Oxygen Species Production In Cells Research Articles

Comparison of the nutritional and phytochemical composition and antioxidant activities of Aralia elata (Miq.) Seem fruits in Northeast China

Aralia elata ( Miq.) Seem is a wild medicinal and culinary plant. The newly grown shoots are edible, and the stem and root bark are traditional materials used for medicinal extraction. However, few studies have examined the quality and biological activity of the fruit. In this study, we compared the nutritional and phytochemical composition and antioxidant activity of A. elata fruit (AF) picked from five regions (Harbin, Benxi, Raohe, Jiaohe, and Mudanjiang) in Northeast China. AF picked from Harbin had the highest saponin, anthocyanin, and flavonoid contents. UHPLC-MS/MS analyses showed that araloside A was one of the main saponin monomers in AF. The highest arloside A concentration was detected in fruits picked from Harbin and the lowest concentration was detected in fruits picked from Benxi. The relative reactive oxygen species production in cells after pretreatment with AFs from Harbin (50 μg/mL) was 0.78 ± 0.01, second only to those from Raohe. AFs (50 μg/mL) from Harbin had the highest catalase activity (43.61 ± 0.44 U/mg protein), followed by that of the AF collected from Jiaohe, Benxi, Mudanjiang and Raohe. The malondialdehyde levels of the AFs (50 μg/mL) obtained from Benxi, Raohe, Jiaohe, and Mudanjiang were significantly higher than those of the AF obtained from Harbin (2.67 ± 0.15 nmol/mg protein). Compared with AFs from Benxi, Raohe, Jiaohe, and Mudanjiang, the strongest H 2 O 2 scavenging ability (144.95 ± 2.89 mmol TE/100 g dm) was observed for AF from Harbin. The ferric reducing antioxidant power of AFs ranged from 7.70 to 4.52 mmol TE/100 g dm in the order of Harbin > Mudanjiang > Raohe > Jiaohe > Benxi. The results provided support for the selection of the best quality AF and the expanded cultivation of A. elata .

Ginsenoside Rg3 exerts a neuroprotective effect in rotenone-induced Parkinson's disease mice via its anti-oxidative properties

Ginsenoside Rg3, extracted from Panax ginseng C.A. Meyer, has been shown to possess neuroprotective properties. The present study aims to investigate the neuroprotective effects of ginsenoside Rg3 on rotenone-induced Parkinson's disease mice. Rotenone, a mitochondrial complex I inhibitor, leads to the augmentation of reactive oxygen species production in cells. Male C57/BL6 mice were intragastrically administered rotenone (30 mg/kg) and then treated with ginsenoside Rg3 (5, 10, or 20 mg/kg). Pole, rotarod, and open field tests were performed to evaluate motor function. Ginsenoside Rg3 decreased the climbing time in the pole test (p < 0.01), whereas it increased the latency in the rotarod test (p < 0.01) and the total distance (p < 0.01) and mean speed in the open field test (p < 0.01). Ginsenoside Rg3 treatment augmented the number of tyrosine hydroxylase-positive neurons in the substantia nigra (p < 0.01), mean density of tyrosine hydroxylase-positive nerve fibers (p < 0.01), and dopamine content (p < 0.01) in the striatum and reduced the reactive oxygen species level in the substantia nigra (p < 0.01). Glutathione cysteine ligase regulatory subunit and glutathione cysteine ligase modulatory subunit expression levels were elevated in the ginsenoside Rg3 groups. Ginsenoside Rg3 also improved motor function in rotenone-induced Parkinson's disease mice. The neuroprotective effects of ginsenoside Rg3 are at least partly associated with its anti-oxidative properties via regulation of glutathione cysteine ligase modulatory subunit and glutathione cysteine ligase regulatory subunit expression.

Understanding the role of neutrophils in chronic inflammatory airway disease.

