Burkholderia cenocepacia is a gram-negative, non-spore-forming bacillus and a member of the Burkholderia cepacia complex. B. cenocepacia can survive intracellularly in phagocytic cells and can produce at least one superoxide dismutase (<em>SOD</em>). The inability of O2- to cross the cytoplasmic membrane, coupled with the periplasmic location of Cu,Zn<em>SODs</em>, suggests that periplasmic <em>SODs</em> protect bacteria from superoxide that has an exogenous origin (for example, when cells are faced with reactive oxygen intermediates generated by host cells in response to infection). In this study, we identified the sodC gene encoding a Cu,Zn<em>SOD</em> in B. cenocepacia and demonstrated that a sodC null mutant was not sensitive to a H2O2, <em>3</em>-morpholinosydnonimine, or paraquat challenge but was killed by exogenous superoxide generated by the xanthine/xanthine oxidase method. The sodC mutant also exhibited a growth defect in liquid medium compared to the parental strain, which could be complemented in trans. The mutant was killed more rapidly than the parental strain was killed in murine macrophage-like cell line RAW 264.7, but killing was eliminated when macrophages were treated with an NADPH oxidase inhibitor. We also confirmed that SodC is periplasmic and identified the metal cofactor. B. cenocepacia SodC was resistant to inhibition by H2O2 and was unusually resistant to KCN for a Cu,Zn<em>SOD</em>. Together, these observations establish that B. cenocepacia produces a periplasmic Cu,Zn<em>SOD</em> that protects this bacterium from exogenously generated O2- and contributes to intracellular survival of this bacterium in macrophages.

OBJECTIVE

Neutrophil accumulation contributes to the pathogenesis of rheumatoid arthritis. This study was undertaken to examine the ability of H2O2 to influence neutrophilic inflammation in a model of antigen-induced arthritis (AIA) in mice.

METHODS

AIA was induced by administration of antigen into the knee joints of previously immunized mice. Neutrophil accumulation was measured by counting neutrophils in the synovial cavity and assaying myeloperoxidase activity in the tissue surrounding the mouse knee joint. Apoptosis was determined by morphologic and molecular techniques. The role of H2O2 was studied using mice that do not produce reactive oxygen species (gp91phox-/- mice) and drugs that enhance the generation or enhance the degradation of H2O2.

RESULTS

Antigen challenge of immunized mice induced neutrophil accumulation that peaked at 12-24 hours after challenge. H2O2 production peaked at 24 hours, after which time, the inflammation resolved. Neutrophil recruitment was similar in wild-type and gp91phox-/- mice, but there was delayed resolution in gp91phox-/- mice or after administration of catalase. In contrast, administration of H2O2 or superoxide dismutase (<em>SOD</em>) resolved neutrophilic inflammation. The resolution of inflammation induced by <em>SOD</em> or H2O2 was accompanied by an increase in the number of apoptotic neutrophils. Apoptosis was associated with an increase in Bax and caspase <em>3</em> cleavage and was secondary to phosphatidylinositol <em>3</em>-kinase (PI<em>3</em>K)/Akt activation.

CONCLUSIONS

Our findings indicate that levels of H2O2 increase during neutrophil influx and are necessary for the natural resolution of neutrophilic inflammation. Mechanistically, enhanced levels of H2O2 (endogenous or exogenous) inhibit p-Akt/NF-κB and induce apoptosis of migrated neutrophils. Modulation of H2O2 production may represent a novel strategy for controlling neutrophilic inflammation in the joints.

OBJECTIVE

Rheumatoid arthritis (RA) is a systemic autoimmune inflammatory disease involving the breakdown of cartilage and juxta-articular bone, which is often accompanied by decreased bone mineral density (BMD) and increased risk of fracture. Anti-inflammatory omega-<em>3</em> fatty acids may prevent arthritis and bone loss in MRL/lpr mice model of arthritis and in humans.

METHODS

In this study, the effect of long term feeding of 10% dietary n-<em>3</em> (fish oil (FO)) and n-6 (corn oil (CO)) fatty acids begun at 6 weeks of age on bone mineral density (BMD) in different bone regions in an MRL/lpr female mouse model of RA was measured at 6, 9, and 12 months of age by dual energy x-ray absorptiometry (DEXA). After sacrificing the mice at 12 months of age, antioxidant enzyme activities were measured in spleen, mRNA for receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG) was measured by RT-PCR in lymph nodes, and synovitis was measured in leg joints.

RESULTS

At 6, 9 and 12 months of age, BMD was significantly higher (p < 0.05) in distal femur, proximal tibia, and lumbar spine of FO fed mice than those of CO fed mice. Spleen catalase (CAT) and superoxide dismutase (SOD) activities were also significantly higher (p < 0.01) in FO fed mice than in CO fed mice. Histology of knee joints revealed mild synovitis in CO fed mice, which was not present in FO fed mice. RT-PCR analysis of lymph nodes revealed decreased RANKL mRNA (p < 0.001) expression and enhanced OPG mRNA expression (p < 0.01) in FO fed mice compared to CO fed mice.

