Latest PUBLICATIONS

  • Solution structure of human beta-endorphin in helicogenic solvents: an NMR study.
    Publication Date: 01/09/1999, on Journal of peptide science : an official publication of the European Peptide Society
    by Saviano G, Crescenzi O, Picone D, Temussi P, Tancredi T
    DOI: 10.1002/(SICI)1099-1387(199909)5:9<410::AID-PSC216>3.0.CO;2-R

    Beta-endorphin is the largest natural opioid peptide. The knowledge of its bioactive conformation might be very important for the indirect mapping of the active site of opioid receptors. We have studied beta-endorphin in a variety of solution conditions with the goal of testing the intrinsic tendency of its sequence to assume a regular fold. We ran NMR experiments in water, dimethylsulfoxide and aqueous mixtures of methanol, ethylene glycol, trifluoroethanol, hexafluoracetone trihydrate and dimethylsulfoxide. The solvent in which the peptide is more ordered is the hexafluoracetone trihydrate/water mixture. The helical structure detected for beta-endorphin in this mixture at 300 K extends for the greater part of its address domain, hinting at a possible mechanism of interaction with opioid receptors: a two-point attachment involving an interaction of the helical part of the address domain (PLVTLFKNAIIKNAY) with one of the transmembrane helices and a classical interaction of the message domain (YGGF) with the receptor subsite common to all opioid receptors.

  • 2-aminoanthracene as an analytical tool with the acetylation reaction catalyzed by arylamine N-acetyltransferase.
    Publication Date: 15/08/1999, on Analytical biochemistry
    by Servillo L, Balestrieri C, Boccellino M, Balestrieri ML, Quagliuolo L, Giovane A
    DOI: 10.1006/abio.1999.4200

    The polynuclear aromatic amine, 2-aminoanthracene, was found to be acetylated with high efficiency in the presence of acetyl-CoA by pigeon liver arylamine N-acetyltransferase (EC 2.3.1.5). As a consequence of acetylation the fluorescence properties of the compound dramatically change and the reaction time course can be easily followed fluorometrically at the emission wavelength of 425 nm upon excitation at 360 nm. When 2-aminoanthracene is employed with pigeon arylamine N-acetyltransferase, as the ultimate acceptor of the acetyl group in coupled fluorometric assays, it is possible to measure enzymatic activities, such as pyruvate dehydrogenase or carnitine acetyltransferase, in continuous assays rapidly and with high sensitivity or to determine with as much sensitivity important metabolites such as acetylcarnitine or acetyl-CoA.

  • Surgical management of the neck in squamous cell carcinoma of the tongue.
    Publication Date: 01/08/1999, on The British journal of oral & maxillofacial surgery
    by Califano L, Zupi A, Mangone GM, Longo F, Coscia G, Piombino P
    DOI: 10.1054/bjom.1999.0076

    If the nodes are involved, survival of patients with squamous cell carcinoma of the tongue is considerably reduced. Surgery remains the treatment of choice and, to define its role, we have reviewed 82 consecutive cases. Sixty-two cases (76%) were T1-2, and 46 patients (56%) had involved nodes. The cervical region II was the most often involved (n=26). Occult nodal metastases were present in 12 cases. The extent of nodal spread and prognosis varies according to whether the body or the base of the tongue is involved. Lesions of the base with involved node should be treated by a selective posterolateral neck dissection, whilst in the case of a lesion of the body of the tongue, the dissection should be selective anterolateral. In lesions of the base, when there are no nodes involved, a prophylactic selective posterolateral neck dissection is recommended, whilst in the case of the lesions of the body, selective supraomohyoid neck dissection in T2-4 lesions is recommended.

