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J Timar

Publications and source records attributed to J Timar.

At least 55 records · Page 3Linked to original sources
PubMed1991

Interaction between elastin and tumor cell lines with different metastatic potential; in vitro and in vivo studies.

Interactions between the extracellular matrix macromolecules and tumor cells are critical in the process of metastasis formation. We show here that elastins (both mature insoluble elastin and a 75-kDa soluble peptide: K-elastin) adhere rapidly to two cell lines with high metastatic capacities: a metastatic lung carcinoma cell line (3LL-HM) and a human amelanotic melanoma cell line (A-2058); by contrast the low-metastatic Lewis lung carcinoma cell line variant as well as a rhabdomyosarcoma cell line with a low metastatic potential bind to elastins to a much lower extent. 3H-labelled K-elastin was used in order to study elastin--3LL-HM interaction. It was found to be saturable (2 ng 3H-labelled K-elastin/10(6) cells), with one class of high-affinity binding sites having Kd equal to 1.3 nM and 16,000 sites/cell. The binding of K-elastin to 3LL-HM cells at its receptor triggered several cell responses; (a) increase of intracellular Ca2+ concentration; (b) induction of 3LL-HM chemotaxis toward the K-elastin gradient; (c) stimulation of the adherence of mature insoluble elastin. In contrast to non-transformed cells such as fibroblasts and smooth muscle cells, the adhesion kinetics of insoluble elastin to 3LL-HM did not exhibit a lag period; the rapid binding of insoluble elastin to the tumor cells was followed by its slow detachment from the cells, which lasted for 6 h. 3LL-HM cells but not human skin fibroblasts were shown to secrete elastinolytic activity inhibitable by metal-chelating agents. In vivo studies were performed in order to evaluate the influence of K-elastin binding to 3LL-HM cells on their ability to form lung colonies in mice. It was shown that pretreatment of 10(4) 3LL-HM cells with 10 microM K-elastin and the simultaneous i.v. injection into mice of 750 micrograms K-elastin together with the highly metastatic cells was able to reduce the number of lung colonies by more than 70% after 12 days.

Adult↗
PubMed1991

Morphological studies on the effect of L-leucine methyl ester on mouse peritoneal macrophages in vitro.

Mouse peritoneal macrophages (MO) were treated in culture with 5 mM L-leucine methyl ester (L-Leu-OMe), for 5, 10, 20, 40, and 60 min. The treatment resulted in rapid vacuolisation of the cytoplasm due to the dilatation and disruption of lysosomes. Autophagy caused by lysosomal enzymes destroyed most of the cytoplasmic organelles by 40 minutes after L-Leu-OMe treatment, but the cell membrane and nucleus were in many MOs resistant to the damage. 60 min after L-Leu-OMe treatment most of the MOs were killed. It is supposed that the disruption of the lysosomes is caused by formaldehyde produced by the hydrolysis of L-Leu-OMe.

Animals↗
PubMed1991

The effect of ricin B chain on the intracellular trafficking of an A chain immunotoxin.

Covalent linkage of the A chain of ricin to the LICR-LOND-Fib75 monoclonal antibody produced an immunotoxin, Fib75-SS-ricin A, which demonstrated immunospecific toxicity to human bladder carcinoma cells in tissue culture (Forrester et al., 1984). The present studies have shown that ricin B chain potentiates the toxicity of the immunotoxin by two orders of magnitude and also significantly increases the rate of protein synthesis inhibition. Using immunoelectron microscopy, the receptor-mediated endocytosis and intracellular routing of the immunotoxin was studied with and without ricin B chain treatment after immunolocalisation of the conjugate. Fib75-SS-ricin A was internalised by the EJ cells predominantly in uncoated pits and vesicles and directed to the endosomes. Some degradation of the complex appeared to take place in multivesicular endosomes at early timepoints and 24 h after internalisation, most of the immunotoxin was found in lysosomes. Some ricin A chain epitopes were detected in Golgi vesicles. Cells treated with immunotoxin and ricin B chain endocytosed the complex predominantly in coated pits and coated vesicles. Using pre-embedding immunoperoxidase techniques, ricin chains were found in the whole Golgi complex and most of the conjugate escaped lysosomal degradation. Internalised immunotoxin was recycled back to the plasma membrane in an active form associated with vesicles which appeared to be derived predominantly from multivesicular endosomes. A similar mode of recycling has recently been reported (McIntosh et al., 1990) for ricin holotoxin in the same cell line. These observations may explain the potentiating effect of toxin B chains in the antibody-directed targeting of toxin A chains.

