Blueberry attenuates liver injury in metabolic dysfunction-associated liver disease by promoting the expression of mitofilin/Mic60 in human hepatocytes and inhibiting the production of superoxide / 细胞与分子免疫学杂志

Objective To study the effect and mechanism of blueberry on regulating the mitochondrial inner membrane protein mitofilin/Mic60 in an in vitro model of metabolic dysfunction-associated liver disease (MAFLD). Methods L02 human hepatocytes were induced by free fatty acids (FFA) to establish MAFLD cell model. A normal group, a model group, an 80 μg/mL blueberry treatment group, a Mic60 short hairpin RNA (Mic60 shRNA) transfection group, and Mic60 knockdown combined with an 80 μg/mL blueberry treatment group were established. The intracellular lipid deposition was observed by oil red O staining, and the effect of different concentrations of blueberry pulp on the survival rate of L02 cells treated with FFA was measured by MTT assay. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), superoxide dismutase (SOD) activity, glutathione (GSH) and malondialdehyde (MDA) contents were measured by visible spectrophotometry. The expression of reactive oxygen species (ROS) in hepatocytes was observed by fluorescence microscopy, and the mRNA and protein expression of Mic60 were detected by real-time quantitative PCR and Western blot analysis, respectively. Results After 24 hours of FFA stimulation, a large number of red lipid droplets in the cytoplasm of L02 cells was observed, and the survival rate of L02 cells treated with 80 μg/mL blueberry was higher. The results of ALT, AST, TG, TC, MDA and the fluorescence intensity of ROS in blueberry treated group were lower than those in model group, while the levels of SOD, GSH, Mic60 mRNA and protein in blueberry treated group were higher than those in model group. Conclusion Blueberry promotes the expression of Mic60, increases the levels of SOD and GSH in hepatocytes, and reduces the production of ROS, thus alleviating the injury of MAFLD hepatocytes and regulating the disorder of lipid metabolism.

Aerobic Swim Training Restores Aortic Endothelial Function by Decreasing Superoxide Levels in Spontaneously Hypertensive Rats

OBJECTIVE: We aimed to determine whether aerobic training decreases superoxide levels, increases nitric oxide levels, and improves endothelium-dependent vasodilation in the aortas of spontaneously hypertensive rats. METHODS: Spontaneously hypertensive rats (SHR) and Wistar Kyoto rats (WKY) were distributed into 2 groups: sedentary (SHRsd and WKYsd, n=10 each) and swimming-trained (SHRtr, n=10 and WKYtr, n=10, respectively). The trained group participated in training sessions 5 days/week for 1 h/day with an additional work load of 4% of the animal’s body weight. After a 10-week sedentary or aerobic training period, the rats were euthanized. The thoracic aortas were removed to evaluate the vasodilator response to acetylcholine (10-10 to 10-4 M) with or without preincubation with L-NG-nitro-L-arginine methyl ester hydrochloride (L-NAME; 10-4 M) in vitro. The aortic tissue was also used to assess the levels of the endothelial nitric oxide synthase and nicotinamide adenine dinucleotide oxidase subunit isoforms 1 and 4 proteins, as well as the superoxide and nitrite contents. Blood pressure was measured using a computerized tail-cuff system. RESULTS: Aerobic training significantly increased the acetylcholine-induced maximum vasodilation observed in the SHRtr group compared with the SHRsd group (85.9±4.3 vs. 71.6±5.2%). Additionally, in the SHRtr group, superoxide levels were significantly decreased, nitric oxide bioavailability was improved, and the levels of the nicotinamide adenine dinucleotide oxidase subunit isoform 4 protein were decreased compared to the SHRsd group. Moreover, after training, the blood pressure of the SHRtr group decreased compared to the SHRsd group. Exercise training had no effect on the blood pressure of the WKYtr group. CONCLUSIONS: In SHR, aerobic swim training decreased vascular superoxide generation by nicotinamide adenine dinucleotide oxidase subunit isoform 4 and increased nitric oxide bioavailability, thereby improving endothelial function.