Airway neutrophilia is a common feature of many chronic inflammatory lung diseases and is associated with disease progression, often regardless of the initiating cause. Neutrophils and their products are thought to be key mediators of the inflammatory changes in the airways of patients with chronic obstructive pulmonary disease (COPD) and have been shown to cause many of the pathological features associated with disease, including emphysema and mucus hypersecretion. Patients with COPD also have high rates of bacterial colonisation and recurrent infective exacerbations, suggesting that neutrophil host defence mechanisms are impaired, a concept supported by studies showing alterations to neutrophil migration, degranulation and reactive oxygen species production in cells isolated from patients with COPD. Although the role of neutrophils is best described in COPD, many of the pathological features of this disease are not unique to COPD and also feature in other chronic inflammatory airway diseases, including asthma, cystic fibrosis, alpha-1 anti-trypsin deficiency, and bronchiectasis. There is increasing evidence for immune cell dysfunction contributing to inflammation in many of these diseases, focusing interest on the neutrophil as a key driver of pulmonary inflammation and a potential therapeutic target than spans diseases. This review discusses the evidence for neutrophilic involvement in COPD and also considers their roles in alpha-1 anti-trypsin deficiency, bronchiectasis, asthma, and cystic fibrosis. We provide an in-depth assessment of the role of the neutrophil in each of these conditions, exploring recent advances in understanding, and finally discussing the possibility of common mechanisms across diseases.

Hyperbaric oxygen affects endothelial progenitor cells proliferation in vitro.

Hyperbaric oxygen is a clinical treatment that contributes to wound healing by increasing fibroblasts proliferation, collagen synthesis, and production of growth factors, inducing angiogenesis and inhibiting antimicrobial activity. It also has been shown that hyperbaric oxygen treatment (HBO), through the activation of nitric oxide synthase promotes an increase in the nitric oxide levels that may improve endothelial progenitor cells (EPC) mobilization from bone marrow to the peripheral blood and stimulates the vessel healing process. However, cellular mechanisms involved in cell proliferation and activation of EPC after HBO treatment remain unknown. Therefore, the present work aimed to analyze the effect of HBO on the proliferation of pre-treated bone marrow-derived EPC with TNF-alpha. Also, we investigated the expression of ICAM and eNOS by immunochemistry, the production of reactive species of oxygen and performed an in vitro wound healing. Although 1h of HBO treatment did not alter the rate of in vitro wound closure or cell proliferation, it increased eNOS expression and decreased ICAM expression and reactive oxygen species production in cells pre-treated with TNF-alpha. These results indicate that HBO can decrease the inflammatory response in endothelial cells mediated by TNF-alpha, and thus, promote vascular recovery after injury.

Ganoderma tsugae induced ROS-independent apoptosis and cytoprotective autophagy in human chronic myeloid leukemia cells

The medicinal fungus Ganoderma, known in Chinese as Lingzhi or Reishi, traditionally has various medicinal uses and has been employed in cancer treatment in Asia for centuries. This study used ethanol-extracted Ganoderma tsugae (GT) and examined its antitumor activities on human chronic myeloid leukemia cells as well as its molecular mechanism of action. Treatment with GT (200–400 μg/mL) significantly reduced cell viability and caused G2/M arrest in K562 cells. In addition, GT induced mitochondrial and death receptor mediated apoptosis, correlated with DNA fragmentation, followed by cytochrome c release, caspase-3/8/9 activation, PARP cleavage, Fas activation, Bid cleavage, and Bax/Bcl-2 dysregulation. Cytoprotective autophagy was found to be induced by GT, as was revealed by increased LC3-II accumulation, Beclin-1/Bcl-2 dysregulation, acidic vesicular organelle formation, and p62/SQSTM1 activation. Notably, pretreatment of cells with the autophagy inhibitors 3-MA and CQ enhanced GT-induced apoptosis. Interestingly, reactive oxygen species production in cells was not triggered by GT administration; equally, the antioxidant N-acetylcysteine was found to be incapable of preventing apoptosis and autophagy induced by GT treatment. Finally, this study discovered that cytoprotective autophagy induced by GT was associated with EGFR and PI3K/AKT/mTOR signaling cascade suppression. In summary, GT demonstrated antitumor activity against human chronic myeloid leukemia.

Future perspective of strategic non-thermal plasma therapy for cancer treatment.

The therapeutic effects of non-thermal plasma are expected in the medical fields, including hemostasis, vascularization, prevention of organ adhesion, and cell proliferation. Cancer is an internal enemy arising from normal tissue in the body. The prognosis of metastatic and recurrent cancers is still poor despite advances in medicine. To apply non-thermal plasma in cancer treatment is now on going. The mechanism of the proliferation-inhibitory effect of plasma is reactive nitrogen oxide species/reactive oxygen species production in cells. There are a number of problems to be overcome, such as existence of intrinsic reactive oxygen species/reactive nitrogen species scavengers and the shallow infiltration of plasma on tumor surface. The current reviews makes referral to the study results of plasma therapy clarified so far, the possibility of its application in the future.

