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M Castagna

Publications and source records attributed to M Castagna.

At least 91 records · Page 5Linked to original sources
PubMed1985

Effects of tumor promoters on the expression of a tumor-related multigenic set in human cells.

We previously found that a minor subfraction of the human genomic DNA, corresponding to 2500-3000 nonrepetitive sequences of 3 kilobases each and designated as tumor-activated DNA (TaDNA) was transcriptionally active in Burkitt's lymphoma cells and almost inactive in normal lymphocytes growing in vitro following integration of the Epstein-Barr virus genome. Furthermore all the neoplastic cells in culture or primary neoplasms (leukemias, sarcomas, carcinomas) studied contained transcripts from most of the TaDNA sequences found in malignant lymphoblasts whereas normal cells growing in vitro contained only a few TaDNA transcripts. It is shown in the present study that treatments of the myeloid leukemia HL60 cells with various inducers of cell differentiation (dimethyl sulfoxide, retinoic acid, mezerein, 12-O-tetradecanoylphorbol-13-acetate, teleocidin) caused a dose-dependent reduction of the level of TaDNA transcripts, correlated with the diminution of c-myc transcripts. The 12-O-tetradecanoylphorbol-13-acetate treatment had this same effect on Burkitt's lymphoma cells (Raji or Namalwa) but the opposite effect on normal cells (Epstein-Barr virus-immortalized lymphocytes or fetal fibroblasts) where it enhanced the formation of Ta-DNA transcripts up to the levels found in untreated malignant cells. These data suggest two conclusions (a) TaDNA corresponds to a multigenic set which seems to be involved in modulation of the malignant phenotype and (b) depending on the origin of the cells, agents like 12-O-tetradecanoylphorbol-13-acetate may operate either as tumor promoters or as differentiation inducers through the control of TaDNA expression.

Actins↗
PubMed1984

Further characterization of tumor-promoter-mediated activation of protein kinase C.

Tumor promoting phorbol esters are able to activate Ca2+-sensitive, phospholipid-dependent protein kinase (protein kinase C) in a reconstituted system. Indol alkaloid teleocidin, a tumor promoter, has been found to be as potent as tumor promoters from the series of phorbol esters and mezerein in activating the mouse brain enzyme. Chemically unrelated tumor promoters such as tetrachlorodibenzo-p-dioxin, anthralin and phenobarbital are devoid of effect. Diacylglycerol 1,2 diolein strongly activated the enzyme whereas 1,3 diolein like 1,2 distearin were poor activators and 1,3 distearin was inactive. Although tumor-promoter-or diacylglycerol-mediated activation of protein kinase C was observed in the presence of 0.5mM EGTA, the reaction requires traces of Ca2+. Tumor promoters did not prevent inhibitory action of antipsychotic phenothiazines and local anesthetics but appear to increase IC50 of these drugs.

Animals↗
PubMed1984

Growth-rate-related and hydroxysterol-induced changes in membrane fluidity of cultured hepatoma cells: correlation with 3-hydroxy-3-methyl glutaryl CoA reductase activity.

3-hydroxy-3-methylglutaryl-coenzyme A reductase (EC 1.1.1.3.4.) activity and cell membrane fluidity measured by fluorescence polarization using 1,6 diphenyl, 1,3,5-hexatriene as fluorescent probe have been concomitantly examined in HTC hepatoma cells, both in relation to growth rate and in response to treatment with hydroxylated sterols. A high level of HMG-CoA reductase activity was observed in cells at log phase of growth which progressively decreased to reach a sustained low level at stationary phase. Similarly, membrane fluidity markedly decreased in relation to growth rate. Hydroxylated sterols such as 7 beta-hydroxycholesterol or 25-hydroxycholesterol strongly inhibited HMG-CoA reductase activity whereas a water-soluble derivative of 7 beta-hydroxycholesterol sodium 3,7-bishemisuccinate had no effect. Within the same range of concentrations 7 beta-hydroxycholesterol and 25-hydroxycholesterol strongly decreased membrane fluidity when the water-soluble derivative was ineffective. Thus, the present results provide evidence for a correlation between the two tested parameters and suggest a dependency of HMG-CoA reductase activity on cell membrane fluidity.

Animals↗
PubMed1984

Cutaneous metastasis of a radiation-associated thyroid medullary carcinoma.

