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Colleretto Giacosa, Italy

Falvo S.,University of Turin | Falvo S.,Bioindustry Park Silvano Fumero SpA | Di Carli M.,ENEA | Desiderio A.,ENEA | And 5 more authors.
Proteomics | Year: 2012

Plants respond to ultraviolet stress inducing a self-defence through the regulation of specific gene family members. The UV acclimation is the result of biochemical and physiological processes, such as enhancement of the antioxidant enzymatic system and accumulation of UV-absorbing phenolic compounds (e.g. flavonoids). Globe artichoke is an attractive species for studying the protein network involved in UV stress response, being characterized by remarkable levels of inducible antioxidants. Proteomic tools can assist the evaluation of the expression patterns of UV-responsive proteins and we applied the difference in-gel electrophoresis (DIGE) technology for monitoring the globe artichoke proteome variation at four time points following an acute UV-C exposure. A total of 145 UV-C-modulated proteins were observed and 119 were identified by LC-MS/MS using a ∼144000 customized Compositae protein database, which included about 19000 globe artichoke unigenes. Proteins were Gene Ontology (GO) categorized, visualized on their pathways and their behaviour was discussed. A predicted protein interaction network was produced and highly connected hub-like proteins were highlighted. Most of the proteins differentially modulated were chloroplast located, involved in photosynthesis, sugar metabolisms, protein folding and abiotic stress. The identification of UV-C-responsive proteins may contribute to shed light on the molecular mechanisms underlying plant responses to UV stress. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Barge A.,University of Turin | Caporaso M.,University of Turin | Cravotto G.,University of Turin | Martina K.,University of Turin | And 6 more authors.
Chemistry - A European Journal | Year: 2013

We report the synthesis and characterization of a water-soluble, star-shaped macromolecular platform consisting of eight β-cyclodextrin (β-CD) units anchored to the narrower rim of a γ-CD core through bis(triazolyl) alkyl spacers. The efficient synthetic protocol is based on the microwave (MW)-promoted Cu-catalyzed 1,3-dipolar cycloaddition of CD monoazides to CD monoacetylenes. The ligandhosting capability of the construct has been assessed by relaxometric titration and nuclear magnetic relaxation dispersion (NMRD) profiling, which showed it to be good, and this was supported by molecular dynamics simulations. To demonstrate the feasibility of obtaining supramolecular structures with high hosting ability, we designed a dimeric platform, formed by joining two nonamers through the γ-CD cores through a bis(lithocholic acid) linker. With a view to the potential biological applications, cytotoxicity and extent of binding to human serum albumin were assessed. The properties of this dendrimeric multicarrier make it suitable for pharmaceutical and diagnostic purposes, ranging from targeted drug delivery to molecular imaging. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Robotti A.,University of Turin | Robotti A.,Bioindustry Park Silvano Fumero SpA | Natale M.,Bioindustry Park Silvano Fumero SpA | Albo A.G.,Bioindustry Park Silvano Fumero SpA | And 5 more authors.
Electrophoresis | Year: 2010

Plasma acute-phase proteins (APPs) glyco-isoforms are important biomarkers of inflammatory processes such as those occurring in multiple sclerosis (MS). Specific analysis of these proteins is often hampered by sample biochemical complexity. The aim of our study was to set up a method to accurately visualize, identify and quantify APPs glyco-isoforms in human serum. An enrichment strategy based on affinity chromatography using the carbohydrate-binding proteins concanavalin A (ConA) and erythrina cristagalli lectin (ECL) was applied to pooled serum samples from 15 patients and 9 healthy individuals. Image analysis of 2-DE detected 30 spots with a fold change higher than 1.5. A total of 14 were statistically significant (p value <0.05): 7 up-regulated and 7 down-regulated in MS samples. ESI LC-Nanospray IT mass spectrometry analysis confirmed that all of them were APPs isoforms supporting the idea that the accurate analysis of differential glycosylation profiles in these biomarkers is instrumental to distinguish between MS patients and healthy subjects. Additionally, overlaps in ConA/ ECL maps protein patterns suggest how the used lectins are able to bind sugars harbored by the same oligosaccharide structure. Among identified proteins, the presence of complex and/or hybrid type N-linked sugar structures is well known. Performing galectin-3 binding and Western blotting, we were able to demonstrate a correlation between hybrid type glyco-isoforms of β-haptoglobin and MS. In conclusion, although the patho-physiological role of the identified species still remains unclear and further validations are needed, these findings may have a relevant impact on disease-specific marker identification approaches. © 2010 Wiley-VCH Verlag GmbH & Co. KGaA.

