Latest PUBLICATIONS

  • The molecular chaperone Hsp90 is a component of the cap-binding complex and interacts with the translational repressor Cup during Drosophila oogenesis.
    Publication Date: 01/03/2009, on Gene
    by Pisa V, Cozzolino M, Gargiulo S, Ottone C, Piccioni F, Monti M, Gigliotti S, Talamo F, Graziani F, Pucci P, Verrotti AC
    DOI: 10.1016/j.gene.2008.11.025

    In metazoa, the spatio-temporal translation of diverse mRNAs is essential to guarantee proper oocyte maturation and early embryogenesis. The eukaryotic translation initiation factor 4E (eIF4E), which binds the 5' cap structure of eukaryotic mRNAs, associates with either stimulatory or inhibitory factors to modulate protein synthesis. In order to identify novel factors that might act at the translational level during Drosophila oogenesis, we have undertaken a functional proteomic approach and isolated the product of the Hsp83 gene, the evolutionarily conserved chaperone Hsp90, as a specific component of the cap-binding complex. Here we report that Hsp90 interacts in vitro with the translational repressor Cup. In addition, we show that Hsp83 and cup interact genetically, since lowering Hsp90 activity enhances the oogenesis alterations linked to diverse cup mutant alleles. Hsp90 and Cup co-localize in the cytoplasm of the developing germ-line cells within the germarium, thus suggesting a common function from the earliest stages of oogenesis. Taken together, our data start elucidating the role of Hsp90 during Drosophila female germ-line development and strengthen the idea that Cup has multiple essential functions during egg chamber development.

  • Different carbon sources affect lifespan and protein redox state during Saccharomyces cerevisiae chronological ageing.
    Publication Date: 01/03/2009, on Cellular and molecular life sciences : CMLS
    by Magherini F, Carpentieri A, Amoresano A, Gamberi T, De Filippo C, Rizzetto L, Biagini M, Pucci P, Modesti A
    DOI: 10.1007/s00018-009-8574-z

    In this study, a proteomic approach that combines selective labelling of proteins containing reduced cysteine residues with two-dimensional electrophoresis/mass spectrometry was used to evaluate the redox state of protein cysteines during chronological ageing in Saccharomyces cerevisiae. The procedure was developed on the grounds that biotin-conjugated iodoacetamide (BIAM) specifically reacts with reduced cysteine residues. BIAM-labelled proteins can then be selectively isolated by streptavidin affinity capture. We compared cells grown on 2% glucose in the exponential phase and during chronological ageing and we found that many proteins undergo cysteine oxidation. The target proteins include enzymes involved in glucose metabolism. Both caloric restriction and growth on glycerol resulted in a decrease in the oxidative modification. Furthermore, in these conditions a reduced production of ROS and a more negative glutathione half cell redox potential were observed.

  • Cognitive neuroscience of drawing: contributions of neuropsychological, experimental and neurofunctional studies.
    Publication Date: 01/03/2009, on Cortex; a journal devoted to the study of the nervous system and behavior
    by Trojano L, Grossi D, Flash T
    DOI: 10.1016/j.cortex.2008.11.015

  • Representational neglect in "invisible" drawing from memory.
    Publication Date: 01/03/2009, on Cortex; a journal devoted to the study of the nervous system and behavior
    by Cristinzio C, Bourlon C, Pradat-Diehl P, Trojano L, Grossi D, Chokron S, Bartolomeo P
    DOI: 10.1016/j.cortex.2008.03.013

    We describe the case of a patient with right hemisphere damage and left unilateral neglect. The patient was asked to draw from memory common objects, either with or without visual feedback. In the conditions without visual feedback the patient was either blindfolded or he made "invisible" drawings using a pen with the cap on, the drawings being recorded with carbon paper underneath. Results showed more neglect without than with visual feedback, contrary to previously published cases. This patient's pattern of performance may result from the contribution of a deficit of spatial working memory. Alternatively or in addition, the patient, who was undergoing cognitive rehabilitation for neglect, may have found easier to compensate for his neglect with visual feedback, which allowed him to visually explore the left part of his drawings.

