Latest PUBLICATIONS

  • Physico-chemical analysis of G-quadruplex containing bunch-oligonucleotides.
    Publication Date: 20/02/2007, on International journal of biological macromolecules
    by Petraccone L, Martino L, Duro I, Oliviero G, Borbone N, Piccialli G, Giancola C
    DOI: 10.1016/j.ijbiomac.2006.07.013

    A growing number of evidences suggest that DNA G-quadruplex structures play an important role in many relevant biological processes. The introduction of chemical modifications in quadruplex structures could enhance the in vivo biological activity. The correlation between the physico-chemical properties and chemical modifications represents an essential step toward the de novo design of quadruplex forming oligonucleotides for biomedical applications. We report the physico-chemical characterisation of a quadruplex formed by a bunch of four d(TG4T) oligonucleotides whose 3'-ends are linked together by a tetra-branched linker. The study was performed by circular dichroism, gel electrophoresis and molecular modelling techniques. The data indicate an high stability for this kind of quadruplex and add some information on the role of the tetra-branched linker on the quadruplex stability.

  • Synaptosomal protein synthesis is selectively modulated by learning.
    Publication Date: 09/02/2007, on Brain research
    by Eyman M, Cefaliello C, Ferrara E, De Stefano R, Crispino M, Giuditta A
    DOI: 10.1016/j.brainres.2006.11.025

    Synaptosomes from rat brain have long been used to investigate the properties of synaptic protein synthesis. Comparable analyses have now been made in adult male rats trained for a two-way active avoidance task to examine the hypothesis of its direct participation in brain plastic events. Using Ficoll-purified synaptosomes from neocortex, hippocampus and cerebellum, our data indicate that the capacity of synaptosomal protein synthesis and the specific activity of newly synthesized proteins were not different in trained rats in comparison with home-caged control rats. On the other hand, the synthesis of two proteins of 66.5 kDa and 87.6 kDa separated by SDS-PAGE and analyzed by quantitative densitometry was selectively enhanced in trained rats. In addition, the synthesis of the 66.5 kDa protein, but not of the 87.6 kDa protein, correlated with avoidances and escapes and inversely correlated with freezings in the neocortex, while in the cerebellum it correlated with avoidances and escapes. The data demonstrate the participation of synaptic protein synthesis in plastic events of behaving rats, and the selective, region-specific modulation of the synthesis of a synaptic 66.5 kDa protein by the newly acquired avoidance response and by the reprogramming of innate neural circuits subserving escape and freezing responses.

  • Lysine 58-cleaved beta2-microglobulin is not detectable by 2D electrophoresis in ex vivo amyloid fibrils of two patients affected by dialysis-related amyloidosis.
    Publication Date: 01/02/2007, on Protein science : a publication of the Protein Society
    by Giorgetti S, Stoppini M, Tennent GA, Relini A, Marchese L, Raimondi S, Monti M, Marini S, Østergaard O, Heegaard NH, Pucci P, Esposito G, Merlini G, Bellotti V
    DOI: 10.1110/ps.062563507

    The lysine 58 cleaved and truncated variant of beta(2)-microglobulin (DeltaK58-beta2m) is conformationally unstable and present in the circulation of a large percentage of patients on chronic hemodialysis, suggesting that it could play a role in the beta2-microglobulin (beta2m) amyloid fibrillogenesis associated with dialysis-related amyloidosis (DRA). However, it has yet to be detected in the amyloid deposits of such patients. Here, we extracted amyloid fibrils, without denaturation or additional purification, from different amyloidotic tissues of two unrelated individuals suffering from DRA, and characterized them by high-sensitivity bidimensional gel electrophoresis (2D-PAGE), immunoblotting, MALDI time-of-flight mass spectrometry, and protein sequencing. To confirm whether or not this species could be identified by our proteomic approaches, we mapped its location in 2D-PAGE, in mixtures of pure DeltaK58-beta2m, and extracts of amyloid fibrils from patients, to a discrete region of the gel distinct from other isoforms of beta2m. Using this approach, the two known principal isoforms found in beta2m amyloid were identified, namely, the full-length protein and the truncated species lacking six N-terminal amino acid residues (DeltaN6-beta2m). In contrast, we found no evidence for the presence of DeltaK58-beta2m.

