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F Facchiano

Publications and source records attributed to F Facchiano.

At least 19 recordsLinked to original sources
PubMed2004

A novel RGDS-analog inhibits angiogenesis in vitro and in vivo.

In this study the anti-angiogenic action of a novel non-peptide RGDS-analog named RAM was tested in vitro and in vivo. RAM inhibited FGF-2-induced chemotaxis by 80% in an adhesion-independent way. Further, it induced HUVEC-apoptosis in collagen-seeded HUVEC, indicating that such pro-apoptotic effect was adhesion-independent. In vivo studies revealed that RAM inhibited FGF-2 induced angiogenesis by 60% in the mouse Matrigel-assay and in the chicken-egg chorion-allantoic membrane assay. Finally, RAM was markedly more stable in serum as compared to the template RGDS and after 24 h incubation in 100% serum was significantly more active than RGDS. Taken together these results show that RAM exerts anti-chemotactic and pro-apoptotic effects, by an unexpected adhesion-independent mechanism, as we have recently shown for the template RGDS molecule [Blood 103 (2004) 4180], and has in vivo relevant anti-angiogenic properties, with marked stability in serum; therefore, RAM represents a novel promising anti-angiogenic molecule.

Angiogenesis Inhibitors↗
PubMed2001

Transglutaminase activity is involved in polyamine-induced programmed cell death.

Natural polyamines, i.e., putrescine, spermidine, and spermine, are ubiquitous molecules essential for cell proliferation and differentiation. In the present study, the effect of polyamines on primary cultures of bovine aortic endothelial cells (BAECs), rat aortic smooth muscle cells (RASMCs), and a human melanoma cell line was examined. While in the absence of fetal calf serum (FCS) polyamines had no effect on viability, in the presence of FCS spermidine and spermine, at concentrations close to physiologic levels, induced a dose-dependent cell death, whereas putrescine was ineffective. RASMCs were significantly more sensitive than other cells. FACS analysis, oligo-nucleosome ELISA, Hoechst nuclear staining, and Annexin V-FITC quantification showed that cell death was likely due to apoptosis. Cells exposed to spermidine showed a marked increase of intracellular transglutaminase (TGase) activity ( approximately 30-fold over control). Inhibitors of polyamine oxidation or inhibitors of TGase activity prevented polyamine-induced apoptosis. Moreover, tissue TGase overexpression significantly increased cell sensitivity to polyamine, suggesting that this effect is likely related to enhanced intracellular TGase activity. These data indicate that polyamines may modulate cell viability through a novel TGase-dependent process.

Animals↗
PubMed2001

Retinoids induce fibroblast growth factor-2 production in endothelial cells via retinoic acid receptor alpha activation and stimulate angiogenesis in vitro and in vivo.

The effect of retinoic acid (RA) on endothelial cells is still controversial and was examined in the present study. In bovine aortic endothelial cells (BAECs), all-trans RA (ATRA) and 9-cis RA (9CRA), but not 13-cis RA (13CRA), induced fibroblast growth factor-2 (FGF-2) production and exhibited a biphasic dose-dependent effect to enhance BAEC proliferation and differentiation into tubular structures on reconstituted basement membrane proteins (Matrigel); both processes were inhibited by FGF-2-neutralizing antibody. The pan RA receptor (RAR)-selective ligand (E)-4-[2-(5,5,8,8,-tetramethyl-5,6,7,8-tetrahydro-2-naphtalenyl)-1-propenyl] benzoic acid and the RARalpha-selective ligand 4-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphtyl)-ethenyl] benzoic acid stimulated the production of FGF-2, whereas the addition of the RARalpha-antagonist RO 41-5253 inhibited this effect. In BAECs, the forced expression of RARalpha, but not RARbeta or RARgamma, enhanced FGF-2 production, whereas the RARalpha-dominant negative, Delta403, blocked this effect. Furthermore, RARalpha overexpression directly stimulated BAEC differentiation on Matrigel and potentiated the effects of ATRA in this assay. Finally, ATRA-treated BAECs coinjected with Matrigel subcutaneously in mice induced neovascularization within the Matrigel plug, and ATRA also enhanced angiogenesis in the chicken chorioallantoic membrane assay. In conclusion, RA can stimulate endothelial cell proliferation and differentiation in vitro via enhanced RARalpha-dependent FGF-2 production, and it can also induce angiogenesis in vivo. The full text of this article is available at http://www.circresaha.org.