  • Closing-in without severe drawing disorders: the "fatal" consequences of pathological attraction.
    Publication Date: 01/03/2009, on Cortex; a journal devoted to the study of the nervous system and behavior
    by Conson M, Salzano S, Manzo V, Grossi D, Trojano L
    DOI: 10.1016/j.cortex.2007.11.013

    The closing-in phenomenon (CIP) is often observed in patients with severe drawing disorders, but its cognitive bases are not well understood. We describe an experimental investigation aimed to clarify the nature of closing-in and its relationships with drawing disorders in a patient with corticobasal degeneration. In copying simple or complex stimuli (Experiment 1), the patient showed adherent and near types of closing-in, not affected by stimulus complexity, and produced distorted and often unrecognisable drawings. On the contrary, in drawing to dictation (without any available model), patients' performances significantly improved with respect to copying (Experiment 2). These data were consistent with the hypothesis that in some patients closing-in may develop from frontal-related release of approach behaviour even in the absence of relevant visuoperceptual impairments. By asking the patient to reproduce given spatial locations within circular frames (Experiment 3), we could further demonstrate the sparing of visuospatial processing and the frontal genesis of closing-in. These findings allowed us to speculate on the heterogeneous nature of closing-in.

  • Screening of ARHSP-TCC patients expands the spectrum of SPG11 mutations and includes a large scale gene deletion.
    Publication Date: 01/03/2009, on Human mutation
    by Denora PS, Schlesinger D, Casali C, Kok F, Tessa A, Boukhris A, Azzedine H, Dotti MT, Bruno C, Truchetto J, Biancheri R, Fedirko E, Di Rocco M, Bueno C, Malandrini A, Battini R, Sickl E, de Leva MF, Boespflug-Tanguy O, Silvestri G, Simonati A, Said E, Ferbert A, Criscuolo C, Heinimann K, Modoni A, Weber P, Palmeri S, Plasilova M, Pauri F, Cassandrini D, Battisti C, Pini A, Tosetti M, Hauser E, Masciullo M, Di Fabio R, Piccolo F, Denis E, Cioni G, Massa R, Della Giustina E, Calabrese O, Melone MA, De Michele G, Federico A, Bertini E, Durr A, Brockmann K, van der Knaap MS, Zatz M, Filla A, Brice A, Stevanin G, Santorelli FM
    DOI: 10.1002/humu.20945

    Autosomal recessive spastic paraplegia with thinning of corpus callosum (ARHSP-TCC) is a complex form of HSP initially described in Japan but subsequently reported to have a worldwide distribution with a particular high frequency in multiple families from the Mediterranean basin. We recently showed that ARHSP-TCC is commonly associated with mutations in SPG11/KIAA1840 on chromosome 15q. We have now screened a collection of new patients mainly originating from Italy and Brazil, in order to further ascertain the spectrum of mutations in SPG11, enlarge the ethnic origin of SPG11 patients, determine the relative frequency at the level of single Countries (i.e., Italy), and establish whether there is one or more common mutation. In 25 index cases we identified 32 mutations; 22 are novel, including 9 nonsense, 3 small deletions, 4 insertions, 1 in/del, 1 small duplication, 1 missense, 2 splice-site, and for the first time a large genomic rearrangement. This brings the total number of SPG11 mutated patients in the SPATAX collection to 111 cases in 44 families and in 17 isolated cases, from 16 Countries, all assessed using homogeneous clinical criteria. While expanding the spectrum of mutations in SPG11, this larger series also corroborated the notion that even within apparently homogeneous population a molecular diagnosis cannot be achieved without full gene sequencing.

  • Photoactive hybrid nanomaterials: indocyanine immobilized in mesoporous MCM-41 for "in-cell" bioimaging.
    Publication Date: 01/03/2009, on ACS applied materials & interfaces
    by Gianotti E, Bertolino CA, Benzi C, Nicotra G, Caputo G, Castino R, Isidoro C, Coluccia S
    DOI: 10.1021/am800196r

