods Forty TM patients who were 41 ± 5 years old were included in the study and divided into two groups according to cardiac magnetic resonance T2* results (Group 1: Τ2* > 25 ms, Group 2: Τ2* ≤ 25 ms). Liver T2* parameters were also measured. Conventional and deformational echocardiographic parameters were measured at baseline and approximately 2 years later. Results Thirty-two patients had Τ2* = 34 ± 4 ms (Group 1), and 8 had Τ2* = 17 ± 9 ms (Group 2). Blood consumption was 185 ± 60 and 199 ± 37 ml/kg/yr (p = 0.64), and liver T2* was 4 ± 5 and 17 ± 21 ms (p = 0.01) in Groups 1 and 2, respectively. At baseline, Group 1 had better left ventricular global longitudinal strain (GLS) (− 22 ± 3 vs. − 18 ± 5, p = 0.01) and similar left ventricular ejection fraction (LVEF) (62 ± 5% vs. 58 ± 10%, p = 0.086) than Group 2. At the 28 ± 11-month follow-up, LVEF, GLS, and T2* values in Group 1 (63 ± 3%, − 21 ± 3%, 34 ± 4 ms) and Group 2 (56 ± 11%, − 17 ± 4%, 17 ± 9 ms) did not change significantly compared to their corresponding baseline values. In 8 patients from Group 1, a worsening (> 15%) in LS (p = 0.001) was detected during follow-up, with a marginal reduction in LVEF. Conclusions GLS seems to be an efficient echocardiographic parameter for detecting hemochromatosis-related cardiac dysfunction earlier than LVEF. It also seems to be affected by other factors (free radical oxygen, immunogenetic mechanisms or viral infections) in a minority of patients, underscoring the multifactorial etiology of cardiomyopathy.

As increasing application of nanoparticles, nanoparticles have been becoming a new emerging environmental pollution that attracts a lot of attention from the scientific community and also regulatory agents. In the past decade, studying the toxicity and environmental impacts of nanoparticles is becoming a hot research field and more and more researches have been published using both plant and animal system. In this chapter, using oxidized metal nanoparticles as an example, we introduce a detailed protocol for performing research on biochemical and physiological toxicity of nanoparticles in plant. We employ a hydroponics system to study phytotoxicity of nanoparticles, which makes it easier to study the impact of nanoparticles. In this chapter, we majorly focus on plant respiration and photosynthesis, root vigor as well as oxidative stress. Oxidative stress is one major physiological response to different environmental pollution, in which we present a detailed method for detecting free radical oxygen species as well as the major molecules and enzymes associating with oxidative stress, including SOD and POD. Although we introduce the methods using cotton as an example, the protocols presented in this chapter can be used almost any plant species to test the biochemical and physiological toxicity of an environmental pollution.

Rationale Oxidative stress is an imbalance between reactive free radical oxygen species and antioxidant defenses. Its consequences can lead to numerous pathologies. Regulating oxidative stress is the complex interplay between antioxidant recycling and thiol-containing regulatory proteins. Understanding these regulatory mechanisms is important for preventing onset of oxidative stress. The aim of this study was to investigae S-thiol protein chemistry associated with oxidized vitamin C (dehydroascorbate, DHA), homocysteine (HcySH) and glutathione (GSH) using mass spectrometry. Methods Glutaredoxin-1 (Grx-1) was incubated with DHA, with and without GSH and HcySH. Disulfide formation was followed by electrospray ionization mass spectrometry (ESI-MS) of intact proteins and by LC/ESI-MS/MS of peptides from protein tryptic digestions. The mechanism of DHA-mediated S-thiolation was investigated using two synthetic peptides: AcFHACAAK and AcFHACE. Three proteins, i.e. human hemoglobin (HHb), recombinant peroxiredoxin 2 (Prdx2) and Grx-1, were S-homocysteinylated followed by S-transthiolyation with GSH and investigated by ESI-MS and ESI-MS/MS. Results ESI-MS analysis reveals that DHA mediates disulfide formation and S-thiolation by HcySH as well as GSH of Grx-1. LC/ESI-MS/MS analysis allows identification of Grx-1 S-thiolated cysteine adducts. The mechanism by which DHA mediates S-thiolation of heptapeptide AcFHACAAK is shown to be via initial formation of a thiohemiketal adduct. In addition, ESI-MS of intact proteins shows that GSH can S-transthiolate S-homocysteinylated Grx-1_ HHb and Prdx2. The GS-S-protein adducts over time dominate the ESI-MS spectrum profile. Conclusions Mass spectrometry is a unique analytical technique for probing complex reaction mechanisms associated with oxidative stress. Using model proteins, ESI-MS reveals the mechanism of DHA-facilitated S-thiolation, which consists of thiohemiketal formation, disulfide formation or S-thiolation. Furthermore, protein S-thiolation by HcySH can be reversed by reversible GSH thiol exchange. The use of mass spectrometry with in vitro models of protein S-thiolation in oxidative stress may provide significant insight into possible mechanisms of action occurring in vivo.

EV Silina,1 VA Stupin,2,3 SB Bolevich,1 NE Manturova2,4 1Department of Human Pathology, I.M. Sechenov First Moscow State Medical University (Sechenov University), Moscow, Russia; 2Institute of Plastic Surgery and Cosmetology, Moscow, Russia; 3Department of Hospital Surgery №1, Pirogov Russian National Research Medical University (RNRMU), Moscow, Russia; 4Department of Plastic and Reconstructive Surgery, Cosmetology and Cell Technologies, Pirogov Russian National Research Medical University (RNRMU), Moscow, Russia Aim: The purpose of this study was to examine the role of free radical oxygen and peroxide–lipid processes along with conducting the study of blood flow level and oxygen saturation of facial tissues in patients of different ages with varying degrees of involutional changes in the skin of the face and neck.Materials and methods: One hundred and fifty-three people (84.3% women and 15.7% men) aged from 26 to 78 years with varying degrees of involutional changes in facial skin were examined. The clinical and laboratory evaluation was carried out dynamically and included various indicators of free radical processes, objective and subjective clinical visualization, and laser Doppler flowmetry (LDF) of the facial skin and transcutaneous oximetry (TcpO2) performed at 10 points on the face. To assess the state of free radical processes, the authors investigated the basal indicator of chemiluminescence intensity (ICb), the intensity of chemiluminescence stimulated (ICs) by zymosan, the activity coefficient (AC) of chemiluminescence, antiperoxide activity of plasma, and malondialdehyde (MDA).Results: With aging, the imbalance of the oxygen constituents of free radical processes grows with the increase in ROS. Proportional to age, the ICs increased 2.1 times on average in people older than 55 years compared to that in people younger than 30 years and ICb decreased by 1.8 times. As a result, the AC increased by 5.6 times. This correlates with involuntary skin changes and with regression of microcirculation and TcpO2. According to LDF, it was established that average total blood flow in people younger than 30 years and people older than 55 years was 8.1 and 6.4 mL/min, respectively The difference between the indicators of TcpO2 in people younger than 30 years and people older than 55 years was 1.6 times (average 56 vs 35 mm Hg). The stability of the indicators of the peroxide–lipid link of oxidative stress in different age groups demonstrated that the activation of ROS formation in mitochondria is not a cause but a consequence of microcirculation and metabolic processes in the face and neck and aging in general.Conclusion: The tissue metabolism and microcirculation parameters naturally regress with aging, which is associated with the increase of ROS. The excess of species leads to the intensification of peroxide processes. This, in turn, is reflected in the aesthetic appearance manifested by aging. Keywords: free radical processes, ROS aging, skin, involution processes, microcirculation&nbsp

Free radical oxygen generated by stress and oxygen deficiency causes problems such as heart disease and stroke. We propose mobile health monitoring system to solve this problem. Based on biofeedback technology used in psychiatry, meaningful parameters were identified by analyzing the autonomic nervous system and breathing patterns that change during deep breathing. As a result, PP-interval was increased (p < 0.05) during deep breathing and the average expiration value of the subject was unconsciously increased (p < 0.05). The deep breathing has been shown to be able to prevent stress and oxygen deficiency. After that, we developed an effective mobile health monitoring system using parameter that have significant result value. If personal health monitoring system that continuously induces deep breathing through the method of this study is developed, it can be used as a way to continuously manage and improve individual health.

