Preparation of mixed micelles carrying folates and stable radicals through PLA stereocomplexation for drug delivery.

Multifunctional mixed micelles possessing targeting folates on the periphery and stable radicals in the core were prepared from the mixture of folate-poly(ethylene glycol)-block-poly(d-lactide) (FA-PEG-b-PDLA) and poly(ethylene glycol) methyl ether-block-poly(l-lactide)- 2,2,6,6-tetramethylpiperidine-1-oxyl (mPEG-b-PLLA-TEMPO). FA-PEG-b-PDLA and mPEG-b-PLLA-TEMPO were prepared by combining ring-opening polymerization (ROP) of lactide with a series of conversion of the end functional groups. The synthesized block copolymers and their intermediates were well characterized by gel permeation chromatography (GPC) and 1H nuclear magnetic resonance (1H NMR) spectroscopy. The mixture of FA-PEG-b-PDLA and mPEG-b-PLLA-TEMPO self-assembled into spherical micelles with the average diameter about 200 nm through PLA stereocomplexation, which were characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM) and differential scanning calorimetry (DSC). The formed micelles were confirmed to emit electron paramagnetic resonance (EPR) signals by EPR spectroscopy. Additionally, the formed micelles exhibited high loading capacity of hydrophobic anticancer drug, controlled in vitro drug release, satisfied biocompatibility and a significantly higher cellular uptake, indicating promising applications in smart drug delivery.

A phosphate-targeted dinuclear Cu(II) complex combining major groove binding and oxidative DNA cleavage.

Free radical generation is an inevitable consequence of aerobic existence and is implicated in a wide variety of pathological conditions including cancer, cardiovascular disease, ageing and neurodegenerative disorder. Free radicals can, however, be used to our advantage since their production is catalysed by synthetic inorganic molecules-termed artificial metallonucleases-that cut DNA strands by oxidative cleavage reactions. Here, we report the rational design and DNA binding interactions of a novel di-Cu2+ artificial metallonuclease [Cu2(tetra-(2-pyridyl)-NMe-naphthalene)Cl4] (Cu2TPNap). Cu2TPNap is a high-affinity binder of duplex DNA with an apparent binding constant (Kapp) of 107 M(bp)-1. The agent binds non-intercalatively in the major groove causing condensation and G-C specific destabilization. Artificial metallonuclease activity occurs in the absence of exogenous reductant, is dependent on superoxide and hydrogen peroxide, and gives rise to single strand DNA breaks. Pre-associative molecular docking studies with the 8-mer d(GGGGCCCC)2, a model for poly[d(G-C)2], identified selective major groove incorporation of the complex with ancillary Cu2+-phosphate backbone binding. Molecular mechanics methods then showed the d(GGGGCCCC)2 adduct to relax about the complex and this interaction is supported by UV melting experiments where poly[d(G-C)2] is selectively destabilized.

In vivo evaluation of different alterations of redox status by studying pharmacokinetics of nitroxides using magnetic resonance techniques.

Free radicals, particularly reactive oxygen species (ROS), are involved in various pathologies, injuries related to radiation, ischemia-reperfusion or ageing. Unfortunately, it is virtually impossible to directly detect free radicals in vivo, but the redox status of the whole organism or particular organ can be studied in vivo by using magnetic resonance techniques (EPR and MRI) and paramagnetic stable free radicals - nitroxides. Here we review results obtained in vivo following the pharmacokinetics of nitroxides on experimental animals (and a few in humans) under various conditions. The focus was on conditions where the redox status has been altered by induced diseases or harmful agents, clearly demonstrating that various EPR/MRI/nitroxide combinations can reliably detect metabolically induced changes in the redox status of organs. These findings can improve our understanding of oxidative stress and provide a basis for studying the effectiveness of interventions aimed to modulate oxidative stress. Also, we anticipate that the in vivo EPR/MRI approach in studying the redox status can play a vital role in the clinical management of various pathologies in the years to come providing the development of adequate equipment and probes.

