Indirect emissions of nitrous oxide in a cropland watershed with contrasting hydrology in central France.

Nitrous oxide (N2O) is an important greenhouse gas. Its atmospheric concentration have increased with the industrialisation and the use of N fertilizer. The contribution of freshwater systems to N2O emissions is still very uncertain, while regional transfer of nitrogen depends on soil and hydrology. Riverine and spring N2O dissolved in water was therefore measured over two years in the 3453 km2 Haut-Loir watershed (France). This temperate cropland watershed is characterized by two different hydrological systems east and west of the Loir River. The eastern rivers, fed by the emergence of the deep Beauce aquifer, exhibited significantly higher dissolved N2O concentrations (Beauce region, mean: 2.93 µg-N L-1) than the western rivers (Perche region, mean: 0.87 µg-N L-1), which were largely influenced by runoff during winter flooding. The eastern rivers had large nitrate concentrations all over the year; in the Perche, nitrate underwent a seasonal cycle with large loads during winter floods, but there were no consistent seasonal patterns in N2O. The ratios of N2O in excess of equilibrium on nitrate, often used as a proxy of emission factor (EF), were much smaller than the default IPCC values, both for rivers (0.014% versus 0.25% for IPCC EF5r) and the Loir spring (0.085% versus 0.6% for the IPCC EF5g for groundwater and springs). EF5r were significantly different between the two parts of the watershed only in winter, because of the seasonal variability of NO3-. Moreover dissolved N2O is controlled not only by NO3-, as it is considered in the calculation of the EF5, but also by water pH and dissolved organic carbon. A good prediction of dissolved N2O was obtained using these physicochemical variables and hydrological regions. Thus, these results suggest that the spatial variability of riverine N2O depends on local hydrology, while further research is needed to understand the seasonal variability.

Xanthine Oxidase-Derived ROS Display a Biphasic Effect on Endothelial Cells Adhesion and FAK Phosphorylation.

In pathological situations such as ischemia-reperfusion and acute respiratory distress syndrome, reactive oxygen species (ROS) are produced by different systems which are involved in endothelial cells injury, ultimately leading to severe organ dysfunctions. The aim of this work was to study the effect of ROS produced by hypoxanthine-xanthine oxidase (Hx-XO) on the adhesion of human umbilical vein endothelial cells (HUVEC) and on the signaling pathways involved. Results show that Hx-XO-derived ROS induced an increase in HUVEC adhesion in the early stages of the process (less than 30 min), followed by a decrease in adhesion in the later stages of the process. Interestingly, Hx-XO-derived ROS induced the same biphasic effect on the phosphorylation of the focal adhesion kinase (FAK), a nonreceptor tyrosine kinase critical for cell adhesion, but not on ERK1/2 phosphorylation. The biphasic effect was not seen with ERK1/2 where a decrease in phosphorylation only was observed. Wortmannin, a PI3-kinase inhibitor, inhibited ROS-induced cell adhesion and FAK phosphorylation. Orthovanadate, a protein tyrosine phosphatase inhibitor, and Resveratrol (Resv), an antioxidant agent, protected FAK and ERK1/2 from dephosphorylation and HUVEC from ROS-induced loss of adhesion. This study shows that ROS could have both stimulatory and inhibitory effects on HUVEC adhesion and FAK phosphorylation and suggests that PI3-kinase and tyrosine phosphatase control these effects.

Antioxidant effect of hydroxytyrosol, a polyphenol from olive oil: scavenging of hydrogen peroxide but not superoxide anion produced by human neutrophils.

Hydroxytyrosol (HT) (also known as dihydroxyphenylethanol (DPE)) is a polyphenol extracted from virgin olive oil. HT is known to exert an antioxidant effect but the mechanism of action and the identity of the reactive oxygen molecule(s) targeted are not known. In this study, we show that HT inhibits luminol-amplified chemiluminescence of human neutrophils stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP), phorbol myristate acetate (PMA) and opsonized zymosan. This effect was dose-dependent and occurred immediately after the addition of HT. However, HT had no effect on lucigenin-amplified chemiluminescence, suggesting that it does not inhibit NADPH oxidase activation or scavenge superoxide anions. Furthermore, HT inhibited H(2)O(2)-dependent-dichlorofuoroscein (DCFH) fluorescence of activated neutrophils, as measured by flow cytometry. Finally, HT inhibited luminol-amplified chemiluminescence in a cell-free system consisting of H(2)O(2) and HRPO. These results suggest that HT, a polyphenol derived from olive oil, could exert its antioxidant effect by scavenging hydrogen peroxide but not superoxide anion released during the respiratory burst.

Measurement of copper(I) formation as a test for the stability of catecholestrogens and methoxyestrogens in solution.