Objectives In peripheral neuropathy, the underlying mechanisms of nerve and muscle degeneration include chronic inflammation and oxidative stress in fibrotic tissues. (‐)‐Epigallocatechin gallate (EGCG) is a major, active component in green tea and may scavenge free radical oxygen and attenuate inflammation. Conservative treatments such as steroid injection only deal with early, asymptomatic, peripheral neuropathy. In contrast, neurolysis and nerve conduit implantation work effectively for treating advanced stages. Materials and methods An EGCG‐loaded polycaprolactone (PCL) porous scaffold was fabricated using an integrated moulding method. We evaluated proliferative, oxidative and inflammatory activity of rat Schwann cells (RSCs) and rat skeletal muscle cells (RSMCs) cultured on different scaffolds in vitro. In a rat radiation injury model, we assessed the morphological, electrophysiological and functional performance of regenerated sciatic nerves and gastrocnemius muscles, as well as oxidative stress and inflammation state. Results RSCs and RSMCs exhibited higher proliferative, anti‐oxidant and anti‐inflammatory states in an EGCG/PCL scaffold. In vivo studies showed improved nerve and muscle recovery in the EGCG/PCL group, with increased nerve myelination and muscle fibre proliferation and reduced macrophage infiltration, lipid peroxidation, inflammation and oxidative stress indicators. Conclusions The EGCG‐modified PCL porous nerve scaffold alleviates cellular oxidative stress and repairs peripheral nerve and muscle structure in rats. It attenuates oxidative stress and inflammation in vivo and may provide further insights into peripheral nerve repair in the future.

Free radical oxygen species cause protein carbonylation, an irreversible oxidative damage associated with diseases such as liver injury, diabetes mellitus, arthritis, cancer and Alzheimer. To determine the inhibitory activity of the protein carbonylation and the hepatoprotective effect of ethanol extract of the fruits of Caesalpinia coriaria Jacq (Fabaceae). Scavenging activity of 2,2-diphenyl-1- picrylhydrazyl radical (DPPH•) and the radical cation 2,2’-azino-bis(3-ethylbenzothiazoline-sulfonic acid) (ABTS•+) of the extract of C. coriaria at 500 mg / L, was evaluated in a preliminary test. Inhibitory activity of the protein carbonylation isolated from mouse liver using Western methodology with dinitrophenylhydrazine (DNPH) probes was evaluated. The hepatoprotective activity of the extract was evaluated in vivo in Wistar rats using carbon tetrachloride (CCl4) as toxic orally. The degree of hepatoprotection was determined by measuring the serum liver enzymes, aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and by histopathological analysis of rat liver. The rate of capture of the radicals DPPH• and ABTS•+ by the extract of C. coriaria was higher than 90 % in both cases. The extract also showed a better inhibitory effect on the carbonylation of the proteins than butyl hydroxytoluene (BHT) used as positive control. The extract of C. coriaria produced a significant reduction of AST concentration (from 1667,67 ± 394,27 to 718,00 ± 24,85 U/L ) and ALT (from 967,67 ± 118,30 to 625.67 ± 60,98 U/L), indicating liver cell damage attenuation. The extract of C. coriaria produced an important antioxidant activity and moderate hepatoprotective effect against CCl4-induced toxicity.

Oxidative stress plays a role in many biological phenomena, but involved mechanisms and individual reactions are not well understood. Correlated electronic structure calculations with the MP2, MP4, and CCSD(T) methods detail thermodynamic and kinetic information for the free radical oxygen protein oxidation pathway studied in a trialanine model system. The pathway includes aerobic, anaerobic and termination reactions. The course of the oxidation process depends on local conditions and availability of specific reactive oxygen species (ROS). A chemical mechanism is proposed for how oxidative stress promotes β-structure formation in the amyloid diseases. The work can be used to aid experimentalists as they explore individual reactions and mechanisms involving oxygen free radicals and oxidative stress in β-structured proteins.

Glucose is the main physiological stimulus of pancreatic β-cells. However, chronic exposure of β-cells to elevated glucose concentrations induces glucotoxicity. In animal models of type 2 diabetes, it has been shown that several days of hyperglycemia impairs glucose-stimulated insulin secretion and increases β-cell apoptosis. In patients with type 2 diabetes, the multiple disorders caused by chronic hyperglycemia in β-cells include elevated basal insulin secretion, increased sensitivity to glucose, diminished response to insulinotropic stimuli and substantial depletion of insulin hoarding [1,2]. These defects associated with insulin resistance lead to a progressive loss of β-cell mass and function and to the onset of diabetes. It is crucial to study the mechanisms by which glucotoxicity induces β-cell failure to develop therapeutic strategies for protecting and recovering a functional β-cell mass. Several mechanisms might explain the glucotoxicity due to prolonged hyperglycemia, such as β-cell exhaustion, oxidative stress induced by free radical oxygen species, endoplasmic reticulum (ER) stress, inflammation caused by proinflammatory cytokines and chemokines, loss of neogenesis, proliferation of β-cells, and so on [3,4,5,6,7,8,9,10]. However, the precise mechanisms of glucotoxicity and its contribution to the pathology of type 2 diabetes mellitus (T2DM) are still not fully understood.

Background: Maternal type 1 diabetes is associated with an increased risk for fetal malformations. The mechanism by which diabetes caused teratogenic disorders is not fully known. Previous studies have demonstrated that many teratogenic diabetic cases were related to free radical oxygen species. This study was conducted to evaluate the effect of maternal diabetes on both embryo and placenta by estimating theoxidative and DNA damage in embryo and placenta of diabetic mellitus- induced rats. The possible role of olive leaves extract of Olea europaea ( O. europaea ) plant in repairing the damage was also assessed. Material and Methods:. Diabetes mellitus was induced by streptozotocin (STZ) by a single intraperitoneal injection (35 mg/kg b wt). O.europaea leaves water extract was administered orally (550 mg/ 100g b wt/ day) for 5days before pregnancy and18 days after. Malondealdehyde (MDA) level, glutathione peroxidase (Gpx) and superoxide dismutase (SOD) activities and glycogen concentration were measured in term embryo and placenta homogenates of diabetic and control rats. Moreover, the evaluation of DNA damage was carried out by the Alkaline Comet Assay usingembryos and placentas taken from STZ-induced diabetic and control pregnant rats. Results: the results showed an elevation in MDA level of the diabetic groups of both embryo and placenta compared to that of the control. This was accompanied by reduction in Gpx and SOD activities indicating oxidative damage. Glycogen level was reduced in diabetic groups of embryo and placenta. Both oxidative and hyperglycemic status were improved in the groups treated with olive leaves water extract. The percentage of tail DNA and tail moment values were also higher in both embryo and placenta of the diabetic -induced rats. DNA damage seems to be partly ameliorated in groups treated with O.europaea leaveswater extract. Conclusion: This study indicated that maternal hyperglycemic condition in diabetic- induced pregnant rats could generate oxidative and DNA damage to embryo and placenta that could be ameliorated by oral doses of olive leaves water extract.

Reactive oxygen species (ROS) play multiple roles in interactions between plants and microbes, both as host defense mechanisms and as mediators of pathogenic and symbiotic associations. One source of ROS in these interactions are photoactivated, ROS-generating perylenequinone pigments produced via polyketide metabolic pathways in plant-associated fungi. These natural products, including cercosporin, elsinochromes, hypocrellins, and calphostin C, are being utilized as medicinal agents, enzyme inhibitors, and in tumor therapy, but in nature, they play a role in the establishment of pathogenic associations between fungi and their plant hosts.Photoactivated perylenequinones are photosensitizers that use light energy to form singlet oxygen (¹O₂) and free radical oxygen species which damage cellular components based on localization of the perylenequinone molecule. Production of perylenequinones during infection commonly results in lipid peroxidation and membrane damage, leading to leakage of nutrients from cells into the intercellular spaces colonized by the pathogen. Perylenequinones show almost universal toxicity against organisms, including plants, mice, bacteria, and most fungi. The producing fungi are resistant, however, and serve as models for understanding resistance mechanisms.Studies of resistance mechanisms by perylenequinone-producing fungi such as Cercospora species are leading to an understanding of cellular resistance to ¹O₂ and oxidative stress. Recent studies show commonalities between resistance mechanisms in these fungi with extensive studies of ¹O₂ and oxidative stress responses in photosynthetic organisms.Such studies hold promise both for improved medical use and for engineering crop plants for disease resistance.

