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G Gaudino

Publications and source records attributed to G Gaudino.

At least 37 records · Page 2Linked to original sources
PubMed1992

Hepatocyte growth factor is a potent angiogenic factor which stimulates endothelial cell motility and growth.

Hepatocyte Growth Factor (HGF, also known as Scatter Factor) is a powerful mitogen or motility factor in different cells, acting through the tyrosine kinase receptor encoded by the MET protooncogene. Endothelial cells express the MET gene and expose at the cell surface the mature protein (p190MET) made of a 50 kD (alpha) subunit disulfide linked to a 145-kD (beta) subunit. HGF binding to endothelial cells identifies two sites with different affinities. The higher affinity binding site (Kd = 0.35 nM) corresponds to the p190MET receptor. Sub-nanomolar concentrations of HGF, but not of a recombinant inactive precursor, stimulate the receptor kinase activity, cell proliferation and motility. HGF induces repairs of a wound in endothelial cell monolayer. HGF stimulates the scatter of endothelial cells grown on three-dimensional collagen gels, inducing an elongated phenotype. In the rabbit cornea, highly purified HGF promotes neovascularization at sub-nanomolar concentrations. HGF lacks activities related to hemostasis-thrombosis, inflammation and endothelial cells accessory functions. These data show that HGF is an in vivo potent angiogenic factor and in vitro induces endothelial cells to proliferate and migrate.

Animals↗
PubMed1991

Scatter factor and hepatocyte growth factor are indistinguishable ligands for the MET receptor.

Scatter Factor (SF) is a fibroblast-secreted protein which promotes motility and matrix invasion of epithelial cells. Hepatocyte Growth Factor (HGF) is a powerful mitogen for hepatocytes and other epithelial tissues. SF and HGF, purified according to their respective biological activities, were interchangeable and equally effective in assays for cell growth, motility and invasion. Both bound with identical affinities to the same sites in target cells. The receptor for SF and HGF was identified as the product of the MET oncogene by: (i) ligand binding and coprecipitation in immunocomplexes; (ii) chemical crosslinking to the Met beta subunit; (iii) transfer of binding activity in insect cells by a baculovirus carrying the MET cDNA; (iv) ligand-induced tyrosine phosphorylation of the Met beta subunit. SF and HGF cDNA clones from human fibroblasts, placenta and liver had virtually identical sequences. We conclude that the same molecule (SF/HGF) acts as a growth or motility factor through a single receptor in different target cells.

Animals↗
PubMed1991

Ligand-independent tyrosine phosphorylation of the receptor encoded by the c-neu oncogene.

Incubation of Swiss 3T3 murine fibroblasts at low temperatures induces phosphorylation on tyrosine of a transmembrane protein of 175 kDa. This phenomenon is time and temperature dependent and reaches a maximum after 2 hr at 4 degrees C. The 175 kDa protein phosphorylated in vivo at low temperatures can be immunoprecipitated by phosphotyrosine antibodies and displays auto-kinase activity in vitro in the presence of radiolabelled ATP. This molecule was found to react with anti-peptide antibodies directed against the product of the HER2/neu proto-oncogene only when immunoprecipitated with phosphotyrosine antibodies from cold-stimulated cells. Activation of protein kinase-C by treatment of the cells with phorbol esters, bombesin or PDGF inhibits the effect of the exposure to low temperatures. Phosphorylation of p175 is not induced by treatment of the cells with the phosphatases inhibitor sodium orthovanadate. These results suggest that, at low temperatures, the tyrosine kinase associated with the putative receptor encoded by c-neu is activated by physico-chemical modifications of the plasma membrane.

3T3 Cells↗
PubMed1991

Phosphotyrosine antibodies as probes for activated oncogene products endowed with tyrosine kinase activity.

