Activities Of Key Antioxidant Enzymes Research Articles

Redox regulation in Atlantic cod (Gadus morhua) embryos developing under normal and heat-stressed conditions

With regard to predicted oceanic warming, we studied the effects of heat stress on the redox system during embryonic development of Atlantic cod (Gadus morhua), with emphasis on the glutathione balance, activities of key antioxidant enzymes, and their mRNA levels. The embryos were incubated at optimal temperature for development (6°C) or slightly above the threshold temperature (10°C). The regulation of all the redox-related parameters measured at optimum development was highly dynamic and complex, indicating the importance of both maternal and zygotic contributions to maintaining redox equilibrium. Development at 10°C caused a significantly higher mortality at the blastula and early gastrula stages, indicating severe stress. Measures of the glutathione redox couple showed a significantly more reduced state in embryos at 10°C compared to 6°C at the post-gastrula stages. Mean normalized expression of nrf2, trxred, g6pd, gclc, nox1, CuZnsod, and mt in embryos kept at 10°C revealed stage-specific significantly reduced mRNA levels. Activities of antioxidant enzymes changed both during ontogenesis and in response to temperature, but did not correlate with mRNA levels. As the embryos need a tightly regulated redox environment to coordinate between growth and differentiation, these findings suggest that the altered redox balance might participate in inducing phenotypic changes caused by elevated temperature.

Maternal Immunization Affects In Utero Programming of Insulin Resistance and Type 2 Diabetes

Maternal immunization with oxidized lipoproteins prior to pregnancy protects against atherogenic in utero programming by gestational hypercholesterolemia and enhances beneficial lymphocyte-dependent immune responses in offspring. To determine whether in utero programming and immunomodulation also affect insulin resistance (IR) and type 2 diabetes, we investigated the effects of immunization on glucose and insulin responses in LDL receptor-deficient mice fed regular or 60% sucrose diets, as well as in offspring fed 0.5% cholesterol or 60% sucrose diets. IR was assessed by fasting glucose and insulin levels, oral glucose tolerance tests, glucose clamps, pancreatic immunohistochemistry and plasma free fatty acid concentrations. Immunizations improved glucose responses in both genders and protected both immunized mice and their offspring against IR and type 2 diabetes. Protection occurred even under euglycemic conditions, but was greatest in obese males exposed to very obesogenic/diabetogenic conditions. Hyperinsulinemic euglycemic clamps confirmed that maternal immunization protected mainly by reducing IR, but pancreatic immunocytochemistry also indicated some protection against beta cell damage. Maternal immunization was associated with marked regulation in offspring of 4 genes relevant to diabetes and 19 genes of importance for oxidative stress, as well as increased hepatic activities of key antioxidant enzymes. These findings establish that targeted immunomodulation may be used to protect immunized subjects and their offspring against IR and type 2 diabetes, and thus to reduce cardiovascular risk. They also support the notion that in utero programming influences offspring disease not by a single mechanism, but by multiple systemic effects.

Mono-(2-ethylhexyl) Phthalate Induces Oxidative Stress and Inhibits Growth of Ovarian Antral Follicles.

