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D Marchetti

Publications and source records attributed to D Marchetti.

At least 37 records · Page 2Linked to original sources
PubMed1994

Abnormal expression of perlecan proteoglycan in metastatic melanomas.

Abnormal expression of proteoglycans has been implicated in cancer and metastasis primarily because these macromolecules are involved in the control of cell growth and matrix assembly. In this report, we have investigated the expression and immunolocalization of perlecan, a major heparan sulfate proteoglycan of basement membranes and pericellular matrices, in human metastatic melanomas. Twenty-six of the 27 tumor samples showed a significant increase (up to 15-fold) in the perlecan mRNA levels when compared with normal tissue. This change correlated with a vast deposition of perlecan protein core in the pericellular matrix of metastatic melanomas. Furthermore, we have established a relationship between perlecan expression in clonal melanoma cells (70W) stimulated with neurotrophins and their increased invasiveness. Interestingly, perlecan mRNA levels were up-regulated within 10 min of neurotrophin stimulation, indicating that perlecan is an early response gene. This upregulation also occurred prior to heparanase production, suggesting that perlecan expression and its regulation might play a pivotal role in the initial onset of invasion.

Blotting, Northern↗
PubMed1994

Malignant melanoma metastasis to brain: role of degradative enzymes and responses to paracrine growth factors.

Mouse and human melanoma cells metastatic to the brain express degradative enzyme activities that are used for invasion of brain basement membrane and parenchyma. Compared to poorly metastatic or lung- or ovary-metastatic murine melanoma lines, the brain-metastatic sublines secreted higher levels of a variety of degradative enzymes. Brain-metastatic murine and human melanoma cells also degraded subendothelial basement membrane and reconstituted basement membrane at rates higher than other metastatic melanoma cells. In some cases these degradative activities in mouse and human melanoma cells can be induced by paracrine factors known to be present in the brain parenchyma, such as nerve growth factor (NGF). NGF stimulates the expression of degradative enzymes, such as the endo-beta-glucuronidase heparanase, that are important in basement membrane penetration but this factor does not stimulate melanoma cell growth. The growth of brain-metastasizing melanoma cells appears to be stimulated by other paracrine growth factors, such as paracrine transferrin. Melanoma cells metastatic to brain express higher numbers of transferrin receptors and respond and proliferate at lower concentrations of transferrin than do melanoma cells metastatic to other sites or poorly metastatic melanoma cells. The results suggest that degradation and invasion of brain basement membrane and responses to paracrine neurotrophins and paracrine transferrins are important properties in brain metastasis of murine and human malignant melanoma cells.

Animals↗
PubMed1994

Tumor metastasis to brain: role of endothelial cells, neurotrophins, and paracrine growth factors.

An important clinical endpoint in patients with cancer is formation of metastases in the brain. Understanding this phenomenon is important in several types of malignancies, including melanoma, lung and breast cancers. Metastatic tumor cells use specific adhesion molecules to home to brain, and there they must attach to microvessel endothelial cells and respond to brain endothelial cell-derived motility factors and brain invasion factors to invade the CNS. Neurotrophins are important invasion factors in this process, and the ability to invade into the brain may well depend on metastatic cell responses to neurotrophins and production of basement membrane-degradative enzymes capable of locally destroying the blood-brain barrier. Brain-metastatic human melanoma cells express low-affinity p75 receptor for neurotrophins such as nerve growth factor, but they do not express the high-affinity-type receptors for nerve growth factor encoded by the protooncogene trkA. Tumor cells can proliferate in the CNS in response to local paracrine growth factors and inhibitors, but their growth also depends on their producing and responding to autocrine growth factors. A major organ-derived (paracrine) growth factor has been isolated that differentially stimulates the growth of cells metastatic to the brain. Characterization of this mitogen demonstrated that it is a transferrin-like glycoprotein; cells that are metastatic to brain express greater numbers of transferrin receptors on their surfaces than cells that are poorly metastatic or metastatic to other sites. Transferrin-like factors are expressed in fetal brain and could represent the transferrin-like factors that stimulate growth of brain-metastatic melanoma and breast cancer cells. These and other factors are probably important in determining whether metastatic cells can successfully invade, colonize, and grow in the CNS.

Animals↗
PubMed1993

Nerve growth factor effects on human and mouse melanoma cell invasion and heparanase production.