  • Structural and membrane-binding properties of saposin D.
    Publication Date: 01/07/1999, on European journal of biochemistry
    by Tatti M, Salvioli R, Ciaffoni F, Pucci P, Andolfo A, Amoresano A, Vaccaro AM
    DOI:

    Saposin D is generated together with three similar proteins, saposins A, B and C, from a common precursor, called prosaposin, in acidic organelles such as late endosomes and lysosomes. Although saposin D has been reported to stimulate the enzymatic hydrolysis of sphingomyelin and ceramide, its physiological role has not yet been clearly established. In the present study we examined structural and membrane-binding properties of saposin D. At acidic pH, saposin D showed a great affinity for phospholipid membranes containing an anionic phospholipid such as phosphatidylserine or phosphatidic acid. The binding of saposin D caused destabilization of the lipid surface and, conversely, the association with the membrane markedly affected the fluorescence properties of saposin D. The presence of phosphatidylserine-containing vesicles greatly enhanced the intrinsic tyrosine fluorescence of saposin D, which contains tyrosines but not tryptophan residues. The structural properties of saposin D were investigated in detail using advanced MS analysis. It was found that the main form of saposin D consists of 80 amino acid residues and that the six cysteine residues are linked in the following order: Cys5-Cys78, Cys8-Cys72 and Cys36-Cys47. The disulfide pattern of saposin D is identical with that previously established for two other saposins, B and C, which also exhibit a strong affinity for lipids. The common disulfide structure probably has an important role in the interaction of these proteins with membranes. The analysis of the sugar moiety of saposin D revealed that the single N-glycosylation site present in the molecule is mainly modified by high-mannose-type structures varying from two to six hexose residues. Deglycosylation had no effect on the interaction of saposin D with phospholipid membranes, indicating that the glycosylation site is not related to the lipid-binding site. The association of saposin D with membranes was highly dependent on the composition of the bilayer. Neither ceramide nor sphingomyelin, sphingolipids whose hydrolysis is favoured by saposin D, promoted its binding, while the presence of an acidic phospholipid such as phosphatidylserine or phosphatidic acid greatly favoured the interaction of saposin D with vesicles at low pH. These results suggest that, in the acidic organelles where saposins are localized, anionic phospholipids may be determinants of the saposin D topology and, conversely, saposin D may affect the lipid organization of anionic phospholipid-containing membranes.

  • Antisense inhibitory effect: a comparison between 3'-partial and full phosphorothioate antisense oligonucleotides.
    Publication Date: 01/07/1999, on Journal of cellular biochemistry
    by Galderisi U, Di Bernardo G, Melone MA, Galano G, Cascino A, Giordano A, Cipollaro M
    DOI:

    Phosphorothioate (PS) antisense oligonucleotides are currently used to inhibit many cell functions both in vivo and in vitro. However, these modified oligos provide reasonable sequence specificity only within a narrow concentration range. To overcome such a limitation we synthesized antisense oligomers, partially phosphorothioated, targeted against the human N-myc mRNA. We utilized such modified oligomers in a human neuroblastoma cell line where the N-myc gene expression was very high, and compared them to full phosphorothioate oligonucleotides. Both full PS and partial PS antisense oligos produced a maximum reduction in target mRNA after 6 h of treatment. They were able to maintain a good level of inhibition for 20 h only at high concentration. While partial PS oligos produced a dose dependent and sequence specific inhibition of N-myc mRNA, full PS molecules suffer from some disadvantages at the highest concentration used. Our results showed that partial PS molecules were capable of reducing gene expression showing a greater sequence specificity over a far broader concentration range. For this reason we conclude that partial PS antisense oligos, with respect to full PS antisense oligos, might be particularly useful for studying gene function.