Phthalates are synthetic plasticizers that are widely used in plastics and many other common consumer products. Exposure to these chemicals can cause developmental and reproductive toxicity. Our previous studies show that di-(2-ethylhexyl) phthalate (DEHP) inhibits follicle growth by inducing oxidative stress. In this study, we tested the hypothesis that mono-(2-ethylhexyl) phthalate (MEHP), the active metabolite of DEHP, also inhibits the growth of antral follicles through an oxidative stress pathway. Antral follicles were isolated from ovaries of adult CD-1 mice (age 30-35 days) and cultured with vehicle control (dimethylsulfoxide, DMSO) or MEHP (0.1-100μg/ml) ± 0.5-1mM N-acetyl cysteine (NAC, an antioxidant) for 96h. During culture, follicle size was evaluated daily as a measurement of follicle growth. At the end of the culture, follicles were collected and processed for in vitro reactive oxygen species (ROS) assays, which measure the presence of free radicals in the samples, an indicator of oxidative stress. The results indicate that MEHP suppresses the growth of follicles and increases ROS levels compared to DMSO controls, whereas NAC blocks the ability of MEHP to inhibit follicle growth and protects the follicles from MEHP-induced ROS levels. To further analyze the mechanism by which MEHP causes oxidative stress and inhibits follicle growth, we measured the expression and activity of various key antioxidant enzymes: Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX), and catalase (CAT). The results suggest that MEHP does not affect Cat expression and that it only decreases Sod1 and Gpx expression at MEHP 100 μg/ml, but not at lower doses. The results from enzyme activity assays indicate that MEHP inhibits GPX activity, but increases CAT activity at 0.1 and 10μg/ml. Interestingly, MEHP increases SOD1 activity at 10μg/ml, but inhibits SOD1 activity at 100μg/ml. Furthermore, NAC blocks the effects of MEHP (100μg/ml) on the expression and activity of SOD1 and GPX. Collectively, these data suggest that MEHP inhibits follicle growth by inducing production of ROS and by altering the expression and activity of key antioxidant enzymes. Supported by: NIHES019178 (JAF), Billie A Field Fellowships (WW and ZRC), and the Environmental Toxicology Scholar Program (MSB).

Impact of phosphine exposure on development in Caenorhabditis elegans: Involvement of oxidative stress and the role of glutathione

Phosphine (PH3) is a widely used and ideal fumigant employed to control insect pests in stored grains and other commodities. Chemically, PH3 is a strong reducing agent and mainly affects the mitochondrial transport system in insects. PH3 toxicity to insects is also associated with oxidative imbalances. Besides inducing mortality, PH3 is shown to delay hatching and adversely affect fecundity/fertility of several stored product insect pests. Recent studies have demonstrated the utility of Caenorhabditis elegans as a model organism to obtain basic insights on the toxic implications of PH3. In the present study, we have examined the impact of PH3 on the development of C. elegans and the involvement of glutathione (GSH) in its developmental toxicity. We exposed eggs of C. elegans to two concentrations of PH3 (0.04 and 0.06mg/L) either in the presence or absence of a GSH depleting agent, diethyl maleate (DEM, 5mM) for 72h. PH3 exposure caused significant delay in the post embryonic development among worms as characterized by the inability of hatched worms to attain gravid adult stage by the end of 72h. Interestingly, among worms co-exposed to DEM+PH3, the decrease in GSH levels was associated with more pronounced developmental delay compared to that of worms exposed to PH3per se. Concomitantly, PH3-induced depletion of glutathione was associated with significant alterations in activities of key antioxidant enzymes. Our data demonstrate the vital role of GSH and antioxidant defenses among worms developing under PH3 exposure. Interestingly, this finding also unfolds newer possibilities for developing strategies to disrupt antioxidant defenses in insect pests to enhance the efficacy of PH3 treatment for the control of stored product insects.

Hepatoprotective effects of antioxidants against non-target toxicity of the bio-insecticide spinosad in rats

The bio-insecticide spinosad (SPD) is increasingly being used in pest management programmes. In order to further assess its toxic effects to non-target species, male rats were exposed sub-chronically to SPD at a dose equivalent to 1/20 LD50 for four weeks. In order to assess the toxicity of SPD, parameters such as the activities of enzymes and concentrations of non-enzymatic antioxidant components, histopathological examination, DNA fragmentation and chromosomal aberrations in liver were analysed. Protection by an antioxidants mixture (AM), containing vitamin C, vitamin E and silymarin, against the effects of SPD was investigated. Exposure to SPD inhibited the activity of acetylcholinesterase, and depleted contents of reduced glutathione and malondialdehyde. SPD caused signification inhibition of activity of key antioxidant enzymes (GST, SOD) and induction in GPx. Treatment with AM attenuated all SPD-mediated effects. Histological examination of the liver revealed that SPD caused focal necrosis and degenerative changes in hepatocytes, along with cytoplasmic vacuolation. All of these lesions were significantly less in rats fed with AM. SPD accelerated formation of internucleosomal DNA fragmentation, which was attenuated by treatment with AM. Similarly, SPD caused significant structural chromosomal aberrations in bone marrow cells, the frequency of which was diminished by AM. Key words: Antioxidants, DNA damage, silymarin, spinosad, vitamin C, vitamin E.

