Histological and biochemical criteria for objective and early selection of grapevine cultivars resistant to Plasmopara viticola

Grapevine breeding is the most effective way to create cultivars resistant to downy mildew (Plasmopara viticola), and to reduce the number of fungicide applications. Four criteria, including histological and biochemical analyses, based on the level of different mechanisms of resistance to grapevine downy mildew, were tested on 42 different cultivars. Plantlets were artificially inoculated with downy mildew and the sporangia density was measured spectrophotometrically 6 d after infection. Callose synthesis in stomata and δ- and ε-viniferin levels at the site of infection were recorded 48 h after inoculation. These observations have allowed the 42 cultivars to be divided into 5 groups: very resistant (VR), resistant (R), less susceptible (LS), susceptible (S) and highly susceptible (HS). All 4 criteria have to be applied to assign the resistance level closer to field conditions. This method allows to rapidly evaluate the level of resistance of seedlings to downy mildew thereby leading to a reduction in duration  of the breeding program by several years.

Compared efficacy of preservation solutions in liver transplantation: A long-term graft outcome study from the european liver transplant registry

International audienceBetween 2003 and 2012, 42 869 first liver transplantations performed in Europe with the use of either University of Wisconsin solution (UW; N = 24 562), histidine-tryptophan-ketoglutarate(HTK; N = 8696), Celsior solution (CE; N = 7756) or Institute Georges Lopez preservation solution (IGL-1; N = 1855) preserved grafts. Alternative solutions to the UW were increasingly used during the last decade. Overall, 3-year graft survival was higher with UW, IGL-1 and CE (75%, 75% and 73%, respectively), compared to the HTK (69%) (p 12 h or grafts used for patients with cancer (p < 0.0001). For partial grafts, 3-year graft survival was 89% for IGL-1, 67% for UW, 68% for CE and 64% for HTK (p = 0.009). Multivariate analysis identified HTK as an independent factor of graft loss, with recipient HIV (+), donor age ≥65 years, recipient HCV (+), main disease acute hepatic failure, use of a partial liver graft, recipient age ≥60 years, no identical ABO compatibility, recipient hepatitis B surface antigen (-), TIT ≥ 12 h, male recipient and main disease other than cirrhosis. HTK appears to be an independent risk factor of graft loss. Both UW and IGL-1, and CE to a lesser extent, provides similar results for full size grafts. For partial deceased donor liver grafts, IGL-1 tends to offer the best graft outcome

Evaluation of Hungarian Wines for Resveratrol by Overpressured Layer Chromatography

A method, including solid phase extraction sample preparation, overpressured layer chromatographic separation and subsequent densitometric evaluation, was developed for measurement of total resveratrol (cis- and trans-isomers) content of wine. The amount of resveratrol was determined in wine samples from different winemaking regions of Hungary. The total resveratrol was high in Hungarian red wines (3.6–11 mg/L), and much lower in white ones (0.04–1.5 mg/L)

Mechanisms underlying the diminished sensitivity to prolactin negative feedback during lactation: Reduced STAT5 signaling and up-regulation of cytokine-inducible SH2 domain-containing protein (CIS) expression in tuberoinfundibular dopaminergic neurons

Hyperprolactinaemia during lactation is a consequence of the sucking stimulus and in part due to reduced prolactin (PRL) negative feedback. To date, the mechanisms involved in this diminished sensitivity to PRL feedback are unknown but may involve changes in PRL signal transduction within tuberoinfundibular dopaminergic (TIDA) neurons. Therefore, we investigated signal transducers and activators of transcription (STAT) 5 signaling in the TIDA neurons of lactating rats. Dual-label confocal immunofluorescence studies were used to determine the intracellular distribution of STAT5 within TIDA neurons in the dorsomedial arcuate nucleus. In lactating rats with pups removed for 16 h, injection of ovine PRL significantly (P < 0.05) increased the STAT5 nuclear/cytoplasmic ratio compared with vehicle-treated mothers. In contrast, ovine PRL injection did not increase the STAT5 nuclear/cytoplasmic ratio in lactating mothers with pups, demonstrating that PRL signal transduction through STAT5 is reduced in TIDA neurons in the presence of pups. To investigate possible mechanisms involved in reduced PRL signaling, we examined the expression of suppressors of cytokine signaling (SOCS) proteins. Northern analysis on whole hypothalamus showed that CIS (cytokine-inducible SH2 domain-containing protein), but not SOCS1 or SOCS3, mRNA expression was significantly (P < 0.01) up-regulated in suckled lactating rats. Semiquantitative RT-PCR on arcuate nucleus micropunches also showed up-regulation of CIS transcripts. Immunofluorescence studies demonstrated that CIS is expressed in all TIDA neurons in the dorsomedial arcuate nucleus, and the intensity of CIS staining in these neurons is significantly (P < 0.05) increased in lactating rats with sucking pups. Together, these results support the hypothesis that loss of sensitivity to PRL-negative feedback during lactation is a result of increased CIS expression in TIDA neurons