N-acetylcysteine downregulates phosphorylated p-38 expression but does not reverse the increased superoxide anion levels in the spinal cord of rats with neuropathic pain

We determined the effect of N-acetylcysteine (NAC) on the expression of the phosphorylated p38 (p-p38) protein and superoxide anion generation (SAG), two important players in the processing of neuropathic pain, in the lumbosacral spinal cord of rats with chronic constriction injury (CCI)-induced neuropathic pain. The sciatic functional index (SFI) was also measured to assess the functional recovery post-nerve lesion. Thirty-six male Wistar rats were divided equally into the following groups: Naive (rats did not undergo surgical manipulation); Sham (rats in which all surgical procedures involved in CCI were used except the ligature), and CCI (rats in which four ligatures were tied loosely around the right common sciatic nerve), which received 2, 4, or 8 intraperitoneal injections of NAC (150 mg·kg-1·day-1) or saline beginning 4 h after CCI. Rats were sacrificed 1, 3, and 7 days after CCI. The SFI was measured on these days and the lumbosacral spinal cord was used for analysis of p-p38 expression and SAG. CCI induced a decrease in SFI as well as an increase in p-p38 expression and SAG in the spinal cord. The SFI showed a partial recovery at day 7 in saline-treated CCI rats, but recovery was improved in NAC-treated CCI rats. NAC induced a downregulation in p-p38 expression at all time-points evaluated, but did not reverse the increased SAG induced by CCI. Since p-p38 is a mediator in neuropathic pain and/or nerve regeneration, modulation of this protein may play a role in NAC-induced effects in CCI rats.

The antioxidative defense system is involved in the premature senescence in transgenic tobacco (Nicotiana tabacum NC89)

BACKGROUND: α-Farnesene is a volatile sesquiterpene synthesized by the plant mevalonate (MVA) pathway through the action of α-farnesene synthase. The α-farnesene synthase 1 (MdAFS1) gene was isolated from apple peel (var. white winterpearmain), and transformed into tobacco (Nicotiana tabacum NC89). The transgenic plants had faster stem elongation during vegetative growth and earlier flowering than wild type (WT). Our studies focused on the transgenic tobacco phenotype. RESULTS: The levels of chlorophyll and soluble protein decreased and a lower seed biomass and reduced net photosynthetic rate (Pn) in transgenic plants. Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) and superoxide radicals (O2._) had higher levels in transgenics compared to controls. Transgenic plants also had enhanced sensitivity to oxidative stress. The transcriptome of 8-week-old plants was studied to detect molecular changes. Differentially expressed unigene analysis showed that ubiquitin-mediated proteolysis, cell growth, and death unigenes were upregulated. Unigenes related to photosynthesis, antioxidant activity, and nitrogen metabolism were downregulated. Combined with the expression analysis of senescence marker genes, these results indicate that senescence started in the leaves of the transgenic plants at the vegetative growth stage. CONCLUSIONS: The antioxidative defense system was compromised and the accumulation of reactive oxygen species (ROS) played an important role in the premature aging of transgenic plants.

The reduction of serum aminotransferase levels is proportional to the decline of the glomerular filtration rate in patients with chronic kidney disease

OBJECTIVE: This study sought to determine the serum aminotransferase levels of patients with predialysis chronic kidney disease and establish their relationships with serum creatinine levels and glomerular filtration rate. METHODS: Patients with chronic kidney disease were evaluated between September 2011 and May 2012. Aminotransferase and creatinine serum levels were measured using an automated kinetic method, and glomerular filtration rates were estimated using the Cockroft-Gault and Modification of Diet in Renal Disease formulas to classify patients into chronic kidney disease stages. RESULTS: Exactly 142 patients were evaluated (mean age: 64±16 years). The mean creatinine serum level and glomerular filtration rate were 3.3±1.2 mg/dL and 29.1±13 mL/min/1.73 m2, respectively. Patients were distributed according to their chronic kidney disease stages as follows: 3 (2.1%) patients were Stage 2; 54 (38%) were Stage 3; 70 (49.3%) were Stage 4; and 15 (10.5%) were Stage 5. The mean aspartate aminotransferase and alanine aminotransferase serum levels showed a reduction in proportion to the increase in creatinine levels (p=0.001 and p=0.05, respectively) and the decrease in glomerular filtration rate (p=0.007 and p=0.028, respectively). Alanine aminotransferase and aspartate aminotransferase serum levels tended to be higher among patients classified as stage 2 or 3 compared with those classified as stage 4 or 5 (p=0.08 and p=0.06, respectively). CONCLUSIONS: The aspartate aminotransferase and alanine aminotransferase serum levels of patients with predialysis chronic kidney disease decreased in proportion to the progression of the disease; they were negatively correlated with creatinine levels and directly correlated with glomerular filtration rate. .