Salinity-induced changes in key anti-oxidant enzyme activities and in the levels of some anti-oxidants, osmo-protectants, inorganic ions, and chlorophyll pigments in okra fruit (Abelmoschus esculentus L.)

SummaryTo study the influence of NaCl stress on various biochemical components in okra (Abelmoschus esculentus L.) fruit, two differential cultivars, ‘Nirali’ and ‘Posa Sawni’, were grown under non-saline and saline conditions until fruit development. Salt treatment (50 mM or 100 mM NaCl) significantly decreased fresh fruit yields (by 47.5 – 67.7%), soluble protein contents (by 62.3 – 138.4%), K+ ion levels (by 17.7 – 29.0%), and peroxidase (POD) activity (by 17.7 – 44.6%) compared to fruit from non-saline treated okra plants. NaCl increased Na+ ion levels (by 47.9 – 118.8%), glycine betaine contents (GB; by 34.3 – 115.2%), H2O2 contents (by 4.14 – 64.4%), and ascorbic acid levels (by 73.0 –109.0%) and superoxide dismutase (SOD) activity in the fruit of both okra cultivars. ‘Nirali’ had higher fresh fruit yields, K+ ion, GB, H2O2, and soluble protein contents, and higher SOD and POD activities, while ‘Posa Sawni’ fruit had higher ascorbic acid and total phenolic compounds contents and catalase (CAT) activity. Overall, the higher fresh yield of ‘Nirali’ could be attributed to enhanced levels of K+ ions, GB, soluble proteins, and SOD and POD activities. Fruit collected from all salt-stressed okra plants (‘Nirali’ and ‘Posa Sawni’) contained higher amounts of some key anti-oxidants and osmo-protectants which are considered beneficial for human health.

Salt stress induction of some key antioxidant enzymes and metabolites in eight Iranian wild almond species

The present work describes the changes in the activities of key antioxidant enzymes and the levels of some metabolites in relation to salt tolerance in eight wild almond species. All the species were exposed to four levels of NaCl (control, 40, 80 and 120 mM). Plant fresh biomass, α-, γ- and δ-tocopherol, total soluble proteins, malondialdehyde (MDAeq), H2O2, total phenolics, and the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) were analyzed in leaves of salt-stressed and non-stressed plants of the eight almond species. In all the species, salt stress significantly enhanced the activities of SOD and POD, levels of total phenolics and γ- and δ-tocopherols. High levels of salt stress significantly depressed the levels of total soluble proteins, MDA and CAT activity, while salt stress did not significantly affect leaf H2O2 contents. Regression analysis showed that the relationship between salt levels and total soluble proteins, CAT, γ-tocopherol, MDAeq, SOD and POD were statistically significant. Principal component analysis discriminated the almond species based on their degree of tolerance/sensitivity to saline conditions: Prunus reuteri and P. glauca were ranked as salt tolerant, P. lycioides and P. scoparia as moderately tolerant, and P. communis, P. eleagnifolia, P. arabica and P. orientalis as salt sensitive. The results could be used for selecting salt tolerant genotypes to be used as rootstocks for almond cultivation.

