Bacteria-derived hydrogen sulfide promotes IL-8 production from epithelial cells

Hydrogen Sulfide (H(2)S) a volatile Sulfur compound, is implicated as a cause of inflammation. especially when it is produced by bacteria colonizing gastrointestinal organs However, It IS Unclear if H(2)S produced by periodontal pathogens affects the inflammatory responses mediated by oral/gingival epithelial cells Therefore. the aims of this Study were (1) to compare the in vitro production of H(2)S among. 14 strains of Oral bacteria and (2) to evaluate the effects of H(2)S on inflammatory response induced in host oral/gingival epithelial cells Porphyromonas gingivalis (Pg) produced the most H(2)S in Culture, Which, in turn resulted in the promotion of proinflammatory cytokine IL-8 from both gingival and Oral epithelial cells The up-regulation of IL-8 expression was reproduced by the exogenously applied H(2)S Furthermore. the Mutant Strains of Pg that do not produce major Soluble Virulent factors. ie gingival, still showed the Production of H(2)S. as well as the promotion of epithelial IL-8 production. which was abrogated by H(2)S scavenging reagents These results demonstrated that Pg produces a concentration of H(2)S capable of Up-regulating-IL-8 expression induced in gingival and oral epithelial cells, revealing a possible mechanism that may promote the inflammation in periodontal disease (C) 2009 Elsevier B.V. All rights reserved

Release of intermediate reactive hydrogen peroxide by macrophage cells activated by natural products

By determining the hydrogen peroxide (H2O2) released in cultures of peritoneal macrophage cells from Swiss mice, we evaluated the action of 27 vegetable compounds (pristimerin, tingenone, jatrophone, palustric acid, lupeol, cladrastin, ocoteine, boldine, tomatine, yohimbine, reserpine, escopoletin, esculine, plumericin, diosgenin, deoxyschizandrin, p-arbutin, mangiferin, and others) using a 2 mg/ml solution of each compound (100 mug/well). Macrophages are cells responsible for the development of the immunological response reaction, liberating more than one hundred compounds into the extracellular environment. Among these are the various cytokines and the intermediate compounds of nitrogen (NO) and oxygen (H2O2). This coordinated sequence of biochemical reactions is known as the oxidative burst. When we compared the results with those obtained with zymosan (an important stimulator of H2O2) we observed that the compounds showing the highest activity were substances 2 (tingenone), 16 (reserpine) and 20. Other substances such as compounds 1, 4, 5, 6, 8, 12, 13, 14, 15, 17, 19, 23, 24, 26, and 27 also showed a certain activity, but with less intensity than the aforementioned ones. Compounds 3, 7, 9, 10, 11, 18, 21, 22 and 25 presented no activity. These results suggest that natural products (mainly tingenone and reserpine and others) with different chemical structures are strong immunological modulators. However, further tests are needed to determine the 'oxidative burst' in future studies.

Indirect cytotoxicity of a 35% hydrogen peroxide bleaching gel on cultured odontoblast-like cells

The aim of this study was to evaluate the trans-enamel and trans-dentinal effects of a 35% hydrogen peroxide (H2O2) bleaching gel on odontoblast-like cells. Enamel/dentin discs obtained from bovine incisors were mounted in artificial pulp chambers (APCs). Three groups were formed: G1- 35% H2O2; G2- 35% H2O2 + halogen light application; G3- control. The treatments were repeated 5 times and the APCs were incubated for 12 h. Then, the extract was collected and applied for 24 h on the cells. Cell metabolism, total protein dosage and cell morphology were evaluated. Cell metabolism decreased by 62.09% and 61.83% in G1 and G2, respectively. The depression of cell metabolism was statistically significant when G1 and G2 were compared to G3. Total protein dosage decreased by 93.13% and 91.80% in G1 and G2, respectively. The cells in G1 and G2 exhibited significant morphological alterations after contact with the extracts. Regardless of halogen light application, the extracts caused significantly more intense cytopathic effects compared to the control group. After 5 consecutive applications of a 35% H2O2 bleaching agent, either catalyzed or not by halogen light, products of gel degradation were capable to diffuse through enamel and dentin causing toxic effects to the cells.