The biological effects of estrogens seem to be divided into three mechanisms of action: (1) the transcriptional action by the estrogen-estrogen receptor (ER) complex, (2) the non-genomic mechanism through ERs in cell membranes, and (3) the ER-independent mechanism. The latter mechanism has been attributed to be mediated by the basic chemical properties of estradiol (E2) metabolites, which seems to include their pro- and anti-oxidant properties. Therefore, in order to study the ER-independent actions of the E2 metabolites, their redox properties must be conserved. In this study, we have developed a test to measure the electron-donating properties of E2 and its metabolites based on the reduction of Cu(II) ion into Cu(I). Our results show that the catechol- and methoxy-metabolites of E2 lose their capability to reduce Cu(II) into Cu(I) after 3 months of storage at -20 degrees C. Thus, we propose this inexpensive and reliable test to verify the electron-donating properties of E2 metabolites in order to study their ER-independent biological effects in vitro.

Blockade of vascular endothelial growth factor receptor I (VEGF-RI), but not VEGF-RII, suppresses joint destruction in the K/BxN model of rheumatoid arthritis.

It was recently shown that vascular endothelial growth factor (VEGF), a growth factor for endothelial cells, plays a pivotal role in rheumatoid arthritis. VEGF binds to specific receptors, known as VEGF-RI and VEGF-RII. We assessed the physical and histological effects of selective blockade of VEGF and its receptors in transgenic K/BxN mice, a model of rheumatoid arthritis very close to the human disease. Mice were treated with anti-mouse VEGF Ab, anti-mouse VEGF-RI and -RII Abs, and an inhibitor of VEGF-RI tyrosine kinase. Disease activity was monitored using clinical indexes and by histological examination. We found that synovial cells from arthritic joints express VEGF, VEGF-RI, and VEGF-RII. Treatment with anti-VEGF-RI strongly attenuated the disease throughout the study period, while anti-VEGF only transiently delayed disease onset. Treatment with anti-VEGF-RII had no effect. Anti-VEGF-RI reduced the intensity of clinical manifestations and, based on qualitative and semiquantitative histological analyses, prevented joint damage. Treatment with a VEGF-RI tyrosine kinase inhibitor almost abolished the disease. These results show that VEGF is a key factor in pannus development, acting through the VEGF-RI pathway. The observation that in vivo administration of specific inhibitors targeting the VEGF-RI pathway suppressed arthritis and prevented bone destruction opens up new possibilities for the treatment of rheumatoid arthritis.

Pharmacological potentiation of natriuretic peptide limits polymorphonuclear neutrophil-vascular cell interactions.

OBJECTIVE: Activated polymorphonuclear neutrophils (PMNs) are the main source of circulating neutral endopeptidase (NEP). We tested the hypothesis that NEP inhibition could potentiate the effect of atrial natriuretic peptide (ANP) on PMN-vascular cell interactions in vitro. METHODS AND RESULTS: ANP alone and its potentiation by retrothiorphan, the NEP inhibitor, significantly inhibited superoxide, lysozyme, and matrix metalloproteinase (MMP)-9 release by N-formyl-Met-Leu-Phe-stimulated PMNs. Activated PMNs degraded exogenous ANP, which was prevented by NEP inhibition. Hypoxia significantly increased the adhesion of PMNs to endothelial cells and their subsequent MMP-9 release by 60% and 150%, respectively (P<0.01). ANP and its potentiation by retrothiorphan limited PMN adhesion to hypoxic endothelial cells and thus decreased their MMP-9 release (P<0.01). Smooth muscle cells (SMCs) incubated with conditioned medium of N-formyl-Met-Leu-Phe-stimulated PMNs exhibited morphological and biochemical changes characteristic of apoptosis (terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling positivity, nuclear condensation/fragmentation, poly ADP-ribose polymerase cleavage, and DNA laddering). SMC detachment and subsequent apoptosis could be related to leukocyte elastase-induced pericellular proteolysis, inasmuch as both events are inhibited by elastase inhibitors. ANP and its potentiation by retrothiorphan were able to limit elastase release, fibronectin degradation, and SMC apoptosis. CONCLUSIONS: ANP potentiation by NEP inhibition could limit PMN activation and its consequences on vascular cells.

Vitamin E uncouples joint destruction and clinical inflammation in a transgenic mouse model of rheumatoid arthritis.

OBJECTIVE: Reactive oxygen species are thought to play a role in rheumatoid arthritis (RA) in humans. We postulated that antioxidant treatment could have a beneficial effect in this disease. We therefore investigated the effects of vitamin E in the transgenic KRN/NOD mouse model of RA. METHODS: Mice were treated by gavage with oral vitamin E (alpha-tocopherol). Clinical, histologic, and biochemical parameters were assessed for 6 weeks. RESULTS: Vitamin E treatment did not modify the clinical features of the disease (date of onset or disease intensity, as measured by the articular index), but it did prevent joint destruction, as measured by qualitative and semiquantitative analyses. Redox status did not differ between treated and control mice. White blood cell chemiluminescence was higher in transgenic KRN/NOD mice than in controls, but vitamin E treatment attenuated this difference. Vitamin E treatment of the transgenic animals led to a significant decrease in the levels of interleukin-(IL-1beta) but not tumor necrosis factor alpha. CONCLUSION: Vitamin E seems to uncouple joint inflammation and joint destruction in this model of RA, with a beneficial effect on joint destruction. Since many investigations are currently in progress to evaluate the benefit of interventions targeted toward anti-IL-1beta, our findings suggest opportunities of therapeutic interest in human RA.