High-Dose Testosterone Propionate Treatment Reverses the Effects of Endurance Training on Myocardial Antioxidant Defenses in Adolescent Male Rats

This study was aimed at evaluation of changes in activities of selected antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase) and contents of key nonenzymatic antioxidants (glutathione, protein thiol groups, and α- and γ-tocopherols) in the left heart ventricle of young male Wistar rats subjected to endurance training (treadmill running, 1 h daily, 5 days a week, for 6 weeks) or/and testosterone propionate treatment (8 or 80 mg/kg body weight, intramuscularly, once a week, for 6 weeks) during adolescence. The training alone increased the activities of key antioxidant enzymes, but lowered the pool of nonenzymatic antioxidants and enhanced myocardial oxidative stress as evidenced by elevation of the lipid peroxidation biomarker malondialdehyde. The lower-dose testosterone treatment showed mixed effects on the individual components of the antioxidant defense system, but markedly enhanced lipid peroxidation. The higher-dose testosterone treatment decreased the activities of the antioxidant enzymes, lowered the contents of the nonenzymatic antioxidants, except for that of γ-tocopherol, reversed the effect of endurance training on the antioxidant enzymes activities, and enhanced lipid peroxidation more than the lower-dose treatment. These data demonstrate the potential risk to cardiac health from exogenous androgen use, either alone or in combination with endurance training, in adolescents.

Lack of PPARα exacerbates lipopolysaccharide-induced liver toxicity through STAT1 inflammatory signaling and increased oxidative/nitrosative stress

Peroxisome proliferator-activated receptor-α (PPARα) has been implicated in a potent anti-inflammatory activity. However, no information is available on whether PPARα can affect signal transducers and activator of transcription proteins (STATs) in acute liver damage. Thus, this study was aimed to investigate the in vivo role of PPARα in elevating STATs as well as oxidative/nitrosative stress in a model of lipopolysaccharide (LPS)-induced acute hepatic inflammatory injury. Using age-matched Ppara-null and wild-type (WT) mice, we demonstrate that the deletion of PPARα aggravates LPS-mediated liver injury through activating STAT1 and NF-κB-p65 accompanied by increased levels of pro-inflammatory cytokines. Furthermore, the activities of key anti-oxidant enzymes and mitochondrial complexes were significantly decreased while lipid peroxidation and protein nitration were elevated in LPS-exposed Ppara-null mice compared to WT. These results indicate that PPARα is important in preventing LPS-induced acute liver damage by regulating STAT1 inflammatory signaling pathways and oxidative/nitrosative stress.

Morphological, functional and gene expression analysis of the hyperoxic mouse retina

This study examined the impact of prolonged (up to 35 day) exposure to hyperoxia on the morphology and function of the retina, in the C57BL/6J mouse, as a basis for interpretation of gene expression changes. Mice of the C57BL/6J strain were raised from birth in dim cyclic illumination (12 h 5 lux, 12 h dark). Adult animals (90–110 days) were exposed to continuous hyperoxia (75% oxygen) for up to 35 d. Retinas were examined after 0 d (controls), 3 d, 7 d, 14 d and 35 d. Spatial and temporal patterns of photoreceptor death were mapped, using the TUNEL technique. Immunohistochemistry and a specific assay were used to assess the expression of a stress-related protein (GFAP) and the activity of key antioxidant enzymes (SOD). The dark-adapted flash electroretinogram was used to assess the function of rods and cones. RNA hybridized to Affymetrix Genechips was used to assess gene expression during the first 3 d of exposure. Photoreceptors were stable during the first 7 d exposure to hyperoxia, but thereafter showed progressive damage and degeneration, which began in a ‘hot-spot’ 0.5 mm inferior to the optic disc, then spread into surrounding retina. SOD activity was upregulated at 14 d, but not at earlier time points. GFAP expression was upregulated in Müller cells from 3 d. Rod and cone components of the ERG were supernormal at 3 d and 7 d, but then fell below control levels. Gene expression changes suggested possible mechanisms for this early supernormality of function. At 14 d exposure, damage to and death of photoreceptors were prominent and spreading, and function was correspondingly degraded. However at 3 d exposure, hyperoxia-induced supernormal functional responses in rods, while leaving their structure apparently undamaged. Variations in early (3 days) gene expression provide a partial insight into the mechanisms involved in this.