Effects of Clinically Relevant MPL Mutations in the Transmembrane Domain Revealed at the Atomic Level through Computational Modeling

BACKGROUND: Mutations in the thrombopoietin receptor (MPL) may activate relevant pathways and lead to chronic myeloproliferative neoplasms (MPNs). The mechanisms of MPL activation remain elusive because of a lack of experimental structures. Modern computational biology techniques were utilized to explore the mechanisms of MPL protein activation due to various mutations. RESULTS: Transmembrane (TM) domain predictions, homology modeling, ab initio protein structure prediction, and molecular dynamics (MD) simulations were used to build structural dynamic models of wild-type and four clinically observed mutants of MPL. The simulation results suggest that S505 and W515 are important in keeping the TM domain in its correct position within the membrane. Mutations at either of these two positions cause movement of the TM domain, altering the conformation of the nearby intracellular domain in unexpected ways, and may cause the unwanted constitutive activation of MPL's kinase partner, JAK2. CONCLUSIONS: Our findings represent the first full-scale molecular dynamics simulations of the wild-type and clinically observed mutants of the MPL protein, a critical element of the MPL-JAK2-STAT signaling pathway. In contrast to usual explanations for the activation mechanism that are based on the relative translational movement between rigid domains of MPL, our results suggest that mutations within the TM region could result in conformational changes including tilt and rotation (azimuthal) angles along the membrane axis. Such changes may significantly alter the conformation of the adjacent and intrinsically flexible intracellular domain. Hence, caution should be exercised when interpreting experimental evidence based on rigid models of cytokine receptors or similar systems

Regulation of the Na,K-ATPase Gamma-Subunit FXYD2 by Runx1 and Ret Signaling in Normal and Injured Non-Peptidergic Nociceptive Sensory Neurons

Dorsal root ganglia (DRGs) contain the cell bodies of sensory neurons which relay nociceptive, thermoceptive, mechanoceptive and proprioceptive information from peripheral tissues toward the central nervous system. These neurons establish constant communication with their targets which insures correct maturation and functioning of the somato-sensory nervous system. Interfering with this two-way communication leads to cellular, electrophysiological and molecular modifications that can eventually cause neuropathic conditions. In this study we reveal that FXYD2, which encodes the gamma-subunit of the Na,K-ATPase reported so far to be mainly expressed in the kidney, is induced in the mouse DRGs at postnatal stages where it is restricted specifically to the TrkB-expressing mechanoceptive and Ret-positive/IB4-binding non-peptidergic nociceptive neurons. In non-peptidergic nociceptors, we show that the transcription factor Runx1 controls FXYD2 expression during the maturation of the somato-sensory system, partly through regulation of the tyrosine kinase receptor Ret. Moreover, Ret signaling maintains FXYD2 expression in adults as demonstrated by the axotomy-induced down-regulation of the gene that can be reverted by in vivo delivery of GDNF family ligands. Altogether, these results establish FXYD2 as a specific marker of defined sensory neuron subtypes and a new target of the Ret signaling pathway during normal maturation of the non-peptidergic nociceptive neurons and after sciatic nerve injury

Brain-Derived Neurotrophic Factor Gene Val66Met Polymorphism Modulates Reversible Cerebral Vasoconstriction Syndromes

