Biomedical subjects

S Giordano

Publications and source records attributed to S Giordano.

At least 37 records · Page 2Linked to original sources

PubMed2001

Gab1 phosphorylation: a novel mechanism for negative regulation of HGF receptor signaling.

Signal transduction by HGF receptor, the tyrosine kinase encoded by the MET oncogene, switches on a genetic program called 'invasive growth' inducing epithelial cell dissociation, migration, growth, and ultimately leading to differentiation into branched tubular structures. Sustained tyrosine phosphorylation of the downstream adaptor protein Gab1 is required for the HGF response. Here we show that serine/threonine phosphorylation of Gab1 provides a control mechanism for negative regulation. Treatment with okadaic acid, a potent inhibitor of the serine/threonine protein phosphatases PP1 and PP2A, was followed by activation of a number of serine/threonine kinases, hyper-phosphorylation in serine and threonine of Gab1 and severe inhibition of the HGF-induced biological responses. Under these conditions, Gab1 was found to be concomitantly hypo-phosphorylated in tyrosine, and thus endowed with reduced ability to recruit SH2 containing signal transducers such as PI3 kinase. Among the serine-threonine kinases activated by PP1 and PP2A inhibition, we found that PKC-alpha and PKC-beta1 are required for negative regulation of Gab1. These data provide a novel negative mechanism for the HGF receptor signaling pathways and highlight a potentially useful target for inhibitors of invasive growth.

Adaptor Proteins, Signal Transducing↗

PubMed2001

MET receptor is overexpressed but not mutated in oral squamous cell carcinomas.

Oral squamous cell carcinoma (SCC) is a neoplasm characterized by a high degree of local invasion and an elevated rate of metastasis to cervical lymph nodes. It has been shown that the Hepatocyte Growth Factor/Scatter Factor Receptor Met is constitutively activated in many human tumors of epithelial origin and that it plays a critical role to confer invasive properties to neoplastic cells. Most frequently, Met activation is due to receptor overexpression, but also point mutations in the tyrosine kinase domain can lead to deregulated activation. Here we show that in all the primary tumors examined this receptor is overexpressed. Direct sequencing of Met mRNAs failed to find any activating mutation in its intracellular domain. Moreover, in cell lines derived from squamous cell carcinomas, HGF-induced activation of Met resulted in the acquisition of invasive properties. All together these data suggest that the MET oncogene is involved in progression of squamous cell carcinoma toward an invasive-metastatic behavior.

Adult↗

PubMed2001

[Epidemiological features of intestinal parasitosis in western Sicily in the period 1993-2000].

To determine the prevalence of intestinal parasitic infections in western Sicily from 1993 to 2000, fecal samples from 3024 patients (subdivided into native Sicilian children, migrants from developing countries and HIV+ patients,) were collected and observed on direct examination or by the formalin-ether sedimentation technique of Ritchie. Lugol, Kinyoun and Gomori's trichromic stain techniques were employed. Trophozoites or mainly protozoan cysts and helminthic eggs were detected in 195 patients (6.4%). The positivity rate was 18.7% in HIV+ patients, 10.3% in migrants and 5.0% in native Sicilian children. Tape test for Enterobius eggs was employed on 1449 patients with a positivity rate of 11%. The importance of parasitology also in new millennium is emphasized.

Adolescent↗

PubMed2000

Somatic mutations of the MET oncogene are selected during metastatic spread of human HNSC carcinomas.

A metastatic cancer develops by accumulation of mutations in genes that control growth, survival and spreading. The latter genes have not yet been identified. In lymph node metastases of head and neck squamous cell carcinomas (HNSCC), we found mutations in the MET oncogene, which encodes the tyrosine kinase receptor for Scatter Factor, a cytokine that stimulates epithelial cell motility and invasiveness during embryogenesis and tissue remodeling. We identified two somatic mutations: the Y1230C, known as a MET germline mutation which predisposes to hereditary renal cell carcinoma, and the Y1235D that is novel and changes a critical tyrosine, known to regulate MET kinase activity. The mutated MET receptors are constitutively active and confer an invasive phenotype to transfected cells. Interestingly, cells carrying the MET mutations are selected during metastatic spread: transcripts of the mutant alleles are highly represented in metastases, but barely detectable in primary tumors. These data indicate that cells expressing mutant MET undergo clonal expansion during HNSCC progression and suggest that MET might be one of the long sought oncogenes controlling progression of primary cancers to metastasis.