CONCLUSIONS

These results suggest beneficial effects of long-term FO feeding in maintaining higher BMD and lower synovitis in this mouse model. These beneficial effects may be due, in part, to increased activity of antioxidant enzymes, decreased expression of RANKL, and increased expression of OPG in FO fed mice thereby altering the RANKL/OPG ratio. These significant beneficial effects on BMD suggest that FO may serve as an effective dietary supplement to prevent BMD loss in patients with RA.

Reactive oxygen species (ROS) are crucial in long-term diabetes complications, including peripheral artery disease (PAD). In this study, we have investigated the potential clinical impact of unacylated ghrelin (UnAG) in a glucose intolerance and PAD mouse model. We demonstrate that UnAG is able to protect skeletal muscle and endothelial cells (ECs) from ROS imbalance in hind limb ischemia-subjected ob/ob mice. This effect translates into reductions in hind limb functional impairment. We show that UnAG rescues sirtuin 1 (SIRT1) activity and superoxide dismutase-2 (<em>SOD</em>-2) expression in ECs. This leads to SIRT1-mediated p5<em>3</em> and histone <em>3</em> lysate 56 deacetylation and results in reduced EC senescence in vivo. We demonstrate, using small interfering RNA technology, that SIRT1 is also crucial for <em>SOD</em>-2 expression. UnAG also renews micro-RNA (miR)-126 expression, resulting in the posttranscriptional regulation of vascular cell adhesion molecule 1 expression and a reduced number of infiltrating inflammatory cells in vivo. Loss-of-function experiments that target miR-126 demonstrate that miR-126 also controls SIRT1 and <em>SOD</em>-2 expression, thus confirming its role in driving UnAG-mediated EC protection against ROS imbalance. These results indicate that UnAG protects vessels from ROS imbalance in ob/ob mice by rescuing miR-126 expression, thus emphasizing its potential clinical impact in avoiding limb loss in PAD.

The second member of the transient receptor potential-melastatin channel family (TRPM2) is expressed in the heart and vasculature. TRPM2 channels were expressed in the sarcolemma and transverse tubules of adult left ventricular (LV) myocytes. Cardiac TRPM2 channels were functional since activation with H2O2 resulted in Ca(2+) influx that was dependent on extracellular Ca(2+), was significantly higher in wild-type (WT) myocytes compared with TRPM2 knockout (KO) myocytes, and inhibited by clotrimazole in WT myocytes. At rest, there were no differences in LV mass, heart rate, fractional shortening, and +dP/dt between WT and KO hearts. At 2-<em>3</em> days after ischemia-reperfusion (I/R), despite similar areas at risk and infarct sizes, KO hearts had lower fractional shortening and +dP/dt compared with WT hearts. Compared with WT I/R myocytes, expression of the Na(+)/Ca(2+) exchanger (NCX1) and NCX1 current were increased, expression of the α1-subunit of Na(+)-K(+)-ATPase and Na(+) pump current were decreased, and action potential duration was prolonged in KO I/R myocytes. Post-I/R, intracellular Ca(2+) concentration transients and contraction amplitudes were equally depressed in WT and KO myocytes. After 2 h of hypoxia followed by <em>3</em>0 min of reoxygenation, levels of ROS were significantly higher in KO compared with WT LV myocytes. Compared with WT I/R hearts, oxygen radical scavenging enzymes (<em>SODs</em>) and their upstream regulators (forkhead box transcription factors and hypoxia-inducible factor) were lower, whereas NADPH oxidase was higher, in KO I/R hearts. We conclude that TRPM2 channels protected hearts from I/R injury by decreasing generation and enhancing scavenging of ROS, thereby reducing I/R-induced oxidative stress.

1-Methylnicotinamide (MNA) is one of the major derivatives of nicotinamide, which was recently shown to exhibit antithrombotic and antiinflammatory actions. However, it is not yet known whether MNA affects gastric mucosal defense. The effects of exogenous MNA were studied on gastric secretion and gastric lesions induced in rats by <em>3</em>.5 h of water immersion and water restraint stress (WRS) or in rats administered 75% ethanol. MNA [6.25-100 mg/kg intragastrically (i.g.)] led to a dose-dependent rise in the plasma MNA level, inhibited gastric acid secretion, and attenuated these gastric lesions induced by WRS or ethanol. The gastroprotective effect of MNA was accompanied by an increase in the gastric mucosal blood flow and plasma calcitonin gene-related peptide (CGRP) levels, the preservation of prostacyclin (PGI(2)) generation (measured as 6-keto-PGF1alpha), and an overexpression of mRNAs for cyclooxygenase (COX)-2 and CGRP in the gastric mucosa. R-<em>3</em>-(4-Fluoro-phenyl)-2-[5-(4-fluoro-phenyl)-benzofuran-2-ylmethoxycarbonylamino]-propionic acid (RO <em>3</em>24479), which is the selective antagonist of IP/PGI(2) receptors, reversed the effects of MNA on gastric lesions and GBF. MNA-induced gastroprotection was attenuated by suppression of COX-1 [5-(4-chlorophenyl)-1-(4-methoxyphenyl)-<em>3</em>-(trifluoromethyl)-1H-pyrazole; SC-560] and COX-2 [4-(4-methylsulfonylphenyl)-<em>3</em>-phenyl-5H-furan-2-one; rofecoxib] activity, capsaicin denervation, and by the pretreatment with CGRP(8-<em>3</em>7) or capsazepine. Addition of exogenous PGI(2) or CGRP restored the MNA-induced gastroprotection in rats treated with COX-1 and COX-2 inhibitors or in those with capsaicin denervation. WRS enhanced MDA content while decreasing superoxide dismutase (<em>SOD</em>) activity in the gastric mucosa, but pretreatment with MNA reversed these changes. MNA exerts potent gastroprotection against WRS damage via mechanisms involving cooperative action of PGI(2) and CGRP in preservation of microvascular flow, antioxidizing enzyme <em>SOD</em> activity, and reduction in lipid peroxidation.

