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Biomedical subjects

M Ruggiero

Publications and source records attributed to M Ruggiero.

At least 73 records · Page 4Linked to original sources
PubMed1989

The 18 kDa cytosolic acid phosphatase from bovine live has phosphotyrosine phosphatase activity on the autophosphorylated epidermal growth factor receptor.

In this paper we demonstrate that the cytosolic low-Mr acid phosphatase purified from bovine liver has phosphotyrosine protein phosphatase activity on 32P-autophosphorylated epidermal growth factor (EGF) receptor. This activity was significantly inhibited by orthovanadate and p-hydroxymercuribenzoate; the latter result indicates that free sulfhydryl groups are required for phosphotyrosine phosphatase activity. The enzyme was active in a broad pH range, with maximum activity between pH 5.5 and 7.5. The apparent Km for 32P-EGF receptor dephosphorylation was 4 nM. The enzyme appeared to be specific for phosphotyrosine in that it dephosphorylated the autophosphorylated EGF receptor and L-phosphotyrosine, but not 32P-Ser-casein, L-phosphoserine or L-phosphothreonine. These data suggest that the cytosolic low-Mr acid phosphatase might play a regulatory role in EGF receptor-dependent transmembrane signalling.

Acid Phosphatase↗
PubMed1989

Hematopoietic bone marrow recovery after radiation therapy: MRI evaluation.

Magnetic resonance imaging (MRI) is able to detect the increase of adipocytes in the hematopoietic bone marrow that occurs as a consequence of radiotherapy and is indicative of the loss of myeloid tissue. By monitoring this process, it is also possible to determine the recovery of the bone marrow. The amount of viable hematopoietic tissue plays a fundamental role in determining whether the patient is able to undergo further antineoplastic therapy, particularly chemotherapy. We examined 35 patients who had been treated with radiotherapy for Hodgkin's lymphoma (12), uterine cervix carcinoma (nine), ovarian dysgerminoma (six), testicular seminoma (four), and non-Hodgkin's lymphoma (four). We observed that radiation-induced modifications of the MRI pattern in the bone marrow are tightly linked to two parameters; the administered radiation dose and the length of time passed after the treatment. Bone marrow recovery was observed only when patients were treated with doses lower than 50 Gy. The earlier radiation-induced modifications of the bone marrow MRI pattern occurred 6 to 12 months after irradiation, and they were most evident 5 to 6 years after the treatment. From 2 to 9 years after radiotherapy, we observed partial recovery. Complete recovery, when it occurred, was observed only 10 to 23 years after the treatment. Our results indicate that MRI studies are likely to be useful in the assessment of radiation-induced injuries.

Bone Marrow↗
PubMed1989

Independent expression of human alpha or beta platelet-derived growth factor receptor cDNAs in a naive hematopoietic cell leads to functional coupling with mitogenic and chemotactic signaling pathways.

Distinct genes encode alpha and beta platelet-derived growth factor (PDGF) receptors that differ in their abilities to be triggered by three dimeric forms of the PDGF molecule. We show that PDGF-receptor mitogenic function can be reconstituted in a naive hematopoietic cell line by introduction of expression vectors for either alpha or beta PDGF receptor cDNAs. Thus, each receptor is independently capable of coupling with mitogenic signal-transduction pathways inherently present in these cells. Activation of either receptor also resulted in chemotaxis, alterations in inositol lipid metabolism, and mobilization of intracellular Ca2+. The magnitude of these functional responses correlated well with the binding properties of the different PDGF isoforms to each receptor. Thus, availability of specific PDGF isoforms and relative expression of each PDGF-receptor gene product are major determinants of the spectrum of known PDGF responses.

Animals↗
PubMed1989

NIH3T3 fibroblasts transformed by the dbl oncogene show altered expression of bradykinin receptors: effect on inositol lipid turnover.