Alleles↗

PubMed2000

Sustained recruitment of phospholipase C-gamma to Gab1 is required for HGF-induced branching tubulogenesis.

A distinctive property of Hepatocyte Growth Factor (HGF) is its ability to induce differentiation of tubular structures from epithelial and endothelial cells (branching tubulogenesis). The HGF receptor directly activates PI3 kinase, Ras and STAT signalling pathways and phosphorylates the adaptator GRB2 Associated Binder-1 (Gab1). Gab1 is also phosphorylated in response to Epidermal Growth Factor (EGF) but is unable to induce tubule formation. Comparison of 32P-peptide maps of Gab1 from EGF- versus HGF-treated cells, demonstrates that the same sites are phosphorylated in vivo. However, while both EGF and HGF induce rapid tyrosine phosphorylation of Gab1 with a peak at 15 min, the phosphorylation persists for over 1 h, only in response to HGF. Nine tyrosines are phosphorylated by both receptors. Three of them (Y307, Y373, Y407) bind phospholipase C-gamma (PLC-gamma). Interestingly, the overexpression of a Gab1 mutant unable to bind PLC-gamma (Gab1 Y307/373/407F) did not alter HGF-stimulated cell scattering, only partially reduced the growth stimulation but completely abolished HGF-mediated tubulogenesis. It is concluded that sustained recruitment of PLCgamma to Gab1 plays an important role in branching tubulogenesis.

Animals↗

PubMed2000

Identification of the Coronal Sources of the Fast Solar Wind.

The present spectroscopic study of the ultraviolet coronal emission in a polar hole, detected on 1996 April 6-9 with the Ultraviolet Coronagraph Spectrometer aboard the Solar and Heliospheric Observatory spacecraft, identifies the interplume lanes and background coronal hole regions as the channels in which the fast solar wind is preferentially accelerated. In interplume lanes, at heliocentric distance 1.7 R middle dot in circle, the corona expands at a rate between 105 and 150 km s-1, that is, much faster than in plumes in which the outflow velocity is between 0 and 65 km s-1. The wind velocity is inferred from the Doppler dimming of the O vi lambdalambda1032, 1037 lines, within a range of values, whose lower and upper limit corresponds to anisotropic and isotropic velocity distribution of the oxygen coronal ions, respectively.

Journal Article↗

PubMed2000

DC-ELF characterization of random mixtures of piecewise nonlinear media.

Biological tissues are ensembles of linear and nonlinear, symmetric and asymmetric constituents. As far as their electromagnetic characterization is concerned, they can be modeled as microscopic mixtures of the corresponding material media. Any medium volume can be properly discretized in a finite number of cells which can be modeled as an equivalent three dimensional network of lumped components, in order to characterize its electromagnetic behavior at wavelengths much longer than the relevant average linear size of the constitutive cells. Therefore, any mixture and the corresponding tissue can be characterized in terms of its effective conductance at extremely low frequency, with respect to a reference set of electrodes (ports of the equivalent network). When the above procedure is implemented for evaluating any of the aforesaid conductances, a resulting nonlinear characteristic should be expected. In reality, it may happen that the effect of the constitutive nonlinearities and the related asymmetries are smeared out by the randomness of the interconnections of the lumped components, leading at a macroscopic level to an isotropic constant equivalent conductance, i.e., to an isotropic constant equivalent conductivity of the mixture. The closed form analysis of a random network of nonlinear (piecewise linear) resistors offers a simple but clear cut example of such a property. This result, if extrapolated to biological media, suggests a new hint for explaining why there is no inconsistency between the typical electric characterization of biological tissues as almost linear macroscopic media, by means of their effective conductivity and permittivity, and the nonlinearities of the biochemical processes occurring in the tissue cells. In fact, the nonlinearities may not be observable by means of macroscopic electrical measurements because of the randomized spatial orientation and location of the processes.

Animals↗

PubMed2000

Expression of functional tyrosine kinases on immortalized Kaposi's sarcoma cells.