Drought is a world-spread problem seriously influencing grain production and quality, the loss of which is the total for other natural disasters, with increasing global climate change making the situation more serious. Wheat is the staple food for more than <em>3</em>5% of world population, so wheat anti-drought physiology study is of importance to wheat production and biological breeding for the sake of coping with abiotic and biotic conditions. Much research is involved in this hot topic, but the pace of progress is not so large because of drought resistance being a multiple-gene-control quantitative character and wheat genome being larger (16,000Mb). On the other hand, stress adaptive mechanisms are quite different, with stress degree, time course, materials, soil quality status and experimental plots, thus increasing the complexity of the issue in question. Additionally, a little study is related to the whole life circle of wheat, which cannot provide a comprehensive understanding of its anti-drought machinery. We selected 10 kinds of wheat genotypes as materials, which have potential to be applied in practice, and measured change of relative physiological indices through wheat whole growing-developmental circle (i.e. seedling, tillering and maturing). Here, we reported the anti-oxidative results of maturation stage (the results of seedling and tillering stage have been published) in terms of activities of POD, <em>SOD</em>, CAT and MDA content as follows: (1) 10 wheat genotypes can be grouped into three kinds (A-C, respectively) according to their changing trend of the measured indices; (2) A group performed better resistance drought under the condition of treatment level 1 (appropriate level), whose activities of anti-oxidative enzymes (POD, <em>SOD</em>, CAT) were higher and MDA lower; (<em>3</em>) B group exhibited stronger anti-drought under treatment level 2 (light-stress level), whose activities of anti-oxidative enzymes were higher and MDA lower; (4) C group expressed anti-drought to some extent under treatment level <em>3</em> (serious-stress level), whose activities of anti-oxidative enzymes were stronger, MDA lower; (5) these results demonstrated that different wheat genotypes have different physiological mechanisms to adapt themselves to changing drought stress, whose molecular basis is discrete gene expression profiling (transcriptom); (6) our results also showed that the concept and method accepted and adopted by most researchers [T.C. Hsiao, Plant response to water stress, Ann. Rev. Plant Physiol. 24 (197<em>3</em>) 519-570], that 75% FC is a proper supply for higher plants, was doubted, because this level could not reflect the true suitable level of different wheat genotypes. The study in this respect is the key to wheat anti-drought and biological-saving water agriculture; (7) our research can provide insights into physiological mechanisms of crop anti-drought and direct practical materials for wheat anti-drought breeding; (8) the physiological study of wheat is more urgent up-to-date and molecular aspects are needed, but cannot substitute this important part. The combination of both is an important strategy and a key and (9) POD, <em>SOD</em> and CAT activities and MDA content of different wheat genotypes had quite different changing trend at different stages and under different soil water stress conditions, which was linked with their origin of cultivation and individual soil water threshold.

Glutathione reductase (GR; EC 1.6.4.2) and superoxide dismutase (<em>SOD</em>; EC 1.15.1.1) are two well-known enzymes involved in the scavenging of reactive oxygen intermediates. However, little is known about the regulation of Gor and <em>Sod</em> genes in plant cells. To obtain information about hypothetical redox regulatory mechanisms controlling Gor and <em>Sod</em> gene expression we artificially enhanced the levels of reduced and oxidized forms of glutathione (GSH and GSSG) in Pinus sylvestris L. needles. Scots pine shoots were placed for 12 h in beakers containing 5 mM GSH, 5 mM GSSG or water. Increased levels of both GSSG and GSH were observed in the GSSG-treated needles after <em>3</em> h. In contrast, only the GSH level was increased by the GSH treatment. Thus, the GSH/GSSG ratio increased up to 15-fold during the GSH treatment and decreased approximately two-fold during the GSSG treatment. The GR activity was significantly higher (60%) when GSSG was applied, without any apparent change in the amount and isoform population of GR or accumulation of Gor gene transcripts. This indicates that the GR activity increased per se in the GSSG treatment. The level of cytosolic CuZn-<em>Sod</em> transcripts was decreased significantly by the GSH treatment without any change in enzyme activity. The chloroplastic CuZn-<em>Sod</em> gene generally showed a more stable transcript level in the different treatments. However, a similarity between the cytosolic and chloroplastic levels of CuZn-<em>Sod</em> transcripts could be observed in different treatments. This suggests that the redox state of glutathione plays an important role in the in vivo regulation of CuZn-<em>Sod</em> gene expression in plants.