The usefulness of phosphotyrosine antibodies for the detection of physiologically regulated or deregulated kinases is shown in this paper. This rather rare enzymatic activity is shared by receptors for some polypeptide growth factors and by the products of class 1 oncogenes. The antibodies are able to detect proteins phosphorylated on tyrosine in fibroblasts stimulated with growth factors, such as EGF and PDGF. The major phosphorylated protein species are the receptors themselves, which undergo phosphorylation only following the addition of the exogenous factor and only transiently. Phosphotyrosine antibodies were able to detect the products of the retroviral class 1 oncogenes, which are endowed of deregulated tyrosine kinase activity. In fact, in these cases a constitutive phosphorylation of the relevant proteins was observed, which occurred continuously and independently of the presence or lack of the growth factor. A tyrosine kinase constitutively activated in human gastric carcinoma cells was detected by P-Tyr antibodies. This molecule has been characterized at molecular level and the mechanisms responsible for its enzymatic activation has been investigated. The question of whether the tyrosine kinase identified is responsible for the induction and the maintenance of the transformed phenotype in gastric carcinomas remains to be answered. It is reasonable to suggest that this might be the case by analogy with other known pathologies, such as class 1 oncogenes activated by transduction by retroviruses, abnormal expression of EGF receptors or deregulated activity of c-abl encoded proteins in CML and ALL. Thus, the search for deregulated kinases by means of phosphotyrosine antibodies seems to be useful for identifying new activated oncogenes in clinical oncology.

Animals↗
PubMed1991

Hepatocyte growth factor (HGF) stimulates the tyrosine kinase activity of the receptor encoded by the proto-oncogene c-MET.

The human proto-oncogene c-MET encodes a heterodimeric 190 kDa transmembrane protein (p190c-met) with structural features of a tyrosine kinase receptor. The ligand for this putative receptor has not yet been identified. By Northern blot hybridization we found that, among a restricted number of human tissues, c-MET is highly expressed in the liver. This prompted us to test the hypothesis of a functional interaction between the c-MET receptor and Hepatocyte Growth Factor (HGF), a heparin-binding polypeptide consisting of heavy and light chains of 65 and 35 kDa. Nanomolar concentrations of highly purified HGF added to GTL-16 cells, which overexpress the c-MET receptor, enhanced the phosphorylation on tyrosine of the p190c-met kinase. Addition of other known growth factors or serum was ineffective. The kinase activity of the c-MET receptor was also stimulated by HGF in an in vitro assay, after detergent solubilization and partial purification of p190c-met. Moreover, elution of immunoprecipitates obtained with anti-MET antibodies from GTL-16 cell lysates yielded an HGF-responsive kinase activity. These results suggest that HGF, or a growth factor structurally related to HGF, is a candidate ligand for the receptor encoded by c-MET.

Adrenal Glands↗
PubMed1990

Tyrosine kinase and control of cell proliferation.

The usefulness of phosphotyrosine antibodies for the detection of physiologically regulated or deregulated tyrosine kinases is discussed in this report. This rather rare enzymatic activity is shared by receptors for some polypeptide growth factors and by the products of Class 1 oncogenes. The antibodies are able to detect proteins phosphorylated on tyrosine in fibroblasts stimulated with growth factors such as EGF and PDGF. The major phosphorylated protein species are the receptors themselves, which undergo phosphorylation only after the addition of the exogenous factor and only transiently. Phosphotyrosine antibodies were able to detect the products of the retroviral Class 1 oncogenes, which are endowed with deregulated tyrosine kinase activity. In fact, in these cases a constitutive phosphorylation of the relevant proteins was observed, which occurred continuously and independently of the presence or lack of exogenous ligands. A tyrosine kinase constitutively activated in human gastric carcinoma cells was detected by P-Tyr antibodies. This molecule has been characterized at the molecular level, and the mechanisms responsible for its enzymatic activation have been investigated. The question of whether the tyrosine kinase identified is responsible for the induction and the maintenance of the transformed phenotype in gastric carcinomas remains to be answered. It is reasonable to suggest that this might be the case by analogy with other situations such as Class 1 oncogenes activated by transduction by retroviruses, abnormal expression of EGF receptors, or deregulated activity of c-abl-encoded proteins in chronic myelogenous leukemia and acute lymphoblastic leukemia. Thus, the search for deregulated kinases by means of phosphotyrosine antibodies seems to be useful for identifying new activated oncogenes in clinical oncology.

Carcinoma↗
PubMed1990

Bombesin receptor antagonists [corrected]. 1. Analogues with deleted, inverted or substituted amino acid residues.