Cold defence responses the role of oxidative stress

Low temperatures provoke increased production of heat accompanied by increased respiration, oxygen consumption and the production of partially reduced oxygen species called ROS. ROS induce different forms of cellular oxidative damage, disturb the redox state and can change the activity of several metabolic enzymes. Organisms have developed a functionally connected set of anti-oxidant enzymes and low molecular mass compounds (together termed the ADS) that metabolise primary ROS. If ROS production within cells overwhelms the ADS, oxidative damage arises and oxidative stress can occur. Short-term cold exposure in endotherms leads to oxidative stress. As cold exposure persists organisms develop adaptive changes toward reducing ROS production and increasing the ADS. In contrast, heterotherms and ectotherms as a normal part of their over-wintering strategy slow down metabolism, oxygen consumption and subsequently cause ROS production. Increased baseline activity of key anti-oxidant enzymes as well as 'secondary' enzymatic defence and/or glutathione levels in preparation for a putative oxidative stressful situation arising from tissue re-oxygenation seems to be the preferred evolutionary adaptation of such animals exposed to low environmental temperatures.

Long-Term Resveratrol Supplementation Mediates Oxidative Stress, but not Sarcopenia Aged Mice

Increases in oxidative stress can have detrimental effects on both muscle mass and force generation. Aging is associated with increases in oxidative stress and a concomitant decrease in muscle size and strength known as sarcopenia. Resveratrol is a naturally occurring polyphenol that has been shown to have both anti-oxidant and anti-aging properties. PURPOSE: To analyze the capacity of resveratrol to protect against aging-induced oxidative stress and subsequent muscle atrophy. METHODS: Middle-aged (18mo) C57/Bl6 mice were randomly assigned to receive either a control diet, or a diet supplemented with.05% trans-resveratrol for 10 months. Additionally, young (6mo) and middle-aged (18mo) mice were used as controls. Antioxidant capacities of muscles were estimated by measuring the activities of key antioxidant enzymes in conjunction with H2O2 levels. Oxidative damage was assessed by analyzing the downstream targets of pro-oxidants, including lipid peroxidation and protein carbonyl formation. RESULTS: Aging resulted in a significant decrease in gastrocnemius muscle wet weight (∼20%). Resveratrol supplementation was unable to attenuate this loss. The activity of CuZnSOD was significantly increased in aged animals compared to both young and middle-aged animals. Resveratrol supplemented animals showed a 34% decrease in CuZnSOD activity compared to non-supplemented animals. There was no change in CuZnSOD protein content within any group. There was a 28% increase in MnSOD activity in aged non-supplemented animals when compared to young, however; MnSOD activity increased by 47% in resveratrol supplemented animals. H2O2 levels were elevated in both middle-aged and aged mice, compared to their young counterparts. Resveratrol supplementation significantly reduced H2O2 levels. CONCLUSIONS: Resveratrol has a protective effect against aging-induced oxidative stress in skeletal muscle, likely through the upregulation of MnSOD activity and an overall decrease in oxidative stress. Resveratrol may be an effective therapeutic agent to improve the redox status of aged skeletal muscle and therefore allow for improved adaptation and recovery following chronic illness and/or injuries during which the capacity of the endogenous antioxidant system may be overwhelmed in aged individuals. Funded by: R01AG021530

NF‐E2 related factor 2 activation induction by decursin and decursinol angelate protect against bete‐amyloid‐peptide‐induced oxidative stress in PC12 cells

The protective effects of decursin(D) and decursinol angelate(DA) purified from Angelica gigas Nakai on beta‐amyloid‐peptide(Aβ)‐induced neurotoxicity and the mechanism was investigated. Aβ is a pathological hallmark of Alzheimer's disease(AD) which can initiate a cascade of oxidative events. It decreased viability of PC12 cells in a concentration‐ and time‐dependent manner. Pretreatment with D and DA resulted in dose‐dependent anti‐Aβ toxicity according to MTT assay and restored the activities of key antioxidant enzymes(superoxide dismutase, glutathione peroxidase, catalase), glutathione(GSH) level and lipid peroxidation altered by Aβ. Furthermore D and DA resulted in up‐regulation of GST activity accompanied by increased transactivation of activator NF‐E2‐related factor‐2(Nrf2) which mediates GST gene transcription. Our results demonstrate activation of Nrf2/ARE pathways in PC12 cells by D and DA treatment protects cells from Aβ‐induced oxidative stress and regulated Aβ‐mediated adaptive responses in PC12 cells via translocation of Nrf2. These findings indicated D and DA might be considered as an adjunct therapeutic strategy to combat neural demise in AD and other oxidative stress‐related disease.This research is supported by Biohealth Products Research Center (BPRC) of Inje University.