Superoxide overproduction and kidney fibrosis: a new animal model / Produção aumentada de superóxido e fibrose renal: novo modelo animal

Objective To establish whether the mutation in the Immp2L gene induces renal fibrosis and whether aging exacerbates renal morphology in mice. Methods Female mutant mice with mutation in the inner mitochondrial membrane peptidase 2-like protein at 3 and 18 months of age were used. Renal fibrosis was analyzed using classic fibrosis score, Masson’s trichrome staining, and analysis of profibrotic markers using real time polymerase chain reaction (superoxide dismutase 1, metalloproteinase-9, erythropoietin, transforming growth factor beta), and immunostaining (fibroblasts and Type IV collagen). Oxidative stress markers were determined by immunohistochemistry. The number of renal apoptotic cells was determined. Renal function was estimated by serum creatinine. Results Young mutant mice had significantly more glomerulosclerosis than age-matched mice (p=0.034). Mutant mice had more tubular casts (p=0.025), collagen deposition (p=0.019), and collagen type IV expression (p<0.001). Superoxide dismutase 1 expression was significantly higher in young mutants (p=0.038). Old mutants exhibited significantly higher expression of the fibroblast marker and macrophage marker (p=0.007 and p=0.012, respectively). The real time polymerase chain reaction of metalloproteinase-9 and erythropoietin were enhanced 2.5- and 6-fold, respectively, in old mutants. Serum creatinine was significantly higher in old mutants (p<0.001). Conclusion This mutation altered renal architecture by increasing the deposition of extracellular matrix, oxidative stress, and inflammation, suggesting a protective role of Immp2L against renal fibrosis. .

Objetivo Estabelecer se a mutação no gene Immp2L induz à fibrose renal e se o envelhecimento exacerba a morfologia renal em camundongos. Métodos Foram usadas fêmeas de camundongos mutantes para proteína semelhante à peptidase 2 da camada interna da mitocôndria, com 3 e 18 meses de idade. Para analisar a fibrose renal, foram usados o escore clássico de fibrose, a coloração com tricrômio de Masson, e a análise de marcadores profibróticos, por meio da reação em cadeia de polimerase em tempo real (superóxido dismutase 1, metalonoproteinase-9, eritropoietina e fator transformador de crescimento beta), e a imunocoloração (fibroblastos e colágeno IV). Marcadores de estresse oxidativo foram determinados por imuno-histoquímica. O número de células apoptóticas renais foi analisado. A função renal foi estimada por creatinina sérica. Resultados Camundongos mutantes jovens apresentaram glomeruloesclerose em quantidade significativamente maior que animais da mesma idade (p=0,034). Os mutantes mostraram maior formação de cilindros tubulares (p=0,025), deposição de colágeno (p=0,019) e maior expressão de colágeno do tipo IV (p<0,001). A expressão de superóxido dismutase 1 foi maior em mutantes jovens (p=0,038). Mutantes idosas exibiram maior expressão dos marcadores de fibroblastos e macrófagos (p=0,007 e p=0,012, respectivamente). As reações da cadeia de polimerase em tempo real da metalanoproteinase-9 e da eritropoietina estavam aumentadas em 2,5 e 6 vezes, respectivamente, em mutantes idosas. A creatinina sérica foi significantemente maior em animais idosos mutantes (p<0,001). Conclusão Essa mutação alterou a arquitetura renal pelo aumento da deposição de matriz extracelular, estresse oxidativo e inflamação, sugerindo papel de proteção de Immp2L contra a fibrose renal. .