The rare event of a single cutaneous metastasis occurred in a 37-year-old white man two years after a subtotal thyroidectomy for sporadic medullary carcinoma is reported. The endocrine nature of the tumor is revealed by the ultrastructural study, and the type of secretion by the biochemical assay of calcitonin (CT) in the serum and in the tumor tissue extract and by the PAP (peroxidase-anti-peroxidase) technique with anti-CT antibodies on paraffin sections. The cytological and histological findings are described. Some peculiar ultrastructural aspects are presented in details. The fact that the patient received a course of Rx therapy (3,000 rads) on the neck for cervical syringomyelia at the age of 20 years is stressed.

Adult↗
PubMed1984

Thermal tolerance by embryos and larvae of the surf clam Spisula solidissima.

Thermal tolerance was tested in three developmental stages of the surf clam Spisula solidissima: cleavage stages, trochophore , and straight hinge larvae. Experiments were designed to simulate larval entrainment in the cooling system of a power plant. Eleven different temperatures were investigated from 20.2 to 43 degrees C. Exposure times varied from 1 min to 6 hr and following each of the (88) different exposures, larvae were returned to the incubation temperature for a further period of development which facilitated differentiation of dead and live animals. Pooled mortality data from triplicate experiments (for each larval state) were subjected to a stepwise regression analysis (percentage mortality on temperature and time). Equations were derived from the regression program comprising first-, second-, and third-order terms for temperature and time, and these were used to generate response surfaces relating mortality to temperature-time exposure. As with earlier work on Mulinia , the term (temperature) provides the best single predictor of larval mortality. Spisula is the only fully marine bivalve species so far subjected to this type of thermal tolerance test and is clearly the most temperature sensitive of all species investigated to date.

Animals↗
PubMed1984

Morphologic assessment of reactivity to monoclonal antibodies generated against breast cancer cells in mammary tissues removed for clinical dysplasia or cancer.

Monoclonal antibodies B 6.2, B 72.3 and B 1.1, reactive against tumor-associated antigens of human breast cancer, have been tested on histologic sections of paraffin blocks from 49 cases of breast cancer and from 13 cases of mammary dysplasia. The results have shown that: (a) the three monoclonal antibodies recognize different antigenic determinants of the cancer cells; (b) monoclonal antibodies B 6.2 and B 1.1 recognize antigens which are frequently expressed in cancer cells; (c) the antigen recognized by monoclonal antibody B 72.3, at the concentrations used, is less frequently expressed in tumors, while it seems to be selective for the apical cell border and luminal secretions of apocrine metaplastic lobules or cysts occurring in mammary dysplasia; (d) intracytoplasmic expression of the antigens is observed only in cancer cells, with rare exceptions; (e) tumor cell populations are heterogeneous in respect to the expression of the antigens detected by the monoclonal antibodies tested; (f) the antigens detected are unrelated to the tumor histotype; (g) the frequency of axillary lymph nodal metastases is not influenced by the presence or intensity of antigenic expression in the primary tumors; but (h) in metastatic tumor cell populations the antigenic expression is higher than in primary tumors. This fact suggests some relationship between tumor metastatization and the expression of antigens recognized by the monoclonal antibodies tested.

Adult↗
PubMed1984

Protein phosphorylation and mechanism of action of tumor-promoting phorbol esters.

The phorbol ester TPA may be intercalated into the membrane phospholipid bilayer and selectively binds to every molecule of C-kinase in exhibiting its full enzymatic activity. Available evidence suggests that C-kinase is the receptive protein of this tumor promoter, and the results presented seem to provide clues for clarifying the mechanism of controlling cell growth and differentiation.

Animals↗
PubMed1983

Synergistic functions of phorbol ester and calcium in serotonin release from human platelets.

In human platelets, thrombin activates Ca2+-activated, phospholipid-dependent protein kinase (protein kinase C) and mobilizes Ca2+ concomitantly, whereas 12-O-tetradecanoylphorbol-13-acetate (TPA) may be intercalated into membranes and directly activates protein kinase C without mobilization of Ca2+ in sufficient quantities. A series of experiments with TPA and Ca2+-ionophore (A23187) indicates that activation of protein kinase C is a prerequisite requirement for release of serotonin, and that this enzyme activation and Ca2+ mobilization act synergistically to elicit a full cellular response. Both cyclic AMP and cyclic GMP inhibit activation of protein kinase C by prohibiting the signal-dependent breakdown of inositol phospholipid to produce diacyl-glycerol, but none of these cyclic nucleotides prevents the TPA-induced activation of this enzyme.

Blood Platelets↗
PubMed1983

Tumor promoter 12-O-tetradecanoylphorbol 13-acetate alters state, fluidity and hydration of 1,2-diacyl-sn-glycero-3-phosphocholine bilayers.