Ischaemia-induced tissue injury has wide-ranging clinical implications including myocardial infarction, stroke, compartment syndrome, ischaemic renal failure and replantation and revascularization. However, the restoration of blood flow produces a 'second hit' phenomenon, the effect of which is greater than the initial ischaemic event and characterizes ischaemia-reperfusion (IR) injury. Some examples of potential settings of IR injury include: following thrombolytic therapy for stroke, invasive cardiovascular procedures, solid organ transplantation, and major trauma resuscitation. Pathophysiological events of IR injury are the result of reactive oxygen species (ROS) production, microvascular vasoconstriction, and ultimately endothelial cell-neutrophil adhesion with subsequent neutrophil infiltration of the affected tissue. Initially thought to increase the amount of free radical oxygen in the system, hyperbaric oxygen (HBO) has demonstrated a protective effect on tissues by influencing the same mechanisms responsible for IR injury. Consequently, HBO has tremendous therapeutic value. We review the biochemical mechanisms of ischaemia-reperfusion injury and the effects of HBO following ischaemia-reperfusion.

Background: The class Tremellomycete (Agaricomycotina) encompasses more than 380 fungi. Although there are a few edible Tremella spp., the only species with current biotechnological use is the astaxanthin-producing yeast Phaffia rhodozyma (Cystofilobasidiales). Besides astaxanthin, a carotenoid pigment with potent antioxidant activity and great value for aquaculture and pharmaceutical industries, P. rhodozyma possesses multiple exceptional traits of fundamental and applied interest. The aim of this study was to obtain, and analyze two new genome sequences of representative strains from the northern (CBS 7918T, the type strain) and southern hemispheres (CRUB 1149) and compre them to a previously published genome sequence (strain CBS 6938). Photoprotection and antioxidant related genes, as well as genes involved in sexual reproduction were analyzed. Results: Both genomes had ca. 19Mb and 6000 protein coding genes, similar to CBS 6938. Compared to other fungal genomes P. rhodozyma strains and other Cystofilobasidiales have the highest number of intron-containing genes and highest number of introns per gene. The Patagonian strain showed 4.4% of nucleotide sequence divergence compared to the European strains which differed from each other by only 0.073%. All known genes related to the synthesis of astaxanthin were annotated. A hitherto unknown gene cluster potentially responsible for photoprotection (mycosporines) was found in the newly sequenced P. rhodozyma strains but was absent in the non-mycosporinogenic strain CBS 6938. A broad battery of enzymes that act as scavengers of free radical oxygen species were detected, including catalases and superoxide dismutases (SODs). Additionally, genes involved in sexual reproduction were found and annotated. Conclusions: A draft genome sequence of the type strain of P. rhodozyma is now available, and comparison with that of the Patagonian population suggests the latter deserves to be assigned to a distinct variety. An unexpected genetic trait regarding high occurrence of introns in P. rhodozyma and other Cystofilobasidiales was revealed. New genomic insights into fungal homothallism were also provided. The genetic basis of several additional photoprotective and antioxidant strategies were described, indicating that P. rhodozyma is one of the fungi most well-equipped to cope with environmental oxidative stress, a factor that has probably contributed to shaping its genome. Fil: Bellora, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto Andino Patagónico de Tecnologías Biológicas y Geoambientales. Universidad Nacional del Comahue. Instituto Andino Patagónico de Tecnologías Biológicas y Geoambientales.; Argentina Fil: Moline, Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto Andino Patagónico de Tecnologías Biológicas y Geoambientales. Universidad Nacional del Comahue. Instituto Andino Patagónico de Tecnologías Biológicas y Geoambientales.; Argentina Fil: David Palma, Márcia. Universidade Nova de Lisboa; Portugal Fil: Coelho, Marco A.. Universidade Nova de Lisboa; Portugal Fil: Hittinger, Chris Todd. University of Wisconsin; Estados Unidos Fil: Sampaio, José P.. Universidade Nova de Lisboa; Portugal Fil: Gonçalves, Paula. Universidade Nova de Lisboa; Portugal Fil: Libkind Frati, Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto Andino Patagónico de Tecnologías Biológicas y Geoambientales. Universidad Nacional del Comahue. Instituto Andino Patagónico de Tecnologías Biológicas y Geoambientales.; Argentina

Tumour hypoxia leads to radio and chemotherapy resistance among cancer patients. The aim of this paper is to formulate a hypothesis on the heterogeneity of hypoxia in solid tumours. Tumour vasculature is known to be significantly variable. The great structural and functional abnormalities of tumour microcirculation cause spatial and temporal heterogeneity in its perfusion. Tumours have constantly been under the influence of pulsatile blood perfusion with variable pressure that initiates inhomogeneous erythrocyte deformation and following impact on oxygen disorder release from red blood cells into plasma within the blood vessel. Furthermore, stochastically released oxygen in tumour vessel, plasma and interstitial fluid may lead to heterogeneity of hypoxia. Under the influence of increased heterogeneity of hemodynamic force, the oxygen molecules dissolved in blood plasma are inclined to form nanobubbles (NBs) in tumour vessels. Considering the fact that tumour interstitial fluid pressure is increased compared to normal tissues, we assume that oxygen NBs may burst under the impact of shear stress. During the course of mechanochemical reaction, when a nanobubble (NB) bursts, both reactive oxygen species and ions form in various charged states. In consequence of a chain reaction, free radical oxygen molecules bind to proteins and lipids, thus reducing oxygen molecules in a chaotic manner within the tumour. The proposed hypothesis should be used as a methodical approach based on the simultaneous ultrasound imaging diagnostic techniques and therapy, regarding the mechanochemical effect on NB conglomerates with drugs in the tumour.

Free radical oxygen species act on biological constituents like lipid, cell membrane and deoxyribonucleic acids by changing their biological functions. These effects are associated with the appearance of anomalies like allergies, cancer, viral infections, inflammations, etc. Antioxidants like flavonoids are abundant in plants and are good scavengers of reactive oxygen species (ROS). Their antioxidant activities are measured through hydrogen atom transfer or charge transfer. These mechanisms depend on the structure of the flavonoid or the nature of the free radical. In this paper, we study the interaction of one of the most reactive flavonoid quercetin with two ROS: hydroxyl radical ( · OH ) and superoxide radical ( · O 2 - ) using B3LYP/6-31++G*level of theory. Correlations between structural changes and antioxidant activity are shown. The potential energy surfaces (PES) are calculated in both cases and indicate a transition state (TS) only for the reaction of quercetin with the hydroxyl radical. This transition state is only localized at the B3LYP/3-21G level of theory. This comparison between the reactants, the TS and the products of the reaction with the · OH radical indicate the nature of the transfer. As far as the reaction of quercetin with the superoxide anion radical is concerned, the PES shows the inexistence of one transition state. The nature of the charge transfer is thus determined using the natural bond orbital (NBO) and the singlet occupied molecular orbital (SOMO) analysis.

The increasing use of micro- and nanostructured silicon-based devices for in vivo therapeutic or sensing applications highlights the importance of understanding the immunogenicity of these surfaces. Four silicon surfaces (nanoporous, microstructured, nanochanneled, and flat) were studied for their ability to provoke an immune response in human blood derived monocytes. The monocytes were incubated with the surfaces for 48 h and the immunogenicity was evaluated based on the viability, shape factors, and cytokine expression. Free radical oxygen formation was measured at 18 h to elicit a possible mechanism invoking immunogenicity. Although no cytokines were significantly different comparing the response of monocytes on the tissue culture polystyrene surfaces to those on the micropeaked surfaces, on average all cytokines were elevated on the micropeaked surface. The monocytes on the nanoporous surface also displayed an elevated cytokine response, overall, but not to the degree of those on the micropeaked surface. The nanochanneled surface response was similar to that of flat silicon. Overall, the immunogenicity and biocompatibility of flat, nanochanneled, and nanoporous silicon toward human monocytes are approximately equivalent to tissue culture polystyrene.