Animals↗
PubMed2000

Acidosis inhibits endothelial cell apoptosis and function and induces basic fibroblast growth factor and vascular endothelial growth factor expression.

Endothelial cells are exposed to an acidotic environment in a variety of pathological and physiological conditions. However, the effect of acidosis on endothelial cell function is still largely unknown, and it was evaluated in the present study. Bovine aortic endothelial cells (BAECs) were grown in bicarbonate buffer equilibrated either with 20% CO(2) (pH 7.0, acidosis) or 5% CO(2) (pH 7.4, control). Acidosis inhibited BAEC proliferation in 10% FCS, whereas by day 7 in serum-free medium, cell number was 3-fold higher in acidotic cells than in control cells. Serum deprivation enhanced BAEC apoptosis, and apoptotic cell death was markedly inhibited by acidosis. Additionally, acidosis inhibited FCS-stimulated migration in a modified Boyden chamber assay and FCS-stimulated differentiation into capillary-like structures on reconstituted basement membrane proteins. Conditioned media from BAECs cultured for 48 hours either at pH 7.0 or pH 7.4 enhanced BAEC proliferation and migration at pH 7.4, and both effects were more marked with conditioned medium from BAECs grown in acidotic than in control conditions. Acidosis enhanced vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) mRNA expression as well as bFGF secretion, and a blocking bFGF antibody inhibited enhanced BAEC migration in response to conditioned medium from acidotic cells. These results show that acidosis protects endothelial cells from apoptosis and inhibits their proangiogenic behavior despite enhanced VEGF and bFGF mRNA expression and bFGF secretion.

Acidosis↗
PubMed2000

Theophylline administration markedly reduces hepatic and pulmonary implantation of B16-F10 melanoma cells in mice.

Theophylline-treated B16-F10 melanoma cells show a lower experimental metastatic potential in vivo. To identify the possible mechanism(s) involved and on the basis of previous reports, we tested the induction of apoptosis in B16-F10 cells. Fluorescence activated cell sorter (FACS) analysis and p53 overexpression in theophylline-treated B16-F10 melanoma cells appeared to suggest enhanced cell death by apoptosis. The in vivo effects of orally administered theophylline in mice were investigated using different treatment schedules in mice that had undergone hepatic or pulmonary colonization with tumour cells. Mice received theophylline in their drinking water according to different protocols: (i) from 3 days before tumour cell inoculation until animal sacrifice ('early treatment'); (ii) from 3 days before until 3 days after tumour cell inoculation ('short treatment'); or (iii) from 3 days after tumour cell inoculation until animal sacrifice ('late treatment'). In the 'early treatment' group, the number of melanoma foci was reduced by 92.3% in the liver and 81.4% in the lung compared with control animals (P < 0.001). In the 'short treatment' group, there was an 80.2% and 72.2% reduction in liver and lung metastases, respectively (P < 0.001). In the 'late treatment' group, the inhibition of metastasis was 59.7% for liver and 45.3% for lung (P < 0.005). Survival studies showed that 50% of the 'early' theophylline-treated animals died 33.2 +/- 2.0 days after intrasplenic injection (control group: 23.1 1.8 days; P < 0.001) and 33.9 +/- 2.5 days after tail vein injection (control group: 24.1 +/- 1.4 days; P < 0.001). Taken together, these observations provide useful information for the potential clinical application of theophylline as a chemotherapeutic agent against malignant melanoma.

Animals↗
PubMed2000

The chemotactic and mitogenic effects of platelet-derived growth factor-BB on rat aorta smooth muscle cells are inhibited by basic fibroblast growth factor.