We have shown that human spermatozoa generate and release reactive oxygen species that can be detected by chemiluminescence techniques. Analysis of the cellular mechanisms responsible for this activity suggests that the probe, luminol, undergoes an intracellular dioxygenation reaction mediated by hydrogen peroxide and a sperm peroxidase located within the acrosome. Support for this model included the following observations: (1) the luminol-dependent signal could be suppressed with peroxidase inhibitors, phenylhydrazine and sodium azide; (2) this suppression could be reversed by the addition of an azide-insensitive peroxidase, horse radish peroxidase (HRP); (<em>3</em>) inhibition of intracellular superoxide dismutase (<em>SOD</em>) with potassium cyanide (KCN) suppressed the luminol signal; (4) peroxidase activity could be detected in purified populations of human spermatozoa with <em>3</em>,<em>3</em>',5,5' tetramethylbenzidine (TMB); (5) this peroxidase was active at the pH prevailing within the acrosomal vesicle; and (6) peroxidase activity and luminol-dependent chemiluminescence were minimal in spermatozoa exhibiting a congenital absence of acrosomes. Human spermatozoa could also generate lucigenin-dependent chemiluminescent signals that could neither be suppressed with peroxidase inhibitors nor enhanced by the addition of peroxidase. However, these signals could be enhanced by suppression of intracellular <em>SOD</em> with KCN or inhibited by exogenous <em>SOD</em>, suggesting that lucigenin was responding to superoxide anion released into the extracellular space. The ability of chemiluminescent techniques to detect and discriminate the production of superoxide and hydrogen peroxide by spermatozoa should facilitate the further analysis of reactive oxygen species as mediators of normal and abnormal human sperm function.

To find effective antiprion compounds, we synthesized and evaluated various pyrazolone derivatives. Seven of 19 compounds showed inhibition of PrP-res accumulation and the remarkably active compound 1<em>3</em> showed an IC50 value of <em>3</em> nM in both ScN2a and F<em>3</em> cell lines. Findings from studies on physicochemical and biochemical properties suggest that the action mechanism of these compounds does not correlate with any antioxidant activities, any of hydroxyl radical scavenging activities, or any <em>SOD</em>-like activities.

Nicotine affects a variety of cellular processes ranging from induction of gene expression to secretion of hormones and modulation of enzymatic activities. The objective of this study was to characterize the toxicity of nicotine enantiomers as well as their ability to induce oxidative stress in an in vitro model using Chinese hamster ovary (CHO) cells. Colony formation assay has demonstrated that (-)-nicotine is the more toxic of the enantiomers. At 6 mM concentrations, (-)-nicotine was found to be approximately 28- and 19-fold more potent than (+)-, and (+/-)-nicotine (racemic), respectively. Results also indicated that the toxicity of (+/-)-nicotine is higher than that of (+)-nicotine. (-)-Nicotine at a 10 mM concentration substantially decreased glutathione (GSH) levels (46% decrease). In addition, a <em>3</em>-fold increase in malondialdehyde (MDA) level was evident in cells after exposure to 10 mM (-)-nicotine. Increased lactate dehydrogenase (LDH) activities in the media demonstrated that cellular membrane integrity was disturbed in nicotine treated cells. In the presence of superoxide dismutase (<em>SOD</em>) and catalase (CAT), the LDH activities returned to control value in 24 h with all concentrations of (-)-, (+)-, and (+/-)-nicotine. The decreases in LDH activities in the presence of the radical scavenging enzymes <em>SOD</em> and CAT suggest that membrane damage may be due to free radical generation.

Studies of the photosensitized oxidation have demonstrated that photodynamic oxidation of methionine is mediated by singlet oxygen ((1)O(2)). In this study, we demonstrated that phagocytosing human polymorphonuclear leukocytes (PMN), but not resting PMN, oxidized both intracellular and extracellular methionine to methionine sulfoxide. N-ethylmaleimide, which inhibits phagocytosis and cellular metabolism, inhibited the oxidation of methionine. Neutrophils from patients with chronic granulomatous disease did not oxidize methionine even in the presence of phagocytosis. The oxidation of methionine by phagocytosing normal PMN was inhibited by (1)O(2) quenchers, (1.4-diazabicyclo-[2,2,2]-octane, tryptophan, NaN(<em>3</em>)), myeloperoxidase (MPO) inhibitors (NaN(<em>3</em>), KCN) and catalase. In contrast, superoxide dismutase, ethanol, and mannitol had no effect. Furthermore, (1)O(2) quenchers did not interfere with the production of superoxide (O(2) (-)) by phagocytosing PMN. The combination of catalase and <em>SOD</em> did not enhance the inhibition of methionine by phagocytosing PMN. On the other hand, deuterium oxide stimulated the oxidation of methionine by PMN almost 200%.H(2)O(2) at high concentrations oxidized methionine to methionine sulfoxide. However, when similar amounts of H(2)O(2) were added to human PMN, they did not oxidize methionine. In contrast, when H(2)O(2), at concentrations too low to oxidize methionine by itself, was added to the granular fraction, but not the soluble fraction, they oxidized methionine to methionine sulfoxide. The oxidation of methionine by the combination of H(2)O(2) and granular fractions was inhibited by (1)O(2) quenchers and MPO inhibitors, but it was stimulated by deuterium oxide. Removal of chloride anion also prevented the oxidation of methionine by the granular fractions. Our results suggest that the oxidation of methionine by phagocytosing PMN is dependent on the MPO-mediated antimicrobial system (MPO-H(2)O(2)-Cl(-)). They also suggest, but do not prove that the oxidation of methionine is mediated by (1)O(2). Oxidation of methionine may be one of the mechanisms that human PMN damage microorganisms.