Indirect cytocompatibility of a low-concentrated hydrogen peroxide bleaching gel to odontoblast-like cells

Aim To assess the initial cytotoxicity and the late phenotype marker expression of odontoblast-like cells (MDPC-23) subjected to less aggressive in-office bleaching therapies. Methodology A 17.5% hydrogen peroxide (H2O2) gel was applied for 45, 15 or 5 min to enamel/dentine discs adapted to trans-wells positioned over cultured MDPC-23 cells. No treatment was performed on the negative control. Immediately after bleaching, the cell viability, gene expression of inflammatory mediators and quantification of H2O2 diffusion were evaluated. The ALP activity, DSPP and DMP-1 gene expression and mineralized nodule deposition (MND) were assessed at 7, 14 or 21 days post-bleaching and analysed statistically with Mann–Whitney U-tests (α = 5%). Results H2O2 diffusion, proportional to treatment time, was observed in all bleached groups. Reductions of approximately 31%, 21% and 13% in cell viability were observed for the 45-, 15- and 5-min groups, respectively. This reduction was significant (P < 0.05) for the 45- and 15-min groups, which also presented significant (P < 0.05) over-expression of inflammatory mediators. The 45-min group was associated with significant (P < 0.05) reductions in DMP-1/DSPP expression at all periods, relative to control. The ALP activity and MND were reduced only in initial periods. The 15-min group had less intense reduction of all markers, with no difference to control at 21 days. Conclusions The 17.5% H2O2 applied to tooth specimens for 5 min caused no alteration in the odontoblast-like cells. When this gel was applied for 45 or 15 min, a slight cytotoxicity, associated with alterations in phenotypic markers, was observed. However, cells were able to recover their functions up to 21 days post-bleaching.

Effect of hydrogen-peroxide-mediated oxidative stress on human dental pulp cells

To evaluate the effect of the oxidative stress on human dental pulp cells (HDPCs) promoted by toxic concentrations of hydrogen peroxide (H2O2) on its odontoblastic differentiation capability through time. Methods HDPCs were exposed to two different concentrations of H2O2 (0.1 and 0.3 μg/ml) for 30 min. Thereafter, cell viability (MTT assay) and oxidative stress generation (H2DCFDA fluorescence assay) were immediately evaluated. Data were compared with those for alkaline phosphatase (ALP) activity (thymolphthalein assay) and mineralized nodule deposition (alizarin red) by HDPCs cultured for 7 days in osteogenic medium. Results A significant reduction in cell viability and oxidative stress generation occurred in the H2O2-treated cells when compared with negative controls (no treatment), in a concentration-dependent fashion. Seven days after H2O2 treatment, the cells showed significant reduction in ALP activity compared with negative control and no mineralized nodule deposition. Conclusion Both concentrations of H2O2 were toxic to the cells, causing intense cellular oxidative stress, which interfered with the odontogenic differentiation capability of the HDPCs. Clinical significance The intense oxidative stress on HDPCs mediated by H2O2 at toxic concentrations promotes intense reduction on odontoblastic differentiation capability in a 7-day evaluation period, which may alter the initial pulp healing capability in the in vivo situation.

Dose-Response and Time-Course of a-Tocoferol Mediating the Cytoprotection Of Dental Pulp Cells Against Hydrogen Peroxide

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Baccharin Prevents Genotoxic Effects Induced by Methyl Methanesulfonate and Hydrogen Peroxide in V79 Cells

Baccharin is one of the major chemical compounds isolated from the aerial parts of Baccharis dracunculifolia DC (Asteraceae), a native plant of South America and the most important botanical source of the Brazilian green propolis that has been used in alternative medicine to treat inflammation, liver disorders, and stomach ulcers. The present study was carried out in V79 cells to determine the possible genotoxic and antigenotoxic activities of baccharin utilizing comet and micronucleus assays, where 2 known mutagenic agents with different mechanisms of DNA damage were used as positive controls. The V79 cells were treated with concentrations of baccharin (0.25, 0.5, 1.0, and 2.0 mu g/mL) and for to investigate the antigenotoxicity these concentrations were associated with methyl methanesulfonate (MMS; 200 mu M-comet assay and 400 mu M-micronucleus assay) or hydrogen peroxide (H2O2; 50 mu M-comet assay and 100 mu M-micronucleus assay). Statistically significant differences in the rate of DNA damage were observed in cultures treated with the highest concentration of baccharin when compared to the control group, but this difference was not found in the micronucleus assay. The results also showed that the frequencies of DNA damage and micronuclei induced by MMS and H2O2 were significantly reduced after treatment with baccharin. The baccharin showed a chemoprevention effect and can be the chemical compound responsible for the antigenotoxicity also demonstrated by the B. dracunculifolia. The antioxidant potential of baccharin may be related to its chemoprevention activity induced against both genomic and chromosomal damages.