    Mesoporous silica nanoparticles are being explored as versatile tools for various biomedical and biotechnological applications including disease diagnosis, drug delivery, and intracellular imaging. In this paper, the synthesis and characterization of a fluorescent hybrid mesoporous silica nanomaterial, which is noncytotoxic and shows great potential for "in-cell" bioimaging applications, will be described. The hybrid mesoporous material has been obtained by confining highly fluorescent organic dyes, belonging to the indocyanine family, within the channels of mesoporous MCM-41. To explore the dispersion of the dye inside the mesoporous channels and the formation of dye aggregates, several hybrid samples with increasing dye/MCM-41 loading (up to 100 mg/g) were prepared. A uniform distribution of monomeric 1,1'-diethyl-3,3,3',3'-tetramethylindocarbocyanine iodide has been achieved at low dye loading (1 mg/g), as evidenced by photoluminescence spectra and lifetime, while a progressive formation of J-aggregates is induced by an increase in the dye loading. To elucidate the properties of the dye immobilized in mesoporous MCM-41, a detailed physical chemical characterization by structural (X-ray diffraction), volumetric and optical (Fourier transform infrared, diffuse-reflectance UV-vis and photoluminescence) techniques has been performed. By ultrasonication of the bulk material, nanoparticles of 2-20 nm diameter were obtained. Biocompatibility, endocytic uptake, and intracellular compartmentalization of such fluorescent nanoparticles were investigated in mammalian cultured cells.

  • Does the default-mode functional connectivity of the brain correlate with working-memory performances?
    Publication Date: 01/03/2009, on Archives italiennes de biologie
    by Esposito F, Aragri A, Latorre V, Popolizio T, Scarabino T, Cirillo S, Marciano E, Tedeschi G, Di Salle F
    DOI:

    The "default-mode" network is an ensemble of cortical regions that are typically deactivated during demanding cognitive tasks in functional magnetic resonance imaging (fMRI) studies. Using functional connectivity analysis, this network can be studied as a "stand-alone" brain system whose functional role is supposed to consist in the dynamic control of intrinsic processing activities like attention focusing and task-unrelated thought generation and suppression. Independent component analysis (ICA) is the method of choice for generating a statistical image of the "default-mode" network (DMN) using a task- and seed-independent distributed model of fMRI functional connectivity without prior specification of node region extent and timing of neural activation. We used a standard graded working-memory task (n-back) to induce fMRI changes in the default-mode regions and ICA to evaluate to DMN functional connectivity in nineteen healthy volunteers. Based on the known spatial variability of the ICA-DMN maps with the task difficulty levels, we hypothesized the ICA-DMN may also correlate with the subject performances. We confirmed that the relative extent of the anterior and posterior midline spots within the DMN were oppositely (resp. positively in the anterior and negatively in the posterior cingulate cortex) correlated with the level of task difficulty and found out that the spatial distribution of DMN also correlates with the individual task performances. We conclude that the working-memory function is related to a spatial re-configuration of the DMN functional connectivity, and that the relative involvement of the cingulate regions within the DMN might function as a novel predictor of the working-memory efficiency.

  • Unilateral left prosopometamorphopsia: a neuropsychological case study.
    Publication Date: 01/02/2009, on Neuropsychologia
    by Trojano L, Conson M, Salzano S, Manzo V, Grossi D
    DOI: 10.1016/j.neuropsychologia.2008.12.015

    We describe a patient who suddenly developed prosopometamorphopsia after a childbirth; she claimed that the left half of well-known and unfamiliar faces looked distorted. Brain MR was normal, whereas SPECT showed hypoperfusion of the left infero-lateral occipital cortex. No visual recognition defects for objects or faces were present. In three matching tasks with half-faces (Experiment 1), chimeric faces (Experiment 2), or chimeric objects (Experiment 3), the patient was impaired only when she matched pairs of chimeric faces differing in their left half; the same results were obtained after 1 year. This is the first behavioural demonstration of selective chronic metamorphopsia for the left side of faces, and provides new insights for models of face processing.