In response to endovascular injury, platelet-derived growth factor-BB (PDGF-BB) and basic fibroblast growth factor (bFGF) are released locally and modulate vascular smooth muscle cells (SMC) proliferation and migration within the vascular wall. The aim of the present in vitro study was to determine how rat aorta SMC respond to the simultaneous exposure to PDGF-BB and bFGF. In a modified Boyden chamber assay bFGF exhibited a dose-dependent effect to inhibit the chemotactic action of PDGF-BB. A comparable result was observed in proliferation assays. In contrast, MIP-1 beta, epidermal growth factor (EGF), fibronectin and acidic FGF (aFGF) did not inhibit the chemotactic effect of PDGF-BB. Denatured bFGF did not exert an inhibitory effect and neutralizing antibodies either to bFGF or to bFGF-receptor abolished the inhibition observed in the presence of bFGF. The role played by PDGF receptor alpha (PDGF-Ralpha) was investigated in PDGF-Ralpha-dominant negative-transfected SMC, by selectively blocking PDGF-BB-binding to PDGF-Ralpha with neomycin, by neutralizing PDGF-Ralpha with a monoclonal antibody and by selectively stimulating PDGF-Ralpha with PDGF-AA; in all cases the effect of bFGF to inhibit PDGF-BB-directed SMC migration was abolished. These in vitro studies show that bFGF significantly inhibits PDGF-BB-induced SMC migration and proliferation and that this effect is mediated by both PDGF-Ralpha and bFGF receptor.

Animals↗
PubMed1999

CtBP/BARS induces fission of Golgi membranes by acylating lysophosphatidic acid.

Membrane fission is essential in intracellular transport. Acyl-coenzyme As (acyl-CoAs) are important in lipid remodelling and are required for fission of COPI-coated vesicles. Here we show that CtBP/BARS, a protein that functions in the dynamics of Golgi tubules, is an essential component of the fission machinery operating at Golgi tubular networks, including Golgi compartments involved in protein transport and sorting. CtBP/BARS-induced fission was preceded by the formation of constricted sites in Golgi tubules, whose extreme curvature is likely to involve local changes in the membrane lipid composition. We find that CtBP/BARS uses acyl-CoA to selectively catalyse the acylation of lysophosphatidic acid to phosphatidic acid both in pure lipidic systems and in Golgi membranes, and that this reaction is essential for fission. Our results indicate a key role for lipid metabolic pathways in membrane fission.

Acyl Coenzyme A↗
PubMed1997

Fas-induced changes in cdc2 and cdk2 kinase activity are not sufficient for triggering apoptosis in HUT-78 cells.

Recent evidence suggested a role for the cell cycle dependent kinases cdc2 and cdk2 in apoptosis. An important mechanism by which many cell types could undergo apoptosis is through the activation of the Fas molecule on the cell membrane. To investigate whether Fas-induced cell death activated cdc2 and cdk2 kinases inappropriately, the human T lymphoma cells HUT-78, which express a high copy number of Fas, and two other previously characterized subclones of the same cell line which express mutant, cell death-deficient dominant-negative forms of Fas, were Fas-challenged and the changes in cdc2 and cdk2 kinase activity monitored. In both wild-type and Fas-mutated HUT-78 cells, apoptosis was associated simultaneously with decreased cdc2 and increased cdk2 activity. This association suggested that changes in cdc2 and cdk2 kinase activity are secondary events in cell death mediated by Fas.

Apoptosis↗
PubMed1997

Ectopic TAL-1/SCL expression in phenotypically normal or leukemic myeloid precursors: proliferative and antiapoptotic effects coupled with a differentiation blockade.

The TAL-1 gene specifies a basic helix-loop-helix domain (bHLH) transcription factor, which heterodimerizes with E2A gene family proteins. tal-1 protein is abnormally expressed in the majority of T-cell acute lymphoblastic leukemias (T-ALLs). tal-1 is expressed and plays a significant role in normal erythropoietic differentiation and maturation, while its expression in early myeloid differentiation is abruptly shut off at the level of late progenitors/early differentiated precursors (G. L. Condorelli, L. Vitelli, M. Valtieri, I. Marta, E. Montesoro, V. Lulli, R. Baer, and C. Peschle, Blood 86:164-175, 1995). We show that in late myeloid progenitors (the phenotypically normal murine 32D cell line) and early leukemic precursors (the human HL-60 promyelocytic leukemia cell line) ectopic tal-1 expression induces (i) a proliferative effect under suboptimal culture conditions (i.e., low growth factor and serum concentrations respectively), via an antiapoptotic effect in 32D cells or increased DNA synthesis in HL-60 cells, and (ii) a total or marked inhibitory effect on differentiation, respectively, on granulocyte colony-stimulating factor-induced granulopoiesis in 32D cells or retinoic acid- and vitamin D3-induced granulo- and monocytopoiesis in HL-60 cells. Furthermore, experiments with 32D temperature-sensitive p53 cells indicate that aberrant tal-1 expression at the permissive temperature does not exert a proliferative effect but causes p53-mediated apoptosis, i.e., the tal-1 proliferative effect depends on the integrity of the cell cycle checkpoints of the host cell, as observed for c-myc and other oncogenes. tal-1 mutant experiments indicate that ectopic tal-1 effects are mediated by both the DNA-binding and the heterodimerization domains, while the N-terminally truncated tal-1 variant (M3) expressed in T-ALL malignant cells mimics the effects of the wild-type protein. Altogether, our results (i) indicate proliferative and antidifferentiative effects of ectopic tal-1 expression, (ii) shed light on the underlying mechanisms (i.e., requirement for the integrity of the tal-1 bHLH domain and cell cycle checkpoints in the host cell, particularly p53), and (iii) provide new experimental models to further investigate these mechanisms.