Antitumor Properties and Modulation of Antioxidant Enzymes Activity by Aloe vera Leaf Active Principles Isolated via Supercritical Carbon Dioxide Extraction

The aim of this study was to evaluate the potential anticancer properties and modulatory effect of selected Aloe vera (A. vera) active principles on antioxidant enzyme activities. Thus, three anthraquinones (Namely: aloesin, aloe-emodin and barbaloin) were extracted from A. vera leaves by supercritical fluid extraction and subsequently purified by high performance liquid chromatography. Additionally, the N-terminal octapeptide derived from verectin, a biologically active 14 kDa glycoprotein present in A. vera, was also tested. In vivo, active principles exhibited significant prolongation of the life span of tumor-transplanted animals in the following order: barbaloin> octapeptide> aloesin > aloe-emodin. A. vera active principles exhibited significant inhibition on Ehrlich ascite carcinoma cell (EACC) number, when compared to positive control group, in the following order: barbaloin> aloe-emodin > octapeptide > aloesin. Moreover, in trypan blue cell viability assay, active principles showed a significant concentration-dependent cytotoxicity against acute myeloid leukemia (AML) and acute lymphocytes leukemia (ALL) cancerous cells. Furthermore, in MTT cell viability test, aloe-emodin was found to be active against two human colon cancer cell lines (i.e. DLD-1 and HT2), with IC(50) values of 8.94 and 10.78 microM, respectively. Treatments of human AML leukemic cells with active principles (100 microg ml(-1)) resulted in varying intensities of internucleosomal DNA fragmentation, hallmark of cells undergoing apoptosis, in the following order: aloe-emodin> aloesin> barbaloin> octapeptide. Intererstingly, treatment of EACC tumors with active principles resulted in a significant elevation activity of key antioxidant enzymes (SOD, GST, tGPx, and LDH). Our data suggest that the tested A. vera compounds may exert their chemo-preventive effect through modulating antioxidant and detoxification enzyme activity levels, as they are one of the indicators of tumorigenesis. These findings are discussed in the light of the potential of A. vera plant extracts for developing efficient, specific and non-toxic anticancer drugs that are affordable for developing countries.

Biochemical and physiological changes in jack bean under mycorrhizal symbiosis growing in soil with increasing Cu concentrations

The influence of jack bean [ Canavalia ensiformis (L.) D.C.] mycorrhization, by the arbuscular mycorrhizal (AM) fungus Glomus etunicatum, in response to increasing Cu concentrations in soil (0, 50, 150 and 450 mg dm −3) was studied. In the highest Cu dose, mycorrhiza decreased Cu concentrations in plant organs and promoted biomass accumulation. In addition, mycorrhizal colonization was not affected by Cu suggesting certain tolerance of the inoculated AM fungus to this metal. Cu-induced proline accumulation and an increase in soluble amino acid contents in leaves, with higher proline contents in AM plants. Marked differences in foliar soluble amino acid composition were also observed in response to Cu in the soil. The activity of key antioxidant enzymes increased mainly in the leaves of non-AM plants, indicating increased production of reactive oxygen species. However, changes in malondialdehyde content were not observed suggesting an effective antioxidant system. Independent of mycorrhization, glutathione content in leaves decreased as Cu increased in the soil. Phytochelatins were detected in the leaves of both Cu treated and untreated plants, and quantitative and qualitative differences were observed due to mycorrhization. In summary, the AM fungus was able to maintain an efficient symbiosis with jack bean plants in soil containing high Cu concentrations. A combination of physiological and nutritional changes caused by the symbiotic association that might be of interest for phytoremediation strategies in Cu-contaminated soils.