Several epidemiological studies suggest that a diet rich in fruits and vegetables, which contain high levels of polyphenols, is associated with a reduced risk of cancer. The aim of the present study was to determine whether a red wine polyphenolic extract (RWPs, a rich source of polyphenols; 2.9g/L) affects the proliferation of human lymphoblastic leukaemia cells (Jurkat cells) and, if so, to determine the underlying mechanism. Cell proliferation and viability were determined by the MTS and trypan blue exclusion assays, respectively. Cell cycle analysis, apoptosis activity and oxidative stress levels were assessed by flow cytometry, and the expression of p7<em>3</em>, UHRF1 and active caspase-<em>3</em> by Western blot analysis. RWPs inhibited the proliferation of Jurkat cells and induced G0/G1 cell cycle arrest in a concentration-dependent manner. Moreover, RWPs triggered apoptosis, which is associated with an increased expression level of the pro-apoptotic protein p7<em>3</em> and the active caspase-<em>3</em>. RWPs induced apoptosis confirmed by DNA fragmentation analysis, and this effect was associated with down-regulation of the antiapoptotic protein UHRF1. Furthermore RWPs significantly increased the formation of reactive oxygen species (ROS). Intracellular scavengers of superoxide anions (MnTMPyP, MnTBAP, PEG-<em>SOD</em>) prevented the RWPs-induced formation of ROS and apoptosis, while native extracellular superoxide dismutase (<em>SOD</em>) was without effect. In addition, the effect of RWPs on the expression levels of p7<em>3</em>, active caspase-<em>3</em> and UHRF1 was also prevented by MnTMPyP. Thus, these findings indicate that RWPs induce apoptosis in Jurkat cells by a redox-sensitive mechanism involving the intracellular formation of superoxide anions and consequently the up-regulation of p7<em>3</em> and down-regulation of UHRF1.

Many cnidarians, such as sea anemones, contain photosynthetic symbiotic dinoflagellates called zooxanthellae. During a light/dark cycle, the intratentacular O(2) state changes in minutes from hypoxia to hyperoxia (<em>3</em>-fold normoxia). To understand the origin of the high tolerance to these unusual oxic conditions, we have characterized superoxide dismutases (<em>SODs</em>) from the three cellular compartments (ectoderm, endoderm and zooxanthellae) of the Mediterranean sea anemone Anemonia viridis. The lowest <em>SOD</em> activity was found in ectodermal cells while endodermal cells and zooxanthellae showed a higher <em>SOD</em> activity. Two, seven and six <em>SOD</em> activity bands were identified on native PAGE in ectoderm, endoderm and zooxanthellae, respectively. A CuZn<em>SOD</em> was identified in both ectodermal and endodermal tissues. Mn<em>SODs</em> were detected in all compartments with two different subcellular localizations. One band displays a classical mitochondrial localization, the three others being extramitochondrial. Fe<em>SODs</em> present in zooxanthellae also appeared in endodermal host tissue. The isoelectric points of all <em>SODs</em> were distributed between 4 and 5. For comparative study, a similar analysis was performed on the whole homogenate of a scleractinian coral Stylophora pistillata. These results are discussed in the context of tolerance to hyperoxia and to the transition from anoxia to hyperoxia.