Kaposi's sarcoma (KS) is the most frequent malignant lesion in patients with AIDS and is characterized by spindle cell proliferation, inflammatory cell infiltration, angiogenesis, edema, and invasiveness. KS origin is still debated. The complex aspect of this disease is probably supported by multiple concomitant pathogenetic factors, among which growth factors and their cognate tyrosine kinase receptors are deeply involved. Here we have investigated the expression status and functional integrity of KDR and Met receptors, as well as of their ligands, in an immortalized KS cell line (KS-IMM). The MET and KDR genes encode the tyrosine kinase receptors for Hepatocyte Growth Factor (HGF) and Vascular Endothelial Growth Factor (VEGF) respectively. Both factors are pleiotropic cytokines controlling growth, survival, motility, invasive migration and differentiation of endothelial cells. We have found that KS-IMM cells, which retain most of the features of the parental tumor and can induce KS-like sarcomas when injected subcutaneously in nude mice, express the Met receptor, but not its ligand. The receptor, which is basally inactive, is functional, being tyrosine phosphorylated in response to ligand stimulation and mediating the expected HGF-dependent biological responses of motility, invasion and proliferation. Moreover, we report that KS-IMM cells coexpress VEGF and KDR and that KDR is constitutively tyrosine phosphorylated, possibly as a consequence of the establishment of an autocrine loop. The receptor, however, maintains responsiveness to exogenously added ligand, by increasing the level of tyrosine phosphorylation and by responding in biological assays of motility, invasion and proliferation. Finally, we have found that the two growth factors synergize in a motility assay. These data show that HGF and VEGF are growth factors active on KS-IMM cells.

Animals↗

PubMed2000

Antibacterial and allelopathic activity of extract from Castanea sativa leaves.

Following the extraction of Castanea sativa with an aqueous solution of sulfuric acid (pH 3.0), the ethyl acetate soluble fraction was tested for its antibacterial and allelopathic activity. The extract was shown to have pronounced antibacterial effects against seven of the eight strains of Gram-positive and Gram-negative bacteria used (MIC in the range of 64-256 microg/ml and MBC in the range of 256-512 microg/ml). The active fraction was analyzed by TLC and HPLC showing the presence of rutin, hesperidin, quercetin, apigenin, morin, naringin, galangin and kaempferol. Standards of the identified flavonoids were tested against the same bacterial strains. The highest activity was shown by quercetin, rutin and apigenin. The allelopathic effect was tested against Raphanus sativus seed germination. The extract, quercetin, rutin and apigenin caused a decrease in the percentage of seed germination and root and epicotyl growth.

Anti-Bacterial Agents↗

PubMed2000

For the protection of others. The value of individual autonomy and the safety of others.

This paper investigates legal and moral justifications of coerced treatment for psychiatric patients who are detained on the grounds that they may harm others. While the general issues concerning compulsory treatment and detention have been widely canvassed, it has seldom, if ever, been noticed that the moral reasons that we may have to detain a person who appears to be dangerous to others are different from the moral reasons we may have to treat him or her. For example, it has not been noticed that compulsory detention and compulsory treatment are supported by two different moral principles, namely the Principle of Harm and the Principle of Beneficence, and, therefore, that the arguments which support compulsory detention do not also support compulsory treatment. The conceptual confusion between legitimacy of compulsory detention and legitimacy of compulsory treatment is exacerbated by the ambiguous wording utilised in S 3 of the UK Mental Health Act, which implies that treatment may be necessary for the protection of others. Failure to pay attention to these distinctions has led to tragic consequences, in terms of violations of individual autonomy and in terms of public safety.

Antisocial Personality Disorder↗

PubMed2000

Different point mutations in the met oncogene elicit distinct biological properties.