Effects of binge ethanol on lipid homeostasis and oxidative stress in a rat model of nonalcoholic fatty liver disease

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Excess fat accumulation renders the liver more vulnerable to ethanol, but it is still unclear how alcohol enhances lipid dysmetabolism and oxidative stress in a pre-existing steatosis condition. The effects produced by binge ethanol consumption in the liver of male Wistar rats fed a standard (Ctrl) or a high-fat diet HFD were compared. The liver status was checked through tissue histology and standard serum parameters. Alteration of hepatic lipid homeostasis and consequent oxidative unbalance were assessed by quantifying the mRNA expression of the lipid-regulated peroxisome proliferator-activated receptors (PPARs), of the cytochromes CYP2E1 and CYP4A1, and of some antioxidant molecules such as the metallothionein isoforms MT1 and MT2 and the enzymes catalase and superoxide dismutase. The number of adipose differentiation-related protein (ADRP)-positive lipid droplets (LDs) was evaluated by immunohistochemical staining. As a response to the double insult of diet and ethanol the rat liver showed: (1) a larger increase in fat accumulation within ADRP-positive LDs; (2) stimulation of lipid oxidation in the attempt to limit excess fat accumulation; (3) induction of antioxidant proteins (MT2, in particular) to protect the liver from the ethanol-induced overproduction of oxygen radicals. The data indicate an increased susceptibility of fatty liver to ethanol and suggest that the synergistic effect of diet and ethanol on lipid dysmetabolism might be mediated, at least in part, by PPARs and cytochromes CYP4A1 and CYP2E1 (AU)

Señalización de la angiotensina II, a través de la producción de radicales libres en estructuras del sistema nervioso: papel en el estres agudo y crónico / Angiotensin II signaling, through the production of free radicals in structures of the nervous system: role in acute and chronic stress

Existen evidencias que apoyan la participación de las especies reactivas de oxígeno en las cascadas de señalización y transducción intracelular de la angiotensina II. La ANG II es importante en el mantenimiento de la homeostasis corporal, regulando la presión arterial y el metabolismo de fluidos y electrolitos. Se sabe que en la periferia, la ANG II es capaz de estimular a la NAD(P)H oxidasa con la subsiguiente producción de ERO. El anión superóxido es metabolizado secuencialmente por las enzimas antioxidantes como la superóxido dismutasa, la catalasa y la glutatión peroxidasa. A su vez, las especies reactivas de oxígeno son capaces de activar a las proteínas kinasas activadas por mitógenos, las cuales se encuentran asociadas al crecimiento y la diferenciación celular. Se evaluó la posible participación de las especies reactivas de oxígeno en el mecanismo de señalización intracelular mediado por el receptorAT1en el hipotálamo, el órgano subfornicaly médula suprarrenal de la rata. Nuestros resultados demostraron que la estimulación del tejido nervioso con ANG II in vitroincrementó la actividad de la enzimas antioxidante. Al evaluar el papel del receptor AT1, la NAD(P)H oxidasa, el anión superóxido y la proteína kinasa C; así como la activación de las ERK1/2 en la señalización de la ANG II en el hipotálamo, OSF y MSR, demostramos que el bloqueo del receptor AT1con losartán, la interferencia del ensamblaje de la NAD(P)H oxidasa con apocinina, el secuestro de anión superóxido empleando un mimético de la SOD, tempol,y la inhibición de la PKC con cheleritrina, bloquearon completamente el efecto que produce la ANG II sobre las enzimas antioxidantes in vitro.Igualmente, la activación de la ERK1/2 inducida por la ANG II fue reducida por APO y LOS a nivel hipotalámico. Adicionalmente, el bloqueo del receptor AT2hipotalámico con PD123319, no bloqueo sino que mas bien potenció la respuesta de las enzimas antioxidantes y la activación de las ERK1/2 inducida por la ANG II, lo que desenmascaró el efecto contra regulatorio del receptor AT2sobre la acción de la ANG II mediada por el receptor AT1. Se sabe que durante el estrés el sistema renina angiotensina circulante y cerebral se encuentra estimulado, por lo tanto el incremento de la ANG II endógena debería desencadenar vías de señalización similares a las reportadas in vitro. Efectivamente, nuestros hallazgos demostraron que tanto,el estrés agudo inducido por la inmovilización forzada,como el estrés crónico en ratas espontáneamente hipertensas incrementaron la actividad de las enzimas antioxidantes en las tres estructuras cerebrales estudiadas. Este efecto es mediado por la vía del receptor AT1, la estimulación de la NAD(P)H oxidasa y la producción de anión superóxido ya que el tratamiento in vivo con LOS, APO y TEM fue capaz de bloquear completamente el incremento de la actividad de las enzimas antioxidantes inducidas por el estrés y por ende por la ANG II endógena.A nivel de la MSR demostramos, por primera vez, que la estimulación del receptor AT2 esta asociada a la estimulación de la NAD(P)H oxidasa, ya que la APOy el PD 123319 fueron capaces de bloquear el incremento de la actividad de las enzimas antioxidantes inducida por la ANG II. Demostrando así, que el receptor AT1en la MSR contrarregula la acción de la ANG II a través del receptor AT2.En conclusión, nuestros resultados indican que a nivel del sistema nervioso las especies reactivas de oxígeno participan en la cascada de señalización intracelular de la ANG II, y ejercen un importante papel en la respuesta al estrés y la hipertensión.

