PubMed Health⌕ Search

Biomedical subjects

D Marmé

Publications and source records attributed to D Marmé.

At least 73 records · Page 4Linked to original sources
PubMed1992

AIDS-associated Kaposi's sarcoma cells in culture express vascular endothelial growth factor.

PCR cloning and cDNA sequencing have been used to identify mRNAs of two splice products of the vascular endothelial growth factor (VEGF) gene, VEGF121 and VEGF165, in cells isolated from Kaposi's sarcomas (KS) of AIDS patients (AIDS-KS). As demonstrated by Northern blot analysis, AIDS-KS cells as well as tumor cells show a high expression level of the VEGF gene as compared to primary human vascular cells like smooth muscle cells or endothelial cells. In addition to the lower expression of the gene, vascular cells express a 3.9 kb band together with a 3.2 kb band instead of a 3.9 kb and a 4.3 kb band in AIDS-KS cells. Our data suggest that the angiogenic properties of AIDS-KS cells might be mediated by the secretion of this growth factor and that this factor alone or in combination with other endothelial mitogens may be involved in endothelial proliferation associated with Kaposi's sarcoma.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Inducible expression of human phospholipase C-gamma 2 and its activation by platelet-derived growth factor B-chain homodimer and platelet-derived growth factor A-chain homodimer in transfected NIH 3T3 fibroblasts.

Growth factors such as platelet-derived growth factor (PDGF) have been shown to activate phospholipase C-gamma 1 (PLC-gamma 1). We have overexpressed the human PLC-gamma 2 (hPLC-gamma 2) cDNA in murine NIH 3T3 fibroblasts using the interferon-type-I inducible murine Mx promoter. Northern blot analysis revealed an induction of hPLC-gamma 2 mRNA by interferon (IFN) alpha of about 25-fold as compared to the uninduced transcript level. Western blot analysis with anti(bovine PLC-gamma 2) antiserum showed increased hPLC-gamma 2 protein levels in hPLC-gamma 2 transfected cells. Induction with IFN alpha resulted only in a slight further increase. After labelling the cells with [35S]methionine an increase of radioactive label in a protein migrating at 148 kDa could be detected in IFN-alpha-stimulated, hPLC-gamma 2 overexpressing cells. PLC activity in homogenates from hPLC-gamma 2 overexpressing cells was increased as compared to control cells transfected with the vector lacking the hPLC-gamma 2 cDNA insert. There was no difference between in vitro PLC activity in homogenates from PDGF B-chain homodimer (BB) treated and untreated cells. PLC activity was mainly present in the soluble fraction. After incubation of hPLC-gamma 2 overexpressing cells with IFN alpha, the in vitro activity of PLC increased significantly in the soluble fraction. Stimulation with PDGF BB increased inositol phosphate production about 3.5-fold in control cells and about 10-fold in hPLC-gamma 2 overexpressing cells. PDGF A-chain homodimer (AA) showed slightly smaller effects. These results demonstrate that human PLC-gamma 2 can be expressed functionally in murine NIH 3T3 fibroblasts and can be activated by both murine PDGF receptors, alpha and beta type.

3T3 Cells↗
PubMed1992

Biochemical characterization of binding sites for dihydropyridine and omega-conotoxin in brain of adult chicken.

Binding studies using the calcium channel blockers omega-conotoxin and dihydropyridine revealed a rather equal amount of binding sites in brain from adult chicken. The omega-conotoxin binding sites could be solubilized using digitonin, without substantial loss, whereas a great decrease in dihydropyridine binding sites was observed, indicating that both types of binding sites have different sensitivity to solubilization. In contrast to ion exchange chromatography where both binding sites comigrated, glycoprotein affinity chromatography led to a different partition of the binding sites in the flow through and eluate fractions. Our results indicate that both types of calcium channel blockers bind to different targets in adult chicken.

Animals↗
PubMed1992

Characterization and partial purification of a membrane protein from rabbit skeletal muscle which inhibits the cAMP-dependent protein kinase.

