Regulation of anthocyanin and sugar accumulation in grape berry through carbon limitation and exogenous ABA application.

To optimize vineyard management practices to adapt viticulture to climate change, knowledge of the regulation mechanism of metabolite accumulation under carbon source limitation and abscisic acid (ABA) application in grapes should be deepened. Here, carbon source limitations were imposed by reducing leaf area from 12 to 2 leaves per vine (at pea sized stage, - 2L-P; or one week prior to veraison - 2L-V) and phloem girdling between the second and third leaf from bottom to top (one week prior to veraison - 12L-girdling) were compared for their effects on berry composition. All three modalities significantly reduced sugar, anthocyanin and ABA content in comparison with berries under sufficient carbon supply (12 leaves per vine - 12L), with 2L-V being the greatest. Allowing leaf area to partially recover (2L-R) or berry ABA application (400 mg. L-1) one week before veraison increased the ratio of anthocyanin to sugar under source limitation. Combined with the analysis of berry metabolites and transcript abundances, our results indicate that source limitation and exogenous ABA co-regulated anthocyanins content through differential gene expression.

Characterization of genetic determinants of the resistance to phylloxera, Daktulosphaira vitifoliae, and the dagger nematode Xiphinema index from muscadine background.

BACKGROUND: Muscadine (Muscadinia rotundifolia) is known as a resistance source to many pests and diseases in grapevine. The genetics of its resistance to two major grapevine pests, the phylloxera D. vitifoliae and the dagger nematode X. index, vector of the Grapevine fanleaf virus (GFLV), was investigated in a backcross progeny between the F1 resistant hybrid material VRH8771 (Vitis-Muscadinia) derived from the muscadine R source 'NC184-4' and V. vinifera cv. 'Cabernet-Sauvignon' (CS). RESULTS: In this pseudo-testcross, parental maps were constructed using simple-sequence repeats markers and single nucleotide polymorphism markers from a GBS approach. For the VRH8771 map, 2271 SNP and 135 SSR markers were assembled, resulting in 19 linkage groups (LG) and an average distance between markers of 0.98 cM. Phylloxera resistance was assessed by monitoring root nodosity number in an in planta experiment and larval development in a root in vitro assay. Nematode resistance was studied using 10-12 month long tests for the selection of durable resistance and rating criteria based on nematode reproduction factor and gall index. A major QTL for phylloxera larval development, explaining more than 70% of the total variance and co-localizing with a QTL for nodosity number, was identified on LG 7 and designated RDV6. Additional QTLs were detected on LG 3 (RDV7) and LG 10 (RDV8), depending on the in planta or in vitro experiments, suggesting that various loci may influence or modulate nodosity formation and larval development. Using a Bulked Segregant Analysis approach and a proportion test, markers clustered in three regions on LG 9, LG 10 and LG 18 were shown to be associated to the nematode resistant phenotype. QTL analysis confirmed the results and QTLs were thus designated respectively XiR2, XiR3 and XiR4, although a LOD-score below the significant threshold value was obtained for the QTL on LG 18. CONCLUSIONS: Based on a high-resolution linkage map and a segregating grapevine backcross progeny, the first QTLs for resistance to D. vitifoliae and to X. index were identified from a muscadine source. All together these results open the way to the development of marker-assisted selection in grapevine rootstock breeding programs based on muscadine derived resistance to phylloxera and to X. index in order to delay GFLV transmission.

Interfacial structure and photocatalytic activity of magnetron sputtered TiO2 on conducting metal substrates.

The photocatalytic behavior of magnetron sputtered anatase TiO2 coatings on copper, nickel, and gold was investigated with the aim of understanding the effect of the metallic substrate and coating-substrate interface structure. Stoichiometry and nanoscale structure of the coating were investigated using X-ray diffraction, Raman spectroscopy, atomic force microscope, and scanning and transmission electron microscopy. Photocatalytic behavior of the coating was explored by using optical spectrophotometry and electrochemical methods via photovoltage, photocurrent, and scanning kelvin probe microscopy measurements. The nature of the metal substrate and coating-substrate interface had profound influence on the photocatalytic behavior. Less photon energy was required for TiO2 excitation on a nickel substrate, whereas TiO2 coating on copper showed a higher band gap attributed to quantum confinement. However, the TiO2 coating on gold exhibited behavior typical of facile transfer of electrons to and from the CB, therefore requiring only a small amount of photon energy to make the TiO2 coating conductive.