In vivo effects of N-benzyloxycarbonyl (Cbz)-Leu–Leu-leucinal (MG132) on chymotryptic-like (ChT-L), tryptic-like, and post-glutamyl peptide hydrolytic-like proteasome activities, protein oxidation, lipid peroxidation (LP), glutathione (GSH) level, as well as on the activity of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), and glutathione-reductase) in the rat liver were studied. The possibility of MG132 provoking the formation of free oxygen radicals was also assayed in primary hepatocytes. The following results were obtained: (1) In vivo, MG132 did not change the spontaneous LP, but increased Fe-induced LP and the amount of oxidized proteins; it decreased the GSH level in liver. From the proteasome activities studied in liver cytosol only ChT-L activity was significantly decreased after MG132 administration. Furthermore, MG132 increased antioxidant enzyme activities of SOD, CAT, and GSH-Px. (2) In vitro, MG132 increased free radical oxygen species in hepatocytes; this effect disappeared in the presence of CAT or mannitol. In conclusion, since nowadays proteasome inhibitors are entering into the swing of laboratory and clinical practice, the present data could provide useful information for MG132 action. Consequently, future in vivo experiments with MG132 could highlight the possibility of its use at different pathological conditions. Copyright © 2008 John Wiley & Sons, Ltd.

The term ‘uncoupling protein’ was originally used for the mitochondrial membrane protein UCP1, which is uniquely present in mitochondria of brown adipocytes, thermogenic cells that regulate body temperature in small rodents, hibernators and mammalian newborns. In these cells, UCP1 acts as a proton carrier activated by free fatty acids and creates a shunt between complexes of the respiratory chain and ATP-synthase resulting in a futile proton cycling and dissipation of oxidation energy as heat. Recent identification of new homologues to UCP1 expressed in brown and white adipose tissue, muscle, brain and other tissues together with the hypothesis that these novel uncoupling proteins (UCPs) may regulate thermogenesis and/or fatty acid metabolism and furthermore may protect against free radical oxygen species production have generated considerable optimism for rapid advances in the identification of new targets for pharmacological management of complex pathological syndromes such as obesity, type 2 diabetes or chronic inflammatory diseases. However, since the physiological and biochemical roles of the novel UCPs are not yet clear, the main challenge today consists first of all in providing mechanistic explanation for their functions in cellular physiology. This lively awaited information may be the basis for potential pharmacological targeting of the UCPs in future.

We studied the cryopreservation of the most common coccolithophore, Emiliania huxleyi which is considered as one of the main global carbon cycle participants. Both stages of this complex life cycle species were submitted to gradual addition of three distinct cryoprotectants: dimethylsulfoxide (7.5% v/v), methanol (5% v/v) and proline (0.5 M). They were then control-rate cooled (−5 °C min−1) to −50 °C before plunging into liquid nitrogen. Free radical oxygen species have been proposed to occur in cells subjected to pre-freezing manipulation or to cooling. Therefore, catalase (preventing accumulation of hydroxyl radicals) was evaluated for its ability to improve cell viability before and after freezing-thawing challenge.

Non syndromic forms of Retinitis Pigmentosa (RP) constitute a collection of clinically and genetically heterogeneous inherited retinal degenerative diseases. They are characterized by a bilateral progressive visual loss susceptible to cause blindness. These diseases are transmitted through pedigrees according to all known modes of inheritance. They are bilateral and usually start during infancy. However, very early clinical presentations exist, such as those observed in children affected by Leber Congenital Amaurosis, as well as late onset autosomal dominant forms of retinitis pigmentosa. The characteristic clinical aspect of the rod-cone RP dystrophies is marked by alterations of the peripheral retina associated with a night blindness and a progressive narrowing of the visual field. The ophthalmoscopic examination of RP patients commonly reveals thin retinal arteries and scattered pigmentary accumulations. In contrast, there are cone rod retinal dystrophies whose onset is marked by a decreased visual acuity before the appearance of any visual field alteration. Some forms of RPs display an ocular fundus devoid of any pigmentary alteration. Syndromic forms of RPs are not uncommon. The association of deafness with RP is detected in nearly 30% of the patients. Other associations with RP can include mental deficiency, facial dysmorphy, microcephaly, obesity, kidney deficiency, immune deficiencies, metabolic disorders. The existence of such syndromic forms of RP localizes RPs at the crossroad of several medical specialties. A long lasting collaboration between our department of ophthalmology and the department of medical genetics of the Necker-Sick Children Hospital has allowed us to establish numerous genotype-phenotype correlations, especially in LCA and Stargardt's disease. ABCR gene mutations cause Stargardt disease. ABCR mutations may also cause some types of Ages Related Macular Degenerations (AMD). Nowadays, there is no known efficient therapy available for patients affected by RP. Gene therapies hold promises of treatment for patients affected by some of these diseases for the next decade. In a not too far future, the use of pharmacological drugs increasing a better intracellular oxygen availability, without triggering any harmful production of free radical oxygen species (ROS), while exerting an anti-apoptotic effect within photoreceptor cells, appears to be a therapeutical strategy deserving to be tested in an appropriately designed clinical trial. For the present time, optical and electronical devices as well as night-vision glasses are the only possible tools allowing to improve the quality of life of some patients.

Investigations of variations in metabolic efficiency and thermogenesis have a short and turbulent history. In small animals, non-shivering thermogenesis and diet-induced thermogenesis have a great impact on overall body weight, and the question is whether mechanisms to waste energy have evolved also in human energy metabolism. The candidate molecules for this adaptive thermogenesis are the uncoupling proteins. This is a newly discovered family of proteins, consisting of at least five proteins, namely UCP1, UCP2, UCP3, UCP4 and UCP5. Although a role for UCP1 in thermogenesis is unequivocal, the physiological function of the newer uncoupling proteins is as yet unclear. UCP1 is present in brown adipose tissue and has a well-documented role in cold-induced thermogenesis. The targeted disruption of the UCP1-gene rendered animals that were cold sensitive, but not obese. UCP2 mRNA has a ubiquitous distribution in tissue, namely, in skeletal muscle, white and brown adipose tissue, the gastro-intestinal tract, the lung and the spleen. By targeting the UCP2-gene there was no effect on whole body energy metabolism, but instead, a reduced ability to protect against free-radical oxygen species. UCP2 has also been shown to act as a negative regulator for insulin secretion. UCP3 is present in skeletal muscle. Targeted disruption of the UCP3-gene gave no effect on whole body energy metabolism, but showed the mitochondria in muscle to be more coupled. In conclusion, the uncoupling proteins may be important in various specific ways, as protectors of free radical oxygen species and as regulators of ATP-dependent processes.

Oxygen is an efficient terminal electron acceptor in respiratory pathways. During aerobic respiration the electron transport chain generates free radical oxygen species as a result of electron leakage; this generation of toxic species is proportional to the oxygen tension (51). In addition, toxic oxygen species (TOS) may be formed exogenously, for example, by chemical processes or through radiation. TOS also result from the oxidative burst of polymorphonuclear leukocytes (PMN). Infection with Helicobacter pylori induces an inflammatory response (gastritis), which leads to an increase in the level of TOS in the gastric mucosa and the gastric juice (4, 24–26, 59). This increase in the level of toxic metabolites is probably the result of the generation of the superoxide anion (O2·− ), a reactive TOS, formed as part of the oxidative burst of PMN and enzymic activities of gastric epithelial cells. There is evidence that H. pylori infection leads to increased production of O2·− via NADPH oxidase in gastric cells, stimulated by lipopolysaccharide as well as xanthine oxidase, another mechanism for the generation of oxygen-derived free radicals (8, 80). In response to increased superoxide anion production in gastric tissue, changes have been detected in the level of expression of human superoxide dismutase (SOD) (12). Human gastric SOD exists as a cytoplasmic copper-zinc-superoxide dismutase (Cu, Zn-SOD) found in gland cells of the gastric body and antral mucosa, and as a manganese-superoxide dismutase (Mn-SOD) within mitochondria (63). An increase in the amount and activity of Mn-SOD has been observed in response to H. pylori infection and gastritis, whereas the amount and activity of the Cu, Zn-SOD remained constant or decreased slightly (39). It has been suggested that the induction of Mn-SOD is in response to increased cytokine production within the inflamed gastric mucosa (39). This situation is reversed following successful treatment of the infection (38). The data suggest that within the gastric environment H. pylori may be exposed to increased levels of TOS. In such an environment it is important for bacterial survival that the impact of such TOS be neutralized.