BACKGROUND: Reversible cerebral vasoconstriction syndrome (RCVS) could be complicated by cerebral ischemic events. Hypothetical mechanisms of RCVS involve endothelial dysfunction and sympathetic overactivity, both of which were reported to be related to brain-derived neurotrophic factor (BDNF). The study investigated the association between functional BDNF Val66Met polymorphism and RCVS. METHODS: Patients with RCVS and controls were prospectively recruited and genotyped for the BDNF Val66Met polymorphism. Magnetic resonance angiography (MRA) and transcranial color-coded Doppler sonography were employed to evaluate cerebral vasoconstriction. Genotyping results, clinical parameters, vasoconstriction scores, mean flow velocities of the middle cerebral artery (V(MCA)), and Lindegaard indices were analyzed. Split-sample approach was employed to internally validate the data. PRINCIPAL FINDINGS: Ninety Taiwanese patients with RCVS and 180 age- and gender-matched normal controls of the same ethnicity completed the study. The genotype frequencies did not differ between patients and controls. Compared to patients with Met/Met homozygosity, patients with Val allele had higher mean vasoconstriction scores of all arterial segments (1.60±0.72 vs. 0.87±0.39, p<0.001), V(MCA) values (116.7±36.2 vs. 82.7±17.9 cm/s, p<0.001), and LI (2.41±0.91 vs. 1.89±0.41, p = 0.001). None of the Met/Met homozygotes, but 38.9% of the Val carriers, had V(MCA) values of >120 cm/s (p<0.001). Split-sample validation by randomization, age, entry time or residence of patients demonstrated concordant findings. CONCLUSIONS: Our findings link BDNF Val66Met polymorphism with the severity of RCVS for the first time and implicate possible pathogenic mechanisms for vasoconstriction in RCVS

Reference gene validation for quantitative RT-PCR during biotic and abiotic stresses in Vitis vinifera

Grapevine is one of the most cultivated fruit crop worldwide with Vitis vinifera being the species with the highest economical importance. Being highly susceptible to fungal pathogens and increasingly affected by environmental factors, it has become an important agricultural research area, where gene expression analysis plays a fundamental role. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is currently amongst the most powerful techniques to perform gene expression studies. Nevertheless, accurate gene expression quantification strongly relies on appropriate reference gene selection for sample normalization. Concerning V. vinifera, limited information still exists as for which genes are the most suitable to be used as reference under particular experimental conditions. In this work, seven candidate genes were investigated for their stability in grapevine samples referring to four distinct stresses (Erysiphe necator, wounding and UV-C irradiation in leaves and Phaeomoniella chlamydospora colonization in wood). The expression stability was evaluated using geNorm, NormFinder and BestKeeper. In all cases, full agreement was not observed for the three methods. To provide comprehensive rankings integrating the three different programs, for each treatment, a consensus ranking was created using a non-weighted unsupervised rank aggregation method. According to the last, the three most suitable reference genes to be used in grapevine leaves, regardless of the stress, are UBC, VAG and PEP. For the P. chlamydospora treatment, EF1, CYP and UBC were the best scoring genes. Acquaintance of the most suitable reference genes to be used in grapevine samples can contribute for accurate gene expression quantification in forthcoming studiesinfo:eu-repo/semantics/publishedVersio

Identification of molecular pathways affected by pterostilbene, a natural dimethylether analog of resveratrol

<p>Abstract</p> <p>Background</p> <p>Pterostilbene, a naturally occurring phenolic compound produced by agronomically important plant genera such as <it>Vitis </it>and <it>Vacciunium</it>, is a phytoalexin exhibiting potent antifungal activity. Additionally, recent studies have demonstrated several important pharmacological properties associated with pterostilbene. Despite this, a systematic study of the effects of pterostilbene on eukaryotic cells at the molecular level has not been previously reported. Thus, the aim of the present study was to identify the cellular pathways affected by pterostilbene by performing transcript profiling studies, employing the model yeast <it>Saccharomyces cerevisiae</it>.</p> <p>Methods</p> <p><it>S. cerevisiae </it>strain S288C was exposed to pterostilbene at the IC₅₀concentration (70 μM) for one generation (3 h). Transcript profiling experiments were performed on three biological replicate samples using the Affymetrix GeneChip Yeast Genome S98 Array. The data were analyzed using the statistical methods available in the GeneSifter microarray data analysis system. To validate the results, eleven differentially expressed genes were further examined by quantitative real-time RT-PCR, and <it>S. cerevisiae </it>mutant strains with deletions in these genes were analyzed for altered sensitivity to pterostilbene.</p> <p>Results</p> <p>Transcript profiling studies revealed that pterostilbene exposure significantly down-regulated the expression of genes involved in methionine metabolism, while the expression of genes involved in mitochondrial functions, drug detoxification, and transcription factor activity were significantly up-regulated. Additional analyses revealed that a large number of genes involved in lipid metabolism were also affected by pterostilbene treatment.</p> <p>Conclusion</p> <p>Using transcript profiling, we have identified the cellular pathways targeted by pterostilbene, an analog of resveratrol. The observed response in lipid metabolism genes is consistent with its known hypolipidemic properties, and the induction of mitochondrial genes is consistent with its demonstrated role in apoptosis in human cancer cell lines. Furthermore, our data show that pterostilbene has a significant effect on methionine metabolism, a previously unreported effect for this compound.</p