  • Solution structure of dynorphin A (1-17): a NMR study in a cryoprotective solvent mixture at 278 K.
    Publication Date: 01/07/1999, on Journal of peptide science : an official publication of the European Peptide Society
    by Spadaccini R, Crescenzi O, Picone D, Tancredi T, Temussi PA
    DOI: 10.1002/(SICI)1099-1387(199907)5:7<306::AID-PSC199>3.0.CO;2-B

    Dynorphin A, the endogenous agonist for the kappa opioid receptor, has been studied by NMR spectroscopy in methanol, acetonitrile, DMSO and in mixtures of hexafluoroacetone/water and DMSO/water. NMR data in the DMSO/water cryomixture at 278 K are consistent with a conformer in which the N-terminal part, like the corresponding message domain of enkephalins, is poorly ordered, whereas the C-terminal part is folded in a loop centred around Pro10. The folded structure of the C-terminal part (address moiety) may shed light on the role of the essential residues Arg7, Lys11 and Lys13.

  • Transformation by oncogenic ras-p21 alters the processing and subcellular localization of the lysosomal protease cathepsin D.
    Publication Date: 01/06/1999, on Journal of cellular biochemistry
    by Démoz M, Castino R, Dragonetti A, Raiteri E, Baccino FM, Isidoro C
    DOI:

    The expression, processing, and intracellular localization of cathepsin D (CD), an endosomal-lysosomal protease involved in malignancy, were studied in rat embryo fibroblasts transformed with an active mutant of c-Ha-ras oncogene. The pattern of the processed molecular forms of CD, comprising two single-chain mature forms of 45 and 43 kDa and two double-chain mature forms of 34 + 9 kDa and 30 + 14 kDa, expressed by the parental cell line was similar to that found in normal rat liver cells. By contrast, in the ras-transfected counterpart this pattern was profoundly altered in that the 45 kDa species was much less represented and the 30 + 14 kDa species virtually absent. In both untransformed and ras-transformed cells the conversion of proCD into mature forms was not inhibited by ammonium chloride, which is known to increase the intravacuolar pH of post-Golgi compartments. Yet, this drug induced the accumulation of the 43 and 45 kDa molecular forms of mature CD in ras-transformed cells and of the 34 kDa molecule in untransformed cells. As compared to controls, in ras-transformed fibroblasts vacuolar compartments containing CD were reduced in number and mostly located toward the periphery of the cell. This contrasted with the perinuclear distribution of CD-positive granules in untransformed cells. Serum deprivation did not affect the growth, nor the intra- and extracellular accumulation of CD activity in ras-transformed cultures, while it blocked the growth and strongly stimulated the accumulation of CD in the medium in cultures of control fibroblasts. Altogether these data are indicative for a crucial role of ras GTPase in the regulation of the transport between post-Golgi organelles.

  • [Pathological fracture of the mandible: a report of a clinical case treated with clodronate].
    Publication Date: 01/06/1999, on Il Giornale di chirurgia
    by Califano L, Zupi A, Maremonti P, Longo F, Mangone GM, Marinelli A
    DOI:

    Oral squamous cell carcinoma often invades the mandible. However, the incidence of pathological fractures of the maxillofacial bones is low and their treatment is rarely satisfactory. A patient, too weak to undergo surgery, affected by squamous cell carcinoma of the mandibular region with bone involvement and pathological fracture of the mandible, underwent chemotherapy with carboplatin associated with diphosphonate.

  • Neglect-associated constructional disorders: a paradoxical phenomenon?
    Publication Date: 01/05/1999, on Neuropsychologia
    by Grossi D, Lepore M, Esposito A, Napolitano A, Serino M, Trojano L
    DOI:

    Five neglect patients without diffuse cognitive impairment or overt constructional disabilities were asked to bisect lines and rectangles and to copy rectangles bisected in their midplane. As a group, patients showed the usual rightward bias in bisecting lines and a milder deviation in bisecting horizontally-aligned rectangles, but showed a leftward deviation of the subjective midline in the copying task. This was due to drawing the left half shorter with respect to normal controls but three patients also drew the right half longer (the total length was the same as that of controls). A possible interpretation of rectangle copying results in these three patients is that they could create a representation of the stimulus to be copied accurately enough to reproduce its total length correctly but the subjective distribution of right and left space within that representation was unbalanced. However, specific experimental work is needed to verify why our patients with mild to moderate unilateral spatial neglect overrepresented the left side in a line bisection task and underrepresented it in a copying task.

