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Biomedical subjects

A Facchiano

Publications and source records attributed to A 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

Primary structure of alpha-globin chains from river buffalo (Bubalus bubalis L.) hemoglobins.

Primary structure analysis of the four river buffalo alpha-globin chains showed that haplotypes A and B differ from each other by a substitution at codon 64 that may encode Ala or Asn. The A haplotype encodes two alpha-globin chains, Ialpha1 and IIalpha3, which differ at positions 129 and 131: Ialpha1 has 64 Ala, 129 Phe, 131 Asn; IIalpha3 has 64 Ala, 129 Leu, 131 Ser. The B haplotype encodes two alpha-globin chains, Ialpha2 and IIalpha4, which differ at positions 10 and 11: Ialpha2 has 10 I1e, 11 Gln, 64 Asn; IIalpha4 has 10 Val, 11 Lys, 64 Asn. Apart from the Ala/Asn polymorphism at position 64, amino acid substitutions in allelic and nonallelic alpha-globin chains seem to have arisen by single point mutations. Detection of electrophoretically silent mutations due to neutral amino acid substitutions and their influence on the isoelectric point are discussed. Furthermore, primary structures of river buffalo alpha-globin chains are compared to other species of the Bovidae family to suggest evolutionary events that have characterized the amino acid substitutions of river buffalo hemoglobin.

Amino Acid Sequence↗
PubMed2000

Rational design of biologically active peptides: inhibition of T cell activation through interference with CD4 function.

In our laboratory we generated one synthetic cyclic peptide (Pep4) and tested it in human mitogen stimulation assays (MSA) and mixed lymphocytes reactions (MLR) generating dose-response curves showing a dose-dependent inhibition of MSA up to 80% and MLR up to 98%. MSA and MLR were repeated after pre incubation of the Pep4 with each separate responder cell subset and subsequent reconstitution: these experiments showed inhibition only when the peptide was present in culture. Pep4 showed species specificity since it was ineffective in inhibiting rat MLR. Combination effect analysis with Pep4 and cyclosporine showed a combination index > 1. This rationally designed peptide (Pep4) shows powerful inhibition of human T cell activation and, although the exact mechanism is still undefined, it seems to exert its major action on the T cell surface, interfering with co receptor interaction and disrupting the same activation signal pathway inhibited by cyclosporine A.

Animals↗
PubMed2000

Rational design of biologically active peptides: inhibition of T cell activation through interference with CD4 function. Transplant Int (2000) 13 [Suppl 1]: S306-S310.

In our laboratory we generated one synthetic cyclic peptide (Pep4) and tested it in human mitogen stimulation assays (MSA) and mixed lymphocytes reactions (MLR) generating dose-response curves showing a dose-dependent inhibition of MSA up to 80% and MLR up to 98%. MSA and MLR were repeated after pre incubation of the Pep4 with each separate responder cell subset and subsequent reconstitution: these experiments showed inhibition only when the peptide was present in culture. Pep4 showed species specificity since it was ineffective in inhibiting rat MLR. Combination effect analysis with Pep4 and cyclosporine showed a combination index > 1. This rationally designed peptide (Pep4) shows powerful inhibition of human T cell activation and, although the exact mechanism is still undefined, it seems to exert its major action on the T cell surface, interfering with co receptor interaction and disrupting the same activation signal pathway inhibited by cyclosporine A.

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

Probing the modelled structure of wheatwin1 by controlled proteolysis and sequence analysis of unfractionated digestion mixtures.

We set up a method to get rapid information on the three-dimensional structure of peptide and proteins of known sequence. Both native and alkylated polypeptide is hydrolyzed with a number of proteases at different digestion times and the resulting mixtures are compared by HPLC analysis to establish the differences in the hydrolysis pathways of the folded and unfolded molecule. Then, the unfractionated digestion mixtures of the native polypeptide are submitted to automatic sequence analysis to identify the hydrolysis sites. The sequence of each fragment present in the mixtures is reconstructed and its amount determined by quantitative data of the sequence analyses. We used this approach to determine the amino acid surface accessibility of wheatwin1, a pathogenesis-related protein from wheat, and constructed a predictive three-dimensional model based on the knowledge of the tertiary structure of barwin, a highly homologous protein from barley. The procedure allowed us to quickly identify and quantify the hydrolysis at the susceptible bonds which could be classified as exposed, partially hidden, or inaccessible. The results were useful to evidentiate and discuss concordances and differences between experimental and model predicted accessibilities of amino acid residues. Proteins 1999;36:192-204.