The role of growth factor networks in regulating the progression of human melanocytes towards tumorigenicity and ultimately the malignant phenotype is poorly understood. In particular, the autocrine and paracrine influences that modulate cellular invasion and extracellular matrix degradative enzymes of melanoma cells remain undefined at the molecular level. We report here that nerve growth factor (NGF) can modify some metastasis-associated cellular properties of human and mouse melanoma cells. Treatment of early-passage human metastatic melanoma cells (MeWo) or their variants (3S5, 70W) with biologically active 2.5S NGF resulted in (a) delayed density-dependent inhibition of melanoma cell growth; (b) increased in vitro invasion through a reconstituted basement membrane; and (c) time- and dose-dependent induction of heparanase, a heparan-sulfate-specific endo-beta-D-glucuronidase associated with human melanoma metastasis. These effects of NGF were most marked in the 70W brain-colonizing cells (70W > MeWo > 3S5). The NGF enhancement of heparanase secretion was not species-specific, since it was also observed in murine B16 melanoma cells; the highest NGF stimulation of heparanase was found in brain-colonizing murine B16-B15b variant (B16-B15b > B16-BL6, B16-F10, B16-F1). NGF also increased the invasive capacity of the human 70W and murine B16-B15b sublines in a chemoinvasion assay performed with filters coated with purified heparan sulfate proteoglycan (HSPG). The enhancement of chemotactic response and heparanase production was detected at NGF concentrations sufficient to fully saturate both low- and high-affinity NGF receptors (NGFR), the neurotrophin receptor (p75) and the trkA gene product, respectively. The results suggest that, in addition to the effects of NGF on cellular development and differentiation within the peripheral and central nervous systems, NGF can exert changes in the invasive properties of neuroectoderm-derived melanoma cells.

Animals↗
PubMed1993

Mediation of NGF-stimulated extracellular matrix invasion by the human melanoma low-affinity p75 neurotrophin receptor: melanoma p75 functions independently of trkA.

Although overexpression of the low-affinity p75 neurotrophin receptor (p75NTR) is frequently associated with advanced stages of human melanoma progression, the functional significance of this finding is unknown. We examined whether the degree of cell surface expression of p75NTR in human melanoma cell variants determines their extent of invasion stimulated by nerve growth factor (NGF). Treatment of MeWo melanoma cells or a metastatic spontaneous wheat germ agglutinin-resistant variant subline (70W) of MeWo cells with 2.5S NGF resulted in a dose-dependent enhancement of invasion through a reconstituted basement membrane. This effect was most pronounced with the 70W subline that exhibits brain-metastasizing potential in nude mice but was not found with a poorly metastatic MeWo variant subline (3S5). The expression of p75NTR as determined by Northern blotting and immunoprecipitation analysis of 125I-labeled cell surface proteins correlated with NGF-stimulated invasion. The MeWo melanoma sublines used in this study did not express p140proto-trkA mRNA or any p140proto-trkA variant transcripts including p70trkA as determined by Northern analysis and RT-PCR analysis. Thus, these melanoma cells would not be expected to form functional p75-p140 heterodimers or p140-p140 homodimers capable of transducing an NGF-generated signal to p140proto-trkA cytoplasmic substrates. These cells did express authentic p145trkC transcripts. However, NGF did not catalytically activate p145trkC receptors via increased tyrosine phosphorylation as would be expected if p145trkC participated in the signaling established by NGF. Furthermore, a NGF-stimulated purine-analogue-sensitive kinase activity was found to coimmunoprecipitate with p75NTR. This p75NTR-associated kinase may coordinate initial signaling events evoked by p75NTR ligand interaction. Addition of 2.5S NGF, at concentrations that should saturate cell surface p75NTR, to matrix-adherent cultures of human MeWo and 70W but not 3S5 melanoma cells suppressed the expression of 92-kDa type IV collagenase and stimulated the production of 72-kDa type IV collagenase in its fully active 68-kDa form. In the absence of p140proto-trkA, the matrix-dependent effects of NGF on metalloproteinase expression of brain-metastatic 70W melanoma cells suggest a signaling role for the low-affinity melanoma p75NTR receptor and its associated purine-analogue-sensitive kinase in signaling enhanced matrix penetration of NGF-rich stromal microenvironments such as the brain.

Base Sequence↗
PubMed1991

Ontogeny of high- and low-affinity nerve growth factor receptors in the lumbar spinal cord of the developing chick embryo.