In vivo high-resolution 3D overhauser-enhanced MRI in mice at 0.2 T.

Overhauser-enhanced MRI (OMRI) offers the potentiality of detecting low-concentrated species generated by specific biological processes. However molecular imaging applications of OMRI need significant improvement in spatial localization. Here it is shown that 3D-OMRI of a free radical injected in tumor-bearing mice can be performed at high anatomical resolution at a constant field. A 30 mm cavity operating at 5.43 GHz was inserted in a C-shaped magnet for proton MRI at 0.194 T. Nude mice with or without brain-implanted C6 rat glioma were positioned in the cavity and injected with TOPCA (1-oxyl-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrole-3-carboxylic acid). OMRI was performed in 3D within several minutes in the brain region without high overheating of the animals. Voxel size was 0.5 × 0.5 × 1 mm³ , providing good delineation of brain regions. Signal amplifications ranged from 2 in tumors to 10 in vessels several minutes after TOPCA injection. Time-course of signal enhancement could be measured by 2D OMRI at 15 s time intervals in a localized thin slice. The method opens the way for molecular imaging of biological activities able to generate OMRI-visible free radicals.

Neuroprotección mediada por diterpenos aislados de Sideritis spp. frente al estrés oxidativo en astrocitos / Results from the application of a quality management system in the community pharmacy

El estrés oxidativo es un proceso de oxidación no controlado de las estructuras biológicas, desencadenado por radicales libres, que produce daño y muerte celular. Numerosas evidencias implican al estrés oxidativo en la patogénesis de enfermedades neurodegenerativas como Alzheimer o Parkinson. Compuestos con capacidad antioxidante pueden prevenir el daño oxidativo al inhibir la producción de radicales libres. Más de 160 diterpenos han sido identificados y aislados de las partes aéreas de Sideritis spp. A ellos se atribuyen en gran medida actividades antiinflamatorias, antioxidantes y digestivas por las cuales se utilizan estas especies en la medicina tradicional mediterránea. El objetivo de este estudio es evaluar el efecto neuroprotector de los diterpenosandalusol, conchitriol y lagascatriol, aislados de las partes aéreas de Sideritis spp. frente al estrés oxidativo inducido por peróxido de hidrógeno en la línea celular U373MG de astrocitoma humano. El posible efecto protector de los diterpenos frente al daño provocado por peróxido de hidrógeno ha sido evaluado mediante el ensayo de MTT. La capacidad de captación de radicales libres de los compuestos se ha estudiado mediante la técnica del DPPH. Los resultados del presente estudio demuestran que los diterpenos andalusol, conchitriol y lagascatriol (5 y 10 μM) protegen a las células U373 MG de la toxicidad del peróxido de hidrógeno, lo que se atribuye en parte a la capacidad antioxidante de dichos compuestos. Se sugiere que estos diterpenos podrían ejercer un efecto beneficioso en la prevención de enfermedades neurodegenerativas asociadas al estrés oxidativo. Estudios futuros irán encaminados a una profundización de su mecanismo de acción(AU)

Oxidative stress is an uncontrolled oxidation process of biological structures, triggering by free radicals, which cause damage and death cell. Many evidences implicate oxidative stress in the pathogenesis of neurodegenerative diseases such as Alzheimer and Parkinson. Compounds with antioxidant capacity may prevent from oxidative damage because of the inhibition of free radical production. More than 160 diterpenes have been identified and isolated from aerial parts of Sideriti sspp. These compounds are largely responsible for the anti-inflammatory, antioxidant and digestive activities, reason why these species are used in traditional Mediterranean medicine. To evaluate the neuroprotective effect of the diterpenes andalusol, conchitriol and lagascatriol, isolated from Sideriti sspp. aerial parts, against oxidative stress induced by hydrogen peroxide in the human astrocytoma U373 MG cell line. The possible protective effect of the diterpenes against the damage induced by hydrogen peroxide has been evaluated using MTT assay. The free radical scavenging capacity of these compounds has been studied using DPPH method. Results of the present study demonstrated that the diterpenes andalusol, conchitriol y lagascatriol (5 and 10 μM) protect U373 MG cells from hydrogen peroxide toxicity, which is due to in part to the antioxidant capacity of these compounds. Itis suggested that these diterpenes could exert a beneficial effect in the prevention of neurodegenerative diseases associated to oxidative stress. These future studies will be focused on a deepening of action mechanism(AU)