Apoptosis↗
PubMed1996

T-cell-directed TAL-1 expression induces T-cell malignancies in transgenic mice.

The TAL-1 gene specifies for a basic domain-helix-loop-helix protein, which is involved in the control of normal hematopoiesis. In human pathology, the TAL-1 gene product is expressed in a high percentage of T-cell acute lymphoblastic leukemias in the pediatric age range; however, it has not been established whether the expression has a causal role in oncogenesis. In this report, we describe the phenotype of mouse transgenic lines obtained by inducing tal-1 protein expression in lymphoid tissues using the LCK promoter. The survival rate of tal-1 transgenic animals was much lower as compared with control mice. Histopathological analysis revealed lymphomas of T-cell type, often comprising a minor B-cell component. Some mice showed marked splenic lymphocyte depletion. Primary lymphocyte cultures showed partial independence from exogenous growth stimuli and increased resistance to low-serum apoptosis. To further unravel the tal-1 oncogenic potential, a strain of tal-1 transgenic mice was crossbred with p53-/- mice; the survival rate in these animals was reduced by more than one-half when compared with that of tal-1 mice, and histopathological analysis revealed exclusively T-cell lymphomas. These data indicate that TAL-1, expressed in T cells, is per se a potent oncogene, which may exert a key leukemogenetic role in the majority of T-cell acute lymphoblastic leukemias.

Adenovirus E2 Proteins↗
PubMed1996

Regulation of constitutive exocytic transport by membrane receptors. A biochemical and morphometric study.

Biochemical and morphometric approaches were combined to examine whether constitutive secretory transport might be controlled by plasma membrane receptors, as this possibility would have significant physiological implications. Indeed, IgE receptor stimulation in rat basophilic leukemia cells potently increased the rate of transport of soluble pulse-labeled 35S-sulfated glycosaminoglycans from distal Golgi compartments to the cell surface. This effect was largely protein kinase C (PKC)-dependent. Direct activation of PKC also stimulated constitutive transport of glycosaminoglycans, as indicated by the use of agonistic and antagonistic PKC ligands. PKC ligands also had potent, but different, effects on the exocytic transport from distal Golgi compartments to the plasma membrane of a membrane-bound protein (vesicular stomatitis virus glycoprotein), which was slightly stimulated by activators and profoundly suppressed by inhibitors of PKC. Morphological analysis showed impressive changes of the organelles of the secretory pathway in response to IgE receptor stimulation and to direct PKC activation (enhanced number of buds and vesicles originating from the endoplasmic reticulum and Golgi and increase in surface and volume of Golgi compartments), suggestive of an overall activation of exocytic movements. These results show that rapid and large changes in constitutive transport fluxes and in the morphology of the exocytic apparatus can be induced by membrane receptors (as well as by direct PKC stimulation).

Animals↗
PubMed1996

Effect of galactose on alpha-crystallin.

Chromatographic separation of alpha-crystallin incubated with [3H]-labelled galactose showed the radioactivity to be concentrated in the low molecular mass subunits (20 and 40 kDa). The effect of glycation on the structural organization of alpha-crystallin was evaluated by FPLC analysis of native (pH 6.8 and 8.2) and glycated protein in dissociating conditions. Results suggest that the glycation acts on the protein surface by altering its charge distribution.

Animals↗
PubMed1993

Tetanus toxin inhibits depolarization-induced [3H]serotonin release from rat brain cortex synaptosomes.