Moderate dependence of reactive oxygen species production on membrane potential in avian muscle mitochondria oxidizing glycerol 3-phosphate.

Mitochondria are a major source of reactive oxygen species production in cells, and the production level is sensitive to the magnitude of the membrane potential (ΔΨ). The present study investigated the level of superoxide production in mitochondria oxidizing glycerol 3-phosphate (GP) and its dependence on ΔΨ in isolated avian muscle mitochondria. The levels of superoxide produced in mitochondria oxidizing GP were lower than those obtained with succinate and were similar to those obtained with NADH-linked substrates (glutamate/malate/pyruvate). The dependence of superoxide production on ΔΨ in mitochondria oxidizing GP was lower than that of mitochondria oxidizing succinate, and a weak dependence of GP-supported superoxide production on ΔΨ was observed in the presence of NADH-linked substrates or succinate. These results suggest that the levels of superoxide generated in response to GP are quantitatively low, but they are unsusceptible to changes in ΔΨ in avian muscle mitochondria.

Effects of biguanides on oxidative phosphorylation

Metformin, and the related biguanides buformin and phenformin, have been proposed to act by inhibiting respiratory complex I, causing the cellular energetic stress that activates AMP-kinase and initiates a multitude of cell-lineage-specific effects, including inhibition of gluconeogenesis in hepatocytes. However, the molecular mechanism of biguanide action on complex I is not known, limiting understanding of pharmacological effects on this proposed primary target. Biguanides have also been proposed to affect reactive oxygen species production in cells, a respiratory-chain linked effect - but there is no consensus even on whether they increase or decrease it. In this talk I will discuss the results of experiments on a hierarchy of systems, from purified respiratory enzymes and membrane preparations, to mitochondria and cells, to define the functional effects of biguanides on the oxidative phosphorylation system. The aim is to define and link the molecular effects of biguanides on specific enzymes to their known effects on cellular and signalling pathways.

A CCL2/ROS autoregulation loop is critical for cancer-associated fibroblasts-enhanced tumor growth of oral squamous cell carcinoma

Cancer-associated fibroblasts (CAFs) have been described to play critical roles in initiation, progression and metastasis of various cancers. However, the involvement of CAFs in oral cancer (OC) has not been well addressed. In this study, we demonstrate that CAFs, when cocultured with OC cells (OCCs), produce high levels of chemokine (C-C motif) ligand 2 (CCL2) and, subsequently, enhance endogenous reactive oxygen species production in cells. Oxidative stress stimulates expression of cell cycle progression proteins in OCCs, leading to promotion of OCC proliferation, migration, invasion and, OC tumor growth. On the other hand, oxidative stress triggered the activation of nuclear factor-kappaB (NF-κB) and STAT3 in CAFs, resulting in accelerating CCL2 expression. In this way, CAFs-OCCs coculture creates a favorable cytokine-rich microenvironment, beneficial for both CAFs and OCCs. In addition, upregulation of CCL2 expression has been observed in oral squamous cell carcinoma tumors and patient plasma. We also showed that inhibition of CCL2 reduced OC tumor burden in mice. Therefore, our data suggested that CCL2 represents a potential therapeutic target for treatment of OC.

Decreased Reactive Oxygen Species Production in Cells with Mitochondrial Haplogroups Associated with Longevity