Antibodies, Monoclonal↗
PubMed1991

Lipoxygenase metabolites of arachidonic and linoleic acids modulate the adhesion of tumor cells to endothelium via regulation of protein kinase C.

12(S)-hydroxyeicosatetraenoic acid (12[S]-HETE) and 13(S)-hydroxyoctadecadienoic acid (13[S]-HODE), lipoxygenase metabolites of arachidonic acid and linoleic acid, respectively, previously have been suggested to regulate tumor cell adhesion to endothelium during metastasis. Adhesion of rat Walker carcinosarcoma (W256) cells to a rat endothelial cell monolayer was enhanced after treatment with 12(S)-HETE and this 12(S)-HETE enhanced adhesion was blocked by 13(S)-HODE. Protein kinase inhibitors, staurosporine, calphostin C, and 1-(5-isoquinoline-sulfonyl)-2-methylpiperazine, inhibited the 12(S)-HETE enhanced W256 cell adhesion. Depleting W256 cells of protein kinase C (PKC) with phorbol 12-myristate-13-acetate abolished their ability to respond to 12(S)-HETE. Treatment of W256 cells with 12(S)-HETE induced a 100% increase in membrane-associated PKC activity whereas 13(S)-HODE inhibited the effect of 12(S)-HETE on PKC translocation. High-performance liquid chromatographic analysis revealed that in W256 cells 12-HETE and 13-HODE were two of the major lipoxygenase metabilites of arachidonic acid and linoleic acid, respectively. Therefore, these two metabolites may provide an alternative signaling pathway for the regulation of PKC. Further, these findings suggest that the regulation of tumor cell adhesion to endothelium by 12(S)-HETE and 13(S)-HODE may be a PKC-dependent process.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1991

The relief of biliary obstruction in experimental rats by diverting the hepatic lymph into the duodenum.

Twelve days after their common bile ducts had been ligated rats were icteric and their hepatic lymph nodes and lymph ducts were two or three times their normal size. Cannulation of the hepatic lymph ducts of these rats yielded bile-stained lymph which flowed at nine times its normal rate. As the lymph flowed, so the concentration of bilirubin in the blood declined; after 5 days the rats were no longer jaundiced, and by day 11 there were normal amounts of bilirubin in the blood. In another series it proved possible on seven occasions to insert the free end of the hepatic lymph cannula into the duodenum at the time that the bile was obstructed. Five of these animals showed no increase in serum bilirubin and remained in good condition until the experiments were terminated up to 61 days later. It seems that the effects of biliary obstruction can be mitigated by shunting the hepatic lymph into the intestine.

Animals↗
PubMed1990

Proteoglycan-targeted antibodies as markers on non-Hodgkin lymphoma xenografts.

A family of mono- and polyclonal antibodies raised against proteoglycans or their "subcomponents" served as novel markers to characterize the phenotypes of three non-Hodgkin lymphoma xenograft lines (HT 58 lymphoblastic, HT 117 centroblastic, HT 130 centrocytic) together with normal, human peripheral blood B lymphocytes. These xenografted NHL lines, maintained by serial transplantations on artificially immunosuppressed mice, expressed very similar B-cell-related antigens and differences on the cell surface (HT 58 greater than HT 117 greater than HT 130 greater than B cells) when they were exposed to monoclonal antibodies (mAb) to cartilage proteoglycans. Anti-proteoglycan antibodies used in this study recognize complex epitopes of core protein segment associated with carbohydrate, shared by human cartilage proteoglycans and certain lymphoma cells. The binding of these antibodies was independent of cell-cycle phase. The results suggest that the anti-proteoglycan mAbs could be used as new phenotypic markers to individualize non-Hodgkin lymphomas.

Animals↗
PubMed1990

Localization and production of proteoglycans by HT1080 cell lines with altered N-ras expression.