The age-related neurodegenerative diseases exemplified by Alzheimerhyp;s disease (AD), Lewy body diseases such as Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), and Huntingtonhyp;s disease are characterized by the deposition of abnormal forms of specific proteins in the brain. Although several factors appear to underlie the pathological depositions, the cause of neuronal death in each disease appears to be multifactorial. In this regard, evidence in each case for a role of oxidative stress is provided by the finding that the pathological deposits are immunoreactive to antibodies recognizing protein side-chains modified either directly by reactive oxygen or nitrogen species, or by products of lipid peroxidation or glycoxidation. Although the source(s) of increased oxidative damage are not entirely clear, the findings of increased localization of redox-active transition metals in the brain regions most affected is consistent with their contribution to oxidative stress. It is tempting to speculate that free radical oxygen chemistry plays a pathogenetic role in all these neurodegenerative conditions, though it is as yet undetermined what types of oxidative damage occur early in pathogenesis, and what types are secondary manifestations of dying neurons. Delineation of the profile of oxidative damage in each disease will provide clues to how the specific neuronal populations are differentially affected by the individual disease conditions.

Background: The risk of gastric carcinoma is increased in persons with a positive family history of the disease. Previous findings suggest that free radicals are involved in both the initiation and promotion of carcinogenesis. Objective: The aim of this study was to assess whether alterations in the antioxidant defense system of gastric mucosal cells in patients with gastric carcinoma are related to a positive family history. Methods: Glutathione peroxidase activity, oxidized glutathione, total glutathione, and malondialdehyde levels were assayed in patients with gastric carcinoma with and without a family history of the disease. A control group consisted of healthy volunteers. Results: The study enrolled 65 patients with gastric carcinoma: 35 with and 30 without a family history of the disease, as well as 30 control subjects. The level of glutathione peroxidase activity in tumor-free tissue samples from patients with a family history of gastric carcinoma was higher than in samples from those without such family history. Glutathione peroxidase activity was also significantly higher in neoplastic than in tumor-free tissue. There was no significant difference in glutathione peroxidase activity between neoplastic tissue samples taken from patients with and without a positive family history. Conclusions: The results appear to support the existence of an enzymatic imbalance caused by cellular initiation in patients with a family history of gastric carcinoma. This imbalance may lead to an altered response to free radical oxygen species and a consequent progressive increase in biologic damage.

Free radical injury has been implicated in the pathogenesis of a wide variety of conditions that may affect newborn infants. This is not surprising since newborns (especially premature infants) are uniquely susceptible to free radical injury. At birth, newborns are exposed to significantly higher oxygen concentrations compared to the intrauterine environment and may develop medical conditions that require treatment with prolonged hyperoxia. In addition, newborns may experience reperfusion injury following periods of hypoxia or ischaemia and may have inadequate antioxidant defences to handle these oxidant challenges. As interest in free radical injury in the newborn has grown, so too has research examining possible methods of prevention and treatment. Antioxidant therapy has great potential to further reduce morbidity and mortality in the newborn period by influencing multiple organ systems and disease states. We will now review current strategies for the prevention and treatment of free radical injury in the newborn.

The skin is one of the largest organs in the body. Skin aging in older adults is manifested by increased numbers of pigmented spots, wrinkles and features of sagging. Because the skin derives much of its oxygen from the atmosphere it also derives free radical oxygen species in the same way and this increases oxidative stress. Other environmental exposures, especially to cigarette smoking and UV light, also predispose to oxidative stress by both increasing free radical oxygen species and reducing the activity of antioxidant enzymes. In addition to these changes, the skin becomes more predisposed to neoplastic changes with age; again, increased oxidative stress may contribute to this. There is some evidence that UV light also impairs the immune function of the skin through altering antioxidant enzyme levels that impact on the number of Langerhans cells in the epidermis.

The development of head and neck cancer may depend not only on exposure to environmental carcinogens but also on a genetically based susceptibility to carcinogen-induced damage. This thesis presents a case-control study that demonstrates the significance of mutagen sensitivity, a measure of an individual's intrinsic DNA repair capacity against free radical damage, as a risk factor for the disease. As part of the case-control analysis, 167 previously untreated patients and 177 age- and sex-matched healthy controls were assessed for various lifestyle factors including tobacco and alcohol habits, occupational exposures, and diet. Mutagen sensitivity expressed by each individual was determined by quantifying bleomycin-induced chromosomal breaks within peripheral blood lymphocytes in vitro. Consistent with our initial observations and those of others, mutagen hypersensitivity was strongly associated with increased risk of head and neck cancer (odds ratio, 4.95; 95% confidence interval, 2.67 to 9.17) after adjusting for age, sex, and race. Low intake of vitamins C and E was also associated with an increased risk of disease and was interactive with mutagen sensitivity in risk estimates. Individuals with both a low intake of various antioxidants and increased chromosomal sensitivity to oxidant-induced DNA damage were at greatest risk. This study supports the concept that the risk of head and neck cancer is determined by a balance of factors that either enhance or protect against free radical oxygen damage, including innate capacities for DNA repair.

It is widely accepted that protein oxidation is involved in a variety of diseases, including neurodegenerative diseases. Especially during aging, a reduction in anti-oxidant defence mechanisms leads to an increased formation of free radical oxygen species and consequently results in a damage of proteins, including mitochondrial and synaptic ones. Even those proteins involved in repair and protein clearance via the ubiquitin proteasome and lysosomal system are subject to damage and show a reduced function. Here, we will discuss a variety of mechanisms and provide examples where cognition is affected and where repair mechanisms are no longer sufficient to compensate for a dysfunction of damaged proteins or even may become toxic. Next to physiological deficits, an accumulation of deficient proteins in aggresomes may occur and result in a formation of pathological hallmark structures typical for aging and disease. A major challenge is how to prevent aberrant oxidation, given that oxidation plays an essential role in aging and neurodegenerative diseases. Particularly interesting are the possibilities to reduce the formation of radical oxygen species leading to a dysfunction of protein repair and protein clearance, or to a formation of toxic byproducts accelerating neurodegeneration.

Several studies have shown that selenium can inhibit tumorigenesis in tissues. However, little is known about the mechanism and the effect of selenium on DNA, especially in brain tumor cells. In this study we examined the biological effect of selenium on human glioma cell lines (A172 and T98G). Selenium exhibited an antiproliferative effect on these cell lines (and induced the typical ladder pattern of DNA fragmentation commonly found in apoptosis), which were prevented by catalase. Few effects of selenium on NT14 fibroblasts were found. These findings demonstrate that selenium may induce, by apoptosis, cell death of human glioma cell lines, which are resulting from free radical oxygen forming.

Objective— The role of endothelial G protein–coupled receptor kinase 2 (GRK2) was investigated in mice with selective deletion of the kinase in the endothelium (Tie2-CRE/GRK2 fl/fl ). Approach and Results— Aortas from Tie2-CRE/GRK2 fl/fl presented functional and structural alterations as compared with control GRK2 fl/fl mice. In particular, vasoconstriction was blunted to different agonists, and collagen and elastic rearrangement and macrophage infiltration were observed. In primary cultured endothelial cells deficient for GRK2, mitochondrial reactive oxygen species was increased, leading to expression of cytokines. Chronic treatment with a reactive oxygen species scavenger in mice corrected the vascular phenotype by recovering vasoconstriction, structural abnormalities, and reducing macrophage infiltration. Conclusions— These results demonstrate that GRK2 removal compromises vascular phenotype and integrity by increasing endothelial reactive oxygen species production.