  • Characterization of low-molecular-mass trypsin isoinhibitors from oil-rape (Brassica napus var. oleifera) seed.
    Publication Date: 01/04/1999, on European journal of biochemistry
    by Ascenzi P, Ruoppolo M, Amoresano A, Pucci P, Consonni R, Zetta L, Pascarella S, Bortolotti F, Menegatti E
    DOI:

    A new low-molecular-mass (6767.8 Da) serine proteinase isoinhibitor has been isolated from oil-rape (Brassica napus var. oleifera) seed, designated 5-oxoPro1-Gly62-RTI-III. The 5-oxoPro1-Gly62-RTI-III isoinhibitor is longer than the Asp2-Pro61-RTI-III and the Ser3-Pro61-RTI-III forms, all the other amino acid residues being identical. In RTI-III isoinhibitors, the P1-P1' reactive site bond (where residues forming the reactive site have been identified as PnellipsisP1 and P1'ellipsisPn', where P1-P1' is the inhibitor scissile bond) has been identified at position Arg21-Ile22. The inhibitor disulphide bridges pattern has been determined as Cys5-Cys27, Cys18-Cys31, Cys42-Cys52 and Cys54-Cys57. The disulphide bridge arrangement observed in the RTI-III isoinhibitors is reminiscent of that found in a number of toxins (e.g. erabutoxin b). Moreover, the organization of the three disulphide bridges subset Cys5-Cys27, Cys18-Cys31 and Cys42-Cys52 is reminiscent of that found in epidermal growth factor domains. Preliminary 1H-NMR data indicates the presence of alphaalphaNOEs and 3JalphaNH coupling constants, typical of the beta-structure(s). These data suggest that the three-dimensional structure of the RTI-III isoinhibitors may be reminiscent of that of toxins and epidermal growth factor domains, consisting of three-finger shaped loops extending from the crossover region. Values of the apparent association equilibrium constant for RTI-III isoinhibitors binding to bovine beta-trypsin and bovine alpha-chymotrypsin are 3.3 x 109 m-1 and 2.4 x 106 m-1, respectively, at pH 8.0 and 21.0 degrees C. The serine proteinase : inhibitor complex formation is a pH-dependent entropy-driven process. RTI-III isoinhibitors do not show any similarity to other serine proteinase inhibitors except the low molecular mass white mustard trypsin isoinhibitor, isolated from Sinapis alba L. seed (MTI-2). Therefore, RTI-III and MTI-2 isoinhibitors could be members of a new class of plant serine proteinase inhibitors.

  • Differentiation and apoptosis of neuroblastoma cells: role of N-myc gene product.
    Publication Date: 01/04/1999, on Journal of cellular biochemistry
    by Galderisi U, Di Bernardo G, Cipollaro M, Peluso G, Cascino A, Cotrufo R, Melone MA
    DOI:

    To clarify the role and function of the N-myc product in cell differentiation and apoptosis, we used the antisense oligonucleotide technique to inhibit N-myc gene expression in neuroblastoma cells with different phenotypes: intermediate (I) and neuronal (N), or Schwann-glia (S), respectively. The results suggest that N-myc operates along different pathways. Inhibiting N-myc gene expression either results in suppression of cell proliferation or in induction of differentiation and/or apoptosis.