Alkylation↗
PubMed1999

Internal repeats of prion protein and A beta PP, and reciprocal similarity with the amyloid-related proteins.

We are currently interested in a group of proteins associated with the dementias characterized by amyloid deposition in the brain: amyloid beta protein precursor (A beta PP) of Alzheimer's disease (AD) and the abnormal isoform of prion protein (PrP) of spongiform encephalopathies such as kuru, Creutzfeldt-Jacob disease (CJD) and Gerstmann-Straussler-Scheinker disease (GSSD). Here we show that both A beta PP and prion protein (PrP) consist of peculiar internal repeats and share regions of sequence similarity. Further analysis revealed that local homology is also shared with the other proteins involved in specific forms of amyloidosis--i.e., the amyloid related proteins (ARP).

Amino Acid Sequence↗
PubMed1997

The murine Tcl1 oncogene: embryonic and lymphoid cell expression.

In human leukemias and lymphomas nonrandom chromosomal rearrangements cause changes in cell growth and/or survival in such a way as to promote malignancy. The detailed study of the biochemical and genetic pathways altered in human cancer requires the identification or development of models to allow the study and manipulation of cancer gene function. Recently, the breakpoint gene TCL1, involved in chromosome translocations observed mostly in mature T-cell proliferations and chronic lymphocytic leukemias (CLL), was isolated and characterized, and showed to be part of a new gene family of proteins involved in these tumors. The murine Tcl1 gene, is similar in sequence to the murine and human MTCP1 gene also involved in T cell leukemias. The murine Tcl1 gene was shown to reside on mouse chromosome 12 in a region syntenic to human chromosome 14. Furthermore, we show that the murine Tcl1 gene is expressed early in mouse embryonic development and demonstrates expression in fetal hematopoietic organs as well as in immature T and B cells. Characterization of the murine Tcl1 gene will help in developing a mouse model of CLL and would provide the best opportunity to study and decipher the role of TCL1 in malignant transformation.

Amino Acid Sequence↗
PubMed1995

Investigating hypothetical products from noncoding frames (HyPNoFs).

Hypothetical Products from Noncoding Frames (i.e., HyPNoFs) are hypothetical, not-coded proteins, translated from alternate reading frames (i.e., coding + 1 and coding + 2) of cDNAs. HyPNoFs of CD4, PKC, oncostatin, bcl-2 proto-oncogene, tumor suppressor p53, cystic fibrosis transmembrane regulator (CFTR), and tumor necrosis factors alpha and beta were searched as query sequences vs the SWISS-PROT data bank. Homology searchers carried out revealed that hypothetical products (i.e., HyPNoFs) may share high similarity with real protein products actually coded. Sequence similarity of hypothetical products to real proteins is sometimes very high, suggesting common conformational features, according to the Sander and Schneider cutoff value. This finding supports the hypothesis that eukaryotic DNA, currently considered to be monocistronic, might occasionally have polycistronic regions, carrying different protein messages on overlapping frames. As yet, polycistronic genes have been observed in viral genomes only. The presence of polycistronic regions in eukaryotic genes is likely reminiscent of an ancient strategy, rather than a present feature of the genome in eukaryotes. These data suggest that thorough investigation of HyPNoFs is likely to improve our ability to trace genes' evolution and to investigate structure-function relationships of protein and DNA sequences.

Amino Acid Sequence↗
PubMed1994

Characterization and localization of the TCL-1 oncogene product.

The TCL-1 gene maps at chromosome 14q32.1 and is activated in T cell leukemias and lymphomas by either chromosome translocations or inversions that juxtapose the TCL-1 gene to the alpha/delta or the beta locus of the T cell receptor. The open reading frame of the TCL-1 gene, coding for a protein of 114 amino acids, was expressed in bacteria and antisera were raised against it. The antibodies recognized the predicted TCL-1 M(r) 14,000 protein product in cells expressing TCL-1 mRNA. Cell fractionation experiments indicated that the TCL-1 protein is present in the microsomal fraction. These results were confirmed by confocal microscopy. The TCL-1 protein has considerable sequence similarities to the product of the MTCP-1 gene on chromosome Xq28, which is involved in T cell lympho-proliferative diseases. Thus, TCL-1 may represent a member of a novel family of genes involved in lymphoid proliferation and/or survival and in T cell malignancies.

Amino Acid Sequence↗
PubMed1994

Homology of calcitonin with the amyloid-related proteins.

