Increasing evidence implicates decreased energy metabolism and mitochondrial dysfunctions among the earliest pathogenic events of Alzheimer's disease (AD). However, the molecular mechanisms underlying bioenergetic dysfunctions in AD remain, to date, largely unknown. In this work, we analyzed transcriptomic changes occurring in the hippocampus and retina of a Tg2576 AD mouse model and wild-type controls, evaluating their functional implications by gene set enrichment analysis. The results revealed that oxidative phosphorylation and mitochondrial-related pathways are significantly down-regulated in both tissues of Tg2576 mice, supporting the role of these processes in the pathogenesis of AD. In addition, we also analyzed transcriptomic changes occurring in Tg2576 mice treated with the 12A12 monoclonal antibody that neutralizes an AD-relevant tau-derived neurotoxic peptide in vivo. Our analysis showed that the mitochondrial alterations observed in AD mice were significantly reverted by treatment with 12A12mAb, supporting bioenergetic pathways as key mediators of its in vivo neuroprotective and anti-amyloidogenic effects. This study provides, for the first time, a comprehensive characterization of molecular events underlying the disrupted mitochondrial bioenergetics in AD pathology, laying the foundation for the future development of diagnostic and therapeutic tools.

Xylella fastidiosa is a xylem-limited plant-pathogenic bacterium causing severe diseases in several crops. Infections in olives cause the olive quick decline syndrome, a disease that compromises the survival of susceptible trees. This represents one of the most challenging pathosystems to study, because infections have a long incubation period (i.e. > 1 year before any visual alteration induced by the bacterium can be detected) and bacterial virulence factors/host response mechanisms are largely unknown. Even so, it is extensively documented that the formation of cell aggregates in the vessels and the degradation of the cell wall cause the hydraulic collapse of the xylem network. Such phenomena are more prominent in the susceptible hosts, while resistant/tolerant genotypes may remain symptomless or show mild symptoms. We aimed to explore the use of different physiological traits, that combined with molecular markers, may identify biomarkers suitable for the early prediction of the host response. Phenotypic measurements likewise stomatal conductance, stem water potential, leaf and canopy temperature, electrical signal changes in the sap flux monitored by electrical impedance spectroscopy and massive transcriptomic data have been generated from plants under different infection conditions. Efforts are directed to integrate transcriptomic and physiological data, in a multiomic approach, for rapid and accurate predictions of resistant vs susceptible phenotype.

Drug-eluting bioresorbable vascular scaffolds (BVSs) have emerged as a potential breakthrough for the treatment of coronary artery stenosis, providing mechanical support and drug delivery followed by complete resorption. Restenosis and thrombosis remain the primary limitations in clinical use. The study aimed to identify potential markers of restenosis and thrombosis analyzing the vascular wall cell transcriptomic profile modulation triggered by BVS at different values of shear stress (SS). Human coronary artery endothelial cells and smooth muscle cells were cultured under SS (1 and 20 dyne cm-2) for 6 h without and with application of BVS and everolimus 600 nM. Cell RNA-Seq and bioinformatics analysis identified modulated genes by direct comparison of SS conditions and Gene Ontology (GO). The results of different experimental conditions and GO analysis highlighted the modulation of specific genes as semaphorin 3E, mesenchyme homeobox 2, bone morphogenetic protein 4, (heme oxygenase 1) and selectin E, with different roles in pathological evolution of disease. Transcriptomic analysis of dynamic vascular cell cultures identifies candidate genes related to pro-restenotic and pro-thrombotic mechanisms in an in-vitro setting of BVS, which are not adequately contrasted by everolimus addition.