We have examined polyphosphoinositide turnover in mouse fibroblasts (NIH3T3) transformed by the dbl oncogene as compared to normal cells. The dbl-transformed fibroblasts did not show alterations of the basal level of inositol polyphosphates, polyphosphoinositides, diacylglycerol or phosphatidic acid. This indicates that the activity of C-type phospholipases, inositol lipid kinases and diacylglycerol kinase is not altered in dbl-induced transformation. However, dbl-transformed NIH3T3 cells exhibited increased inositol lipid turnover in response to bradykinin. Further analysis revealed significantly higher number of bradykinin receptors in dbl transfectants as compared to control NIH3T3. When several clonally-derived dbl NIH3T3 transfectants were analyzed, we observed a large variation of their bradykinin receptor number. Cell lines exhibiting increased bradykinin binding, however, failed to show augmented mitogenic response to the peptide agonist. Among other oncogenes, only ras showed a similar effect. We conclude that increased bradykinin receptor number is a phenomenon observed with several cell lines transformed by different oncogenes, and it does not correlate with either enhanced mitogenic responsiveness of transformed cells to the peptide, or with the presence of a specific oncogene in the transformant.

Animals↗
PubMed1989

A new filtration system for bone marrow transplantation. Clinical and laboratory studies.

A new method of filtering marrow obtained from marrow donors during marrow transplantation is described. The technique involves a small reusable filter unit and provides a simple, inexpensive, and fast means of filtering marrow in a predominantly closed system prior to infusion into the marrow recipient. In vitro studies of marrow from 10 donors showed no reduction in granulocytemacrophage or erythroid progenitors following filtration. Clinical studies of 75 patients receiving marrow filtered by this system showed no untoward effects or interference with engraftment. We conclude that this predominantly closed filtration system is efficient and should be further evaluated in marrow transplantation.

Adolescent↗
PubMed1988

Effect of exogenously added acylphosphatases on inositol lipid metabolism in human platelets.

In this paper we demonstrate that human platelets contain an acylphosphatase isoenzyme. We then investigated the effect of exogenously added human muscle and erythrocyte acylphosphatases on inositol lipid content in human platelets permeabilized with saponin. Alterations in the level of the polyphosphoinositides were observed: in particular, the levels of phosphatidylinositol 4,5-bisphosphate, and of phosphatidylinositol 4-monophosphate were decreased, whereas the level of phosphatidylinositol was increased. These results suggest that acylphosphatases promote polyphosphoinositide dephosphorylation, possibly through intracellular Ca2+ mobilization.

Acid Anhydride Hydrolases↗
PubMed1988

Morphine withdrawal in vitro: potentiation of agonist-dependent polyphosphoinositide breakdown.

Naloxone (10(-5) -10(-9) M) significantly increased the K+ (30 mM)-induced release of [3H[noradrenaline when it was applied to cortical slices taken from morphine-dependent rats but did not change the release of transmitter when applied to slices prepared from non-dependent animals. Therefore, this preparation was considered suitable to study withdrawal-related events and was used to monitor the agonist-induced changes of phospholipase C activity in the withdrawal state. Noradrenaline (1-100 microM) and carbachol (50-500 microM), when applied to cortical slices preincubated with [3H]inositol or with [32P]orthophosphate, dose dependently increased the formation of labeled inositol phosphates or of phosphatidic acid. This confirmed that noradrenaline and carbachol increase phospholipase C activity. This increase was significantly enhanced by naloxone (10(-6) M) when the slices were taken from dependent animals. The results now reported show for the first time in mammalian tissues that opioid withdrawal is associated with changes of phosphoinositide metabolism.

Animals↗
PubMed1988

Signal transduction through the EGF receptor transfected in IL-3-dependent hematopoietic cells.