We present evidence for a structural homology between the amino acid sequence of calcitonin (CT)--the fibrillar protein of the amyloid deposits of medullary thyroid cancer--and that of other 12 amyloid-related proteins (ARP). Seven of the 32 residues of CT are conserved in at least five ARP, and five of these seven amino acids belong to the stretch Gly2-Gln14. Gln14 is conserved in all 12 ARP and Cys7 in all eight ARP containing cysteine. The concentration of the homology in the N-terminal half of CT goes along with the knowledge that is the C-terminal region the one more important for the hormonal action of CT. Since an imbalance between synthesis and catabolism of a given ARP is believed to be the general pathogenetic mechanism of amyloidosis, the intratumoral deposition of CT in the form of amyloid fibrils would be due to the overproduction of a protein structurally similar to the ARP.

Amino Acid Sequence↗
PubMed1994

Thyroid hormone binding to isolated human apolipoproteins A-II, C-I, C-II, and C-III: homology in thyroxine binding sites.

Thyroid hormone binding to lipid-free apolipoprotein (apo) A-II, C-I, C-II, and C-III isolated from human plasma was investigated by photoaffinity labeling with [125I]T4 and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Both the monomeric and polymeric forms were specifically labeled. Inhibition by 10 microM unlabeled L-T4 was > or = 50%, suggesting affinity constants in the nM to microM range; the least inhibition was seen with apoA-II. Unlabeled D-T4 and reverse T3 (rT3) gave the same inhibition as unlabeled L-T4. Inhibitors of thyroid hormone binding to plasma proteins showed a different inhibitor potency with each apolipoprotein and a pattern different from that seen with T4 binding globulin (TBG) and transthyretin (TTR). Also in contrast to TBG, where only unsaturated nonesterified fatty acids (NEFA) are effective inhibitors, both unsaturated and saturated NEFA as well as other lipids inhibited T4 labeling. The flavonoid EMD 21388 was ineffective, confirming that it is a selective inhibitor of T4 binding to TTR. T4 binding to the apoCs was confirmed by the quenching of tryptophan fluorescence by unlabeled L-T4. (ApoA-II was not studied since it lacks tryptophan). Since the self-association of apolipoproteins involves interaction between amphipathic alpha-helices, and since the polymeric forms show specific T4 binding properties as in the parent monomer, the T4-binding domain appears to be outside the alpha-helical domain, as previously seen with apoA-I.(ABSTRACT TRUNCATED AT 250 WORDS)

Affinity Labels↗
PubMed1993

Inhibition of interleukin-1 release and activity by the rat seminal vesicle protein SV-IV.

SV-IV, a 9.8-kd protein isolated from rat seminal vesicle secretion, has been shown to have strong non-species-specific immunosuppressive, anti-inflammatory, antithrombotic, and antiphospholipase A2 properties. The present investigation was undertaken to determine the mechanism of action of its immunosuppressive effects. It was found that SV-IV is a potent inhibitor of interleukin-1 (IL-1) release from lipopolysaccharide (LPS)-activated human adherent monocytes and an effective inhibitor of IL-1-induced thymocyte proliferation. The ability of SV-IV to form a noncovalent dimeric association with IL-1 alpha but not with IL-1 beta, its ability to induce a marked decrease of IL-1 binding to its own receptors on the thymocyte surface, and its capacity to bind specifically to the macrophage plasma membrane might play an important role in the molecular mechanism of these inhibitory effects.

Animals↗
PubMed1993

The amino acid sequence of a protein from wheat kernel closely related to proteins involved in the mechanisms of plant defence.

The amino acid sequence of wheatwin1, a monomeric protein of 125 residues isolated from wheat kernel (variety S. Pastore), is reported. Wheatwin1 is highly homologous (95%) to barwin, a protein from barley seed, which was shown to be related to the C-terminal domain of two proteins encoded by the wound-induced genes win1 and win2 in potato and to a protein encoded by the same domain of the hevein gene (hev1) in rubber tree. Similarly to barwin, wheatwin1 contains six cysteine residues all linked in disulfide bridges and the N-terminal residue is pyroglutamate. Moreover, structural studies performed on wheatwin1 and win1 protein by predictive methods demonstrated that these proteins and barwin are closely related in the secondary structure also. The high level of homology found with the product of win1, win2, and hev1 genes strongly suggests that barwin and wheatwin1 play a common role in the mechanism of plant defence.

Amino Acid Sequence↗
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↗