The alterations in the production of proteoglycans in relation to the expression of the malignant phenotype - controlled by the level of expression of activated N-ras gene - was studied in HT1080 human fibrosarcoma cells and in its revertant variants rev.1c and rev.10a. A decreased production of radiolabelled PGs - especially HSPGs - was observed in HT1080 cells, compared to the revertant lines. By immunofluorescence, the HSPG epitopes were localized mainly into the putative endoplasmic reticulum and/or Golgi zone in HT1080 cells, and to the diffuse cytoplasmic and membrane localization in the revertant lines. It is suggested, that the altered expression of PGs represents an important aspect of the transformed phenotype of HT1080 fibrosarcoma cells.

Cell Division↗
PubMed1990

The intracellular movement and cycling of ricin.

The binding, internalization and recycling of the plant toxin ricin, was studied using electron microscopy and biochemical techniques. For the electron microscope study, ricin was visualized using a gold-labeled second antibody, in the cells of the EJ human bladder carcinoma line growing in monolayer culture. The labeled antibody/toxin complex was found to enter the cell in coated pits and to accumulate in endosomes and to a lesser extent in vesicles associated with the Golgi system. The complex recycled to the cell surface partly in uncoated vesicles, but largely in multivesicular bodies which appeared to exocytose their contents to the extracellular space. Twenty hours after the initial contact with ricin as much as 50% of the cellular label was found on the cell surface mainly associated with shed vesicles. When cells were treated with unlabeled ricin holotoxin and then after 20 h stained post-fixation, ricin molecules, partly associated with vesicles, were present on the cell surface. Biochemical studies showed that ricin was internalized by cells and then released in an intact form to the extracellular space. It was found that less than 10% of the released material had been degraded during its passage through the cells, which is in accord with the low level of label found in the lysosomal system during the morphological study.

Antitoxins↗
PubMed1990

Interactions of exogenous heparan sulfate with tumor cells of different metastatic phenotype.

Heparan sulfate (HS) enhanced the growth of the highly metastatic (HM) 3LL cell line, but not that of the low metastatic (LM) counterpart, in a dose-dependent way. Heparin, chondroitin sulfate, hyaluronate did not produce this effect. At 4 degrees C both cell lines exhibited high affinity binding sites (Kd 10(-8) M) for exogenous HS. Unlike LM cells, the HM ones lost half of the surface-bound HS during the 24-hour incubation at 37 degrees C. HM cells in the exponential growth phase took up the bound HS at lower rate than the LM counterparts. Both cell lines fragmented the exogenous HS intracellularly, but only the HM cells were able to degrade it at the cell surface. The HM cells contained much more heparin-binding proteins--especially in the cell membrane and nuclear fraction--than the LM ones. These results clearly demonstrate that the interactions of tumor cells with exogenous HS are influenced by the invasive phenotype. We suggest also that there could be a correlation between the surface degradation, the slow uptake and the growth-promoting effect of the exogenous HS.

Animals↗
PubMed1989

Elastonectin and the elastin receptor.

Two different types of interactions were described between cells and elastin: 1) the adhesion of cells to insoluble fibrous elastin and 2) the binding of soluble elastin derived peptides by cells. We could show that these are two different mechanisms underlying those two types of interactions. The adhesion of cells to elastin fibers is mediated by a cell-membrane complex with a 120 kD protein as the main adhesive compound which we proposed to designate elastonectin. Three other proteins (67, 60 and 45 kD) were coisolated with elastonectin. Adhesion of insoluble elastin to mesenchymal cells (fibroblasts, smooth muscle cells) is inducible with soluble elastin peptides. Highly metastatic Lewis lung carcinoma cells and human melanoma cells also exhibited the adhesion mechanism, but without a lag phase (constitutive adhesion). Binding curve (Scatchard plots) obtained with radiolabelled elastin peptides indicated the presence of high affinity elastin receptors on mesenchymal cells and human white blood cells (monocytes and PMNs) with Kd-s in the nanomolar range. The binding of elastin peptides triggers several cellular reactions such as chemotaxis, Ca++ influx (increase of Ca++i) and with mononuclear blood cells release of lytic enzymes and oxygen radicals. All these cells which exhibited elastin receptor function also exhibited adhesion although the two processes could be inhibited selectively. The receptor action is mediated by a G-protein-phospholipase-IP3 mechanism, involved in the increase of intracellular Ca++. It appears that this action also triggers the biosynthesis and membrane localization of elastonectin. The coupling of these 2 mechanisms between receptor and adhesion appears to be modified in transformed cells.

Cell Adhesion↗
PubMed1987

Ultrastructure and immunoelectron microscopy of human T-cell leukaemia virus type I-producing lymphoid and non-lymphoid human tumour cells.