Trehalose: Protector of antioxidant enzymes or reactive oxygen species scavenger under heat stress?

Trehalose is known to protect membranes and macromolecules. Its accumulation has been implicated in allowing plants to tolerate stress, including heat-shock. However, under heat-shock, it is not clear whether trehalose eliminates reactive oxygen species (ROS) directly or indirectly by protecting antioxidant enzymes. In this study, we initially examined the effects of trehalose on the activities of key antioxidant enzymes, including superoxide dismutases (SODs), ascorbate catalases (CATs), and ascorbate peroxidases (APX) from wheat ( Triticum aestivum L.), and then measured the ability of trehalose to scavenge hydrogen peroxide (H 2O 2) and superoxide anions (O 2 − ). Our results indicated that trehalose protected SOD activity slightly. However, it inhibited CAT and APX activities under heat stress, with a little protection of CAT activity (only about 7% promotion) at 22 °C. Moreover, trehalose scavenged H 2O 2 and O 2 − greatly in a concentration-dependent manner, reaching the maximal scavenging H 2O 2 rate of 95% and O 2 − rate of 78%, respectively, at 50 mM trehalose. These results suggest that trehalose plays a direct role in eliminating H 2O 2 and O 2 − in wheat under heat stress.

Study of the Synergistic Effect on Hepatoma 22 Tumor Cells by Focused Ultrasound Activation of Hematoporphyrin

The synergistic effect of ultrasound and drugs on tumor cells is known as sonodynamic therapy (SDT). The purpose of this study was to evaluate the effects of SDT on lipid peroxidation and the activity of antioxidative enzymes in isolated hepatoma 22 (H-22) cells to better understand the bioeffects of SDT. The viability of cells was evaluated by the Trypan blue dye exclusion test. The morphologic changes of H-22 cells were observed by a scanning electron microscope immediately after treatment. The intracellular reactive oxygen species levels were detected by 2',7'-dichlorofluorescein diacetate. Colorimetry and enzymatic chemical methods were used to measure the lipid peroxidation levels and activities of key antioxidant enzymes (ie, superoxide dismutase, selenium-dependent glutathione peroxidase, and catalase) in H-22 tumor cells. Our experiments indicated that the ultrasonically induced cell damage rate was increased with 100-microg/mL hematoporphyrin, whereas no cell damage was observed with hematoporphyrin alone. Generation of reactive oxygen species in cell suspensions after SDT treatment was remarkably higher than in controls. The malondialdehyde content was remarkably enhanced, and antioxidative enzyme activities were obviously decreased compared with controls. This study suggests that oxygen free radicals may play an important role in improving membrane lipid peroxidation and decreasing the activities of key antioxidant enzymes in cells. It was speculated that this biological mechanism might be involved in mediating the killing effect of H-22 cells in SDT.

Seasonal changes in photoprotective mechanisms of leaves from shaded and unshaded field-grown coffee (Coffea arabica L.) trees

Coffee is native to shady environments, but often grows and yields better without shade. Thus, it may be reasoned that coffee leaves should display enough plasticity to acclimate themselves to contrasting light environments. However, little is known about mechanisms associated with such plasticity in coffee. This work aimed, therefore, to explore differences in leaf photoprotective mechanisms. Plants were grown in the field and received either 48 or 100% natural light. Evaluations were made using outer leaves from the sun-facing sides of the coffee hedgerow in Viçosa (Brazil) in August and October, when growth and photosynthetic rates are expected to be minimal and maximal, respectively, and in December, when temporary depressions in those variables are common. Regardless of light treatments, coffee leaves showed: (1) very low photosynthetic rates (generally below 2.5 μmol m−2 s−1), (2) chronic photoinhibition in August (dry, cool season) that was accompanied by strong loss of pigment concentration, and (3) discrete, dynamic photoinhibition in October and December (rainy, warm season). Compared with shaded leaves, sunlit leaves generally exhibited lower pigment concentration, lower quantum yield of electron transport, steeper inclinations and similar electron transport rate. Total ascorbate pool tended to be larger in sunlit than in shaded leaves (but with similar redox state), whereas activities of key antioxidant enzymes, as well as malondialdehyde accumulation and electrolyte leakage, were similar between those leaf types. As a whole, the photosynthetic apparatus of the coffee tree showed a low phenotypic plasticity to varying irradiance.