The effects of tetanus toxin on depolarization-induced [3H]serotonin release from superfused rat brain cortex synaptosomes was investigated. Two hours' preincubation of the synaptosomes with tetanus toxin resulted in a concentration-dependent decrease of K(+)-stimulated release, with an IC50 of about 30 nM (4.5 micrograms/ml); this inhibitory effect was blocked by a previous incubation of the tetanus toxin with antitoxin serum. Tetanus toxin had no effect on reserpine-induced release, a model of Ca(2+)-independent release. These results indicate that tetanus toxin is able to alter the exocytotic machinery of serotoninergic terminals, in agreement with results obtained with other neurotransmitters. They also indicate that serotoninergic terminals possess the receptor for tetanus toxin. These findings are in line with in vivo observations suggesting a role for serotoninergic system in tetanus intoxication.

Animals↗
PubMed1993

Covalent modification of synapsin I by a tetanus toxin-activated transglutaminase.

The synapsins are neuronal phosphoproteins that bind to small synaptic vesicles and to actin filaments and are believed to play a regulatory role in neurotransmitter release. Here we show that synapsin I is covalently modified with remarkable affinity and selectivity by the enzyme transglutaminase. Transglutaminase catalyzes the formation of covalent bonds between protein glutamine residues and primary amines and has been found recently to be potently activated by tetanus toxin, a dichain clostridial protein that selectively blocks neurotransmitter secretion. We also report the presence of two species of immunoreactive transglutaminases in nerve endings, one cytosolic and one located on synaptic vesicles; they are potently activated by tetanus toxin and, when activated, covalently modify synaptic vesicle-bound synapsin I. These results suggest a role for transglutaminase in the control of neurotransmitter secretion and provide evidence for synapsin I being a molecular target of tetanus toxin.

Animals↗
PubMed1993

Divergent evolution may link human immunodeficiency virus GP41 to human CD4.

A local sequence similarity of HIV envelope proteins (gp120 and gp41) to immunoglobulins suggests that a mimicry phenomenon may form the basis of the HIV-cell membrane interaction and of HIV-induced autoimmune reaction. We explored the hypothesis of any deeper relationship between HIV env proteins and immunoglobulin family members. An overall DNA sequence similarity between gp41 coding region of env gene and the HIV-receptor CD4 gene was observed and a 14-base-long oligonucleotide, almost unique in the GenBank, was found in gp41 and CD4 genes. The alignment of env gene to CD4 gene and to 84 different sequences showed a significantly higher homology score and a nonrandom similarity in the CD4-env alignment. A significant similarity was also found between the env protein and the sequence encoded by an alternate reading frame of CD4 gene. Our observations suggest that gp41 coding region might have a different origin than the gp120 coding region of the env gene, and that a divergent evolution might link gp41 to CD4 or immunoglobulin family members. In this study the analysis of alternate-reading-frame products is also proposed as a novel approach to investigate evolutionary links and structure-function relationships.

Amino Acid Sequence↗
PubMed1993

The transglutaminase hypothesis for the action of tetanus toxin.

Tetanus toxin potently and almost irreversibly inhibits the release of neurotransmitters from nerve terminals. The toxin binds to and activates transglutaminase, a Ca(2+)-dependent enzyme that can form stable crosslinks between substrate proteins. Transglutaminase is present in nerve terminals and recognizes synapsin I, an abundant synaptic vesicle phosphoprotein involved in neurotransmission, as an excellent substrate. The neuroparalytic action of tetanus toxin might be due, at least in part, to the stimulation of synaptic transglutaminase and the consequent crosslinking of synapsin I.

Animals↗
PubMed1992

Tetanus toxin potently stimulates tissue transglutaminase. A possible mechanism of neurotoxicity.

The observation that tetanus toxin (TT) contains two sequences that show homology to known transglutaminase (TGase) substrate sites suggested that the toxin and TGase might interact. This prediction was confirmed by two pieces of evidence. First, TT potently stimulated the enzymatic activity of TGase. The effect was maximal at physiological (micromolar) concentrations of the endogenous TGase regulators calcium and GTP. Second, TT and TGase displayed marked variations of their intrinsic fluorescence properties when they were coincubated, indicating the occurrence of binding between them. TT-TGase binding and TGase activation occurred at similar concentrations of TT and are probably causally related. The activation of TGase, an enzyme present in nerve endings that, when activated, can irreversibly cross-link cellular proteins, might mediate the neurotoxic action of TT.

Amino Acid Sequence↗