The ultrastructure and distribution of some viral and cellular antigens was examined in lymphoid C91/PL and fibroblastic HOS/PL cells infected with human T-cell leukaemia virus type I. Tubuloreticular structures in the endoplasmic reticulum, characteristic of virus-infected or interferon-treated cells, were found in both cell types. Virions were observed particularly in the HOS/PL cells in budding and mature forms and in coated pits, coated vesicles and lysosomes, indicative of receptor-mediated endocytosis. Using immunogold indirect labelling, viral antigens, recognized by sera of infected patients and by monoclonal antibody, were detected in patches on the cell surface and in material loosely attached to the envelope of some virions. Beta 2-microglobulin was associated with virion envelopes from C91/PL and HOS/PL cells, but major histocompatibility complex class 1 (HLA) antigens were not associated with virions even when produced by HLA-positive HOS/PL cells.

Antigens, Surface↗
PubMed1986

Long-term administration of (-)deprenyl (selegiline), a compound which facilitates dopaminergic tone in the brain, leaves the sensitivity of dopamine receptors to apomorphine unchanged.

The effect of repeated administration of the MAO-B enzyme blocker (-)deprenyl on the apomorphine (APO) sensitivity of dopamine (DA) receptors was investigated in rats, and compared to the effect of other drugs influencing the dopaminergic system. APO was given either in a high dose (0.1-0.6 mg/kg), which induces stereotyped behaviour or in a smaller one (0.02 mg/kg) causing sedation. Repeated administration of all the other drugs investigated (except (-)deprenyl), i.e. haloperidol, d-amphetamine, (1 mg/kg s.c., respectively) and the MAO-A blocker clorgyline (0.25 mg/kg s.c.) altered the efficiency of APO on the stereotypy. Haloperidol, clorgyline (0.5 mg/kg s.c.) and imipramine (10 mg/kg i.p.) attenuated the APO-sedation. The long-lasting administration of (-)deprenyl (0.25 mg/kg s.c., daily for 42 days) however, left the effects of APO unchanged, demonstrating that (-)deprenyl facilitates the dopaminergic tone in the rat brain without altering the sensitivity of DA receptors.

Animals↗
PubMed1985

Flow cytometric measurements and electron microscopy of cell surface glycosaminoglycans using acridine orange.

Cell surface glycosaminoglycans (GAGs) were measured, after various treatments, by their binding to Acridine Orange using flow cytometry. Using a critical electrolyte concentration and combining it with specific degradation of individual GAG elements, it was found possible to differentiate between GAG components. The technique was adapted for electron microscopy level to reveal characteristics of membrane-associated GAG. By this means, the cell membrane of the human leukaemic cell line K562 was shown to contain a large amount of GAG; 75% of it was highly sulphated GAG, mostly heparan sulphate. This component was evenly distributed in the outer plasma membrane layer. In the presence of other GAGs, the appearance of complex proteoglycan granules was detected.

Acridine Orange↗
PubMedDate not supplied

Epitopes of cartilage core proteins and GAG pattern in human non-Hodgkin lymphoma xenografts.

Glycosaminoglycan and core protein components of proteoglycans (PGs) have been studied in three human non-Hodgkin lymphoma xenografts of B cell origin. Lymphomas showed similar GAG content, but different composition of GAG subtypes. This variability was accompanied by an individual capacity to adhere to extracellular matrix elements. The core proteins identified by monoclonal antibodies raised against human cartilage chondroitin sulfate PG were also distinctly expressed and released. These proteins shared by different cell types may have biological significance.

Aggrecans↗
PubMedDate not supplied

Serotonin-mediated behavior in rats chronically treated with (-) deprenyl.

(-) Deprenyl given in small (0.25 mg/kg) daily doses for 30 days to rats left the serotonin (5-HT) and the 5-hydroxyindoleacetic acid (5-HIAA) levels in the striatum unchanged. The postsynaptic 5-HT receptor responsiveness, as measured with 5-methoxy-N,N-dimethyl-tryptamine (5-MeODMT), remained also unaltered. In (-) deprenyl-treated rats the p-chloroamphetamine (PCA)-induced 5-HT syndrome was significantly increased and this enhancement of the PCA effect was not abolished by the reserpine-induced depletion of the 5-HT stores.

Animals↗