Sodium/hydrogen exchanger NHA2 is critical for insulin secretion in β-cells

NHA2 is a sodium/hydrogen exchanger with unknown physiological function. Here we show that NHA2 is present in rodent and human β-cells, as well as β-cell lines. In vivo, two different strains of NHA2-deficient mice displayed a pathological glucose tolerance with impaired insulin secretion but normal peripheral insulin sensitivity. In vitro, islets of NHA2-deficient and heterozygous mice, NHA2-depleted Min6 cells, or islets treated with an NHA2 inhibitor exhibited reduced sulfonylurea- and secretagogue-induced insulin secretion. The secretory deficit could be rescued by overexpression of a wild-type, but not a functionally dead, NHA2 transporter. NHA2 deficiency did not affect insulin synthesis or maturation and had no impact on basal or glucose-induced intracellular Ca(2+) homeostasis in islets. Subcellular fractionation and imaging studies demonstrated that NHA2 resides in transferrin-positive endosomes and synaptic-like microvesicles but not in insulin-containing large dense core vesicles in β-cells. Loss of NHA2 inhibited clathrin-dependent, but not clathrin-independent, endocytosis in Min6 and primary β-cells, suggesting defective endo-exocytosis coupling as the underlying mechanism for the secretory deficit. Collectively, our in vitro and in vivo studies reveal the sodium/proton exchanger NHA2 as a critical player for insulin secretion in the β-cell. In addition, our study sheds light on the biological function of a member of this recently cloned family of transporters.

Microfluidic hydrogen generator : featuring passive on-demand gas generation and self-circulated reactant for integration with micro fuel cell

A microfluidic hydrogen generator is presented in this work. Its fabrication, characterization, and integration with a micro proton exchange membrane (PEM) fuel cell are described. Hydrogen gas is generated by the hydrolysis of aqueous ammonia borane. Gas generation, as well as the circulation of ammonia borane from a rechargeable fuel reservoir, is performed without any power consumption. To achieve this, directional growth and selective venting of hydrogen gas is maintained in the microchannels, which results in the circulation of fresh reactant from the fuel reservoir. In addition to this self-circulation mechanism, the hydrogen generator has been demonstrated to self-regulate gas generation to meet demands of a connected micro fuel cell. All of this is done without parasitic power consumption from the fuel cell. Results show its feasibility in applications of high-impedance systems. Lastly, recommendations for improvements and suggestions for future work are described

Determination of the hoop fracture properties of unirradiated hydrogen-charged nuclear fuel cladding from ring compression tests

In this work, a new methodology is devised to obtain the fracture properties of nuclear fuel cladding in the hoop direction. The proposed method combines ring compression tests and a finite element method that includes a damage model based on cohesive crack theory, applied to unirradiated hydrogen-charged ZIRLOTM nuclear fuel cladding. Samples with hydrogen concentrations from 0 to 2000 ppm were tested at 20 �C. Agreement between the finite element simulations and the experimental results is excellent in all cases. The parameters of the cohesive crack model are obtained from the simulations, with the fracture energy and fracture toughness being calculated in turn. The evolution of fracture toughness in the hoop direction with the hydrogen concentration (up to 2000 ppm) is reported for the first time for ZIRLOTM cladding. Additionally, the fracture micromechanisms are examined as a function of the hydrogen concentration. In the as-received samples, the micromechanism is the nucleation, growth and coalescence of voids, whereas in the samples with 2000 ppm, a combination of cuasicleavage and plastic deformation, along with secondary microcracking is observed.

Escherichia coli exhibits negative chemotaxis in gradients of hydrogen peroxide, hypochlorite, and N-chlorotaurine: products of the respiratory burst of phagocytic cells.

Escherichia coli can respond to gradients of specific compounds, moving up gradients of attractants and down gradients of repellents. Stimulated phagocytic leukocytes produce H2O2, OCl-, and N-chlorotaurine in a response termed the respiratory burst. E. coli is actively repelled by these compounds. Catalase in the suspending medium eliminated the effect of H2O2. Repulsion by H2O2 could be demonstrated with 1 microM H2O2, which is far below the level that caused overt toxicity. Strains with defects in the biosynthesis of glutathione or lacking hydroperoxidases I and II retained this response to H2O2, and 2.0 mM CN- did not interfere with it. Mutants with defects in any one of the four known methyl-accepting chemotaxis proteins also retained the ability to respond to H2O2, but a "gutted" mutant that was deleted for all four methyl-accepting chemotaxis proteins, as well as for CheA, CheW, CheR, CheB, CheY, and CheZ, did not respond to H2O2. Hypochlorite and N-chlorotaurine were also strongly repellent. Chemotaxis down gradients of H2O2, OCl-, and N-chlorotaurine may contribute to the survival of commensal or pathogenic microorganisms.