NADPH oxidase activation contributes to native low-density lipoprotein-induced proliferation of human aortic smooth muscle cells

Elevated plasma concentration of native low-density lipoprotein (nLDL) is associated with vascular smooth muscle cell (VSMC) activation and cardiovascular disease. We investigated the mechanisms of superoxide generation and its contribution to pathophysiological cell proliferation in response to nLDL stimulation. Lucigenin-induced chemiluminescence was used to measure nLDL-induced superoxide production in human aortic smooth muscle cells (hAoSMCs). Superoxide production was increased by nicotinamide adenine dinucleotide phosphate (NADPH) and decreased by NADPH oxidase inhibitors in nLDL-stimulated hAoSMC and hAoSMC homogenates, as well as in prepared membrane fractions. Extracellular signal-regulated kinase 1/2 (Erk1/2), protein kinase C-theta (PKCtheta) and protein kinase C-beta (PKCbeta) were phosphorylated and maximally activated within 3 min of nLDL stimulation. Phosphorylated Erk1/2 mitogen-activated protein kinase, PKCtheta and PKCbeta stimulated interactions between p47phox and p22phox; these interactions were prevented by MEK and PKC inhibitors (PD98059 and calphostin C, respectively). These inhibitors decreased nLDL-dependent superoxide production and blocked translocation of p47phox to the membrane, as shown by epifluorescence imaging and cellular fractionation experiments. Proliferation assays showed that a small interfering RNA against p47phox, as well as superoxide scavenger and NADPH oxidase inhibitors, blocked nLDL-induced hAoSMC proliferation. The nLDL stimulation in deendothelialized aortic rings from C57BL/6J mice increased dihydroethidine fluorescence and induced p47phox translocation that was blocked by PD98059 or calphostin C. Isolated aortic SMCs from p47phox-/- mice (mAoSMCs) did not respond to nLDL stimulation. Furthermore, NADPH oxidase 1 (Nox1) was responsible for superoxide generation and cell proliferation in nLDL-stimulated hAoSMCs. These data demonstrated that NADPH oxidase activation contributed to cell proliferation in nLDL-stimulated hAoSMCs.

Effects of Antioxidant on Reduction of Hindlimb Muscle Atrophy Induced by Cisplatin in Rats

PURPOSE: The purpose of this study was to examine the effects of Cu/Zn SOD on reduction of hindlimb muscular atrophy induced by cisplatin in rats. METHODS: Forty-two rats were assigned to three groups; control group, Cisplatin (CDDP) group and cisplatin with Cu/Zn SOD (CDDP-SOD) group. At day 35 hindlimb muscles were dissected. Food intake, activity, withdrawal threshold, muscle weight, and Type I, II fiber cross-sectional area (CSA) of dissected muscles were measured. Relative SOD activity and expression of MHC and phosphorylated Akt, ERK were measured after dissection. RESULTS: Muscle weight and Type I, II fiber CSA of hindlimb muscles in the CDDP group were significantly less than the control group. Muscle weight and Type I, II fiber CSA of hindlimb muscles, food intake, activity, and withdrawal thresholds of the CDDP-SOD group were significantly greater than the CDDP group. There were no significant differences in relative SOD activities of hindlimb muscles between the CDDP-SOD and CDDP groups. MHC expression and phosphorylated Akt, ERK of hindlimb muscles in the CDDP-SOD group were significantly greater than the CDDP group. CONCLUSION: Cu/Zn SOD attenuates hindlimb muscular atrophy induced by cisplatin through increased food intake and activity. Increment of phosphorylated Akt, ERK may relate to attenuation of hindlimb muscular atrophy.

Interval training attenuates the metabolic disturbances in type 1 diabetes rat model / Treinamento intervalado atenua os distúrbios metabólicos em modelo de ratos diabéticos do tipo 1

OBJECTIVE: This study investigated the effect of interval training on blood biochemistry and immune parameters in type 1 diabetic rats. MATERIALS AND METHODS: Male Wistar rats were divided into four groups: sedentary (SE, n = 15), interval training (IT, n = 17), diabetic sedentary (DSE, n = 17), diabetic interval training (DIT, n = 17). Diabetes was induced by i.v. injection of streptozotocin (60 mg/kg). Swimming Interval Training consisted of 30-s exercise with 30-s rest, for 30 minutes, during 6 weeks, four times a week, with an overload of 15% of body mass. Plasma glucose, lactate, triacylglycerol and total cholesterol concentrations, phagocytic capacity, cationic vesicle content, and superoxide anion and hydrogen peroxide production by blood neutrophils and peritoneal macrophages were evaluated. Proliferation of mesenteric lymphocytes was also estimated. RESULTS: Interval training resulted in attenuation of the resting hyperglycemic state and decreased blood lipids in the DIT group. Diabetes increased the functionality of blood neutrophils and peritoneal macrophages in the DSE group. Interval training increased all functionality parameters of peritoneal macrophages in the IT group. Interval training also led to a twofold increase in the proliferation of mesenteric lymphocytes after 6 weeks of exercise in the DIT group. CONCLUSION: Low-volume high-intensity physical exercise attenuates hyperglycemia and dislipidemia induced by type 1 diabetes, and induces changes in the functionality of innate and acquired immunity.