Bombesin is a 14-residue peptide hormone which serves a variety of biological functions, including cell growth control in Swiss 3T3 cultured fibroblasts. It has been also involved in an autocrine system regulating the growth of small cell lung carcinoma. A series of bombesin analogues were developed by amino acid deletion, inversion or substitution in the carboxy-terminal region, identified by the target cell receptor. Their properties were examined for: i) competitive binding assays; ii) mitogenic induction and inhibition assays; iii) effects on early cellular events (inositol phosphates production and protein tyrosine phosphorylation). Inversion of the Trp residue, or deletion of the C-terminal tripeptide amide, induced complete loss of receptor affinity and biological effects. Deletion of the His residue, or its derivatization as His (Dnp) in conjunction with Met deletion or modification, gave rise to compounds with reduced receptor affinity and weak antagonistic properties. Other modifications in the C-terminal tripeptide affected the potency but not the biological profile of the parent peptide. These results indicate the basis for the eventual design of improved, specific bombesin receptor antagonists and stimulate further studies on their possible application in the therapy of human small cell lung cancer.

Amino Acid Sequence↗
PubMed1989

Organization of human class I homeobox genes.

We report the genomic organization of 20 human class I homeoboxes and the predicted primary sequence of the encoded homeodomains. These homeoboxes are clustered in four complex HOX loci on chromosomes 2, 7, 12, and 17. The homeoboxes of one HOX locus can be aligned to the homeoboxes of the other HOX loci so that corresponding homeodomains in all loci can share the maximal peptide sequence identity. This correspondence of individual homeoboxes in different chromosomal loci suggests the hypothesis of large-scale duplications of a single complex locus. The existence of an ancestral complex locus might have predated the divergence of vertebrates and invertebrates.

Amino Acid Sequence↗
PubMed1988

Activation of the protein-tyrosine kinase associated with the bombesin receptor complex in small cell lung carcinomas.

It has been hypothesized that bombesin-like peptides produced by small cell lung carcinomas may sustain deregulated proliferation through an autocrine mechanism. We have shown that the neuropeptide bombesin leads to the activation of a protein-tyrosine kinase that phosphorylates a 115-kDa protein (p115) associated with the bombesin receptor complex in mouse Swiss 3T3 fibroblasts. We now report that phosphotyrosine antibodies recognize a 115-kDa protein, phosphorylated on tyrosine, in four human small cell lung carcinoma cell lines producing bombesin but not in a nonproducer "variant" line. p115 from detergent-treated small cell lung carcinoma cells binds to bombesin-Sepharose and can be phosphorylated on tyrosine in the presence of radiolabeled ATP and Mn2+. As for the p115 immunoprecipitated from mouse fibroblast, the small cell lung carcinoma p115 can be phosphorylated in an immunocomplex kinase assay. However, the latter does not require the presence of exogenous bombesin for activity. Binding data, obtained by using radiolabeled ligand, suggest receptor occupancy in the cell lines producing bombesin. These observations are consistent with the hypothesis that proliferation in some human small cell lung carcinoma lines is under autocrine control, regulated through activation of bombesin receptors.

Animals↗
PubMed1988

A tyrosine protein kinase activated by bombesin in normal fibroblasts and small cell carcinomas.

In Swiss 3T3 fibroblasts, antibodies which recognize a phosphotyrosine residue (P-Tyr antibodies) identify a 115-kDa cell surface protein (p115) that becomes phosphorylated on tyrosine as a response to bombesin stimulation of quiescent cells. The extent of phosphorylation is dose-dependent and correlates with the mitogenic effect induced by bombesin, measured by [3H]thymidine incorporation. Tyrosine phosphorylation of p115 is detectable minutes after addition of bombesin and precedes the activation of c-fos and c-myc gene transcription. Immunocomplexes of phosphorylated p115 with P-Tyr antibodies bind 125I-labeled [Tyr4]bombesin in a specific and saturable manner and display an associated tyrosine protein kinase activity enhanced by bombesin. P-Tyr antibodies also recognize a protein of 115 kDa, phosphorylated at tyrosine, in four human SCLC lines producing bombesin but not in a non-producer "variant" line. Phosphorylation of SCLC p115 does not require the addition of exogenous bombesin. As in the case of the p115 immunoprecipitated from mouse fibroblasts, the SCLC p115 is phosphorylated in an immunocomplex kinase assay. These observations are in agreement with the hypothesis of autocrine activation of bombesin receptors in human small cell lung carcinoma cells.