The MET proto-oncogene, encoding the tyrosine kinase receptor for HGF, controls genetic programs leading to cell growth, invasiveness, and protection from apoptosis. Recently, MET mutations have been identified in hereditary and sporadic forms of papillary renal carcinoma (PRC). Introduction of different naturally occurring mutations into the MET cDNA results in the acquisition of distinct biochemical and biological properties of transfected cells. Some mutations result in a high increase in tyrosine kinase activity and confer transforming ability in focus forming assays. These mutants hyperactivate the Ras signaling pathway. Other mutations are devoid of transforming potential but are effective in inducing protection from apoptosis and sustaining anchorage-independent growth. These Met(PRC) receptors interact more efficiently with the intracellular transducer Pi3Kinase. The reported results show that MET(PRC) mutations can be responsible for malignant transformation through different mechanisms, either by increasing the growth ability of cells or by protecting cells from apoptosis and allowing accumulation of other genetic lesions.-Giordano, S., Maffe, A., Williams, T. A., Artigiani, S., Gual, P., Bardelli, A., Basilico, C., Michieli, P., Comoglio, P. M. Different point mutations in the met oncogene elicit distinct biological properties.

Apoptosis↗

PubMed1999

Mutant Met-mediated transformation is ligand-dependent and can be inhibited by HGF antagonists.

Mutations in the genes encoding for Met, Ret and Kit receptor tyrosine kinases invariably result in increased kinase activity and in the acquisition of transforming potential. However, the requirement of receptor ligands for the transformation process is still unclear. We have investigated the role of hepatocyte growth factor (HGF), the high-affinity ligand for Met, in mutant Met-mediated cell transformation. We provide evidence that the transforming potential displayed by mutant forms of Met found in human cancer is not only sensitive but entirely dependent on the presence of HGF, by showing that mutant Met transforms NIH3T3 fibroblasts, which produce endogenous HGF, but is not able to transform epithelial cells, unless exogenous HGF is supplied. Accordingly, mutant Met-induced transformation of NIH3T3 cells can be inhibited by HGF antagonists and increased by HGF stimulation. We also show that an engineered Met receptor which contains an oncogenic mutation but is impaired in its ability to bind HGF completely loses its transforming activity, which can be rescued by causing receptor dimerization using a monoclonal antibody. These results indicate that point mutations resulting in Met kinase activation are necessary but not sufficient to cause cell transformation, the latter being dependent on ligand-induced receptor dimerization. They also suggest that mutant Met-driven tumour growth depends on the availability and tissue distribution of active HGF, and provide proof-of-concept for the treatment of mutant-Met related pathologies by HGF-antagonizing drugs.

3T3 Cells↗

PubMed1999

Concomitant activation of pathways downstream of Grb2 and PI 3-kinase is required for MET-mediated metastasis.

The Met tyrosine kinase - the HGF receptor - induces cell transformation and metastasis when constitutively activated. Met signaling is mediated by phosphorylation of two carboxy-terminal tyrosines which act as docking sites for a number of SH2-containing molecules. These include Grb2 and p85 which couple the receptor, respectively, with Ras and PI 3-kinase. We previously showed that a Met mutant designed to obtain preferential coupling with Grb2 (Met2xGrb2) is permissive for motility, increases transformation, but - surprisingly - is impaired in causing invasion and metastasis. In this work we used Met mutants optimized for binding either p85 alone (Met2xPI3K) or p85 and Grb2 (MetPI3K/Grb2) to evaluate the relative importance of Ras and PI 3-kinase as downstream effectors of Met. Met2xPI3K was competent in eliciting motility, but not transformation, invasion, or metastasis. Conversely, MetP13K/Grb2 induced motility, transformation, invasion and metastasis as efficiently as wild type Met. Furthermore, the expression of constitutively active PI 3-kinase in cells transformed by the Met2xGrb2 mutant, fully rescued their ability to invade and metastasize. These data point to a central role for PI 3-kinase in Met-mediated invasiveness, and indicate that simultaneous activation of Ras and PI 3-kinase is required to unleash the Met metastatic potential.

Adaptor Proteins, Signal Transducing↗

PubMed1999

Antibacterial activity of pure flavonoids isolated from mosses.

Seven pure flavonoids were isolated and identified from five moss species. The flavonoids were the flavones apigenin, apigenin-7-O-triglycoside, lucenin-2, luteolin-7-O-neohesperidoside, saponarine and vitexin; and the biflavonoid bartramiaflavone. Some of these flavonoids were shown to have pronounced antibacterial effects against Enterobacter cloaceae, E. aerogenes and Pseudomonas aeruginosa (minimal bacteriostatic concentration MIC in the range of 4-2048 micrograms/ml). Because of their antibacterial spectrum mainly active against Gram negative bacterial strains, responsible for severe opportunistic infections and resistant to common antibacterial therapy, these flavonoids may be important tools in antibacterial strategies.