OBJETIVO: Este estudo investigou os efeitos do treinamento intervalado sobre parâmetros bioquímicos e imunológicos em ratos diabéticos do tipo 1. MATERIAIS E MÉTODOS: Ratos Wistar machos foram divididos em quatro grupos: sedentário (SE, n = 15), treinamento intervalado (TI, n = 17), sedentário diabético (SED, n = 17) e treinamento intervalado diabético (TID, n = 17). O diabetes foi induzido por uma injeção intravenosa de estreptozotocina (60 mg/kg). O treinamento intervalado de natação consistiu de 30s de exercício com 30s de recuperação, 30 minutos, durante 6 semanas, 4 vezes por semana, com sobrecarga de 15% da massa corporal. Foram avaliados glicemia, lactato sanguíneo, concentração de triacilglicerol e colesterol total, capacidade fagocítica, conteúdo de vesículas catiô­nicas, produção de ânion superóxido e peróxido de hidrogênio por neutrófilos sanguíneos e macrófagos peritoneais. A proliferação de linfócitos mesentéricos também foi avaliada. RESULTADOS: O treinamento intervalado resultou em atenuação do estado hiperglicêmico e diminuiu os lipídeos sanguíneos no grupo TID. O diabetes aumentou a funcionalidade dos neutrófilos sanguíneos e macrófagos peritoneais do grupo SED. O treinamento intervalado aumentou todos os parâmetros funcionais dos macrófagos peritoneais do grupo TI. O treinamento intervalado também aumentou duas vezes a proliferação dos linfócitos mesentéricos após seis semanas de exercício do grupo TID. CONCLUSÃO: O treinamento intervalado atenua a hiperglicemia e a dislipidemia induzida pelo diabetes do tipo 1 e induz mudanças na funcionalidade da imunidade inata e adquirida.

Aluminum and ethanol induce alterations in superoxide and peroxide handling capacity (SPHC) in frontal and temporal cortex.

Aluminum is an omnipresent neurotoxicant and has been associated with several neuropathological disorders. Cerebrum and cerebellum have been shown to face augmented oxidative stress when animals are exposed to aluminum and high doses of ethanol. To establish the link between oxidative stress and neurobehavioral alterations, the present study was conducted to determine the extent of oxidative stress in low levels of pro-oxidant (ethanol exposure) status of the functionally discrete regions of the cerebrum. Male Wistar rats were exposed to aluminum (10 mg/kg body wt) and ethanol (0.2-0.6 g/kg body wt) for 4 weeks. Spontaneous motor activity (SMA) and Rota-Rod performances (RRP) were recorded weekly during the period of exposure. At the end of 4th week, oxidative stress parameters were determined from the homogenized cerebral tissue. GSH-independent superoxide peroxide handling capacity (GI-SPHC) and GSH-dependent superoxide peroxide handling capacity (GD-SPHC) were determined for FC and TC upon exposure to ethanol in the absence and presence of aluminum exposure. Aluminum was found to augment the oxidative stress at higher doses (0.6 g Ethanol/kg body wt) of ethanol, particularly in FC. The SPHC of FC was also found to be compromised significantly in aluminum-ethanol co-exposed animals. It was concluded that even though the manifestation of oxidative stress was not observed as revealed by assaying the widely used oxidative stress biochemical markers (indices), aluminum and ethanol (low doses) exposure induced alterations in the handling capacity of oxidant imbalance that could be recognized by studying the SPHC of FC. Comparison of GD-SPHC and GI-SPHC offered a possible mechanism of compromised SPHC in FC. This observation is likely to offer insights into the mechanism of association between aluminium exposure and behavioral changes in neurodegenerative disorders towards therapeutic strategies for these disorders.

Evaluation of changes in lipid peroxidation, ROS production, surface structures, secondary metabolites and yield of linseed (Linum usitatissimum L.) under individual and combined stress of ultraviolet-B and ozone using open top chambers.