  • An effective method for adenoviral-mediated delivery of small interfering RNA into mesenchymal stem cells.
    Publication Date: 01/02/2007, on Journal of cellular biochemistry
    by Forte A, Napolitano MA, Cipollaro M, Giordano A, Cascino A, Galderisi U
    DOI: 10.1002/jcb.21025

    Mesenchymal stem cells (MSCs) promise as a main actor of cell-based therapeutic strategies, due to their intrinsic ability to differentiate along different mesenchymal cell lineages, able to repair the diseased or injured tissue in which they are localized. The application of MSCs in therapies requires an in depth knowledge of their biology and of the molecular mechanisms leading to MSC multilineage differentiation. The knockdown of target genes through small interfering RNA (siRNA) carried by adenoviruses (Ad) represents a valid tool for the study of the role of specific molecules in cell biology. Unfortunately, MSCs are poorly transfected by conventional Ad serotype 5 (Ad5) vectors. We set up a method to obtain a very efficient transduction of rat MSCs with low doses of unmodified Ad5, carrying the siRNA targeted against the mRNA coding for Rb2/p130 (Ad-siRNA-Rb2), which plays a fundamental role in cell differentiation. This method allowed a 95% transduction rate of Ad-siRNA in MSC, along with a siRNA-mediated 85% decrease of Rb2/p130 mRNA and a 70% decrease of Rb2/p130 protein 48 h after transduction with 50 multiplicities of infection (MOIs) of Ad5. The effect on Rb2/p130 protein persisted 15 days after transduction. Finally, Ad-siRNA did not compromise the viability of transduced MSCs neither induced any cell cycle modification. The effective Ad-siRNA-Rb2 we constructed, together with the efficient method of delivery in MSCs we set up, will allow an in depth analysis of the role of Rb2/p130 in MSC biology and multilineage differentiation.

  • Autonomic neuropathy in mixed cryoglobulinemia.
    Publication Date: 01/02/2007, on Journal of neurology
    by Ammendola A, Sampaolo S, Migliaresi S, Ambrosone L, Ammendola E, Ciccone G, Di Iorio G
    DOI: 10.1007/s00415-006-0333-6

    A retrospective, cross-sectional study was performed on a series of HCV-related mixed cryoglobulinemia (HCV-MC) patients to assess autonomic neuropathy (AN) and its relation to peripheral neuropathy (PN). Thirty consecutive patients affected by HCV-MC underwent clinical, neurological and electrodiagnostic examinations. Autonomic nervous system (ANS) involvement was assessed by functional cardiovascular tests and sympathetic skin response (SSR) evaluation. Sural nerve biopsy was performed in 10 patients with PN. All patients received steroids, 15 also received recombinant interferon-alpha2b (RIfn-alpha2b). PN occurred in 27 patients (90.0%) and AN in 4 (13.3 %) all with signs of PN. SSR was the autonomic test more frequently altered. Biopsy disclosed axonal degeneration more evident in the 4 patients with AN. Three out of 4 patients with AN received steroids and rIFN-alpha2b and 1 steroids alone. In our study on HCV-MC, it was concluded that AN can occur also without dysautonomic symptoms, SSR appears to be one of the optional tests to use together with dysautonomic tests to identify AN and finally PN and AN do not seem to be positively influenced by addition of rIFN-alpha2b to steroid treatment.

  • Neurofibromatosis type 1 and optic pathway glioma. A long-term follow-up.
    Publication Date: 01/02/2007, on Minerva pediatrica
    by Lama G, Esposito Salsano M, Grassia C, Calabrese E, Grassia MG, Bismuto R, Melone MA, Russo S, Scuotto A
    DOI:

    Optic pathway gliomas (OPG) are the predominant intracranial tumours associated with neurofibromatosis type 1 (NF1). The aim of this study was to evaluate the prevalence and the outcome of OPG in 200 NF1 patients (122 males and 78 females, aged 1-25 years) followed up to 16 years (mean of 6 years).

  • Chronic cocaine administration modulates the expression of transcription factors involved in midbrain dopaminergic neuron function.
    Publication Date: 01/02/2007, on Experimental neurology
    by Leo D, di Porzio U, Racagni G, Riva MA, Fumagalli F, Perrone-Capano C
    DOI: 10.1016/j.expneurol.2006.08.024

    Chronic cocaine use leads to pronounced alterations in neuronal functions in brain circuits associated with reward. In the present study, we examined in the rat midbrain the effects of acute, subchronic (5 days) and chronic cocaine treatments (14 days) on the gene expression of transcription factors involved in the development and maintenance of dopaminergic neurons. We show that chronic, but not acute or subchronic, cocaine administration downregulates Nurr1 and Pitx3 transcripts whereas En1 transcripts are upregulated. Conversely, Lmx1b and En2 transcripts are not affected by the drug treatment, indicating that the modulation of the midbrain transcription factors analyzed is highly selective. Interestingly, modification of the gene expression for these transcription factors persists in midbrain as long as two weeks after the last drug administration, suggesting that it may account for some of the enduring alterations in midbrain dopaminergic circuits associated with chronic cocaine use.