An expression vector for the epidermal growth factor (EGF) receptor was introduced into the 32D myeloid cell line, which is devoid of EGF receptors and absolutely dependent on interleukin-3 (IL-3) for its proliferation and survival. Expression of the EGF receptor conferred the ability to utilize EGF for transduction of a mitogenic signal. When the transfected cells were propagated in EGF, they exhibited a more mature myeloid phenotype than was observed under conditions of IL-3-directed growth. Moreover, exposure to EGF led to a rapid stimulation of phosphoinositide metabolism, while IL-3 had no detectable effect on phosphoinositide turnover either in control or EGF receptor-transfected 32D cells. Although the transfected cells exhibited high levels of functional EGF receptors, they remained nontumorigenic. In contrast, transfection of v-erbB, an amino-terminal truncated form of the EGF receptor with constitutive tyrosine kinase activity, not only abrogated the IL-3 growth factor requirement of 32D cells, but caused them to become tumorigenic in nude mice. These results show that a naïve hematopoietic cell expresses all of the intracellular components of the EGF-signaling pathway necessary to evoke a mitogenic response and sustain continuous proliferation.

Animals↗
PubMed1988

Bone marrow transplantation for the treatment of acute nonlymphoblastic leukemia in children aged less than 2 years.

Eleven children with acute nonlymphoblastic leukemia in first remission who were less than 2 years of age at diagnosis were treated with 120 mg/kg of cyclophosphamide, 12-Gy fractionated total-body irradiation, and marrow transplantation. Seven patients remain in complete remission from 3.5 to 13.8 years posttransplant, four for more than 6.75 years. The immediate posttransplant course was relatively uncomplicated in surviving patients. No child developed severe graft-v-host disease. The major long-term side effect has been a slowing in growth. Although the prognosis for such children with conventional chemotherapy remains poor, intensive cytotoxic therapy and marrow transplantation offers an alternative therapy with a chance for cure.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1987

Molecular mechanism underlying impaired platelet responsiveness in liver cirrhosis.

We have studied platelet function in 10 patients with severe liver cirrhosis, compared to healthy subjects. Using washed platelets, we have investigated the molecular mechanism underlying the defect in platelet aggregation frequently observed in these patients. We have found that platelets from cirrhotic patients have a reduced responsiveness to thrombin and collagen in terms of aggregation, and receptor-dependent activation of phospholipase C, A2 and cyclooxygenase/thromboxane synthetase. We thus suggest that this impairment in transmembrane signalling is responsible for the defective platelet function observed in cirrhosis.

Adult↗
PubMed1987

GABA-receptor stimulation enhances norepinephrine-induced polyphosphoinositide metabolism in rat hippocampal slices.

The effect of gamma-aminobutyric acid (GABA)receptor stimulation on norepinephrine (NE)-induced metabolism of polyphosphoinositides (PIPs) was studied in rat hippocampal slices. Inositol phosphates (IPs), PIPs, and phosphatidic acid were measured. NE induced formation of IPs and phosphatidic acid in a dose-dependent manner with an EC50 of 4.5 microM. GABA, 3-aminopropanesulphonic acid (3APS) and muscimol did not affect PIPs breakdown, but they strongly increased (greater than 100%) PIPs metabolism induced by 1 microM NE. Their action was antagonized by bicuculline (10 microM). We discuss the implications of these findings in hippocampal neurotransmission.

Animals↗
PubMed1987

Phospholipase C activation induced by noradrenaline in rat hippocampal slices is potentiated by GABA-receptor stimulation.

We have studied the effect of gamma-aminobutyric acid (GABA) and other GABA-receptor agonists (3-aminopropanesulphonic acid and muscimol) on the noradrenaline-induced stimulation of polyphosphoinositide metabolism in rat hippocampal slices. Formation of water-soluble inositol phosphates, and polyphosphoinositide metabolism were studied in hippocampal slices prelabelled with [3H]myoinositol. Noradrenaline induced formation of inositol mono-, bis- and trisphosphate during 10 min incubation in the presence of lithium; activation of phospholipase C by noradrenaline was also reflected by the hydrolysis of polyphosphoinositides and by the increased metabolism of phosphatidylinositol. GABA-receptor agonists were unable to activate per se phospholipase C; however, when added together with a low concentration of noradrenaline, they greatly potentiated the noradrenaline-stimulated polyphosphoinositide metabolism. We conclude that GABA-receptor agonists potentiate the effect of noradrenaline on polyphosphoinositide turnover and we discuss the role of this neurotransmitter interaction in the physiology of the hippocampus.