Strontium ranelate decreases receptor activator of nuclear factor-ΚB ligand-induced osteoclastic differentiation in vitro: involvement of the calcium-sensing receptor.

Strontium ranelate exerts both an anticatabolic and an anabolic effect on bone cells. To further investigate the mechanism by which strontium ranelate inhibits bone resorption, the effects of varying concentrations of Sr(o)(2+) on osteoclastic differentiation were studied using RAW 264.7 cells and peripheral blood monocytic cells (PBMCs). We report that increasing concentrations of Sr(o)(2+) down-regulate osteoclastic differentiation and tartrate-resistant acid phosphatase activity, leading to inhibition of bone resorption (-48% when PBMCs were cultured for 14 days in the presence of 2 mM Sr(o)(2+)). Using a dominant-negative form of the calcium-sensing receptor (CaR) and a small interfering RNA approach, we provide evidences that the inhibition of osteoclast differentiation by Sr(o)(2+) is mediated by stimulation of the CaR. Moreover, our results suggest that the effects of Sr(o)(2+) on osteoclasts are, at least in part, mediated by inhibition of the receptor activator of nuclear factor-κB ligand (RANKL)-induced nuclear translocation of nuclear factor-κB and activator protein-1 in the early stages of osteoclastic differentiation. In conclusion, our data indicate that Sr(2+) directly inhibits the formation of mature osteoclasts through down-regulation of RANKL-induced osteoclast differentiation and decreases osteoclast differentiation through the activation of the CaR.

Raman imaging and Kelvin probe microscopy for the examination of the heterogeneity of doping in polycrystalline boron-doped diamond electrodes.

The issue of the heterogeneity of boron doping in microcrystalline diamond films was addressed by four different methods: micro-Raman spectroscopy and Raman imaging, Kelvin probe force microscopy, conducting atomic force microscopy, and scanning electrochemical microscopy. The samples were commercially available films from Windsor Scientific, with an average boron concentration of about 5 x 10(20) cm(-3). In agreement with previous works, all of the methods showed that the boron uptake was nonuniform across the surface of the electrode. Two different types of regions were evidenced, with metallic or semiconducting properties that were characterized with different types of Raman spectra. The line shape of these spectra was strongly dependent on the excitation wavelength. Local variations in electroactivity were evidenced by the SECM curves, which are related to the electronic properties of the individual grains, which, in turn, are governed by the boron content of the individual crystallites. In this study, two different micro-Raman imaging techniques were used that reveal the grain structure of the films: the images constructed from the diamond line intensity perfectly reproduced the optical image obtained by illuminating the sample in reflection. The method also allows detection of the presence of nondiamond carbon, especially in the metallic parts of the samples. Other spectral features (intensity of the boron-related broad lines, as well as the frequency and width of the diamond line) were used to construct images. In every case, the grain structure of the film was revealed, as well as twinning within individual crystallites. All approaches revealed that no enhanced doping or boron depletion occurred at the grain boundaries.

Flavonoid quercetin decreases osteoclastic differentiation induced by RANKL via a mechanism involving NF kappa B and AP-1.