Anti-Bacterial Agents↗

PubMed1998

Uncoupling signal transducers from oncogenic MET mutants abrogates cell transformation and inhibits invasive growth.

The assumption that genes encoding tyrosine kinase receptors could play a role in human cancers has been confirmed by the identification of oncogenic mutations in the kinase domain of RET and KIT. Recently, homologous residues were found mutated in MET, in papillary renal carcinomas (PRCs). The link coupling these genetic lesions to cellular transformation is still unclear. METPRC mutations result in increased kinase activity and-in some instances, i.e., M1250T substitution-in changes in substrate specificity. A direct correlation occurs between the transforming potential of METPRC mutants and their ability to constitutively associate with signal transducers through two phosphorylated tyrosines (Y1349VHVNATY1356VNV) located in the receptor tail. Substitution of these "docking tyrosines" with phenylalanines leaves unaffected the altered properties of the kinase but abrogates transformation and invasiveness in vitro. Uncoupling the receptor from signal transducers with a tyrosine-phosphorylated peptide derivative (YpVNV) inhibits invasive growth induced by METPRC mutants. These data indicate that constitutive receptor coupling to downstream signal transducers is a key mechanism in neoplastic transformation driven by mutated MET and suggest a therapeutic strategy to target neoplastic diseases associated with this oncogene.

Amino Acid Sequence↗

PubMed1998

Synthesis and comparison of the antiinflammatory activity of manoalide and cacospongionolide B analogues.

We have synthesized analogues of two naturally occurring antiinflammatory marine compounds, manoalide and cacospongionolide B, containing a pyranofuranone moiety which is considered the pharmacophoric group. The two compounds, and hence their analogues, differ in the presence or absence in the dihydropyran ring of an hemiacetal function which was considered essential to irreversibly inactivate phospholipase A2 (PLA2). The two series of compounds were tested for their inhibitory effects on secretory PLA2 belonging to the groups I, II, and III, and the activities were found to be similar in both series, irrespective of the presence or absence of the additional hemiacetal function. In addition, the PLA2 inhibitory activity increases with the increasing hydrophobic character of the side chain linked to the pyranofuranone moiety. The most active compounds, FCA and FMA, carry a farnesyl residue linked to the pyranofuranone substructure. The most potent PLA2 inhibitor, FMA, was tested in the mouse carrageenan paw edema at the oral dose of 10 mg/kg and showed an activity similar to the reference antiinflammatory drug, indomethacin.

4-Butyrolactone↗

PubMed1998

Antibacterial activity in Pleurochaete squarrosa extract (Bryophyta).

An acetone extract of the moss Pleurochaete squarrosa was tested against eleven bacterial strains, some of which are pathogenous for man. The extract was active on some Gram-negative strains. Antibacterial activity of the extract, expressed as MICs, was compared with three reference antibiotics. Acute toxicity assay was performed in Balb C mice.

Acetone↗

PubMed1998

C-met activation is necessary but not sufficient for liver colonization by B16 murine melanoma cells.

Metastasis to the liver is a frequent event in clinical oncology, the molecular mechanisms of which are not fully understood. We have recently reported a consistent overexpression of c-met in B16 melanoma cells selected in vivo for enhanced liver metastatic ability. In this study we address the question as to whether constitutive activation of c-met is a necessary and sufficient condition for enhanced liver colonization by B16 melanoma cells. Different levels of c-met expression and/or activation in B16 cells were achieved by subcloning, or by c-DNA transfection with either HGF/SF or the oncogenic form of c-met (tpr-met). Metastatic ability of the different populations was then evaluated in vivo by the lung colonization (experimental metastasis) assay. Results indicate that c-met (but not tpr-met) activation in B16 melanoma cells may increase their liver colonizing potential, probably by enhancing motility and invasion in response to paracrine interactions with its ligand. C-met expression per se, however, is not able to change the organ specificity of the cells. C-met activation appears instead to be required at later stages of liver colonization by B16 melanoma cells, in order to enhance their site-specific metastatic ability.

Animals↗