Non-phenolic radical-trapping antioxidants.

OBJECTIVES: The aim of this review article is to introduce the reader to the mechanisms, rates and thermodynamic aspects of the processes involving the most biologically relevant non-phenolic radical-trapping antioxidants. KEY FINDINGS: Antioxidant defences in living organisms rely on a complex interplay between small molecules and enzymes, which cooperate in regulating the concentrations of potentially harmful oxidizing species within physiological limits. The noxious effects of an uncontrolled production of oxygen- and nitrogen-centered radicals are amplified by chain reactions (autoxidations), sustained mainly by peroxyl radicals (ROO(*)), that oxidize and alter essential biomolecules such as lipids, lipoproteins, proteins and nucleic acids. SUMMARY: Non-phenolic antioxidants represent an important and abundant class of radical scavengers in living organisms. These compounds react with peroxyl radicals through various mechanisms: (i) formal H-atom donation from weak X-H bonds (X = O, N, S), as in the case of ascorbic acid (vitamin C), uric acid, bilirubin and thiols; (ii) addition reactions to polyunsaturated systems with formation of C-radicals poorly reactive towards O(2), for example beta-carotene and all carotenoids in general; (iii) co-oxidation processes characterized by fast cross-termination reactions, for example gamma-terpinene; and (iv) catalytic quenching of superoxide (O(2)(*-)) with a superoxide dismutase-like mechanism, for example di-alkyl nitroxides and FeCl(3). Kinetic data necessary to evaluate and rationalize the effects of these processes are reported. The mechanisms underlying the pro-oxidant effects of ascorbate and other reducing agents are also discussed.

Kinetic study of the prooxidant effect of alpha-tocopherol. Hydrogen abstraction from lipids by alpha-tocopheroxyl radical.

A kinetic study of the prooxidant effect of alpha-tocopherol was performed. The rates of allylic hydrogen abstraction from various unsaturated fatty acid esters (ethyl stearate 1, ethyl oleate 2, ethyl linoleate 3, ethyl linolenate 4, and ethyl arachidonate 5) by alpha-tocopheroxyl radical in toluene were determined, using a double-mixing stopped-flow spectrophotometer. The second-order rate constants (k (p)) obtained are <1 x 10(-2) M(-1 )s(-1) for 1, 1.90 x 10(-2) M(-1 )s(-1) for 2, 8.33 x 10(-2 )M(-1 )s(-1) for 3, 1.92 x 10(-1) M(-1 )s(-1) for 4, and 2.43 x 10(-1 )M(-1 )s(-1) for 5 at 25.0 degrees C. Fatty acid esters 3, 4, and 5 contain two, four, and six -CH(2)- hydrogen atoms activated by two pi-electron systems (-C=C-CH(2)-C=C-). On the other hand, fatty acid ester 2 has four -CH(2)- hydrogen atoms activated by a single pi-electron system (-CH(2)-C=C-CH(2)-). Thus, the rate constants, k (abstr)/H, given on an available hydrogen basis are k (p)/4 = 4.75 x 10(-3 )M(-1 )s(-1) for 2, k (p)/2 = 4.16 x 10(-2) M(-1 )s(-1) for 3, k (p)/4 = 4.79 x 10(-2 )M(-1 )s(-1) for 4, and k (p)/6 = 4.05 x 10(-2 )M(-1 )s(-1) for 5. The k (abstr)/H values obtained for 3, 4, and 5 are similar to each other, and are by about one order of magnitude higher than that for 2. From these results, it is suggested that the prooxidant effect of alpha-tocopherol in edible oils, fats, and low-density lipoproteins may be induced by the above hydrogen abstraction reaction.

pH-sensitive radical-containing-nanoparticle (RNP) for the L-band-EPR imaging of low pH circumstances.