Rabbit skeletal muscle membranes contain a protein which inhibits the cAMP-dependent protein kinase. The activity of the partially purified membrane protein is characterized by an IC-50 of 10 to 30 nM with respect to the inhibition of the activity of the catalytic subunit of the cAMP-dependent protein kinase and is sensitive to treatment with heat, acid, alkali and trypsin. The active fractions contain proteins ranging from 40 to 120 kDa, analysed by SDS-gel electrophoresis.

Animals↗
PubMed1992

Inducible overexpression of human protein kinase C alpha in NIH 3T3 fibroblasts results in growth abnormalities.

We have stably overexpressed the human protein kinase C alpha (hPKC alpha) in NIH 3T3 fibroblasts under the control of the interferon (IFN) type I inducible murine Mx promoter. These cells showed a 10-fold increase in the transcription of hPKC alpha mRNA after induction with interferon alpha. The increase in the amount and activity of protein kinase C (PKC)-protein in these cells was only about 3-fold after induction with interferon alpha. Compared to control cells which were transfected with the vector only, the NIH 3T3 fibroblasts transfected with the hPKC alpha cDNA showed already a slightly increased PKC-activity and amount of PKC-protein in the absence of interferon alpha. The hPKC alpha overexpressing cells had an altered, "transformed-like" morphology, which was reversed by staurosporine, an increased growth rate and a higher saturation density. The growth rate was further increased by treating the cells with interferon alpha. The hPKC alpha overexpressing cells were able to grow in soft agarose after treatment with phorbol ester such as TPA (12-O-tetradecanoylphorbol 13-acetate). After phorbol ester and interferon treatment a stronger expression of the protooncogene c-jun was detectable in the hPKC alpha overexpressing cells, whereas expression of c-fos and c-myc was not affected. Since these cells show a specific response pattern due to induced PKC alpha expression they might be useful as an assay system for the development of PKC isozyme-specific inhibitors and activators.

3T3 Cells↗
PubMed1992

Dihydropyridine binding of the calcium channel complex from skeletal muscle is modulated by subunit interaction.

The dihydropyridine-binding subunit alpha 1 of the calcium channel complex from rabbit skeletal muscle can be partially depleted from the alpha 2 delta beta-complex using wheat germ agglutinin-affinity chromatography. This depletion of the alpha 1 from the other subunits leads to a loss of dihydropyridine-binding, which can be fully reconstituted by repletion of the alpha 1 with the other subunits. Reassembly of these subunits results in an increase in the Kd and Bmax of the dihydropyridine-binding indicating that the non-dihydropyridine-binding subunits influence dihydropyridine-binding. The affinity of the alpha 1 subunit for the other subunits was determined to be approximately 35 nM. Since the free alpha 1 subunit will not bind to the beta subunit alone, there is evidence, given the selective partitioning of the beta subunit to the lectin-bound subunit pool, that either beta binds with higher affinity to the alpha 2 delta-complex than to the free alpha 1 subunit or that the bound alpha 1 creates or modulates beta-binding. This indicates a functional high affinity interaction between the dihydropyridine-binding alpha 1 subunit and the alpha 2 delta beta-complex.

Animals↗
PubMed1990

High-efficiency expression of rat protein kinase C-gamma in baculovirus-infected insect cells.

We have expressed rat protein kinase C-gamma in insect cells using a baculovirus vector. The yield of expressed protein kinase C-gamma is about 4% of total protein. The recombinant protein shows a prominent band at about 80 kDa on SDS-polyacrylamide gels, which can be identified as protein kinase C-gamma by Western blotting using monoclonal antibodies against protein kinase C-gamma. Upon incubation with [gamma-32P]ATP and in the presence of Ca2+, phosphatidylserine and diacylglycerol this protein autophosphorylates. Its enzyme activity shows the characteristic properties of mammalian protein kinase C.

Animals↗
PubMed1988

Pertussis toxin inhibits angiotensin II-mediated phosphatidylinositol breakdown and ADP-ribosylates a 40 Kd protein in cultured smooth muscle cells.