Mitochondrial DNA (mtDNA) is highly polymorphic, and its variations in humans may contribute to individual differences in function. Zhang and colleagues found a strikingly higher frequency of a C150T transition in the D-loop of mtDNA from centenarians and twins of an Italian population, and also demonstrated that this base substitution causes a remodeling of the mtDNA 151 replication origin in human leukocytes and fibroblasts [1]. The C150T transition is a polymorphism associated with several haplogroups. To determine whether haplogroups that carry the C150T transition display any phenotype that may be advantageous for longevity, we analyzed cybrids carrying or not the C150T transition. These cybrids were obtained by fusing cytoplasts derived from human fibroblasts with human mtDNA-less cells (ρ0 cells). We chose for cybrid construction and analysis haplogroup-matched pairs of fibroblast strains containing or not the C150T transition. In particular, we used, as one pair of mtDNA donors, a fibroblast strain of the U3a haplogroup, carrying the C150T transition and a strain of the U-K2 haplogroup, without the C150T transition, and as another pair, fibroblasts of the J2b haplogroup, carrying the C150T transition and of the J1c haplogroup, without the C150T transition. We have found no association of respiratory capacity, mtDNA level, mitochondrial gene expression level, or growth rate with the presence of the C150T transition. However, we have found that the cybrids with haplogroups that include the C150T transition have in common a lower reactive oxygen species (ROS) production rate than the haplogroup-matched cybrids without that transition. Thus, the lower ROS production rate may be a factor in the increased longevity associated with the U and the J2 haplogroups. Of further interest, we found that cybrids with the U3a haplogroup exhibited a higher respiration rate than the other cybrids examined.

Phytoglycoprotein (75 kDa) isolated from Cudrania tricuspidata Bureau inhibits expression of interleukin‐4 in the presence of di (2‐ethylhexyl) phthalate via modulation of p38 mitogen‐activated protein kinase in primary‐cultured mouse thymocytes

The purpose of this study is to determine the inhibitory effect of a glycoprotein isolated from Cudrania tricuspidata Bureau (CTB glycoprotein) on di(2-ethylhexyl) phthalate (DEHP)-induced differentiation of T helper (Th) type 2 cells in primary cultured thymocytes. The results obtained from this study revealed that the CTB glycoprotein in the presence of DEHP produces an antioxidative activity against intracellular reactive oxygen species production in cells. In addition, the activities of p38 mitogen-activated protein kinase and GATA-binding protein 3 were decreased by treatment with the CTB glycoprotein (100 microg ml(-1)). The CTB glycoprotein also has an inhibitory effect on the expressions of interleukin (IL)-4 and IL-10 induced by DEHP in thymocytes. Hence, we speculate that the CTB glycoprotein might be one component for preparation of health supplements for prevention of Th2 cell response-related immune diseases.

Differential regulation of CuZnSOD expression in rat brain by acute and/or chronic stress.

Neuroendocrine stress (NES) causes increase of glucocorticoids and alters physiological levels of reactive oxygen species production in cells, which might involve modifications in the antioxidant defense system. We investigated the hypothesis that acute, chronic, or combined stress alters copper-zinc superoxide dismutase (CuZnSOD) expression pattern at both, mRNA and subcellular protein level in the cerebral cortex and hippocampus of rats and that there may be a relationship between stress-induced corticosterone and CuZnSOD expression. The most effective stress model which led to the most pronounced changes in CuZnSOD expression patterns was also investigated. Our results demonstrated that acute stress immobilization up-regulates mRNA expression of hippocampal CuZnSOD, while cytosolic protein expression of this enzyme was increased in both brain structures. Chronic stress isolation had no effect on either mRNA and protein expression level and caused a lack of significant up-regulation to a novel acute stressors. The presence of this protein in nuclear fractions of both brain structures was also confirmed. The elevated cytosolic CuZnSOD protein levels following acute immobilization might reflect on the defense system against oxidative stress. Chronic isolation compromises CuZnSOD protein expression, which may lead to the inefficient defense against reactive oxygen species (ROS). The stress-triggered CuZnSOD protein expression was not correlated by the corresponding mRNA. The results suggest that different stress models exert a different degree of influence on mRNA and protein level of CuZnSOD in both brain structures as well as serum corticosterone.

Phosphorylation and Kinetics of Mammalian Cytochrome c Oxidase

The influence of protein phosphorylation on the kinetics of cytochrome c oxidase was investigated by applying Western blotting, mass spectrometry, and kinetic measurements with an oxygen electrode. The isolated enzyme from bovine heart exhibited serine, threonine, and/or tyrosine phosphorylation in various subunits, except subunit I, by using phosphoamino acid-specific antibodies. The kinetics revealed slight inhibition of oxygen uptake in the presence of ATP, as compared with the presence of ADP. Mass spectrometry identified the phosphorylation of Ser-34 at subunit IV and Ser-4 and Thr-35 at subunit Va. Incubation of the isolated enzyme with protein kinase A, cAMP, and ATP resulted in serine and threonine phosphorylation of subunit I, which was correlated with sigmoidal inhibition kinetics in the presence of ATP. This allosteric ATP-inhibition of cytochrome c oxidase was also found in rat heart mitochondria, which had been rapidly prepared in the presence of protein phosphatase inhibitors. The isolated rat heart enzyme, prepared from the mitochondria by blue native gel electrophoresis, showed serine, threonine, and tyrosine phosphorylation of subunit I. It is concluded that the allosteric ATP-inhibition of cytochrome c oxidase, previously suggested to keep the mitochondrial membrane potential and thus the reactive oxygen species production in cells at low levels, occurs in living cells and is based on phosphorylation of cytochrome c oxidase subunit I.