Ginseng, one of the oldest traditional Chinese medicinal herbs, has been used widely in China and Asia for thousands of years. Ginsenosides extracted from ginseng, which is derived from the roots and rhizomes of Panax ginseng C. A. Meyer, have been used in China as an adjuvant in the treatment of diabetes mellitus. Owing to the technical complexity of ginsenoside production, the total ginsenosides are generally extracted. Accumulating evidence has shown that ginsenosides exert antidiabetic effects. In vivo and in vitro tests revealed the potential of ginsenoside Rg1, Rg<em>3</em>, Rg5, Rb1, Rb2, Rb<em>3</em>, compound K, Rk1, Re, ginseng total saponins, malonyl ginsenosides, Rd, Rh2, F2, protopanaxadiol (PPD) and protopanaxatriol (PPT)-type saponins to treat diabetes and its complications, including type 1 diabetes mellitus, type 2 diabetes mellitus, diabetic nephropathy, diabetic cognitive dysfunction, type 2 diabetes mellitus with fatty liver disease, diabetic cerebral infarction, diabetic cardiomyopathy, and diabetic erectile dysfunction. Many effects are attributed to ginsenosides, including gluconeogenesis reduction, improvement of insulin resistance, glucose transport, insulinotropic action, islet cell protection, hepatoprotective activity, anti-inflammatory effect, myocardial protection, lipid regulation, improvement of glucose tolerance, antioxidation, improvement of erectile dysfunction, regulation of gut flora metabolism, neuroprotection, anti-angiopathy, anti-neurotoxic effects, immunosuppression, and renoprotection effect. The molecular targets of these effects mainly contains GLUTs, SGLT1, GLP-1, FoxO1, TNF-α, IL-6, caspase-<em>3</em>, bcl-2, MDA, <em>SOD</em>, STAT5-PPAR gamma pathway, PI<em>3</em>K/Akt pathway, AMPK-JNK pathway, NF-κB pathway, and endoplasmic reticulum stress. Rg1, Rg<em>3</em>, Rb1, and compound K demonstrated the most promising therapeutic prospects as potential adjuvant medicines for the treatment of diabetes. This paper highlights the underlying pharmacological mechanisms of the anti-diabetic effects of ginsenosides.

OBJECTIVE

Liver fibrosis is a reversible wound-healing response that occurs following liver injury. In this study, we aimed to investigate the possible protective effects of ursolic acid in liver fibrosis induced by carbon tetrachloride (CCl4).

METHODS

ICR mice were randomly divided into six groups (Group 1: normal; Group 2: CCl4-treated group; Group <em>3</em>: CCl4 plus ursolic acid 25mg/kg group; Group 4: CCl4 plus ursolic acid 50mg/kg group; Group 5: CCl4 plus colchicine 1mg/kg group; Group 6: ursolic acid 50mg/kg group). Mice were administered with CCl4 (2 mL of CCl4 in olive oil (1:1, v/v) per kg body weight twice weekly) by intraperitoneal injection and oral injection of colchicine (1mg/kg) or ursolic acid (25, 50mg/kg) daily. After six weeks, serum aminotransferase activity, hepatic reactive oxygen species (ROS) production, thiobarbituric acid reactive substances (TBARS), antioxidase (<em>SOD</em>, CAT, GPx) activity and histopathological analysis were performed. The levels of nuclear factor E2-related factor 2 (Nrf2), NAD(P)H: quinone oxidoreductase-1 (NQO1), glutathione S-transferase (GST) and heme oxygenase-1 (HO-1), tumor necrosis factor-alpha (TNF-α), prostaglandin E2 (PGE2) and inducible nitric oxide synthase (iNOS), Bcl-2 and caspase-<em>3</em> were measured.

RESULTS

Ursolic acid significantly prevented CCl4-induced hepatotoxicity and fibrosis, indicated by both diagnostic indicators and histopathological analysis. CCl4-induced profound elevations of oxidative stress, inflammation and apoptosis in liver were suppressed by ursolic acid.

CONCLUSIONS

These results suggest that ursolic acid has the hepatoprotective actions. The inhibition of CCl4-induced liver fibrosis, inflammation and apoptosis by ursolic acid is due at least in part to its ability to modulate the Nrf2/ARE signalling pathway.

This study investigated the antioxidative effect of S-propargyl-cysteine (SPRC) on nonalcoholic fatty liver (NAFLD) by treating mice fed a methionine and choline deficient (MCD) diet with SPRC for four weeks. We found that SPRC significantly reduced hepatic reactive oxygen species (ROS) and methane dicarboxylic aldehyde (MDA) levels. Moreover, SPRC also increased the superoxide dismutase (<em>SOD</em>) activity. By Western blot, we found that this protective effect of SPRC was importantly attributed to the regulated hepatic antioxidant-related proteins, including protein kinase B (Akt), heme oxygenase-1 (HO-1), nuclear factor erythroid 2-related factor 2 (Nrf2), and cystathionine γ-lyase (CSE, an enzyme that synthesizes hydrogen sulfide). Next, we examined the detailed molecular mechanism of the SPRC protective effect using oleic acid- (OA-) induced HepG2 cells. The results showed that SPRC significantly decreased intracellular ROS and MDA levels in OA-induced HepG2 cells by upregulating the phosphorylation of Akt, the expression of HO-1 and CSE, and the translocation of Nrf2. SPRC-induced HO-1 expression and Nrf2 translocation were abolished by the phosphoinositide <em>3</em>-kinase (PI<em>3</em>K) inhibitor LY294002. Moreover, the antioxidative effect of SPRC was abolished by CSE inhibitor DL-propargylglycine (PAG) and HO-1 siRNA. Therefore, these results proved that SPRC produced an antioxidative effect on NAFLD through the PI<em>3</em>K/Akt/Nrf2/HO-1 signaling pathway.