  • Protein synthesis in presynaptic endings from squid brain: modulation by calcium ions.
    Publication Date: 15/03/1999, on Journal of neuroscience research
    by Benech JC, Crispino M, Kaplan BB, Giuditta A
    DOI: 10.1002/(SICI)1097-4547(19990315)55:6<776::AID-JNR12>3.0.CO;2-1

    Previous biochemical, autoradiographic, and ultrastructural data have shown that, in the synaptosomal fraction of the squid optic lobe, protein synthesis is largely due to the presynaptic terminals of the retinal photoreceptor neurons (Crispino et al. [1993a] Mol. Cell. Neurosci. 4:366-374; Crispino et al. [1993b] J. Neurochem. 61:1144-1146; Crispino et al. [1997] J. Neurosci. 17:7694-7702). We now report that this process is close to its maximum at the basal concentration of cytosolic Ca++, and is markedly inhibited when the concentration of this ion is either decreased or increased. This conclusion is supported by the results of experiments with: 1) compounds known to increase the level of cytosolic Ca++, such as A23187, ionomycin, thapsigargin, and caffeine; 2) compounds sequestering cytosolic calcium ions such as BAPTA-AM; and 3) agents that block the role of Ca++ as second messenger, such as TFP and W7, which inhibit calmodulin, and calphostin, which inhibits protein kinase C. We conclude that variations in the level of cytosolic Ca++ induced in presynaptic terminals by neuronal activity may contribute to the modulation of the local synthesis of protein.

  • Ribosomal RNAs synthesized by isolated squid nerves and ganglia differ from native ribosomal RNAs.
    Publication Date: 01/03/1999, on Journal of neurochemistry
    by Perrone-Capano C, Crispino M, Menichini E, Kaplan BB, Giuditta A
    DOI:

    The large rRNA of the squid comprises two chains that may be dissociated by heating at 65 degrees C. A single chain constitutes the small rRNA. Surprisingly, the RNAs synthesized by dissected squid fin nerves and stellate nerves and ganglia differed in size from native rRNAs and did not manifest thermal instability. Nonetheless, they resembled native rRNAs in relative abundance, subcellular distribution, lack of poly(A), and metabolic stability. In addition, newly synthesized RNA was localized in nerve and glial cells, as shown by autoradiographic analysis, and was assembled into 80S ribosomes, which supported the synthesis of neuron-specific neurofilament proteins. Following incubation of nerves and ganglia for >10 h, native rRNAs started to disappear, while two major newly synthesized RNAs progressively accumulated. As a result, after 20 h, native rRNAs were substituted by the two novel RNAs. With use of 32P-cDNA synthesized from the latter RNAs as a probe, the novel RNAs demonstrated a considerable degree of homology with native rRNA in northern analysis. Taken together, the data suggest that in dissected squid nerves and ganglia, the synthesis of native rRNAs is gradually terminated while two novel rRNAs are being synthesized, presumably as a correlate of reactive gliosis and/or neuronal degeneration/regeneration.

  • Craniofacial pain followed by scalp necrosis and stroke. An unusual presentation of the primary antiphospholipid syndrome.
    Publication Date: 01/02/1999, on Journal of neurology
    by Sanges G, Sampaolo S, Di Iorio G
    DOI:

  • Neuroendocrine (Merkel cell) carcinoma of the oral mucosa: report of a case with immunohistochemical study and review of the literature.
    Publication Date: 01/02/1999, on Journal of oral pathology & medicine : official publication of the International Association of Oral Pathologists and the American Academy of Oral Pathology
    by Longo F, Califano L, Mangone GM, Errico ME
    DOI:

    Merkel cell carcinoma (Mcc) is an uncommon and aggressive tumour with neuroendocrine features that occur predominantly in the head and neck region. The rarity of this tumour, especially when it arises in the oral mucosa, makes both early identification and standardisation of treatment difficult, particularly as regards complementary treatment. The availability of monoclonal antibodies with restricted specificity for some antigens thought to be related to neuroendocrine carcinomas, such as Merkel cell carcinoma, and ultrastructural studies offer some new leads to investigation. This has allowed, a greater number of these tumours to be discovered, thereby increasing the chances of effective management. A case of Mcc of the floor of the mouth is reported, together with the results of cytokeratin, neuron specific enolase and chromogranin immunohistochemistry.