Angiotensin II causes an increase of inositol phosphate production in cultured vascular smooth muscle cells from rat aorta. Pretreatment of the cells with pertussis toxin attenuates this effect. Pertussis toxin ADP-ribosylates a protein of about 40 kD in a crude membrane fraction. These data demonstrate the possible involvement of a GTP-binding protein (G-protein) in the angiotensin II-induced activation of phosphoinositidase in vascular smooth muscle cells.

Adenosine Diphosphate Ribose↗
PubMed1987

Synthetic diacylglycerols induce a rise of quin2-detectable free intracellular calcium in human platelets.

The two activators of protein kinase C, oleoylacetylglycerol (OAG) and dioctanoylglycerol (DOG), are able to induce a concentration-dependent rise in cytoplasmic free Ca2+ concentration in gel-filtered human platelets, detected as an increase in quin2 fluorescence. The phorbol ester phorbol-12-myristate-13-acetate (PMA) has no effect. The OAG-induced increase of intracellular Ca2+ is not influenced by forskolin, in contrast to the effect of the diterpene on the thrombin-stimulated increase in cytoplasmic Ca2+. It is concluded that the increase in intracellular free Ca2+ concentration induced by synthetic diacylglycerols and their activation of protein kinase C are two different and independent processes.

Aminoquinolines↗
PubMed1987

Phorbol 12,13-dibutyrate-induced contraction of isolated rabbit vascular smooth muscle.

The contractile effects of the phorbol-12,13-dibutyrate (PDBu) on rabbit basilar, renal, and saphenous arterial rings were investigated under various bathing conditions. At millimolar concentrations of Ca2+, the contractile responses with respect to both maximal contraction and sensitivity to PDBu were in the rank order basilar artery greater than renal artery greater than saphenous artery. Contractions were induced by PDBu also at micromolar concentrations of Ca2+. Depolarization of the cell membrane in the presence of micromolar Ca2+, which did not result in contraction of the vascular rings, caused a shift of the concentration response curves towards lower PDBu concentrations. These results are compatible with the assumption that PDBu induces contraction in vascular smooth muscle by activating protein kinase C.

Animals↗
PubMed1987

Pertussis toxin inhibits the angiotensin II and serotonin-induced rise of free cytoplasmic calcium in cultured smooth muscle cells from rat aorta.

Angiotensin II, serotonin and K+-depolarization cause an increase in free cytoplasmic Ca2+ in cultured smooth muscle cells. The involvement of a guanine nucleotide-binding protein has been investigated by using pertussis toxin. When smooth muscle cells were pretreated with pertussis toxin angiotensin II and serotonin-induced rise of cytosolic Ca2+ was found to be significantly reduced whereas the Ca2+ influx mediated by K+-depolarization remained unchanged. These results suggest the participation of a guanine nucleotide-binding protein in the receptor-mediated rise of intracellular Ca2+.

Angiotensin II↗
PubMed1987

Stereoselective inhibition of calmodulin-dependent cAMP phosphodiesterase from bovine heart by (+)- and (-)-nimodipine.

The inhibitory effects of racemic (+/-)-nimodipine and of optically pure (+)- and (-)-nimodipine on the basal and calmodulin-dependent activity of a cAMP phosphodiesterase from bovine heart were investigated. The inhibition by (+/-)-nimodipine could not be overcome by an excess of calmodulin. However, increase of the cAMP concentration in the assay from 2 X 10(-4) mol/l to 2 X 10(-2) mol/l caused a shift of the IC50 for the inhibition by (+/-)-nimodipine from 2.8 X 10(-6) mol/l to 6 X 10(-5) mol/l. Dixon-plot analysis revealed an inhibitory constant of Ki = 2.3 mumol/l. Experiments with the two enantiomers showed that (+)-nimodipine is by about one order of magnitude more potent than (-)-nimodipine. This contrasts with the stereoselectivity of the Ca2+ channel inhibitory activity on isolated rings of the rabbit basilar artery where (-)-nimodipine is more effective than (+)-nimodipine in relaxing the smooth muscle contracted by K+-depolarisation. It is concluded that cAMP phosphodiesterase may be an intracellular target for nimodipine and its inhibition may contribute to the pharmacological activity of this 1,4-dihydropyridine.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1986

3,4,5-Triiodobenzoic acid affects [3H]verapamil binding to plant and animal membrane fractions and smooth muscle contraction.