The individual and interactive effects of supplemental UV-B (sUV-B) (ambient + 7.2 kJ m-2 d-1) and elevated O3 (ambient + 10 ppb) were evaluated under field conditions using open top chambers on two cultivars, Padmini and T-397 of linseed (Linum usitatissimum L.). Mean monthly surface level of O3 concentrations varied from 27.7 ppb to 59.0 ppb during the experimental period. Both UV-B and O3 induced the production of ROS (H2O2 and O2.-), resulting in significant damage of membranes due to lipid peroxidation and electrolyte leakage. Synthesis of secondary metabolites (flavonoids, anthocyanin, lignin and wax) was also enhanced in all the treatments, whereas biomass and yield were reduced. Alterations in frequency of stomata and wax distribution were also observed through scanning electron microscopy (SEM). Cultivar Padmini was found to be more sensitive because of higher damage of membrane vis-a-vis reduction in biomass and seed yield. However, concentrations of flavonoids, anthocyanin, lignin and wax were higher in T-397, suggesting its relative resistance against applied stress. Combined exposure of sUV-B and O3 was less harmful, as compared to their individual treatment. Among the three treatments, O3 was found to be more detrimental for overall growth and sUV-B for economic yield.

Role of ERK1/2 Kinase in the expression of iNOS by NDMA in human neutrophils.

Potential role of ERK1/2 kinase in conjunction with p38 in the regulation of inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production, and superoxide anion generation by human neutrophils (PMNs) exposed to N-nitrosodimethylamine (NDMA) was determined. Increased synthesis of NO due to the involvement of iNOS in neutrophils exposed to NDMA was observed. In addition, intensified activation of ERK1/2 and p38 kinases was determined in these cells. Inhibition of kinase regulated by extracellular signals (ERK1/2) pathway, in contrast to p38 pathway, led to an increased production of NO and expression of iNOS in PMNs. Moreover, as a result of inhibition of ERK1/2 pathway, a decreased activation of p38 kinase was observed in neutrophils, while inhibition of p38 kinase did not affect activation of ERK1/2 pathway in these cells. An increased ability to release superoxide anion by the studied PMNs was observed, which decreased after ERK1/2 pathway inhibition. In conclusion, in human neutrophils, ERK1/2 kinase is not directly involved in the regulation of iNOS and NO production induced by NDMA; however, the kinase participates in superoxide anion production in these cells.

Gemmomodification: an emerging source of natural antioxidants from Silybum marianum

Gemmomodification is a form of herbal medicine in which young freshly growing buds of plants are used. At germinating stage, plants metabolic activities are maximum and various nutrients, hormones enzymes and bioactive phytoconstituents are released and available at this stage. Plants may be promising source of natural antioxidants at growing stage. Oxidative stress leads to many chronic and degenerative diseases. Antioxidants are very essential for human body; they can protect the body from damage caused by free radical induced oxidative stress. This research project had been designed to investigate the antioxidant potential of gemmo modified and native [dry leaves] extract of Silybum marianum. Total phenolic contents was calculated by using Folin-Ciocalteu reagent and antioxidant potential was evaluated by using four radical assays [DPPH, ABTS, Super oxide and nitric oxide], reducing power assay and lipid peroxidation assay spectrophotometrically. Gemmo modified extract showed significantly higher [p<0.050] TPC [830 mg GAE/g of plant extract] as compared to native extract [800 mg GAE/g.] Results of this study revealed that gemmo modified extract demonstrated better antioxidant potential than natively used parts of Silybum marianum

Superoxide Anion Production by Human Neutrophils Activated by Trichomonas vaginalis

Neutrophils are the predominant inflammatory cells found in vaginal discharges of patients infected with Trichomonas vaginalis. In this study, we examined superoxide anion (O2(.-)) production by neutrophils activated by T. vaginalis. Human neutrophils produced superoxide anions when stimulated with either a lysate of T. vaginalis, its membrane component (MC), or excretory-secretory product (ESP). To assess the role of trichomonad protease in production of superoxide anions by neutrophils, T. vaginalis lysate, ESP, and MC were each pretreated with a protease inhibitor cocktail before incubation with neutrophils. Superoxide anion production was significantly decreased by this treatment. Trichomonad growth was inhibited by preincubation with supernatants of neutrophils incubated for 3 hr with T. vaginalis lysate. Furthermore, myeloperoxidase (MPO) production by neutrophils was stimulated by live trichomonads. These results indicate that the production of superoxide anions and MPO by neutrophils stimulated with T. vaginalis may be a part of defense mechanisms of neutrophils in trichomoniasis.