  • The epsilon isoform of 14-3-3 protein is a component of the prion protein amyloid deposits of Gerstmann-Sträussler-Scheinker disease.
    Publication Date: 01/02/2007, on Journal of neuropathology and experimental neurology
    by Di Fede G, Giaccone G, Limido L, Mangieri M, Suardi S, Puoti G, Morbin M, Mazzoleni G, Ghetti B, Tagliavini F
    DOI: 10.1097/nen.0b013e3180302060

    The 14-3-3 proteins are highly conserved, ubiquitous molecules involved in a variety of biologic events, such as transduction pathway modulation, cell cycle control, and apoptosis. Seven isoforms have been identified that are abundant in the brain, preferentially localized in neurons. Remarkable increases in 14-3-3 are seen in the cerebrospinal fluid of patients with Creutzfeldt-Jakob disease (CJD), and it has been found in pathologic inclusions of several neurodegenerative diseases. Moreover, the zeta isoform has been detected in prion protein (PrP) amyloid deposits of CJD patients. To further investigate the cerebral distribution of 14-3-3 in prion-related encephalopathies, we carried out an immunohistochemical and biochemical analysis of brain tissue from patients with Gerstmann-Sträussler-Scheinker disease (GSS) and sporadic, familial and acquired forms of CJD, using specific antibodies against the seven 14-3-3 isoforms. The study showed a strong immunoreactivity of PrP amyloid plaques of GSS patients for the 14-3-3 epsilon isoform, but not for the other isoforms. The epsilon isoform of 14-3-3 was not found in PrP deposits of CJD. These results indicate that the epsilon isoform of 14-3-3 is a component of PrP amyloid deposits of GSS and suggest that this is the sole 14-3-3 isoform specifically involved in the neuropathologic changes associated with this disorder.

  • Upper motor neuron involvement in X-linked recessive bulbospinal muscular atrophy.
    Publication Date: 01/02/2007, on Clinical neurophysiology : official journal of the International Federation of Clinical Neurophysiology
    by Pachatz C, Terracciano C, Desiato MT, Orlacchio A, Mori F, Rocchi C, Bernardi G, Massa R
    DOI: 10.1016/j.clinph.2006.10.006

    Clinicopathological findings of X-linked recessive bulbospinal muscular atrophy (SBMA) are indicative of lower motor neuron and primary sensory neuron involvement. The aim of our study was to investigate the presence of subclinical upper motor neuron (UMN) dysfunction in this disease.

  • Giant intracranial chordoma: neuroradiological and radiotherapeutic aspects.
    Publication Date: 31/01/2007, on The neuroradiology journal
    by Conforti R, Taglialatela G, Scuotto A, D'agostino V, Cirillo M, Cirillo L, Barone A, Giordano A, Parlato C, Cirillo S
    DOI:

    We describe a rare case of giant intracranial chordoma, emphasizing the patient's long survival and his excellent response to radiotherapy that led to a progressive regression of neurological symptomatology up to disappearance, in the absence of cerebral white matter damages.

  • Non-Inferential Multi-Subject Study of Functional Connectivity during Visual Stimulation.
    Publication Date: 31/01/2007, on The neuroradiology journal
    by Esposito F, Cirillo M, Aragri A, Caranci F, Cirillo L, Di Salle F, Cirillo S
    DOI:

    Independent component analysis (ICA) is a powerful technique for the multivariate, non-inferential, data-driven analysis of functional magnetic resonance imaging (fMRI) data-sets. The non-inferential nature of ICA makes this a suitable technique for the study of complex mental states whose temporal evolution would be difficult to describe analytically in terms of classical statistical regressors. Taking advantage of this feature, ICA can extract a number of functional connectivity patterns regardless of the task executed by the subject. The technique is so powerful that functional connectivity patterns can be derived even when the subject is just resting in the scanner, opening the opportunity for functional investigation of the human mind at its basal "default" state, which has been proposed to be altered in several brain disorders. However, one major drawback of ICA consists in the difficulty of managing its results, which are not represented by a single functional image as in inferential studies. This produces the need for a classification of ICA results and exacerbates the difficulty of obtaining group "averaged" functional connectivity patterns, while preserving the interpretation of individual differences. Addressing the subject-level variability in the very same framework of "grouping" appears to be a favourable approach towards the clinical evaluation and application of ICA-based methodologies. Here we present a novel strategy for group-level ICA analyses, namely the self-organizing group-level ICA (sog-ICA), which is used on visual activation fMRI data from a block-design experiment repeated on six subjects. We propose the sog-ICA as a multi-subject analysis tool for grouping ICA data while assessing the similarity and variability of the fMRI results of individual subject decompositions.