<em>SOD</em>-4, a cytosolic form of superoxide dismutase in maize, originally was defined as a single band of activity by zymogram analysis. The protein was purified to "homogeneity" as shown by a single band on native or denaturing polyacrylamide gels and a single spot on two dimensional gels. The N-terminal amino acid sequence for the first 20 residues was determined for the purified <em>SOD</em>-4 protein. All residues were clearly determined except for residue twelve, where both glutamic and aspartic acids were found. A maize lambda gt11 cDNA library was constructed from scutellar poly(A)+ RNA. Two cDNAs were isolated, restriction mapped, and their DNA sequences determined. The amino acid sequence deduced from both cDNAs matched perfectly the N-terminal sequence of the purified protein except for the residue at position 12. Significantly, at the twelfth codon, one cDNA was found to code for glutamic acid and the other cDNA had a codon for aspartic acid. Both cDNAs contained similar but not identical 5' and <em>3</em>' untranslated sequences. Both cDNAs contained polyadenylation signals and tails. cDNA isolations, RNA, and genomic DNA blots confirm the existence and expression of two genes that produce indistinguishable <em>SOD</em>-4 proteins.

We employed next-generation RNA sequencing (RNA-Seq) technology to determine the extent to which exercise training alters global gene expression in skeletal muscle feed arteries and aortic endothelial cells of obese Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Transcriptional profiles of the soleus and gastrocnemius muscle feed arteries (SFA and GFA, respectively) and aortic endothelial cell-enriched samples from rats that underwent an endurance exercise training program (EndEx; n = 12) or a interval sprint training program (IST; n = 12) or remained sedentary (Sed; n = 12) were examined. In response to EndEx, there were <em>3</em>9 upregulated (e.g., MANF) and 20 downregulated (e.g., ALOX15) genes in SFA and 1 upregulated (i.e., Wisp2) and 1 downregulated (i.e., Crem) gene in GFA [false discovery rate (FDR) < 10%]. In response to IST, there were <em>3</em>05 upregulated (e.g., MANF, HSPA12B) and <em>3</em>24 downregulated genes in SFA and 101 upregulated and 66 downregulated genes in GFA, with an overlap of <em>3</em>2 genes between arteries. Furthermore, in aortic endothelial cells, there were 18<em>3</em> upregulated (e.g., eNOS, <em>SOD</em>-<em>3</em>) and 141 downregulated (e.g., ATF<em>3</em>, Clec1b, npy, leptin) genes with EndEx and 71 upregulated and 69 downregulated genes with IST, with an overlap of <em>3</em>5 between exercise programs. Expression of only two genes (Tubb2b and Slc9a<em>3</em>r2) was altered (i.e., increased) by exercise in all three arteries. The finding that both EndEx and IST produced greater transcriptional changes in the SFA compared with the GFA is intriguing when considering the fact that treadmill bouts of exercise are associated with greater relative increases in blood flow to the gastrocnemius muscle compared with the soleus muscle.

In our study, we evaluated the neuroprotective effects of dexmedetomidine on oxidant-antioxidant systems, pro-inflammatory cytokine TNF-alpha and number of apoptotic neurons on hippocampus and dentate gyrus after transient global cerebral I/R injury. Eighteen rats divided into <em>3</em> groups, equally. Group I rats were used as shams. For group II and III rats, they were prepared for transient global cerebral ischemia using a four-vessel-occlusion model. 5 mL/kg/h 0.9% sodium chloride was infused to the Group II and <em>3</em> microg/kg/h/5 ml dexmedetomidine was infused to the Group III for 2 h after I/R injury. The levels of MDA and NO and activities of <em>SOD</em> and CAT were measured in the left hippocampus tissue. The levels of TNF-alpha concentration were measured in the plasma. The number of apoptotic neurons was counted by TUNNEL method in histological samples of right hippocampus tissue. MDA and NO levels increased in Group II compared with Group I rats (p=0.002, p=0.002, respectively). In group III, MDA and NO levels decreased as compared to Group II (p=0.015, p=0.002, respectively). <em>SOD</em> and CAT activities increased in Group III as compared to Group II rats (p=0.002, p=0.002, respectively). The decrease in TNF-alpha levels of group III was significant as compared to group II (p=0.016). The number of apoptotic neurons in group III was lower than Group II rats. Our study showed that dexmedetomidine has a neuroprotective effect on hippocampus and dentate gyrus of rats after transient global cerebral I/R injury.

Fumonisins (FB) are mycotoxins produced by Fusarium verticillioides, frequently associated with corn. It produces toxicity, including teratogenicity, equine leukoencephalomalacia, porcine pulmonary edema, hepatic or renal damage in most animal species and perturb sphingolipid metabolism. The aim of the present study was to evaluate the protective effects of royal jelly (RJ) against FB toxicity. Sixty male Sprague-Dawley rats were divided into six treatment groups including the control group; group fed FB-contaminated diet (200mg/kg diet) and the groups treated orally with RJ (100 or 150mg/kg body weight) with or without FB for <em>3</em> weeks. FB alone decreased body weight gain, feed intake, GPX and <em>SOD</em>. Whereas it increased in ALT, AST, triglycerides, cholesterol, HDL, LDL, createnine and uric acid levels. Animals received FB showed severe histological and histochemical changes in liver and kidney tissues. Cotreatment with FB plus RJ resulted in a significant improvement in all the tested parameters and the histological and histochemical pictures of the liver and kidney. These improvements were pronounced in animals fed FB-contaminated diet plus the high dose of RJ. It could be concluded that RJ have a protective effects against FB toxicity and this protection was dose dependent.