While cataract surgery is generally recognized as being one of the safest operations, there is still a significant complication rate. From 30 to 50% of all patients in the United States having cataract extraction develop opacification of the posterior lens capsule within two years and require laser treatment with its own significant risk of complications. Of the patients having cataract surgery, 0.8% develop retinal detachments, from 0.6% to 1.3% were rehospitalized for corneal edema or required corneal transplantation and about 0.1% presented with endophthalmitis . Thus, aside from secondary cataract, about 2% of 1.3 million people, or 26,000 individuals in the United States annually develop serious complications as a result of cataract surgery. The aim of this investigation was to increase the safety and effectiveness of an individual intraocular lens (IOL) preventing an impairment in peroxide metabolism of the mature human cataractous lenses compared to normal lenses employing the specific nanotechnology coating which substitutes the inhibitory effect of the implantable device towards the active species of oxygen and the ability of IOL to regulate the H2O2 and lipid hydroperoxides levels in the surrounding medium. The implantation of IOLs with metabolic activity improves the capability of the surrounding ocular tissues to withstand oxidative stress induced in ocular humors by the photochemical and other metabolic reactions. The coated implantable medical device with thin film of platinum applied with magnetron sputtering, reacts as a body enzyme with deleterious peroxide compounds and free radical oxygen species in body fluids and tissue when said device is implanted into human body. The IOL having haptics coated with thin film of platinum, catalyzes the reduction of peroxide compounds to decrease their levels within the aqueous humor. Further, the coatings also scavenge toxic free radicals of oxygen, thus preventing cellular dysfunction resulting from oxidative attack. Coated IOLs according to the patented nanotechnology can address the vast majority of cataract surgery-induced complications, such as secondary cataract, intraocular inflammation (endophthalmitis) and foreign body reactions, cystoid macular oedema, corneal edema. The nanotechnology offers physicians and surgeons to develop and commercialize costeffective therapeutic medical implantable devices, products and support systems with metabolic activities for the treatment of ophthalmic diseases and of a wide range of pathological states and disorders which are treated by insertion of the implantable and prosthetic (polymeric) devices.

In premature babies birth an high oxygen level exposure can occur and newborn hyperoxia exposure can be associated with free radical oxygen release with impairment of myocardial function, while in adult animal models short exposure to hyperoxia seems to protect heart against ischemic injury. Thus, the mechanisms and consequences which take place after hyperoxia exposure are different and related to animals age. The aim of our work has been to analyze the role played by HIF-1α in the occurrence of the morphological modifications upon hyperoxia exposure in neonatal rat heart. Hyperoxia exposure induces connective compartment increase which seems to allow enhanced blood vessels growth. An increased hypoxia inducible factor-1α (HIF-1α) translocation and vascular endothelial growth factor (VEGF) expression has been found upon 95% oxygen exposure to induce morphological modifications. Upstream pPKC-α expression increase in newborn rats exposed to 95% oxygen can suggest PKC involvement in HIF-1α activation. Since nitric oxide synthase (NOS) are involved in heart vascular regulation, endothelial NOS (e-NOS) and inducible NOS (i-NOS) expression has been investigated: a lower eNOS and an higher iNOS expression has been found in newborn rats exposed to 95% oxygen related to the evidence that hyperoxia provokes a systemic vasoconstriction and to the iNOS pro-apoptotic action, respectively. The occurrence of apoptotic events, evaluated by TUNEL and Bax expression analyses, seems more evident in sample exposed to severe hyperoxia. All in all such results suggest that in newborn rats hyperoxia can trigger oxygen free radical mediated membrane injury through a pPKCα mediated HIF-1α signalling system, even though specificity of such response could be obtained by in vivo administration to the rats of specific inhibitors of PKCα. This intracellular signalling can switch molecular events leading to blood vessels development in parallel to pro-apoptotic events due to an immature anti-oxidant defensive system in newborn rat hearts.

Mitochondria are the major producers of free radical oxygen species (ROS) as well as the major target of oxidative damage. Defects in the mitochondrial respiratory chain complexes can increase ROS production and reduce ROS removal, leading to oxidative modification of proteins, lipids, and DNA. AAA proteases of the inner mitochondrial membrane, paraplegin and AFG3L2, participate in the biogenesis and maintenance of respiratory chain complexes. These proteins form hetero-oligomeric paraplegin/ AFG3L2 and homo-oligomeric AFG3L2 complexes named m-AAA proteases. Inactivation of m-AAA proteases causes respiratory defects and altered mitochondrial morphology both in yeast and in mammals. In fact, mouse models defective for Afg3l2 display a very severe neurological syndrome and die within two weeks after birth. They display widespread morphological alterations of mitochondria in the central and peripheral nervous system and deficiencies in respiratory chain complex I and in complex III, which are major producers of ROS in physiological and especially in pathological conditions. Therefore, an efficient and reliable methodology to monitor the effect of increased ROS production is useful for accurately phenotyping cellular and animal models mutants in m-AAA. By measuring carbonyl formation as marker of protein oxidation, we have shown that respiratory defects cause oxidative damage in Afg3l2 mutants, indicating that oxidative stress is crucial in the pathogenesis of m-AAA deficiency.

Conductive polymeric films have been of great interest recently, although one major drawback to their use is environmental stability. Polypyrroles doped with p-toluenesulfonate, tetrafluoroborate and perchlorate anions were examined under elevated temperatures and environments. The use of d.c. electrical conductivity decay measurements and fitting of kinetic models allows changes in bulk properties to be examined. Surface characterisation performed by X-ray photoelectron spectroscopy (XPS) enables detailed investigation into changes in chemical states of surface atoms. The mechanisms described for the change in electrical and structural properties may be attributed to polymer backbone oxidation through free radical oxygen insertion and electrolyte dopant degradation.

Factors that potentially affect the generation of excess low molecular weight DNA (LMW-DNA) in cultured phytohemagglutinin (PHA)-stimulated lymphocytes of patients with systemic lupus erythematosus (SLE) were studied because this species of DNA is consistently found and this DNA may play a role in the pathogenesis of the disease. Superoxide dismutase (SOD; 0.05 mg/mL), a scavenger of free radical oxygen, decrease LMW-DNA formation in lymphocytes by 22%. Co-cultivation with cysteamine, a second scavenger of free radical oxygen and a sulfhydryl radioprotective agent, resulted in a 32% decrease in the generation of excess LMW-DNA at a concentration of 0.5 x 10(-3) mol/L and largely prevented its formation at 1.0 x 10(-3) mol/L. Other free radical scavengers (catalase, mannitol, vitamins C and E), cyclooxygenase inhibitors (ibuprofen and aspirin), a xanthine oxidase inhibitor (allopurinol), and an iron chelator (desferoxamine) did not affect excess LMW-DNA formation. Glutathione (1 x 10(-3) mol/L) had no effect and cysteine was toxic. Because scavengers of free radicals might be useful in the therapy of lupus, a trial of cysteamine (30 to 60 mg/kg/d) was administered to six acutely ill patients with SLE. A therapeutic benefit was not demonstrated, and some patients had exacerbation of disease. Lymphocyte cell growth from control and lupus subjects was stimulated when cysteamine, 1 x 10(-5) to 1 x 10(-4) mol/L was added to the media, but inhibited at concentrations of 2 x 10(-4) mol/L or greater. These studies suggest that the autooxidation and toxicity of high-dose cysteamine preclude its therapeutic use as a free radical scavenger.