Study of cell killing effect on S180 by ultrasound activating protoporphyrin IX

The present study was initiated to investigate the potential biological mechanism of cell killing effect on isolate sarcoma 180 (S180) cells induced by ultrasound activating protoporphyrin IX (PPIX). S180 cells were exposed to ultrasound for 30s duration, at a frequency of 2.2MHz and an acoustic power of 3W/cm2 in the presence of 120μM PPIX. The viability of cells was evaluated using trypan blue staining. The generation of oxygen free radicals in cell suspensions was detected immediately after treatment using a reactive oxygen detection kit. A copper reagent colorimetry method was used to measure the level of FFAs released into cell suspensions by the process of cell damage induced by ultrasound and PPIX treatment. Oxidative stress was assessed by measuring the activities of key antioxidant enzymes (i.e., SOD, CAT, GSH-PX) in S180 tumor cells. Treatment with ultrasound and PPIX together increased the cell damage rate to 50.91%, while treatment with ultrasound alone gave a cell damage rate to 24.24%, and PPIX alone kept this rate unchanged. Colorimetry and enzymatic chemical methods showed that the level of FFAs in cell suspension increased significantly after the treatment, while the activity of all the above enzymes decreased in tumor cells at different levels, and were associated with the generation of oxygen free radicals in cell suspension after treatment. The results indicate that oxygen free radicals may play an important role in improving the membrane lipid peroxidation, degrading membrane phospholipids to release FFAs, and decreasing the activities of the key antioxidant enzymes in cells. This biological mechanism might be involved in mediating the effects on S180 cells and resulting in the cell damage seen with SDT.

CURCUMIN, THE ACTIVE PRINCIPLE OF TURMERIC (CURCUMA LONGA), AMELIORATES DIABETIC NEPHROPATHY IN RATS

Chronic hyperglycaemia in diabetes leads to the overproduction of free radicals and evidence is increasing that these contribute to the development of diabetic nephropathy. Among the spices, turmeric (Curcuma longa) is used as a flavouring and colouring agent in the indian diet every day and is known to possess anti-oxidant properties. The present study was designed to examine the effect of curcumin, a yellow pigment of turmeric, on renal function and oxidative stress in streptozotocin (STZ)-induced diabetic rats. Diabetes was induced by a single intraperitoneal injection of STZ (65 mg/kg) in rats. Four weeks after STZ injection, rats were divided into four groups, namely control rats, diabetic rats and diabetic rats treated with curcumin (15 and 30 mg/kg, p.o.) for 2 weeks. Renal function was assessed by creatinine, blood urea nitrogen, creatinine and urea clearance and urine albumin excretion. Oxidative stress was measured by renal malonaldehyde, reduced glutathione and the anti-oxidant enzymes superoxide dismutase and catalase. Streptozotocin-injected rats showed significant increases in blood glucose, polyuria and a decrease in bodyweight compared with age-matched control rats. After 6 weeks, diabetic rats also exhibited renal dysfunction, as evidenced by reduced creatinine and urea clearance and proteinuria, along with a marked increase in oxidative stress, as determined by lipid peroxidation and activities of key anti-oxidant enzymes. Chronic treatment with curcumin significantly attenuated both renal dysfunction and oxidative stress in diabetic rats. These results provide confirmatory evidence of oxidative stress in diabetic nephropathy and point towards the possible anti-oxidative mechanism being responsible for the nephroprotective action of curcumin.