Previous results (Castagna et al. (1979) FEBS Lett. 100, 62-66; Fisher et al. (1979) Biochem. Biophys. Res. Commun. 86, 1063-1068) indicated us that the active tumor promoter TPA (12-O-tetradecanoylphorbol 13-acetate) decreased fluorescence polarisation of diphenylhexatriene in lymphoblastoid and rat embryo cells. In the present study, experiments aimed at examining the molecular interactions of tumor promoters with cell membrane components are performed with fully hydrated multibilayers of 1,2-diacyl-sn-glycero-3-phosphocholine (DPPC) into which increasing amounts of TPA are inserted. The thermotropic behaviour of both the phospholipid bilayers and the interbilayer water was investigated using the differential scanning calorimetry (DSC) and the approach of Ter-Minassian-Saraga et al. ((1982) J. Colloïd Interface Sci. 81, 369-383). The major effects of the tumor promoter are confined to concentrations up to 20% mol fractions of TPA. In this range of concentrations the incorporation of TPA into liposomes decreases the phase-transition temperature but did not affect delta HDPPC. Furthermore TPA increases the hydration of the multibilayers. Above 20% mol fractions of TPA, a different thermal behaviour of the system which might suggest morphological rearrangements was observed. The lipid state in TPA-treated liposomes was monitored by fluorescence polarisation using diphenylhexatriene as a lipophilic fluorescent probe and the phase-transition temperature was calculated. The phase transition temperatures determined by both methods were in good agreement. The lowering of this temperature and the decay of fluorescence anisotropy of diphenylhexatriene were parallel. Those effects are consistent with the "fluidising' effect of TPA on DPPC.

Calorimetry, Differential Scanning↗
PubMed1983

Tumor promoters enhance cap formation in mouse thymocytes.

Potent tumor promoters such as 12-O-tetradecanoylphorbol 13-acetate (TPA) and teleocidin, rapidly evoked a dose-dependent stimulation of concanavalin A (Con A)-induced cap formation in mouse T lymphocytes. The effect was reversible upon removal of the drugs. Weaker tumor promoters, phorbol didecanoate, phorbol dibenzoate and iodoacetic acid stimulated capping to a lower extent. Mezerein, a phorbol-related macrocyclic diterpene derivative, which acts as a second-stage promoter was also active in increasing the number of caps. In contrast, 4 alpha-phorbol didecanoate and phorbol which are devoid of tumor promoting activity, did not affect capping. Anti-promoting glucocorticoids inhibited capping stimulation. Flow cytofluorometric analysis of Con A binding has shown that TPA did not modify the lectin binding to surface receptors. TPA-facilitated capping was energy-dependent. Cytochalasin B prevented the TPA-induced response whereas colchicine was ineffective. Phenothiazines fully inhibited the TPA effect, thus suggesting that tumor-promoter-mediated lectin receptor redistribution may be ascribed to the facilitation of a Ca2+-dependent process involving the submembrane actin filaments.

Animals↗
PubMed1983

Transmembrane signalling systems.

The three major strategies for transmembrane signalling at the plasma membrane level involve either second messengers, ion channels or receptor-mediated endocytosis. Two second-messenger types of transmembrane signalling systems have been analyzed: the adenylate cyclase and protein kinase C systems. Both are triggered by a variety of biological signals and eventually activate a specific protein kinase. The pathogenic stimulation of adenylate cyclase and protein kinase C systems by cholera toxin and tumor promoters, respectively, is described.

Adenylyl Cyclases↗
PubMed1982

Direct activation of calcium-activated, phospholipid-dependent protein kinase by tumor-promoting phorbol esters.

Tumor-promoting phorbol esters such as 12-O-tetradecanoylphorbol-13-acetate (TPA) directly activate in vitro Ca2+-activated, phospholipid-dependent protein kinase (protein kinase C), which normally requires unsaturated diacylglycerol. Kinetic analysis indicates that TPA can substitute for diacylglycerol and greatly increases the affinity of the enzyme for Ca2+ as well as for phospholipid. Under physiological conditions, the activation of this enzyme appears to be linked to the receptor-mediated phosphatidylinositol breakdown which may be provoked by a wide variety of extracellular messengers, eventually leading to the activation of specific cellular functions or proliferation. Using human platelets as a model system, TPA is shown to enhance the protein kinase C-specific phosphorylation associated with the release reaction in the total absence of phosphatidylinositol breakdown. Various phorbol derivatives which have been shown to be active in tumor promotion are also capable of activating this protein kinase in in vitro systems.

Animals↗
PubMed1981

Phorbol esters parallel effects on tumor promotion, insulin release and calcium ionophoresis.