  • Small animal imaging facility: new perspectives for the radiologist.
    Publication Date: 01/02/2009, on La Radiologia medica
    by Grassi R, Cavaliere C, Cozzolino S, Mansi L, Cirillo S, Tedeschi G, Franchi R, Russo P, Cornacchia S, Rotondo A
    DOI: 10.1007/s11547-008-0352-8

    In recent years, new technologies have become available for imaging small animals. The use of animal models in basic and preclinical sciences, for example, offers the possibility of testing diagnostic markers and drugs, which is becoming crucial in the success and timeliness of research and is allowing a more efficient approach in defining study objectives and providing many advantages for both clinical research and the pharmaceutical industry. The use of these instruments offers data that are more predictive of the distribution and efficacy of a compound. The mouse, in particular, has become a key animal model system for studying human disease. It offers the possibility of manipulating its genome and producing accurate models for many human disorders, thus resulting in significant progress in understanding pathologenic mechanisms. In neurobiology, the possibility of simulating neurodegenerative diseases has enabled the development and validation of new treatment strategies based on gene therapy or cell grafting. Noninvasive imaging in small living animal models has gained increasing importance in preclinical research, itself becoming an independent specialty. The aim of this article is to review the characteristics of these systems and illustrate their main applications.

  • Enzymatically active fibrils generated by the self-assembly of the ApoA-I fibrillogenic domain functionalized with a catalytic moiety.
    Publication Date: 01/02/2009, on Biomaterials
    by Guglielmi F, Monti DM, Arciello A, Torrassa S, Cozzolino F, Pucci P, Relini A, Piccoli R
    DOI: 10.1016/j.biomaterials.2008.10.036

    Enzymatically active fibrils were produced by self-assembly of a bifunctional chimeric protein, made up of a fibrillogenic and a catalytic moiety. For this purpose, the fibrillogenic domain of Apolipoprotein A-I (ApoA-I), a 93-residue polypeptide named [1-93]ApoA-I, was functionalized with the enzyme glutathione S-transferase (GST). The fusion protein GST-[1-93]ApoA-I was expressed, isolated to homogeneity and characterized. In the soluble form, GST-[1-93]ApoA-I was found to be fully active as a GST enzyme, and to have high propensity to self-aggregate. Upon incubation for 3 weeks at pH 6.4, insoluble aggregates were generated. Analyzed by AFM, they were found to contain fibrillar structures often organized into large fiber networks. Fibrils were loaded on the membrane of a microfiltration unit and tested for enzymatic activity by filtering the substrate through the fibrillar network. Fibrils were shown to be catalytically active, stable over time and reusable, as no loss of activity was detected when fibrils were repeatedly tested. Our findings suggest that catalytically active fibrils may be of interest for biocatalytic applications in nanobiotechnology.

  • Injury to rat carotid arteries causes time-dependent changes in gene expression in contralateral uninjured arteries.
    Publication Date: 01/01/2009, on Clinical science (London, England : 1979)
    by Forte A, Finicelli M, De Luca P, Nordström I, Onorati F, Quarto C, Santè P, Renzulli A, Galderisi U, Berrino L, De Feo M, Hellstrand P, Rossi F, Cotrufo M, Cascino A, Cipollaro M
    DOI: 10.1042/CS20080080

    Vascular surgery aimed at stenosis removal induces local reactions often leading to restenosis. Although extensive analysis has been focused on pathways activated in injured arteries, little attention has been devoted to associated systemic vascular reactions. The aim of the present study was to analyse changes occurring in contralateral uninjured rat carotid arteries in the acute phase following unilateral injury. WKY (Wistar-Kyoto) rats were subjected to unilateral carotid arteriotomy. Contralateral uninjured carotid arteries were harvested from 4 h to 7 days after injury. Carotid arteries were also harvested from sham-operated rats and uninjured rats. Carotid morphology and morphometry were examined. Affymetrix microarrays were used for differential analysis of gene expression. A subset of data was validated by real-time RT-PCR (reverse transcription-PCR) and verified at the protein level by Western blotting. A total of 1011 genes were differentially regulated in contralateral uninjured carotid arteries from 4 h to 7 days after arteriotomy (P<0.0001; fold change, >or=2) and were classified into 19 gene ontology functional categories. To a lesser extent, mRNA variations also occurred in carotid arteries of sham-operated rats. Among the changes, up-regulation of members of the RAS (renin-angiotensin system) was detected, with possible implications for vasocompensative mechanisms induced by arteriotomy. In particular, a selective increase in the 69 kDa isoform of the N-domain of ACE (angiotensin-converting enzyme), and not the classical somatic 195 kDa isoform, was observed in contralateral uninjured carotid arteries, suggesting that this 69 kDa isoenzyme could influence local AngII (angiotensin II) production. In conclusion, systemic reactions to injury occur in the vasculature, with potential clinical relevance, and suggest that caution is needed in the choice of controls during experimental design in vivo.