Perga S.,San Luigi University Hospital | Perga S.,C o San Luigi University Hospital | Albo A.G.,Bioindustry Park Silvano Fumero SpA | Albo A.G.,Bioindustry Park Silvano Fumero SpA | And 17 more authors.
PLoS ONE | Year: 2015

Background: Multiple sclerosis (MS) is a multifactorial autoimmune disease of the central nervous system with a heterogeneous and unpredictable course. To date there are no prognostic biomarkers even if they would be extremely useful for early patient intervention with personalized therapies. In this context, the analysis of inter-individual differences in cerebrospinal fluid (CSF) proteome may lead to the discovery of biological markers that are able to distinguish the various clinical forms at diagnosis. Methods: To this aim, a two dimensional electrophoresis (2-DE) study was carried out on individual CSF samples from 24 untreated women who underwent lumbar puncture (LP) for suspected MS. The patients were clinically monitored for 5 years and then classified according to the degree of disease aggressiveness and the disease-modifying therapies prescribed during follow up. Results: The hierarchical cluster analysis of 2-DE dataset revealed three protein spots which were identified by means of mass spectrometry as Apolipoprotein E (ApoE) and two isoforms of vitamin D binding protein (DBP). These three protein spots enabled us to subdivide the patients into subgroups correlated with clinical classification (MS aggressive forms identification: 80%). In particular, we observed an opposite trend of values for the two protein spots corresponding to different DBP isoforms suggesting a role of a post-translational modification rather than the total protein content in patient categorization. Conclusions: These findings proved to be very interesting and innovative and may be developed as new candidate prognostic biomarkers of MS aggressiveness, if confirmed. © 2015 Perga et al.

Acquadro E.,Bioindustry Park Silvano Fumero SpA | Caron I.,Mario Negri Institute for Pharmacological Research | Tortarolo M.,Mario Negri Institute for Pharmacological Research | Bucci E.M.,Bioindustry Park Silvano Fumero SpA | And 3 more authors.
Journal of Proteome Research | Year: 2014

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal neurodegenerative disease caused by the degeneration of motor neurons. The transgenic mouse model carrying the human SOD1G93A mutant gene (hSOD1G93A mouse) represents one of the most reliable and widely used model of this pathology. In the present work, the innovative technique of matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) was applied in the study of pathological alterations at the level of small brain regions such as facial and trigeminal nuclei, which in rodents are extremely small and would be difficult to analyze with classical proteomics approaches. Comparing slices from three mice groups (transgenic hSOD1G93A, transgenic hSOD1WT, and nontransgenic, Ntg), this technique allowed us to evidence the accumulation of hSOD1G93A in the facial and trigeminal nuclei, where it generates aggregates. This phenomenon is likely to be correlated to the degeneration observed in these regions. Moreover, a statistical analysis allowed us to highlight other proteins as differentially expressed among the three mice groups analyzed. Some of them were identified by reverse-phase HPLC fractionation of extracted proteins and mass spectrometric analysis before and after trypsin digestion. In particular, the 40S ribosomal protein S19 (RPS19) was upregulated in the parenkyma and reactive glial cells in facial nuclei of hSOD1G93A mice when compared to transgenic hSOD1WT and nontransgenic ones. © 2014 American Chemical Society.

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