Nitrogen-use efficiency (NUE) is a complex trait of great interest in breeding programs because through its improvement, high crop yields can be maintained whilst N supply is reduced. In this study, we report a transcriptomic analysis of four NUE-contrasting eggplant (Solanum melongena) genotypes following short- and long-term exposure to low N, to identify key genes related to NUE in the roots and shoots. The differentially expressed genes in the high-NUE genotypes are involved in the light-harvesting complex and receptor, a ferredoxin-NADP reductase, a catalase and WRKY33. These genes were then used as bait for a co-expression gene network analysis in order to identify genes with the same trends in expression. This showed that up-regulation of WRKY33 triggered higher expression of a cluster of 21 genes and also of other genes, many of which were related to N-metabolism, that were able to improve both nitrogen uptake efficiency and nitrogen utilization efficiency, the two components of NUE. We also conducted an independent de novo experiment to validate the significantly higher expression of WRKY33 and its gene cluster in the high-NUE genotypes. Finally, examination of an Arabidopsis transgenic 35S::AtWRKY33 overexpression line showed that it had a bigger root system and was more efficient at taking up N from the soil, confirming the pivotal role of WRKY33 for NUE improvement.

This Deliverable reports the activity of Task 3.3: "In vitro models and molecular pathways of thrombosis and restenosis (protein)". This task is devoted to obtain a panel of molecular markers potentially involved in restenosis and in-stent thrombosis of drug-eluting Bioresorbable Vascular Scaffold (BVS) and to supply it for the development of the silico module implemented in Work Package 6. In order to achieve this objective, gene expression profile is obtained in cultured vascular wall cells, namely human coronary artery endothelial cells (HCAECs) and human coronary artery smooth muscle cells (HCASMCs) submitted to static or dynamic conditions (corresponding to shear stress values of 1 and 20 dyne/cm2) in absence or in presence of a BVS - a PLLA prototype provided by Boston Scientific Corporation - and at a constant drug (Everolimus) concentration: Ibidi µ-Slide systems were adopted as laminar flow 2D bioreactors to evaluate the impact of hemodynamic forces in the presence of drug-eluting BVS on gene expression in human vascular cells.

The infection of Arabidopsis thaliana plants with avirulent pathogens causes the accumulation of cGMP with a biphasic profile downstream of nitric oxide signalling. However, plant enzymes that modulate cGMP levels have yet to be identified, so we generated transgenic A. thaliana plants expressing the rat soluble guanylate cyclase (GC) to increase genetically the level of cGMP and to study the function of cGMP in plant defence responses. Once confirmed that cGMP levels were higher in the GC transgenic lines than in wild-type controls, the GC transgenic plants were then challenged with bacterial pathogens and their defence responses were characterized. Although local resistance was similar in the GC transgenic and wild-type lines, differences in the redox state suggested potential cross-talk between cGMP and the glutathione redox system. Furthermore, large-scale transcriptomic and proteomic analysis highlighted the significant modulation of both gene expression and protein abundance at the infection site, inhibiting the establishment of systemic acquired resistance. Our data indicate that cGMP plays a key role in local responses controlling the induction of systemic acquired resistance in plants challenged with avirulent pathogens.

The economical success of the grape and wine-related agribusiness depends on quality of the grapes produced, which is heavily influenced by genetic diversity (clones) and environment (terroir). Among the important Italian varieties, 'Nebbiolo' (with the renowned wines made with this grape: Barolo and Barbaresco), is one of the best examples of this connection. The behavior of three 'Nebbiolo' clones (CVT71, CVT185, CVT423) grafted on the same rootstock (Kober 5BB) and grown in three different experimental vineyards in typical growing areas of Piedmont (Italy), was analyzed in relation to the different soil and environmental conditions. In 2013 the performances of those vines were monitored from winter pruning to harvest recording phenological dates, vigour, potential fertility, yield, juice composition and anthocyanin profile. RNA-Seq analyses were performed on berries collected in three different developmental stages during the vegetative season: berry pea size stage, veraison and at harvest (24 °Brix). The analysis of agronomical data revealed many cases of statistically significant effects linked to genotype x environment interactions. The environmental component appears to be important for some quality parameters, such as sugar and acidity, whose levels are often similar within individual vineyards. The clone CVT423 showed higher extractability of anthocyanins with high levels of peonidin, a di-substituted anthocyanidin, therefore less stable but more easily extractable. These interactions between genetic and environmental component will be further studied during the 2014 vintage. Within the next few months the completion of RNA-Seq analyses on berries will allow to relate the phenotypical characteristics of different clones to the transcriptomic changes. In addition, it will allow to identify genes differentially modulated during grapevine plastic adjustments, as well as genes, transcripts, and splicing variants influencing the clones fitness under diverse growing environments.