The tumor promoters 12-O-tetradecanoylphorbol-13-acetate (TPA) and phorbol 12,13-didecanoate (PDD) both stimulated insulin release from rat pancreatic islets and facilitated ionophore-mediated calcium transport in liposomes, the latter effect not resulting from a change in viscosity of the liposomial matrix. Phorbol and 4 alpha-phorbol 12,13-didecanoate (4 alpha-PDD), which fail to cause tumor promotion, also failed to stimulate insulin release and ionophore-mediated calcium transport. An alteration of calcium transport may represent a fundamental event in both the early (insulin release) and late (tumor promotion) biological responses to phorbol esters.

Animals↗
PubMed1981

Variation with embryonic development and regional localization of specific [3H]phorbol 12,13-dibutyrate binding to brain.

We have previously characterized specific binding of the phorbol ester tumor promoter [20-3H]phorbol 12,13-dibutyrate to several tissues, including mouse skin and brain. We report here that specific binding activity in chicken brain increases dramatically during development. In whole embryonic chicken brain, binding increased from 2.0 to 6.8 to 10.6 pmol/mg of protein at 7, 14, and 20 days, respectively. Adult chicken brain bound 15 pmol/mg of protein. In contrast to specific binding activity, binding affinity remained constant. Substantial regional localization of binding activity within the brain was found. For calf brain, representative values ranged from 4.6 pmol/mg for medulla to 38.1 pmol/mg for frontal lobe. Binding affinity did not vary. Inhibition of binding by postulated neurotransmitters and antagonists was examined. D-Propranolol and quinidine, membrane-stabilizing drugs, inhibited binding competitively, although at mM concentrations.

Animals↗
PubMed1981

Kinetics and subcellular localization of specific [3H]phorbol 12, 13-dibutyrate binding by mouse brain.

The specific binding of [3H]phorbol 12,13-dibutyrate ([3H]-PDBU) to particulate preparations from mouse brain has been further characterized. Kinetic analysis, using a filtration assay to measure binding, yielded a second-order rate constant at 23 degrees of 3.75 X 10(7) M-1 min-1 and a first-order dissociation rate constant of 0.21 min-1. The Kd of 5.6 nM calculated from the kinetic data agreed well with the value determined previously in equilibrium binding studies. The Kd for [3H]PDBU binding varied only slightly with temperature. From its temperature dependence, [3H]PDBU binding appeared to be associated with a small increase in enthalpy (delta H degrees = +0.4 kcal/mol) and a large increase in entropy (delta S degrees = +38 e.u.). Such values are characteristic for hydrophobic interactions. The dissociation rate constant for binding, in contrast to the Kd, varied dramatically with temperature. The half-time for release ranged from 1.75 min at 30 degrees to 62 min at 4 degrees. The Kd for binding was Ca2+ sensitive; chelation of Ca2+ by ethyleneglycolbis(beta-aminoethyl ether)N,N'-tetraacetic acid increased the Kd 2.4-fold. Upon subcellular fractionation, the specific [3H]PDBU binding activity was exclusively particulate; no binding to cytosol was detectable. Binding clearly did not correlate with nuclear or mitochondrial markers. On the other hand, a broader distribution of binding activity was seen on sucrose density gradients than for either Na+-K+-adenosine triphosphatase activity or binding of quinuclidinyl benzilate (a muscarinic cholinergic antagonist). The localization of specific [3H]PDBU binding to the plasma membrane therefore remains uncertain.

Animals↗
PubMed1980

Correlation between growth rate, cell density, and intracellular concentrations of cyclic nucleotides in chemostat cultures of mouse L1210 cells.

Constant intracellular concentrations of both adenosine 3',5' cyclic monophosphate (cyclic AMP) and guanosine 3',5' cyclic monophosphate (cyclic GMP) were obtained when mouse leukemia L1210 cells were cultivated under steady-state conditions in the chemostat. When L1210 cells were maintained at a constant growth rate in the chemostat, the mean steady-state intracellular concentration of cyclic AMP decreased with increasing cell density, while the concentration of cyclic GMP remained unchanged. When cell growth rate was increased by three-fold, independently of cell density, in the chemostat, then the mean steady-state intracellular concentration of cyclic GMP increased by 3.6-fold, whereas that of cyclic AMP increased by only 22%. The ratio of cyclic AMP to cyclic GMP was found to decrease with increasing cell growth rate in the chemostat. Our results show that a close correlation exists between the steady-state intracellular concentrations of cyclic nucleotides and changes in both cell growth rate and cell density in chemostat cultures of L1210 tumour cells.

Animals↗