For the imaging of low pH circumstances in vivo, a pH-sensitive radical-containing-nanoparticle (RNP), which has an intense electron paramagnetic resonance (EPR) signal, was designed and developed using a self-assembling amphiphilic block copolymer (PEG-b-PCTEMPO) composed of a hydrophilic poly(ethylene glycol) (PEG) segment and a hydrophobic poly(chloromethylstyrene) (PCMS) segment in which the chloromethyl groups were converted to 2,2,6,6-tetramethylpiperidinyloxys (TEMPOs) via the amination of PEG-b-PCMS block copolymer with 4-amino-TEMPO. This RNP formed core-shell-type micelles in the physiological environment, and the cumulant average diameter of the RNP was about 50 nm. The cytotoxicity and acute toxicity studies for the RNP revealed that the median inhibitory concentration (IC(50)) of TEMPO radicals in RNP core and median lethal dose (LD(50)) of RNP were >8 mmol N(TEMPO)/L and >600 mg/kg (>960 mumol N(TEMPO)/kg), respectively, indicating fairly low toxicity. The blood circulation of the RNP was evaluated using ICR mice. Contrary to the rapid clearance of low-molecular-weight TEMPO derivatives such as 4-hydroxy-TEMPO (TEMPOL) from the bloodstream, the EPR signal of the RNP remained for a fairly long period of time. Actually, the signal was observed in the blood for more than 2 h, as monitored by EPR spectroscopy. The compartmentalization of the TEMPO radicals in the RNP core improved the stability in the bloodstream. Since an amino group was introduced in each repeating unit of the PCTEMPO segment, the disintegration of the RNP was caused by the protonation of the amino groups in response to the acidic pH environment (pH < 6.0), as confirmed by the dynamic light scattering (DLS) measurements. In addition, a drastic change in the EPR spectra from broad to sharp triplet was observed, accompanying the disintegration. This change was based upon the mobility of the TEMPO moieties covalently conjugated in the hydrophobic segment, which was confirmed by the rotational correlation time of the TEMPO moieties on the PCTEMPO segment. Note that the peak intensity of the EPR signal increased at around the phase transition point (ca. pH = 6.0). When pH-sensitive RNP solutions at pH values 5.6 and 7.4 were visualized using an L-band EPR imaging system, the phantom images showed a remarkable on-off regulation in response to the acidic pH environment. These results demonstrate that pH-sensitive RNPs are expected to serve as nanoprobes for the in vivo EPR imaging of low pH circumstances.

Nano-emulsions of fluorinated trityl radicals as sensors for EPR oximetry.

This article reports the development and evaluation of two nano-emulsions (F45T-03/HFB and F15T-03/PFOB) containing fluorinated trityl radicals dissolved in perfluorocarbons. Preparation with a high-pressure homogenizer conferred sub-micronic size to both nano-emulsions. In vitro and in vivo EPR spectroscopy showed that the nano-emulsions had much greater oxygen sensitivity than the hydrophilic trityl, CT-03. In vivo experiments in rodents confirmed the ability of the nano-emulsions to follow the changes in oxygen concentration after induced ischemia. Histological evaluation of the tissue injected with the nano-emulsions revealed some acute toxicity for the F45T-03/HFB nano-emulsion but none for the F15T-03/PFOB nano-emulsion. These new formulations should be considered for further EPR oximetry experiments in pathophysiological situations where subtle changes in tissue oxygenation are expected.

Reaction between the anesthetic agent propofol and the free radical DPPH in semiaqueous media: kinetics and characterization of the products.

The reaction of the free radical diphenylpicrylhydrazyl (DPPH ) with the anesthetic agent 2,6-diisopropylphenol (propofol, PPF) was investigated in buffered hydroalcoholic media. The kinetics was followed using a stopped-flow system. DPPH was reduced to the hydrazine analogue DPPH-H with a measured stoichiometry (DPPH /PPF) of 2. The main product of the reaction, 3,5,3',5'-tetraisopropyl-(4,4')-diphenoquinone (PPFDQ) was isolated by chromatography and its structure was fully characterized. The reaction mechanism was inferred from the stoichiometry, kinetics, and product identification. The first step, which primarily determines the kinetics, is the reaction of DPPH with PPF to produce DPPH-H and the PPF radical. The rate constant was found to be 31.8, 207, and 908 M(-1) s(-1) at pH 6.4, 7.4, and 8.4, respectively. The pH dependence is indicative of a higher reactivity of the phenolate form of PPF. Then, PPF radicals combine to form dipropofol, which is quickly oxidized to PPFDQ by the remaining DPPH . This reaction scheme is corroborated by numerical simulations of the kinetics. In the course of this study we also disclosed an unexpected effect, the photochemical degradation of PPFDQ. The need to compare antioxidants on a kinetics basis is again emphasized. In our hands, PPF presents a significantly weaker reactivity than Trolox.