Endogenous intracellular glutathionyl radicals are generated in neuroblastoma cells under hydrogen peroxide oxidative stress.

We report the detection of endogenous intracellular glutathionyl (GS.) radicals in the intact neuroblastoma cell line NCB-20 under oxidative stress. Spin-trapping and electron paramagnetic resonance (EPR) spectroscopic methods were used for monitoring the radicals. The cells incubated with the spin trap 5,5-dimethyl-1-pyrroline 1-oxide (DMPO) were challenged with H2O2 generated by the enzymic reaction of glucose/glucose oxidase. These cells exhibit the EPR spectrum of the GS. radical adduct of DMPO (DMPO-.SG) without exogenous reduced glutathione (GSH). The identity of this radical adduct was confirmed by observing hyperfine coupling constants identical to previously reported values in in vitro studies, which utilized known enzymic reactions, such as horseradish peroxidase and Cu/Zn superoxide dismutase, with GSH and H2O2 as substrates. The formation of the GS. radicals required viable cells and continuous biosynthesis of GSH. No significant effect on the resonance amplitude by the addition of a membrane-impermeable paramagnetic broadening agent indicated that these radicals were located inside the intact cell. N-Acetyl-L-cysteine (NAC)-treated cells produced NAC-derived free radicals (NAC.) in place of GS. radicals. The time course studies showed that DMPO-.SG formation exhibited a large increase in its concentration after a lag period, whereas DMPO-NAC. formation from NAC-treated cells did not show this sudden increase. These results were discussed in terms of the limit of antioxidant enzyme defenses in cells and the potential role of the GS. radical burst in activation of the transcription nuclear factor NF-kappa B in response to oxidative stress.

Amorphous silicon solar cells by hydrogen implantation : final report, January 1, 1979-August 31, 1980 /

"August 1980."

Azurin of pathogenic Neisseria spp. is involved in defense against hydrogen peroxide and survival within cervical epithelial cells

Laz, a lipid-modified azurin of the human pathogens Neisseria gonorrhoeae and Neisseria meningitidis, is involved in defense against oxidative stress and copper toxicity; laz mutant strains are hypersensitive to hydrogen peroxide and copper. The N. gonorrhoeae laz mutant also has decreased survival in an ex vivo primary human ectocervical epithelial assay.

Activation of ERK1/2, JNK and PKB by hydrogen peroxide in human SH-SY5Y neuroblastoma cells:role of ERK1/2 in H2O2-induced cell death

Reactive oxygen species including H2O2 activate an array of intracellular signalling cascades that are closely associated with cell death and cell survival pathways. The human neuroblastoma SH-SY5Y cell line is widely used as model cell system for studying neuronal cell death induced by oxidative stress. However, at present very little is known about the signalling pathways activated by H2O2 in SH-SY5Y cells. Therefore, in this study we have investigated the effect of H2(O2 on extracellular signal-regulated kinase 1/2 (ERK1/2), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38 MAPK) and protein kinase B (PKB) activation in undifferentiated and differentiated SH-SY5Y cells. H2O2 stimulated time and concentration increases in ERK1/2, JNK and PKB phosphorylation in undifferentiated and differentiated SH-SY5Y cells. No increases in p38 MAPK phosphorylation were observed following H2O2 treatment. The phosphatidylinositol 3-kinase (PI-3K) inhibitors wortmannin and LY 294002 ((2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one) inhibited H2O2-induced increases in ERK1/2 and PKB phosphorylation. Furthermore, H2O2-mediated increases in ERK1/2 activation were sensitive to the MAPK kinase 1 (MEK1) inhibitor PD 98059 (2'-amino-3'-methoxyflavone), whereas JNK responses were blocked by the JNK inhibitor SP 600125 (anthra[1-9-cd]pyrazol-6(2H)-one). Treatment of SH-SY5Y cells with H2O2 (1 mM; 16 h) significantly increased the release of lactate dehydrogenase (LDH) into the culture medium indicative of a decrease in cell viability. Pre-treatment with wortmannin, SP 600125 or SB 203580 (4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole; p38 MAPK inhibitor) had no effect on H2O2-induced LDH release from undifferentiated or differentiated SH-SY5Y cells. In contrast, PD 98059 and LY 294002 significantly decreased H2O2-induced cell death in both undifferentiated and differentiated SH-SY5Y cells. In conclusion, we have shown that H2O2 stimulates robust increases in ERK1/2, JNK and PKB in undifferentiated and differentiated SH-SY5Y cells. Furthermore, the data presented clearly suggest that inhibition of the ERK1/2 pathway protects SH-SY5Y cells from H2O2-induced cell death.