  • Nutrition, physical activity, and cardiovascular disease: an update.
    Publication Date: 15/01/2007, on Cardiovascular research
    by Ignarro LJ, Balestrieri ML, Napoli C
    DOI: 10.1016/j.cardiores.2006.06.030

    Many epidemiological studies have indicated a protective role for a diet rich in fruits and vegetables against the development and progression of cardiovascular disease (CVD), one of the leading causes of morbidity and mortality worldwide. Physical inactivity and unhealthy eating contribute to these conditions. This article assesses the scientific rationale of benefits of physical activity and good nutrition on CVD, especially on atherosclerosis-related diseases. Compelling evidence has accumulated on the role of oxidative stress in endothelial dysfunction and in the pathogenesis of CVD. Reduced nitric oxide (NO) bioavailability due to oxidative stress seems to be the common molecular disorder comprising stable atherosclerotic narrowing lesions. Energy expenditure of about 1000 kcal (4200 kJ) per week (equivalent to walking 1 h 5 days a week) is associated with significant health benefits. Such benefits can be achieved through structured or nonstructured physical activity, accumulated throughout the day (even through short 10-min bouts) on most days of the week. Some prospective studies showed a direct inverse association between fruit and vegetable intake and the development of CVD incidents such as acute plaque rupture causing unstable angina or myocardial infarction and stroke. Many nutrients and phytochemicals in fruits and vegetables, including fiber, potassium, and folate, could be independently or jointly responsible for the apparent reduction in CVD risk. Novel findings and critical appraisal regarding antioxidants, dietary fibers, omega-3 polyunsaturated fatty acids (n-3 PUFAs), nutraceuticals, vitamins, and minerals, are presented here in support of the current dietary habits together with physical exercise recommendations for prevention and treatment of CVD.

  • Structural insight into the hTERT intron 6 sequence d(GGGGTGAAAGGGG) from 1H-NMR study.
    Publication Date: 01/01/2007, on Nucleosides, nucleotides & nucleic acids
    by Virno A, Mayol L, Ramos A, Fraternali F, Pagano B, Randazzo A
    DOI: 10.1080/15257770701521854

    The interest in DNA quadruplex structures has been fueled by the recognition that telomeres, the 3' single stranded guanine-rich overhangs found at the termini of chromosomes, are likely to form G-tetrads type structures important in cell senescence and cancer. In addition to their presence in telomeres, where they may play a role in maintaining the stability and integrity of chromosomes, guanine-rich regions are found in other region of the genome, amongst these is intron 6 of hTERT a gene codifying for the enzyme telomerase. Interestingly, the formation of G-quadruplexes in this region is involved in the down-regulation of telomerase activity caused by an alteration of the hTERT splicing pattern. Therefore, we have analyzed several sequences of that intron by (1)H-NMR and CD spectroscopy, and we have found that the sequence d(GGGGTGAAAGGGG) is able to fold in a single well-defined antiparallel quadruplex structure consisting of four G-tetrads, possessing a twofold symmetry, and containing four Gs in a syn glycosidic conformation.

  • Folding, activity and targeting of mutated human cathepsin D that cannot be processed into the double-chain form.
    Publication Date: 01/01/2007, on The international journal of biochemistry & cell biology
    by Follo C, Castino R, Nicotra G, Trincheri NF, Isidoro C
    DOI: 10.1016/j.biocel.2006.11.010

    The precursor of human cathepsin D (CD) is converted into the single-chain and the double-chain active polypeptides by subsequent proteolysis reactions taking place in the endosomal-lysosomal compartment and involving specific aminoacid sequences. We have mutagenized the region of aminoacids (comprising the beta-hairpin loop) involved in the latter proteolytic maturation step and generated a mutant CD that cannot be converted into the mature double-chain form. This mutant CD expressed in rodent cells reaches the lysosome and is stable as single-chain polypeptide, bears high-mannose type sugars, binds to pepstatin A and is enzymatically active, indicating that it is correctly folded. The present work provides new insights on the aminoacid region involved in the terminal processing of human CD and on the function of the processing beta-hairpin loop.

  • Frequency of the LRRK2 G2019S mutation in Italian patients affected by Parkinson's disease.
    Publication Date: 01/01/2007, on Journal of human genetics
    by Squillaro T, Cambi F, Ciacci G, Rossi S, Ulivelli M, Malandrini A, Mencarelli MA, Mari F, Renieri A, Ariani F
    DOI: 10.1007/s10038-006-0105-2

    Mutations in the gene Leucine-Rich Repeat Kinase 2 (LRRK2) have been identified in both dominant and sporadic cases affected by Parkinson's disease (PD). The LRRK2 G2019S mutation (c.6055G>A) is the most frequent substitution in Caucasians, accounting for approximately 5-6% of familial and 0.5-2.0% of apparently sporadic PD cases. We investigated the frequency of the LRRK2 G2019S mutation in 98 unrelated Italian PD patients, including 12 probands belonging to families compatible with autosomal dominant inheritance (12%) and 86 sporadic cases (88%). We detected the G2019S mutation in one sporadic female patient (1.2%). These results confirm that the G2019S mutation is a relevant cause of sporadic PD cases in the Italian population and stress the importance of performing this genetic test, which has important implications for genetic counselling.