Increasing evidence suggests a critical role for oxidative and nitrosative stress in the pathogenesis of most important neurodegenerative disorders. Parkinson's disease (PD) is a neurodegenerative disease characterized by a severe depletion in number of dopaminergic cells of the substantia nigra (SN). Administration of L-DOPA (LD) is the more effective treatment for patients with PD. However, the vast majority of patients suffer LD-related complications, which represent the major problem in the clinical management of PD. In the present study, LD administration to rats resulted in a significant dose-dependent increase in Hsp70 synthesis which was specific for the SN. The amount of 70 kDa protein increased after 6 h treatment reaching the maximal induction after 24-48 h. Induction of Hsp70 in the SN was associated with a significant increase in constitutive Hsc70 and mitochondrial Hsp60 stress proteins, and with increased expression of mitochondrial complex I whereas no significant changes were found in the activity of complex IV. In the same experimental conditions, a significant decrease in reduced glutathione was observed, which was associated with an increased content of oxidized glutathione content as well as nitric oxide (NO) synthase activity, NO metabolites and nitrotyrosine immunoreactivity. Interestingly, Hsp70 induction, iNOS up-regulation and nitrotyrosine formation have been confirmed also in SN and striatum of rats treated with LD and carbidopa, this latter being an inhibitor of the peripheral DOPA decarboxylase. Our data are in favor of the importance of the heat shock signal pathway as a basic mechanism of defense against neurotoxicity elicited by free radical oxygen and nitrogen species produced in aging and neurodegenerative disorders.

In the present study the potential neurotoxicity of diphenyl diselenide, as measured by the manifestation of seizures in rat pups (postnatal days, PND, 12-14) was evaluated. The results suggest that the latency for the appearance of tonic-clonic seizures, characterized by rearing and falling of rat pups body, was dependent of the dose tested. Diphenyl diselenide at high doses induced seizure episodes in rat pups. The highest dose of diphenyl diselenide (500 mg/kg) increased the levels of lipid peroxidation and catalase activity as well as decreased delta-ALA-D (delta-aminolevulinate dehydratase) and Na(+), K(+) ATPase activity in the brain of rat pups. Our results indicate the possible involvement of free radical oxygen injury in diphenyl diselenide-induced seizures. The data obtained with the dose of 150 mg/kg in the brain of rats that exhibited seizures are: an increase in lipid peroxidation levels; the lack of effect on catalase activity; an inhibition of delta-ALA-D activity, supporting that the enzyme activity is more sensitive than other parameters analyzed as an indicator of oxidative stress. The lowest dose of diphenyl diselenide emphasizes the relationship between the appearance of seizures and the latency for the onset of the first episode. Taken together, this paper could add to our understanding of diphenyl diselenide neurotoxic effect demonstrated by the appearance of seizures which are, at least in part, related to the oxidative stress.

Mitochondria play central roles in cellular metabolism and apoptosis and are a major source of reactive oxygen species (ROS). We investigated the role of ROS and mitochondria in radiation-induced apoptosis in multiple myeloma cells. Two distinct levels of ROS were generated following irradiation: a small increase observed early, and a pronounced late increase, associated with depletion of reduced glutathione (GSH) and collapse of mitochondrial membrane potential (deltapsi(m)). Exogenous ROS and caspase-3 induced deltapsi(m) drop and cytochrome c release from mitochondria, which could be prevented by molecular (dominant-negative caspase-9) and pharmacologic (zVAD-fmk) caspase inhibitors and overexpression of Bcl-2. Exogenous ROS also induced mitochondrial permeability transition (PT) pore opening and cytochrome c release in isolated mitochondria, which could be blocked by inhibition of PT with cyclosporin A. These results indicate that the late ROS production is associated with increased PT pore opening and decreased deltapsi(m), and GSH, events associated with caspase activation and cytochrome c release.

Cardiolipin, a lipid of the mitochondrial inner membrane, is lost from many types of cells during apoptotic death. Here we show that the cardiolipin content of nerve growth factor (NGF)-deprived rat sympathetic neurons undergoing apoptotic death in cell culture decreased before extensive loss of mitochondria from the cells. By 18-24 h after NGF deprivation, many neurons did not stain with the cardiolipin-specific dye, Nonyl Acridine Orange, suggesting complete loss of cardiolipin. Gas chromatography confirmed the decline of cardiolipin content in NGF-deprived neurons. Electron microscopy and immunoblots for the mitochondrial-specific protein, heat shock protein 60 (HSP60), revealed that there was only a slight decrease in mitochondrial mass at this time. Cardiolipin loss after NGF deprivation was concurrent with increased production of mitochondrial-derived reactive oxygen species [Kirkland, R.A., Franklin, J.L., 2001. J. Neurosci. 21, 1949-1963] and increased lipid peroxidation. Compounds having antioxidant effects blocked peroxidation, loss of cardiolipin, and the decrease of mitochondrial mass in NGF-deprived neurons. These compounds also blocked an increase in the number of lysosomes and autophagosomes in NGF-deprived cells. The findings reported here show that the important mitochondrial inner membrane lipid, cardiolipin, is lost from mitochondria during neuronal apoptosis and that this loss occurs before significant loss of mitochondria from cells. They suggest that the loss of cardiolipin is mediated by free radical oxygen.

Uncoupling proteins are inner mitochondrial membrane proteins, which dissipate the proton gradient, releasing the stored energy as heat. Five proteins have been cloned, named UCP1, UCP2, UCP3, UCP4 and UCP5/BMCP1. These proteins are structurally related but differ in tissue expression. UCP1 is expressed uniquely in the brown adipose tissue, while UCP2 is widely distributed, UCP3 is mainly restricted to skeletal muscle and UCP4 and UCP5/BMCP1 expressed in the brain. The properties and regulation of the uncoupling proteins and their exact function has been the focus of an intense research during recent years. This review briefly summarizes the actual knowledge of the properties and function of this new family of proteins. While UCP1 has a clear role in energy homeostasis, the newcomers UCP2-UCP5 may have more delicate physiological importance acting as free radical oxygen scavengers and in the regulation of ATP-dependent processes, such as secretion.

[Arg6, D-Trp7,9, NmePhe8]-substance P (6–11) (antagonist G) inhibits small cell lung cancer (SCLC) growth and is entering Phase II clinical investigation for the treatment of SCLC. As well as acting as a neuropeptide receptor antagonist, antagonist G stimulates c-jun-N-terminal kinase (JNK) activity and apoptosis in SCLC cells. We extend these findings and show that the stimulation of JNK and apoptosis by antagonist G is dependent upon the generation of reactive oxygen species (ROS) being inhibited either by anoxia or the presence of N-acetyl cysteine (n-AC). Antagonist G is not intrinsically a free radical oxygen donor but stimulates free radical generation specifically within SCLC cells (6.2-fold) and increases the activity of the redox-sensitive transcription factor AP-1 by 61%. In keeping with this, antagonist G reduces cellular glutathione (GSH) levels (38% reduction) and stimulates ceramide production and lipid peroxidation (112% increase). At plasma concentrations achieved clinically in the phase I studies, antagonist G augments, more than additively, growth inhibition induced by etoposide. Our results suggest that antagonist G may be particularly effective as an additional treatment with standard chemotherapy in SCLC. These novel findings will be important for the clinical application of this new and exciting compound and for the future drug development of new agents to treat this aggressive cancer. © 2000 Cancer Research Campaign

It is believed that the mechanism underlying vascular endothelial cell damage in preeclampsia is associated with the formation of lipid peroxides and other reactive free radical oxygen species generated from persistent placental underperfusion. Support for the role of oxidative stress (free radical-mediated lipid peroxidation) in the pathophysiology of preeclampsia is provided by biochemical evidence that the plasma serum of patients with the disorder has a high concentration of lipid peroxidation products and is deficient in endogenous antioxidants. One clinical trial showed that oral administration of lycopene, a strong antioxidant and free radical scavenger, significantly reduced the risk of preeclampsia and intrauterine growth retardation in low-risk primigravidas. Because of this and other evidence, it was expected that lycopene would prevent preeclampsia in a similar low-risk population. This randomized double blind placebo-controlled trial evaluated the effectiveness of antioxidant supplementation with lycopene for the prevention of preeclampsia in 159 healthy singleton primigravidas between 12 and 20 weeks of gestation. The study subjects were randomized to receive oral daily doses of lycopene 2 mg (n = 77) or a placebo (n = 82) and were followed until delivery for the development of preeclampsia. There was no difference between the 2 groups in the percentage of patients who developed preeclampsia (18% in both groups). However, women who received lycopene had a significantly higher incidence of preterm labor than the placebo (10.4% vs. 1.2%, P = 0.02), and more low birth weight (