  • Technical advances in proteomics mass spectrometry: identification of post-translational modifications.
    Publication Date: 01/01/2009, on Clinical chemistry and laboratory medicine
    by Amoresano A, Carpentieri A, Giangrande C, Palmese A, Chiappetta G, Marino G, Pucci P
    DOI: 10.1515/CCLM.2009.154

    The importance of post-translational modifications (PTMs) of proteins has become evident in the proteomic era as it plays a critical role in modulating cellular function, and can vary in response to different stimuli thereby tuning cellular mechanisms. Assessment of PTMs on a proteomic scale is a challenging task since they are substoichiometric, transient and reversible. Moreover, the amount of post-translationally modified proteins is generally very small when compared to their unmodified counterparts. Existing methodologies for identification of PTMs essentially relies on enrichment procedure to selectively increase the amount of modified peptides. These procedures need to be integrated with sophisticated mass spectrometric methods to enable the identifications of PTMs. Although the strategies developed so far are not optimal, a number of examples will be given where the combination of innovative separation methods along with advanced mass spectrometric analyses provide positive results. These experiences are leading the way for the next generation of proteomic approaches for identification of a wide range of PTMs.

  • The analysis of phosphoproteomes by selective labelling and advanced mass spectrometric techniques.
    Publication Date: 01/01/2009, on Methods in molecular biology (Clifton, N.J.)
    by Amoresano A, Cirulli C, Monti G, Quemeneur E, Marino G
    DOI: 10.1007/978-1-60327-834-8_13

    This chapter focuses on the development of new proteomic approaches based on classical biochemical procedures coupled with new mass spectrometry methods to study the phosphorylation, the most important and abundant PTMs in modulating protein activity and propagating signals within cellular pathways and networks. These phosphoproteome studies aim at comprehensive analysis of protein phosphorylation by identification of the phosphoproteins, exact localization of phosphorylated residues, and preferably quantification of the phosphorylation. Because of low stoichiometry, heterogeneity, and low abundance, enrichment of phosphopeptides is an important step of this analysis. The first section is focused on the development of new enrichment methods coupled to mass spectrometry. Thus, improved approach, based on simple chemical manipulations and mass spectrometric procedures, for the selective analysis of phosphoserine and phosphothreonine in protein mixtures, following conversion of the peptide phosphate moiety into DTT derivatives, is described. However the major aim of this work is devoted to the use of isotopically labelled DTT, thus allowing a simple and direct quantitative MS analysis. The final part of the work is focused on the development of a strategy to study phosphorylation without preliminary enrichment but using the high performance of a novel hybrid mass spectrometer linear ion trap.

  • Inhibition of prostaglandin synthesis reduces the induction of MyoD expression in rat soleus muscle.
    Publication Date: 01/01/2009, on Journal of muscle research and cell motility
    by Monda M, Vicidomini C, Viggiano A, Sampaolo S, Di Iorio G, Viggiano A, Viggiano E, De Luca B
    DOI: 10.1007/s10974-009-9182-0

    MyoD is a myogenic regulatory factor with a critical role in skeletal muscle development and regeneration. As muscle regeneration comes with an inflammatory process, it has been proposed that the inflammatory cells can play an important role in the induction of muscle fibres regeneration. The aim of the present work was to verify if a cyclooxygenase inhibitory drug (ketoprofen) would alter the normal expression of MyoD in a regenerating rat soleus muscle after an over-load lesion. Using immunohistochemical techniques, the numbers of m-cadherin-positive cells, a selective marker of satellite cells, and MyoD-positive cells were evaluated in functionally overloaded rat soleus muscles 4 days after a gastrocnemius tendon cut. The same study was conducted either with four rats injected with ketoprofen (100 mg/kg b.w./day) or with four rats injected with saline solution. The data obtained showed a very large decrease in the number of MyoD positive/m-cadherin positive cells in the ketoprofen injected group compared to the control group. Although further studies are needed to elucidate the sequence of biochemical events that induce a reduction of MyoD expression due to ketoprofen, the results demonstrate that prostaglandin synthesis is required for the induction of MyoD expression and that ketoprofen can affect this expression, with possible adverse effects on muscle regeneration.