3,4,5-Triiodobenzoic acid, known as auxin transport inhibitor, stimulates specific [3H]verapamil binding to zucchini microsomes by 100% (EC50 = 1 microM). This stimulatory effect is due to a decrease of the apparent equilibrium dissociation constant KD for verapamil from 60 nM to 33 nM without significantly changing the maximum number of binding sites. 3,4,5-Triiodobenzoic acid also increases specific [3H]verapamil binding to rabbit skeletal muscle membranes (EC50 greater than or equal to 20 microM) without affecting [3H]nitrendipine and [3H]-d-cis-diltiazem binding. If 3,4,5-triiodobenzoic acid is added to isolated rings of rabbit A. saphena contracted by potassium depolarization, a dose-dependent relaxation is observed with an IC50 value of about 8 microM. Contractions initiated by the addition of 3 microM norepinephrine can also be abolished by 3,4,5-triiodobenzoic acid with half maximal inhibition at 40 microM.

Animals↗
PubMed1986

Microsomal CA2+ uptake in human platelets is stimulated by calmodulin.

Calcium accumulating vesicles were prepared from washed human platelets. Ca2+ uptake could be stimulated with calmodulin. The stimulation of Ca2+ uptake was not abolished by the adenylate cyclase inhibitor 2',5'-dideoxyadenosine, thus excluding enhanced cAMP formation as a possible cause for the calmodulin effect. Our findings suggest that, in addition to cAMP, calmodulin is also involved in the regulation of platelet free Ca2+ concentrations, i.e. Ca2+ homeostasis in platelets is under dual control.

Adenylyl Cyclases↗
PubMed1986

Stereospecific inhibition of 5-HT-induced increase of intracellular free calcium by (+)- and (-)-desmethoxyverapamil in human platelets.

The concentration of intracellular free calcium [Ca2+]i in human platelets was measured by the quin-2 method. 5-Hydroxytryptamine (5-HT) at 10(-5) M induced a rapid transient increase of [Ca2+]i which was antagonized by 10(-7) M ketanserin or cyproheptadine. The verapamil derivative, desmethoxyverapamil (D888), showed stereospecific inhibition of the 5-HT-induced [Ca2+]i increase. The IC50 for (-)-D888 was approx. 2 X 10(-8) M; (+)-D888 was almost 50 times less potent.

Blood Platelets↗
PubMed1985

Specific binding of the calcium antagonist [3H]verapamil to membrane fractions from plants.

Specific binding of the Ca2+ channel blocker [3H] verapamil to a membrane fraction from plants has been characterized. Binding to zucchini membranes was saturable and reversible. The apparent equilibrium dissociation constant is KD = 102 nM and the maximum number of binding sites is Bmax = 60 pmol/mg of protein. The KD determined from the association and dissociation rate constants is 130 nM. [3H]Verapamil binding to zucchini membranes could not be inhibited by the Ca2+ antagonists nifedipine and diltiazem. However, [3H]verapamil could be displaced by diltiazem but not by nifedipine from corn membranes. Sucrose density fractionation of zucchini membrane preparations revealed that [3H]verapamil binding sites are located primarily at the plasma membrane.

Binding Sites↗
PubMed1985

[Calcium. Molecular mechanisms of cellular function].

Calcium plays a pivotal role in cellular signal transduction. Precise control and regulation of cytoplasmic free ionic calcium concentration is a prerequisite for intracellular calcium-mediated physiology. Calmodulin is the ubiquitous calcium receptor protein which conveys the information to the calcium-dependent enzymes. Protein kinase C and receptor-stimulated phosphatidylinositol metabolisms are components of another regulatory mechanism where calcium plays an important regulatory role, too. Oncogene products have been found to possess enzymatic activities involved in phosphatidylinositol metabolism.

Animals↗