Animals↗
PubMed1987

Polyphosphoinositide metabolism is rapidly stimulated by activation of a temperature-sensitive mutant of Rous sarcoma virus in rat fibroblasts.

We have examined polyphosphoinositide turnover in a Rat-1 fibroblast line infected with a temperature-sensitive mutant (ts LA24) of the Rous sarcoma virus (RSV). When ts LA24-infected cells are shifted from the non-permissive to the permissive temperature, a rapid and sustained activation of phospholipase C (PLC) is observed. Normal and wild-type RSV-infected Rat-1 cells do not show any PLC activation upon temperature shiftdown. Pre-treatment of ts LA24-infected fibroblasts with tetrodotoxin (a Na+ channel inhibitor) or incubation in Na+-free medium significantly prevent temperature shiftdown-induced PLC activation. Therefore, we conclude that PLC activation occurs concomitantly with pp60v-src expression, and hypothesize that pp60v-src-related membrane depolarization is the causal link between pp60v-src tyrosine kinase activity and stimulation of polyphosphoinositide metabolism. Finally, we discuss the relationship between the phenomena we have observed and the mechanism of action of the ras oncogene.

Animals↗
PubMed1986

Point-mutated p21ras couples a muscarinic receptor to calcium channels and polyphosphoinositide hydrolysis.

A high-affinity muscarinic receptor is detectable both in normal 3T3 mouse fibroblasts and in their transformed counterpart obtained by transfection with the oncogene EJ/T24-H-ras. However, only the transformed cell line is responsive to muscarinic agonist carbamylcholine in terms of Ca2+ influx and polyphosphoinositide hydrolysis, whereas the normal cell line is unresponsive. Using a point-mutated p21ras protein and monoclonal antibodies anti-p21ras, we provide evidences that p21ras couples to receptor-operating calcium channels and to polyphosphoinositide hydrolysis a muscarinic receptor which is uncoupled in normal mouse fibroblasts.

Animals↗
PubMed1986

Internalization and metabolism of endogenous heparin by cultured endothelial cells.

We have studied the ability of bovine adrenal capillary cells cultured in vitro to uptake and metabolize heparin. We have previously demonstrated that endogenous heparin can be extracted from human plasma (Vannucchi, S. et al., (1985) Biochem. J. 227, 57-65), and here we show that also endothelial cells contain heparin. However, experiments with (35S)sodium sulfate labeling indicate that these cells do not synthesize de novo heparin, but they uptake it from culture serum. Bovine adrenal capillary endothelial cells are able to bind and uptake (3H)heparin added to culture medium and they also release its low molecular weight degradation products, thus indicating a metabolism of heparin. We discuss about the role of endothelial cell-mediated uptake and metabolism of endogenous heparin in relationship with circulating heparin. We also discuss about these events as related to some of the antithrombogenic properties of the endothelium.

Animals↗
PubMed1986

Inositol 1,4,5-trisphosphate induces aggregation and release of 5-hydroxytryptamine from saponin-permeabilized human platelets.

Inositol 1,4,5-trisphosphate induces aggregation and the release of [3H]5-hydroxytryptamine from human platelets rendered permeable with saponin. This action of inositol 1,4,5-trisphosphate is associated with a significant formation of thromboxane B2, activation of phospholipase C, and phosphorylation of 20,000- and 40,000-dalton proteins, which are the substrates for myosin light chain kinase and protein kinase C, respectively. All of these responses are blocked by the cyclooxygenase inhibitors indomethacin and aspirin and the dual cyclooxygenase and lipoxygenase inhibitor 3-amino-1-[m-(trifluoromethyl)phenyl]-2-pyrazoline (BW 755C). These data indicate that platelet activation by inositol 1,4,5-trisphosphate is initiated by the mobilization of Ca2+, which leads to phospholipase A2 activation. The thromboxanes and endoperoxides that are subsequently generated then induce activation via cell surface receptors.

Adenosine Triphosphate↗