Knockdown of apoptosis-inducing factor disrupts function of respiratory complex I

Recent findings suggest that apoptotic protein apoptosis-inducing factor (AIF) may also play an important non-apoptotic function inside mitochondria. AIF was proposed to be an important component of respiratory chain complex I that is the major producer of superoxide radical. The possible role of AIF is still controversial. Superoxide production could be used as a valuable measure of complex I function, because the majority of superoxide is produced there. Therefore, we employed superoxide-specific mitochondrial fluorescence dye for detection of superoxide production. We studied an impact of AIF knockdown on function of mitochondrial complex I by analyzing superoxide production in selected cell lines. Our results show that tumoral telomerase-positive (TP) AIF knockdown cell lines display significant increase in superoxide production in comparison to control cells, while a non-tumoral cell line and tumoral telomerase-negative cell lines with alternative lengthening of telomeres (ALT) show a decrease in superoxide production. According to these results, we can conclude that AIF knockdown disrupts function of complex I and therefore increases the superoxide production in mitochondria. The distinct effect of AIF depletion in various cell lines could result from recently discovered activity of telomerase in mitochondria of TP cancer cells, but this hypothesis needs further investigation.

Effect of aluminum exposure on superoxide and peroxide handling capacities by liver, kidney, testis and temporal cortex in rat.

Oxidant imbalance is one of the causative mechanisms of aluminum-induced neurotoxicity. In this study, we investigated aluminum-induced oxidant imbalance in non-neuronal tissues (liver, kidney and testis) and temporal cortex in rats. The differences in adaptations to superoxide and peroxide handling capacities (SPHC) of studied organs due to aluminum insult were also evaluated. Male Wistar rats were exposed to aluminum (10 mg/Kg body wt/day) for 4 weeks through orogastric intubation. Liver showed significant decrease in reduced glutathione level, while significant alteration in lipid peroxidation was observed in temporal cortex in aluminium-exposed animals. Superoxide dismutase activity was significantly altered in liver and temporal cortex and catalase activity significantly reduced in the liver due to aluminum exposure, while glutathione reductase and glutathione peroxidase activities were altered in all the tested organs. Among the organs, glutathione-independent SPHC was relatively higher in liver and kidney, while glutathione-dependent SPHC was relatively higher in testis and temporal cortex. As compared to control, aluminum-exposed rats demonstrated reduction in glutathione-dependent SPHC in temporal cortex and increment of the same in testis, while increment in glutathione-independent SPHC was observed in liver. In conclusion, aluminum-induced alteration in oxidant handling capacity could be the cause of oxidative stress both in the neuronal and non-neuronal tissues.

Effect of ozone on response to ovalbumin & its modulation by vitamins C & E in sensitized guinea pigs.

Background & objectives: Exposure to ozone and asthma are both associated with increased oxidative stress. Exposure to ozone therefore, may potentiate the airway response to allergens. We undertook this study to investigate the effect of ozone exposure on airway response to ovalbumin in sensitized guinea pigs and its modulation by dietary supplementation with antioxidant vitamins C and E. Methods: After in vivo measurements of specific airways conductance (SGaw) and airway hyperresponsiveness (AHR) to inhaled histamine, guinea pigs were sensitized to ovalbumin and divided into three groups: (i) sensitized; (ii) sensitized and exposed daily to ozone; and (iii) sensitized, exposed daily to ozone and given dietary supplementation with vitamin C, 2 mg/kg body wt and E, 7 IU/kg body wt. A control group of nonsensitized animals was included. After 4 wk, AHR was measured again and animals were challenged with inhaled ovalbumin. Changes in SGaw were followed for early and late airway bronchoconstrictive responses. The following measurements were obtained: (i) parameters of oxidative stress - plasma malonaldehyde (MDA) as marker of lipid peroxidation and superoxide anion generation by leukocytes and bronchoalveolar lavage (BAL) cells; (ii) antioxidant status: red cell superoxide dismutase (SOD); and (iii) glutathione peroxidase (GPx). BAL cytology was studied. Results: Ozone exposure resulted in an increase in AHR and early and late bronchoconstrictive responses after ovalbumin challenge, greater superoxide anion generation in BAL cells, higher plasma MDA levels and decrease in red cell SOD activity. Dietary supplementation with vitamin C and E prevented or ameliorated these responses. Interpretation & conclusions: Exposure to ozone at concentrations of 0.12 ppm for 2 h daily for 4 wk enhances the airway response to allergens in sensitized guinea pigs. Dietary supplementation with antioxidant vitamins E and C, affords variable degree of protection against this enhancement.