Cranial neural crest cells give rise to a large part of the facial structures, and disturbed development of these cells may therefore cause congenital malformations affecting the head and face. We studied the effects of increased glucose concentration on the migration and development of cranial neural crest cells, maintained in vitro for 48 h. Pre-migratory cranial neural crest cells were removed from embryos of normal and diabetic rats on gestational day 9. After 24 h in 10 mmol/l glucose the cells were exposed to glucose concentrations of 10, 30, or 50 mmol/l for another 24 h. The cultures were photographed at 24 h and 48 h in a phase-contrast microscope to evaluate cell morphology, cell number, and cell migration. Exposure to 50 mmol/l glucose reduced the total number of neural crest cells, their mean migratory distance and migratory area expansion compared to cells cultured in 10 mmol/l glucose. To investigate the effect of antioxidant agents, high glucose cultures were studied after addition of N-acetylcysteine (NAC), or superoxide dismutase (SOD). Addition of NAC diminished the inhibitory effect of high glucose, whereas SOD did not offer any improvement in cell development. Neural crest cell culture from embryos of diabetic rats showed reduced cell migration in vitro at all glucose concentrations compared to normal cells. In addition, the cells from embryos of diabetic rats showed reduced migratory area expansion after culture in the basal 10 mmol/l glucose concentration, indicating that maternal diabetes permanently influences the future development of premigratory cranial neural crest cells. These findings indicate that high glucose concentration inhibits cranial neural crest development in vitro, and that antioxidant therapy may diminish this inhibition. Free radical oxygen species may be involved in the induction of malformations and antioxidants may therefore have a role in future attempts to block the teratogenic effects of diabetic pregnancy.

Objective: Polymorphonuclear neutrophil (PMN) granulocytes play an important role in trauma, shock, and sepsis. In these acute illness situations, multiple parameters can be used to describe functional and structural changes of PMN granulocytes like, for example, cell count, multiple surface adhesion molecules, free radical oxygen production, activity of phagocytosis, and cytokine production. Nevertheless, up to now, in most of the daily clinical situations, only the general white blood cell count is used to receive information, ignoring the fact that the leukocyte-cell system is a mix of different complex cell-subsystems with totally different functions. Considering that complex systems show nonlinear behavior, we could expect heterogeneous reactions of PMN granulocytes to near-identical stimulations. Therefore, functional and structural parameters of this cell type were assessed during an ischemiareperfusion situation due to an identical extremity trauma impact. Material and Methods: Twenty male patients; elective anterior cruciate ligament reconstruction with tourniquet-induced ischemia; structural parameters: CD11b, CD16, CD18, CD62L, CD55, CD63, CD95; functional parameters: cell count, respiratory burst (free radical oxygen production) with and without stimulation by tumor necrosis factor, PMA, or fMLP. Analysis by fluorescence cytometry. Results: Each parameter showed uniformed changes in all patients. This could be interpreted as a homogenous reaction to homogenous stimulations. Nevertheless, in the analysis of all 20 patients intraindividually, it can be demonstrated that summarizing all functional and structural parameters each patient showed a total different pattern of PMN granulocyte reactions to a homogenous ischemia-reperfusion trauma. Conclusions: To describe cell behavior in acute illness situations, it is not adequate to use the routine, for example, white blood cell count. Even the analysis of single bhigh-techQ parameters can describe the principle change of the complex cell system only. It is necessary to analyze the pattern of cell behavior intraindividually to get deeper insights for a better understanding of reactions to, for example, trauma situations.

The reactivity of double bond type lattice oxygen Mo6+O in supported molybdena catalyst has been investigated by means of ESR and ir spectroscopy. It has been shown that the double bond oxygen is similar to a free radical oxygen in nature, in contrast to σ-bond oxygen MoOMo. This character increases as a result of electron transfer from an adsorbed olefin species to a nonbonding atomic orbital of Mo6+ with some decrease in the strength of MoO bond during vapor-phase oxidation of lower olefins. The reactivity of the double bond oxygen as a radical increases inversely with the electronegativity of the molybdenum ion as modified with an oxide of a VA group element or of an alkali metal. This was supported by studies of the catalysis of the oxidation of propylene to acrolein, of butadiene to maleic anhydride and the oxidative dehydrogenation of trans-2-butene to butadiene. On the other hand, the activity for these reactions of catalyst modified with a high concentration of alkali metal oxide increases with the electronegativity of the alkali metal. This different effect due to the concentration of the alkali metal oxide is also discussed.

Data is presented showing that one commercial preparation of cytochome c, used to trap and measure free radical superoxide anion, can be contaminated with cytochrome c oxidase activity. This activity can vary from lot to lot, can introduce variability into the measurement of superoxide anion and can result in falsely low estimations of free radical formation. This cytochrome c oxidase activity can be inhibited by low (0.2 mM ) concentrations of KCN. Blockade of the cytochrome c oxidase activity allows reproducible measurement of superoxide anion formation at low levels by red cells.

Increasing evidence suggests a role for reactive free radical oxygen species in the multi-stage events of chemical carcinogenesis. We hypothesized that variations in the level of superoxide dismutase (SOD), a major endogenous antioxidant enzyme, may account in part for variations in susceptibility to cancer induced by polycyclic aromatic hydrocarbons (PAH). The SOD activity of mammary epithelial cells from rats with varying susceptibility to dimethylbenz[a]anthracene (DMBA)-induced breast cancer was assayed. Ageing, pregnancy and previous multiple pregnancies reduce susceptibility of Sprague--Dawley female rats to DMBA. These decreases in susceptibility were correlated with increased levels of SOD activity. Only minor differences in SOD activity was observed in mammary epithelium of genetic strains of rats with differences in susceptibility to DMBA. These data suggest that, in models where physiological differences may account for variations in effectiveness of PAH to induce mammary cancer, SOD activity is inversely correlated with breast cancer susceptibility and support the hypothesis that cancer susceptibility may be partially mediated through reactive free radical oxygen intermediates.

Daunorubicin administration to mice produces a marked stimulation of lipid peroxidation in both liver and heart 48 hours following administration. In direct contrast 1,4-Bis{2-[(2-hydroxyethyl)amino]ethylamino}-9, 10-anthracenedione (HAQ) does not induce lipid peroxidation in the liver and actually inhibits this event in the heart. In addition, neither daunorubicin nor HAQ deplete reduced glutathione in liver or heart 48 hours after drug administration. Daunorubicin induced glutathione (GSH) depletion was observed 2.5 hours following administration. These results correlate with daunorubicin increased microsomal oxygen consumption whilst HAQ produced no measurable effect on the rate of microsomal oxygen utilisation. It would appear that redox cycling to produce free radical oxygen involved in lipid peroxidation and GSH depletion, an established action of daunorubicin, does not occur with HAQ. This apparent lack of HAQ reactivity may help explain the relatively low cardiotoxicity of this novel antitumour agent.

The amount and activity of superoxide dismutase (SOD) (EC 1.15.1.1) were measured in red cells collected from 50 white controls, 101 black controls, 50 patients with sickle hemoglobin (SS Hb), 12 with sickle trait, and 11 with other sickling hemoglobinopathies. Red cells from normal black subjects had more SOD amount and activity than normal whites (1.77 U/mg Hb and 2.96 micrograms/mg Hb vs. 1.47 U/mg Hb and 2.64 micrograms/mg Hb, respectively) or blacks with SS Hb or other sickling hemoglobinopathies. Patients with more severe manifestations of SS Hb had lower levels of SOD activity than those with milder symptoms but had the same amount of enzyme protein. Individuals with sickle trait had amounts and activities of SOD comparable to black controls. An alteration in defense to free radical oxygen may play a role in the severity of symptoms experienced by patients with homozygous sickle cell disease.