Metformin Prevents Glucose-Induced Protein Kinase C-β2 Activation in Human Umbilical Vein Endothelial Cells Through an Antioxidant Mechanism

Hyperglycemia determines the vascular complications of diabetes through different mechanisms: one of these is excessive activation of the isoform beta2 of protein kinase C (PKC-beta2). Metformin, a widely used antidiabetic agent, is associated with decreased cardiovascular mortality in obese type 2 diabetic patients. Therefore, we assessed the role of metformin in glucose-induced activation of PKC-beta2 and determined the mechanism of its effect in human umbilical venous endothelial cells grown to either normo- (5 mmol/l) or hyperglycemia (10 mmol/l) and moderately and acutely exposed to 25 mmol/l glucose. We studied PKC-beta2 activation by developing adenovirally expressed chimeras encoding fusion protein between green fluorescent protein (GFP) and conventional beta2 isoform (PKC-beta2-GFP). Glucose (25 mmol/l) induced the translocation of PKC-beta2-GFP from the cytosol to the membrane in cells grown to hyperglycemia but not in those grown in normal glucose medium. Metformin (20 micromol/l) prevented hyperglycemia-induced PKC-beta2-GFP translocation. We also assessed oxidative stress under the same conditions with a 4-((9-acridine-carbonyl)amino)-2,2,6,6-tetramethylpiperidin-oxyl,free radical (TEMPO-9-AC) fluorescent probe. We observed significantly increased radical oxygen species production in cells grown in hyperglycemia medium, and this effect was abolished by metformin. We show that in endothelial cells, metformin inhibits hyperglycemia-induced PKC-beta2 translocation because of a direct antioxidant effect. Our data substantiate the findings of previous large intervention studies on the beneficial effect of this drug in type 2 diabetic patients.

Fatty Aldehyde Dehydrogenase: POTENTIAL ROLE IN OXIDATIVE STRESS PROTECTION AND REGULATION OF ITS GENE EXPRESSION BY INSULIN

Phosphatidylinositol 3-kinase signaling regulates the expression of several genes involved in lipid and glucose homeostasis; deregulation of these genes may contribute to insulin resistance and progression toward type 2 diabetes. By employing RNA arbitrarily primed-PCR to search for novel phosphatidylinositol 3-kinase-regulated genes in response to insulin in isolated rat adipocytes, we identified fatty aldehyde dehydrogenase (FALDH), a key component of the detoxification pathway of aldehydes arising from lipid peroxidation events. Among these latter events are oxidative stresses associated with insulin resistance and diabetes. Upon insulin injection, FALDH mRNA expression increased in rat liver and white adipose tissue and was impaired in two models of insulin-resistant mice, db/db and high fat diet mice. FALDH mRNA levels were 4-fold decreased in streptozotocin-treated rats, suggesting that FALDH deregulation occurs both in hyperinsulinemic insulin-resistant state and hypoinsulinemic type 1 diabetes models. Moreover, insulin treatment increases FALDH activity in hepatocytes, and expression of FALDH was augmented during adipocyte differentiation. Considering the detoxifying role of FALDH, its deregulation in insulin-resistant and type 1 diabetic models may contribute to the lipid-derived oxidative stress. To assess the role of FALDH in the detoxification of oxidized lipid species, we evaluated the production of reactive oxygen species in normal versus FALDH-overexpressing adipocytes. Ectopic expression of FALDH significantly decreased reactive oxygen species production in cells treated by 4-hydroxynonenal, the major lipid peroxidation product, suggesting that FALDH protects against oxidative stress associated with lipid peroxidation. Taken together, our observations illustrate the importance of FALDH in insulin action and its deregulation in states associated with altered insulin signaling.