Spatially resolved time-course studies of free radical reactions with an EPRI/MRI fusion technique.

Electron paramagnetic resonance imaging (EPRI) using nitroxyl radicals is a useful technique for visualizing reactive oxygen species (ROS) and the pharmacokinetics of probes. To unambiguously identify anatomical locations, coregistration of EPRI with images obtained by MRI is necessary. In this study the feasibility of performing reliable EPRI/MRI fusion imaging using nitroxyl radical fiducial markers was tested. The pharmacokinetics of the nitroxyl radicals were observed after oral or intravenous administration in C57BL6 mice. To fuse both images, the nitroxyl radical was used as fiducial markers. The EPR and MR images corresponded well and clearly illustrated minimal changes in pharmacokinetics between carbamoyl-PROXYL and carboxy-PROXYL. These results demonstrate that the EPRI/MRI fused imaging technique is useful for investigating in vivo pharmacokinetics and provides unambiguous anatomic details.

Diferential gene expression and adiposity reduction induced by ascorbic acid supplementation in a cafeteria model of obesity

Obesity is considered as an inflammatory disease, in which free radical-induced oxidative stress and excessive intake of macronutrients exacerbate their symptoms. In this context, we assessed in rats the possible preventive effect of the supplementation with an antioxidant molecule, ascorbic acid, in order to reduce the adiposity induced by the intake of a high-fat diet. For this purpose, during 56 days, three groups of male Wistar rats were fed on: a) standard pelleted diet, b) Cafeteria diet, c) ascorbate-supplemented (750 mg/kg of body weight) Cafeteria diet. At the end of the experimental period, microarray analysis was used to identify genes transcriptionally induced or repressed by both experimental dietary models (Cafeteria diet supplemented or not with ascorbic acid) in subcutaneous adipose tissue. Dietary ascorbic acid was able to protect against high fat diet effects, reducing the increase of body weight, total body fat and enlargement of different adipose depots induced by the Cafeteria diet without affecting food intake. An association analysis accurately and differentially allowed the detection of gene expression changes related with adiposity and insulin resistance. The genes that more strongly correlated with body fat and HOMA insulin resistance index were involved in adipocyte differentiation, lipid and glucocorticoid metabolism, cell cycle regulation, as well as in several insulin-induced processes. Some other transcripts are regulated by the vitamin C-mediated reduction of adiposity, such as genes that participate in glucocorticoid metabolism, adipogenesis, pentose phosphate pathway, or tricarboxylic acid cycle. This strategy was able to link variations in adipose tissue gene expression with markers of diet-induced obesity in rats, such as insulin resistance and body fat content (AU)

La obesidad está considerada como una enfermedad inflamatoria, en la que el estrés oxidativo, inducido por los radicales libres, y la excesiva ingesta de macronutrientes incrementan sus síntomas. En este trabajo se estudia el posible efecto preventivo de la suplementación con la molécula antioxidante ácido ascórbico, para reducir la adiposidad inducida por la ingesta de una dieta rica en grasas en rata. Para este propósito, tres grupos de ratas macho de estirpe Wistar se alimentaron durante 56 días con: a) dieta estándar, b) dieta de cafetería, c) dieta de cafetería suplementada con ácido ascórbico (750 mg/kg de peso corporal). Al final del período experimental, se aplicó un análisis por microarray en el tejido adiposo subcutáneo para identificar los genes inducidos o reprimidos por ambos modelos dietéticos experimentales. El ácido ascórbico pudo proteger contra los efectos proadipogénicos de la dieta, previniendo del aumento en el peso corporal, del contenido total de grasa corporal y del peso de los diversos depósitos adiposos inducidos por la dieta de cafetería. Un análisis de asociación permitió además la detección de los cambios en la expresión de los genes más relacionados con la resistencia a insulina y la adiposidad. Los genes que correlacionaron más fuertemente con el porcentaje de grasa corporal y con el índice HOMA están implicados en la diferenciación adipocitaria, el metabolismo de lípidos y de glucocorticoides y la regulación del ciclo celular, así como en varios (..) (AU)