Flavonoids are micronutrients widely present in food of plant origin. They have been attributed pharmacological properties such as anticancer and prevention of age-related pathologies. It has been recently hypothesized that flavonoids increase bone mass and prevent osteoporosis. However, little is known about the in vitro effects of flavonoids on osteoclast activities. We investigated the effects of quercetin, one of the most commonly occurring flavonoids, on osteoclast differentiation which is a critical determinant step of in vivo bone resorption. Two in vitro models of osteoclast differentiation were used in this study: a murine one, involving the culture of RAW 264.7 cells in presence of receptor activator of NF kappa B ligand (RANKL), and a human model consisting of differentiating peripheral blood monocytic cells (PBMC) isolated from peripheral blood in presence of RANKL and macrophage-colony stimulating factor (M-CSF). Osteoclastogenesis was assessed by osteoclast-like number, tartrate resistant acid phosphatase (TRAP) activity, and bone resorbing activity. We showed that quercetin (0.1-10 microM) decreased osteoclastogenesis in a dose dependent manner in both models with significant effects observed at low concentrations, from 1 to 5 microM. The IC(50) value was about 1 microM. Analysis of protein-DNA interaction by electrophoretic mobility shift assay (EMSA) performed on RAW cells showed that a pre-treatment with quercetin inhibited RANKL-induced nuclear factor kB (NF kappa B) and activator protein 1 (AP-1) activation. NF kappa B and AP-1 are transcription factors highly involved in osteoclastic differentiation and their inhibition could play an important role in the decrease of osteoclastogenesis observed in the presence of quercetin. In conclusion, the present results demonstrate for the first time that quercetin, a flavonoid characterized by antioxidant activities, is a potent inhibitor of in vitro osteoclastic differentiation, via a mechanism involving NF kappa B and AP-1.

Nucleosides and ODN electrochemical detection onto boron doped diamond electrodes.

Boron doped diamond (BDD) is a promising material for electroanalytical chemistry due mainly to its chemical stability, its high electrical conductivity and to the large amplitude of its electroactive window in aqueous media. The latter feature allowed us to study the direct oxidation of the two electroactive nucleosides, guanosine and adenosine. The BDD electrode was first activated by applying high oxidizing potentials, allowing to increase anodically its working potential window through the oxidation of CH surface groups into hydroxyl and carbonyl terminations. Guanosine (1.2 V vs. Ag/AgCl) and adenosine (1.5 V vs. Ag/AgCl) could then be detected electrochemically with an acceptable signal to noise ratio. The electrochemical signature of each oxidizable base was assessed using differential pulse voltammetry (DPV), in solutions containing one or both nucleosides. These experiments pointed out the existence of adsorption phenomena of the oxidized products onto the diamond surface. Scanning electrochemical microscopy (SECM) was used to investigate these adsorption effects at the microscopic scale. The usefulness of BDD electrodes for the direct electrochemical detection of synthetic oligonucleotides is also evidenced.

Biotin grafting on boron-doped diamond.

Grafting of biotin on top of a polycrystalline boron-doped diamond layer was achieved by surface oxidation followed by an esterification reaction and revealed by fluorescently labelled streptavidin.

Potent inhibitory effect of naturally occurring flavonoids quercetin and kaempferol on in vitro osteoclastic bone resorption.

Several recent studies have suggested that flavonols, a class of phytochemicals with many biological activities, might exert a protective effect against post-menopausal bone loss. In the present study, we investigated the effects of quercetin and kaempferol, two of the major naturally occurring flavonols on the in vitro bone resorbing activity of osteoclasts. Our results indicate that both compounds, at concentrations ranging from 0.1 to 100 microM reduce bone resorption in a time and dose-dependent manner. Significant inhibitory effects were observed at concentrations as low as 0.1 microM especially with kaempferol. The IC(50)s, or concentration inhibitory of 50% of basal resorption, calculated for quercetin and kaempferol were 1.6 and 5.3 microM, respectively. Using highly purified rabbit osteoclasts, we showed that both flavonols directly induce apoptosis of mature osteoclasts in the same dose-range effective for inhibiting bone resorption. When osteoclasts were treated with 50 microM of quercetin and kaempferol, intracellular reactive oxygen species levels decreased significantly by 75 and 25%, respectively, indicating these molecules keep their antioxidant properties at this concentration. However, at concentrations below 50 microM, neither quercetin nor kaempferol exerted antiradical action, suggesting that antioxidant properties cannot fully explain the inhibitory effect on bone resorption. Finally, we report that kaempferol-, but not the quercetin-induced inhibition of bone resorption was partially abolished by the presence of the pure anti-estrogen ICI 182780 suggesting that kaempferol's estrogenic effect could be involved in the inhibition of bone resorption. The present study demonstrates that flavonols widely distributed in human diet such as quercetin and kaempferol, exert a potent inhibitory effect on in vitro bone resorption.