Suppression of ROS generation by 4,4'-diaminodiphenylsulfone in non-phagocytic human diploid fibroblasts

The action mode of 4,4'-diaminodiphenylsulfone (DDS) is still under debate, although it has long been used in treatment of several dermatologic diseases including Hansen's disease. In this study, we tested the effect of DDS as an antioxidant on paraquat-induced oxidative stress in non-phagocytic human diploid fibroblasts (HDFs). Overall, preincubation of HDFs with DDS prevented the oxidative stress and the resulting cytotoxic damages caused by paraquat in these cells. The specific effects of DDS in paraquat-treated HDFs are summarized as follows: a) reducing the expression of NADPH oxidase 4 (NOX4) by inhibiting paraquat-induced activation of PKC; b) inhibiting paraquat-induced decreases in mitochondrial complex protein levels as well as in membrane potentials; c) consequently, inhibiting the generation of cytosolic and mitochondrial superoxide anions. Taken together, these findings suggest that DDS would suppress the radical generation in non-phagocytic HDFs during oxidative stress, and that DDS might have the extended potential to be used further in prevention of other oxidative stress-related pathologies.

Purine nucleotides reduce superoxide production by nitric oxide synthase in a murine sepsis model

Sepsis involves a systemic inflammatory response of multiple endogenous mediators, resulting in many of the injurious and sometimes fatal physiological symptoms of the disease. This systemic activation leads to a compromised vascular response and endothelial dysfunction. Purine nucleotides interact with purinoceptors and initiate a variety of physiological processes that play an important role in maintaining cardiovascular function. The purpose of the present study was to investigate the effects of ATP on vascular function in a lipopolysaccharide (LPS) model of sepsis. LPS induced a significant increase in aortic superoxide production 16 h after injection. Addition of ATP to the organ bath incubation solution reduced superoxide production by the aortas of endotoxemic animals. Reactive Blue, an antagonist of the P2Y receptor, blocked the effect of ATP on superoxide production, and the nonselective P2Y agonist MeSATP inhibited superoxide production. Nitric oxide synthase (NOS) inhibition by L-NAME blocked vascular relaxation and reduced superoxide production in LPS-treated animals. In the presence of L-NAME there was no ATP effect on superoxide production. A vascular reactivity study showed that ATP increased maximal relaxation in LPS-treated animals compared to controls. The presence of ATP induced increases in Akt and endothelial NOS phosphorylated proteins in the aorta of septic animals. ATP reduces superoxide release resulting in an improved vasorelaxant response. Sepsis may uncouple NOS to produce superoxide. We showed that ATP through Akt pathway phosphorylated endothelial NOS and “re-couples” NOS function.

Role of bilirubin as antioxidant in neonatal jaundice and effect of ethanolic extract of sweet lime peel on experimentally induced jaundice in rat.

Bilirubin above a threshold level is toxic to human system and is excreted in urinary and through gastrointestinal tract. The role of bilirubin as antioxidant is debatable. This paper aims at elucidating the role of bilirubin as an antioxidant in neonatal jaundice patients. It is observed that bilirubin up to 6 mg/dl in blood acts as an antioxidant and above 12.5 mg/dl is strongly prooxidant. Phototherapy is the accepted therapeutic management of neonatal jaundice and has been shown to enhance the oxidative stress. Approaches have been taken to formulate a herbal medication which will reduce bilirubin level in the neonates without inducing additional damages. The ethanolic extract of sweet lime peel, administered orally at a dose of 72 microg is found to reduce the oxidative stress in erythrocytes of phenylhydrazine-induced jaundiced rats treated with phototherapy.