Radicales libres de origen mitocondrial y longevidad / Mitochondrial free radicals and longevity

Desde los inicios del siglo XX se han propuesto muchas teorías diferentes sobre las causas del envejecimiento. Hoy en día, la teoría de los radicales libres de origen mitocondrial es la que disfruta de más apoyos a su favor en la literatura científica. En el presente artículo se revisan los trabajos publicados sobre la relación entre la longevidad máxima de las distintas especies animales y sus niveles endógenos de antioxidantes y de generación mitocondrial de radicales de oxígeno. La mayoría de los estudios sobre suplementación experimental con antioxidantes indican que pueden aumentar la esperanza de vida pero no cambian la longevidad máxima. Además, los antioxidantes endógenos correlacionan de forma negativa con la longevidad máxima. Sin embargo, la intensidad de producción mitocondrial de radicales de oxígeno y el daño oxidativo al ADN mitocondrial son menores en los animales longevos que en los de vida corta. Los animales longevos también muestran un menor grado de insaturación de los ácidos grasos de sus membranas tisulares que las especies de vida corta. Por otra parte, la restricción calórica, la manipulación experimental mejor conocida que disminuye la velocidad del envejecimiento, también disminuye la producción mitocondrial de radicales libres y el daño oxidativo al ADN mitocondrial. Este descenso ocurre en el complejo I. Estos resultados sugieren que se han utilizado mecanismos similares para aumentar la longevidad tanto en la restricción calórica como durante la evolución de especies animales con distinta velocidad de envejecimiento, y que dichos mecanismos incluyen un descenso en la intensidad de generación de radicales libres en la mitocondria

Since the beginning of the XXth century many theories have been proposed to explain aging. Nowadays, the mitochondrial free radical theory is strongly supported by the available scientific evidence. In this article published studies on the relationship between the maximum longevity of animals and their levels of endogenous antioxidants and mitochondrial rates of oxygen radical generation are reviewed. Most studies of antioxidant supplementation indicate that these chemicals can increase mean but not maximum longevity. In addition, endogenous antioxidant levels negatively correlate with maximum longevity. However, the rate of production of oxygen radicals at mitochondria and the steady-state levels of oxidative damage in mitochondrial DNA are lower in long-lived than in short-lived animal species. Long-lived species also have lower levels of fatty acid unsaturation in their cellular membranes. On the other hand, caloric restriction, the best know manipulation that decreases the rate of aging, also lowers mitochondrial free radical production and oxidative damage to mitochondrial DNA. This decrease occurs at complex I. These results suggest that common mechanisms have been used to increase longevity in caloric restriction and during the evolution of animal species with different aging rates. These mechanisms include a decrease in the rate of generation of free radicals at mitochondria

Relación entre el estrés oxidativo mitocondrial y la velocidad del envejecimiento / Relationship between mitochondrial oxidative stress and rate of ageing

Aunque se han propuesto muchas teorías diferentes sobre las causas del envejecimiento, la teoría de los radicales libres es la que disfruta de más apoyos a su favor en la bibliografía científica. En el presente artículo se revisan los trabajos publicados sobre la relación entre la longevidad máxima de las distintas especies animales y sus valores endógenos de antioxidantes y de generación de radicales de oxígeno. La mayoría de los estudios sobre suplementación experimental con antioxidantes indican que pueden aumentar la esperanza de vida pero no cambian la longevidad máxima. Además, los antioxidantes endógenos correlacionan de forma negativa con la longevidad máxima. Sin embargo, la intensidad de producción mitocondrial de radicales de oxígeno y el daño oxidativo al ADN mitocondrial son menores en los animales longevos que en los de vida corta. Los animales longevos también muestran un menor grado de insaturación de los ácidos grasos de sus membranas tisulares que las especies de vida corta. Por otra parte, la restricción calórica, la única manipulación experimental que disminuye la velocidad del envejecimiento, también disminuye la producción mitocondrial de radicales libres y el daño oxidativo al ADN mitocondrial. Este descenso ocurre en el complejo I. Estos resultados sugieren que se han utilizado mecanismos similares para aumentar la longevidad en la restricción calórica y durante la evolución de especies animales con distinta velocidad de envejecimiento, y que dichos mecanismos incluyen un descenso en el estrés oxidativo mitocondrial