The binding of 125I-labeled nerve growth factor-beta (NGF) to soluble extracts of intact or dissociated embryonic chick lumbar cords was used to investigate the kinetic properties and to quantify the levels of NGF receptors (NGFRs) in the developing chick between Embryonic Day 6 (E6) and E10. Both high-affinity (type I; Kd = 7.4 x 10(-11) M) and low-affinity (type II; Kd = 2.4 x 10(-9) M) NGFRs were detected by Scatchard analysis of 125I-NGF binding to E6 spinal cord extracts. A total of 4 x 10(9) type I and 5 x 10(10) type II receptors/cord were found in extracts of E6 cords. As development progressed there was a decline of both types of NGFRs; however, the decline of type I receptors occurred more rapidly than that of type II. Between E6 and E8 greater than 90% of the type I but only 25% of the type II receptors were lost. These relative rates of loss were maintained over the next week of development, with type I receptors no longer detectable by E12, and type II receptors reduced to 0.025% of their E6 numbers by E15. Analyses of NGFR levels in subpopulations of E6 and E8 lumbar cord cells, prepared by metrizamide density gradient centrifugation, showed that during this period there is an enrichment of both types of NGFRs in the motoneuron-containing subpopulation, relative to other cell populations. The loss of NGFRs does not appear to be influenced by those peripheral-trophic interactions which control other aspects of motoneuron development: curarization of the embryos between E6 and E9 increased motoneuron number in E10 embryos by 30%, but did not significantly affect the loss of NGFRs. These results provide the first quantitative evidence that type I and type II NGFRs are differentially regulated in the spinal cord during embryonic development and raise the possibility that distinct cellular mechanisms may govern their expression.

Animals↗
PubMed1990

Characterization of nerve growth factor binding to embryonic rat spinal cord neurons.

The binding of iodinated beta-nerve growth factor, [125I]-NGF, to embryonic (E16) rat spinal cord cells, was investigated to characterize the binding properties and cellular distribution of nerve growth factor receptors. Spinal cord cells prepared without trypsin yielded two classes of NGF binding sites with Kd's of 3 x 10(-11) M and 4 x 10(-9) M. Fractionation of the cells by discontinuous gradients composed of 8%, 12%, and 17% metrizamide was used to separate motoneurons from other cell types. The motoneuron enriched fraction (8% metrizamide) contained approximately 10% of the cells and 64% of the choline acetyltransferase (ChAT) activity. In contrast, the 12% metrizamide fraction contained most (51%) of the cells and 36% of the ChAT activity, while the 17% metrizamide fraction contained the remainder of the cells and negligible amounts of ChAT activity. Characterization of [125I]-NGF binding to each metrizamide fraction showed that the motoneuron-enriched fraction exhibited both high and low affinity binding sites, while the other metrizamide fractions exhibited only the low affinity binding sites. These findings indicate that although low affinity NGF receptors appear to be relatively evenly distributed amongst embryonic rat spinal cord cells, high affinity NGF receptors are found primarily on motoneurons.

Animals↗
PubMed1990

Rescue of motoneurons from cell death by a purified skeletal muscle polypeptide: effects of the ChAT development factor, CDF.

Rat skeletal muscle contains a 22 kd polypeptide that increases the level of choline acetyltransferase (ChAT) activity in cultures of embryonic rat spinal cord neurons and has been purified to homogeneity. The application of this factor, ChAT development factor or CDF, to developing chick embryos during the period of naturally occurring motoneuron cell death significantly increased the survival of motoneurons but did not affect the survival of dorsal root ganglion neurons or sympathetic preganglionic neurons (column of Terni). These results provide the first demonstration that an isolated, skeletal muscle-derived molecule can selectively enhance the survival of motoneurons in vivo and suggest that CDF may function in vivo to regulate the survival and development of motoneurons.

Animals↗
PubMed1990

Nerve growth factor activity and aging in CNS.

This is a discussion of aging in CNS and the influence of the nerve growth factor (NGF) protein. The paper considers neuronal plasticity and neuronotrophic substances, neuronal cell death and the nerve growth factor protein, including its effects, receptors, and model systems for the study of CNS aging.

Aging↗
PubMed1989

[Early postoperative enteral nutrition and total parenteral nutrition. Technics compared in patients undergoing radical surgery for carcinoma of the large intestine. Preliminary report].

A preliminary study has been carried out on 10 patients suffering from cancer of the colon and rectum, all subjected to radical surgery. The clinical effectiveness of two alternative techniques of postoperative artificial nutrition, EPEN and TPN, are compared. The results confirm that enteral nutrition is technically applicable and well tolerated in the immediate post-operative period in these patients; it also presents a lower cost, it is more manageable and present a lower incidence of complications compared to TPN while offering similar metabolic and nutritional results.

Adult↗
PubMed1989

Flow cytometric analysis of glucocorticoid receptor using monoclonal antibody and fluoresceinated ligand probes.