OBJECTIVE

To provide evidence that free radical damage is a component of postischemic retinal injury; to determine whether antioxidant enzymes, superoxide dismutase (SOD) and catalase, can protect the retina from ischemic injury.

METHODS

Total retinal ischemia for 60 or 75 min was produced in Dutch rabbits by raising intraocular pressure. Retinal recovery was monitored with the electroretinogram. Enzymes were administered as an intravenous bolus dose 2-3 min before restoration of circulation.

RESULTS

In eyes subjected to 60 min ischemia, the amplitude of the a-wave 4 hours after reperfusion averaged 114.9% of baseline value in control rabbits and 126.5% in SOD-treated animals. The b-wave amplitude at this time was 79.3% and 106.8% in control rabbits and SOD-treated rabbits, respectively. After an ischemic insult of 75 min, at 4 hours the a-wave amplitude was 89.2% of baseline in control eyes, 108.8% in SOD-treated eyes, 159.6% in eyes that received a combination of SOD and catalase, and 149.8% in catalase-treated eyes. The amplitude of the b-wave was reduced to 47.8% in control eyes and 44.8% in SOD-treated eyes, but recovered to 92.3% in rabbits that received the combination therapy and 98.8% in animals that received catalase alone.

CONCLUSIONS

These findings suggest that free radical generation is involved in ischemic tissue damage. The fact that antioxidant enzymes can be protective has implications for the treatment of acute ischemic diseases of the retina.

OBJECTIVE

To investigate the effect of formaldehyde (FA) on testes and the protective effect of vitamin E (VE) against oxidative damage by FA in the testes of adult rats.

METHODS

Thirty rats were randomly divided into three groups: (1) control; (2) FA treatment group (FAt); and (<em>3</em>) FAt + VE group. FAt and FAt + VE groups were exposed to FA by inhalation at a concentration of 10 mg/m(<em>3</em>) for 2 weeks. In addition, FAt + VE group were orally administered VE during the 2-week FA treatment. After the treatment, the histopathological and biochemical changes in testes, as well as the quantity and quality of sperm, were observed.

RESULTS

The testicular weight, the quantity and quality of sperm, the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and glutathione (GSH) were significantly decreased whereas the level of malondialdehyde (MDA) was significantly increased in testes of rats in FAt group compared with those in the control group. VE treatment restored these parameters in FAt + VE group. In addition, microscopy with hematoxylin-eosin (HE) staining showed that seminiferous tubules atrophied, seminiferous epithelial cells disintegrated and shed in rats in FAt group and VE treatment significantly improved the testicular structure in FAt + VE group.

CONCLUSIONS

FA destroys the testicular structure and function in adult rats by inducing oxidative stress, and this damage could be partially reversed by VE.

We have recently shown that superoxide and hydrogen peroxide are putative inducers of angiogenesis in vivo, possibly through up regulation of inducible nitric oxide synthase (NOS) and increased production of endogenous nitric oxide (NO). The aim of the present work was to elucidate the implication of reactive oxygen species in endothelial cell functions, using cultures of human umbilical vein endothelial cells (HUVEC). Superoxide dismutase (<em>SOD</em>), tempol (membrane permeable <em>SOD</em> mimetic) and the NADPH oxidase inhibitors, 4-(2-aminoethyl)-benzenesulfonyl fluoride and apocynin, but not allopurinol, inhibited HUVEC proliferation and migration, as well as activity of endothelial NOS (eNOS). Catalase and the intracellular hydrogen peroxide scavenger sodium pyruvate decreased, while hydrogen peroxide increased HUVEC proliferation, migration and activity of eNOS. Dexamethasone induced the proliferation and migration of HUVEC and activated eNOS. Nomega-nitro-L-arginine methyl ester (L-NAME), but not Nomega-nitro-D-arginine methyl ester, decreased endothelial cell functions and reversed the effects of dexamethasone and hydrogen peroxide. N5-(1-iminoethyl)-L-ornithine dihydrochloride, but not the inducible NOS specific inhibitor N-[[<em>3</em>-(aminomethyl)phenyl]methyl]-ethanimidamide dihydrochloride also decreased endothelial cell functions, similarly to L-NAME. The guanylate cyclase inhibitor 1H-[1,2,4]Oxadiazole[4,<em>3</em>-a]quinoxalin-1-one inhibited HUVEC proliferation in a concentration-dependent manner and completely reversed hydrogen peroxide-induced proliferation, migration and cGMP accumulation. In conclusion, superoxide and hydrogen peroxide seem to play a significant role in promoting endothelial cell proliferation and migration, possibly through regulation of eNOS activity.