Although many theories have been put forward on the causes of ageing, the explanation that has received greatest support in the scientific literature is the free radical theory. The present article reviews the studies published on the relationship between maximal longevity of the distinct animal species and their endogenous antioxidant levels and oxygen radical generation. Most studies on experimental supplementation with antioxidants indicate that they can increase life expectancy but do not change maximal longevity. Moreover, endogenous antioxidants are negatively correlated with maximal longevity. However, the intensity of mitochondrial production of oxygen radicals and oxidative damage to mitochondrial DNA are lower in longevous animals than in short-lived animals. Longevous animals also show a lower grade of fatty acid unsaturation in tissue membranes than short-lived species. Caloric restriction, the only experimental procedure that reduces the rate of ageing, also reduces mitochondrial production of free radicals and oxidative damage to mitochondrial DNA. This decrease occurs in complex I. These results suggest that similar mechanisms are used to increase longevity in caloric restriction and during the evolution of animal species with different rates of ageing and that these mechanisms include a decrease in mitochondrial oxidative stress

Actividad antioxidante in vitro del extracto de metanol de los rizomas de Curculigo orchioides Gaertn / In vitro antioxidant activity of methanol extract of rhizomes of Curculigo orchioides Gaertn

Se analizó la actividad antioxidante in vitro del extracto de metanol de Curculigo orchioides mediante ensayo DPPH, barrido de superóxido, barrido de óxido nítrico, determinación del poder reductor y peroxidación lipídica in vitro vitro. El extracto de metanol resultó ser extremadamente eficaz en el barrido del radical superóxido (IC 50 29.28 ìg/ml), mientras que la actividad fue moderada en el barrido del radical DPPH (IC 50 105.94 µg/ml), el radical de óxido nítrico (IC 50 90.96 µg/ml) y la inhibición de la peroxidación lipídica (IC 50 94.78 µg/ml). El extracto de metanol presentó distintos niveles de actividad antioxidante en los modelos anali- analizados. También apoyó la actividad antioxidante al presentar un poder reductor significativo. Para facilitar litar su identificación y caracterización, se estableció un perfil de huella de cromatografía de capa fina de alto rendimiento (HPTLC) adicional del extracto de metanol. El presente estudio ha demostrado la actividad antioxidante in vitro del extracto de metanol de Curculigo orchioides

Methanol extract of Curculigo orchioides was screened for in-vitro antioxidant activity using DPPH assay, superoxide scavenging, nitric oxide scavenging, determination of reducing power and in-vitro lipid peroxidation. Methanol extract was found to be extremely effective in scavenging superoxide radical (IC 50 29.28 µg/ml) whereas activity was moderate in scavenging DPPH radical (IC 50 105.94 µg/ml), nitric oxide radical(IC 50 90.96 µg/ml) and in inhibition of lipid peroxidation (IC 50 94.78 ìg/ml). Methanol extract showed different levels of antioxidant activities in tested models. It also supported anti oxidant activity by showing significant reducing power. Further HPTLC finger print profile of the methanol extract was established to facilitate its identification and characterization. Present study demonstrated antioxidant activity of methanol extract of Curculigo orchioides in-vitro

A quantitative approach to the recycling of alpha-tocopherol by coantioxidants.

A systematic investigation is reported on the regeneration of alpha-tocopherol (alpha-TOH) in homogeneous solution by coantioxidants in order to better understand the mechanism and the factors responsible for the effectiveness of this process. The current availability of thermochemical data concerning the reactants involved in the regeneration reactions, as well as a large number of the kinetic constants for the various reactions involved, allowed us to rationalize the experimental observations collected so far. Three limiting cases have been considered. The first case is that of a coantioxidant irreversibly regenerating alpha-TOH, where the effectiveness of the recycling process depends on the magnitude of the rate constant k(r). The second case is that of a coantioxidant reversibly recycling alpha-TOH, where regeneration can only be observed if the bond dissociation enthalpy value of the coantioxidant is lower or at least close to that of the O-H bond of alpha-tocopherol. The third case is that of a catechol derivative (chosen as a model compound for polyphenolic antioxidants), where recycling of alpha-TOH is feasible even though the BDE value is significantly higher than that of vitamin E. In this case, the driving force for the recycling process is the removal of the semiquinone radical from the catechol derivative by the alpha-tocopheroxyl radical, which makes the regeneration of alpha-TOH practically irreversible.