Conditions were established for single cell analysis of glucocorticoid receptor (GR) content by flow cytometry using several clones of a human leukemic cell line (CCRF-CEM). These included CEM-7A, 7R, C1, and ICR 27 Tk.3 cells which were examined both by standard [3H]dexamethasone radiometric binding and by two independent flow cytometry assays. The latter involved either mouse monoclonal antibody against GR (GR-MoAb) or fluoresceinated cortisol ligand probes. For CEM-7A, 7R, and C1 cells, there was a correlation between GR-MoAb and radiometrically defined GR values. However, clone ICR-27 Tk.3 with low [3H]dexamethasone binding exhibited the highest GR-MoAb fluorescence. The fluoresceinated cortisol assay correlated with dexamethasone binding values in all four clones. Thus, GR-MoAb identifies the total immunologically reactive GR present, while the fluoresceinated cortisol assay quantifies only the functionally intact GR in terms of its initial binding. Their combined use may reveal the cellular heterogeneity of GR expression and function also in human tumor samples, to which they have been successfully applied. When coupled with DNA counterstaining, GR expression can be related directly to frequently DNA-aneuploid tumor cells and cell cycle distribution.

Antibodies, Monoclonal↗
PubMed1989

Expression of nerve growth factor receptors by human peripheral blood mononuclear cells.

In the rat, nerve growth factor (NGF) has been shown to affect immune reactivity by binding to cell surface receptors on a subpopulation of splenic mononuclear cells. This binding occurs in a specific and saturable fashion to what appear to be low-affinity (type II) NGF receptors (NGFR). Immunofluorescence studies here showed that NGFR are also present on a proportion of human peripheral blood mononuclear cells (PBMC). Equilibrium binding studies demonstrated that the binding of NGF to its receptors on PBMC occurs with a single equilibrium binding constant (mean) of 2.11 X 10(-9) M. The number of receptors per cell was determined to be approximately 6.94 X 10(3) receptors/cell. These results would suggest a role for NGF in the regulation of immune function in man, as well as in animals.

Cell Membrane↗
PubMed1988

Effects of gonadal steroids on nerve growth factor receptors in sympathetic and sensory ganglia of neonatal rats.

The numbers of neurons in the rat superior cervical sympathetic ganglion (SCG) differ in males and females, with the males having 30% more SCG neurons than females at 60 days of age. This sex difference arises during the early postnatal period, when testosterone administration increases the numbers of neurons and alters the nerve growth factor (NGF) content of the rat SCG. In contrast, there is no gender difference in number of neurons in the L1 dorsal root ganglion. In both males and females, the amount of NGF bound per ganglion increased between postnatal days 5 and 15 (P5 and P15) in both dorsal root ganglia (DRGs) and the SCG. There is also a gender difference in NGF binding: SCGs and DRGs of female rats at both P5 and P15 bind more NGF per ganglion than do those of males. This effect was more marked in DRGs than in the SCG. Treatment of neonatal females with testosterone reduced NGF binding in both SCGs and DRGs to levels comparable to males at P5, and in DRGs at P15. In contrast, treatment of males with testosterone from birth resulted in a 2-3 fold increase of NGF binding in both SCGs and DRGs as compared to controls at P15. At P15, testosterone treatment of females increased NGF binding in the SCG. Males and females had opposing responses to neonatal exposure to estradiol. Treatment with estradiol from birth increased NGF binding in SCGs and DRGs of females, but had no effect on NGF binding of SCGs, and reduced NGF binding in DRGs of males.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Analysis of idiotypic and anti-idiotypic antibodies as models of receptor and ligand.

Antibodies to small bioactive ligands and peptides may mimic the binding characteristics of the natural receptor; in turn, the anti-idiotypic antibodies generated against the binding sites of such anti-ligand antibodies may mimic some aspects of small bioactive ligands and peptides. Among the several levels of investigation of such antibody-receptor networks are (a) the quantitative structure-activity relationships of ligand binding to antibody as compared with natural receptor; (b) the molecular modeling of antibody-receptor binding sites and the genomic basis for such structures; and (c) the characteristics of the molecular mimicry exhibited by "mimetopes" on anti-idiotypic antibodies. To illustrate the analysis encountered at each of these levels, we discuss here antibody and anti-idiotypic systems that are directed to small neuroactive ligands and their receptors.

Amino Acid Sequence↗
PubMed1987

Binding constants of soluble NGF-receptors in rat oligodendrocytes and astrocytes in culture.

The neuronotrophic factor NGF binds to peripheral neurons of the dorsal root ganglion and the sympathetic nervous system. NGF binds to a cell surface receptor, NGFR, on these cells and displays Kd's of 10(-9) and 10(-11)M. NGF receptors have also been reported for basal forebrain magnocellular neurons. In addition, NGF specifically binds to NGFR on Schwann cells although the biological significance of this binding is not known. Here we report that NGF binds in a saturable and specific fashion to receptors on cultured isolated populations of rat astrocytes but not to oligodendrocytes. The binding to astrocytes in culture displayed a Kd of 2.7 +/- 1.0 nM with 36,000 receptors per cell.

Animals↗