Animals↗
PubMed1986

An immunoreactive peptide in milk contains bombesin-like bioactivity.

Parallel in vitro bioassays using rat uterus and guinea pig large intestine tissues specific for the bombesin family of peptides, demonstrated that the bombesin-like peptides present in bovine milk can produce a dose-related response similar to bombesin and litorin. The bioactivity of this type of milk peptide appeared to be approximately 20-50% as active as the amphibian peptides. These data support the proposal that a bombesin immunoreactive peptide in milk contains bombesin bioactivity.

Animals↗
PubMed1986

The organization of serotonin-immunoreactive neuronal systems in the brain of the crested newt, Triturus cristatus carnifex Laur.

The distribution of serotonin (5-HT) immunoreactive structures has been investigated in the brain of the crested newt by means of indirect immunofluorescence, and unlabeled antibody peroxidase-antiperoxidase-complex (PAP) or biotin-avidin-system (BAS) techniques. In the newt, the bulk of the serotoninergic system extends from the raphe region of the medulla oblongata, through the isthmus, toward the mesencephalic tegmentum, and is characterized by pyriform neurons mainly located in a subependymal position, close to the midline. Also in the caudal hypothalamus, in addition to some 5-HT-positive adenohypophysial cells, many immunoreactive CSF-contacting neurons are found lining the paraventricular organ and the nucleus infundibularis dorsalis. A rich serotoninergic innervation was observed in the preoptic area and in the habenular complex. Concerning the telencephalon, immunopositive nerve fibers are encountered in the dorsal pallium, primordium hippocampi, striatum and olfactory bulbs. The general organization of serotoninergic systems in the newt brain exhibit close similarities to that described in higher vertebrates.

Animals↗
PubMed1986

Receptor for bombesin with associated tyrosine kinase activity.

The neuropeptide bombesin is known for its potent mitogenic activity on murine 3T3 fibroblasts and other cells. Recently it has been implicated in the pathogenesis of small cell lung carcinoma, in which it acts through an autocrine loop of growth stimulation. Phosphotyrosine (P-Tyr) antibodies have been successfully used to recognize the autophosphorylated receptors for known growth factors. In Swiss 3T3 fibroblasts, phosphotyrosine antibodies identified a 115,000-Mr cell surface protein (p115) that became phosphorylated on tyrosine as a specific response to bombesin stimulation of quiescent cells. The extent of phosphorylation was dose dependent and correlated with the mitogenic effect induced by bombesin, measured by [3H]thymidine incorporation. Tyrosine phosphorylation of p115 was detectable minutes after the addition of bombesin, and its time course paralleled that described for the binding of bombesin to its receptor. Immunocomplexes of phosphorylated p115 and phosphotyrosine antibodies bound 125I-labeled [Tyr4]bombesin in a specific and saturable manner and displayed an associated tyrosine kinase activity enhanced by bombesin. Furthermore, the 125I-labeled bombesin analog gastrin-releasing peptide, bound to intact live cells, was coprecipitated with p115. These data strongly suggest that p115 participates in the structure and function of the surface receptor for bombesin, a new member of the family of growth factor receptors with associated tyrosine kinase activity.

Amino Acids↗
PubMed1985

[Primary empty sella. Clinico-radiologic considerations in 18 cases].

18 patients with "primary empty sella" were reviewed for this study. In 3 of them the sellar enlargement was discovered occasionally by performing skull radiographs for other reasons. The galattorrhea-dismenorrhea or amenorrhea syndrome and obesity were the most common clinical features. Endocrinological tests were normal in ten patients and abnormal in eight. Slight elevation of serum PRL was the most common record. 12 patients had enlarging of the sella turcica; in 4, only the floor was asymmetric and in 2 the sella was quite normal. In 5 patients C.T. without intra-thecal contrast was sufficient to discover the E.S. In 13 patients we performed C.T. cysternography by injecting in the lumbar subarachnoid space 8-10 ml of Iopamidolo 200. This is an excellent and safe technique to perform C.T. cysternography.

Adolescent↗