Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0017638 (glioma)
 30,880 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Intact and permeabilized glioma C6 cells were incubated with [14C]serine in media containing low (100 nM) or high (2 mM) [Ca2+] and serine incorporation into phosphatidylserine was examined. In all cases thapsigargin, a blocker of the endoplasmic reticulum Ca(2+)-ATPase, diminished this process, whereas the action of the ionophore A23187 was dependent on the external calcium concentration and time of incubation. In permeabilized cells incubated at 100 nM Ca2+, serine incorporation into phosphatidylserine was diminished when the endoplasmic reticulum Ca2+ levels were lowered by the ionophore. In intact cells incubated at 2 mM CaCl2, addition of A23187 had no effect for the first 30 min and later decreased [14C]serine incorporation. This result seems to be not connected with the degradation of already formed phosphatidylserine, or with an enhanced metabolic conversion of this phospholipid, but with the decrease of its synthesis. The mechanism of this last process appears to be involved in the ionophore-induced perturbation of cellular Ca2+ homeostasis. Our results indicate that phosphatidylserine synthesis is a Ca(2+)-regulated process.
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PMID:Effect of ionophore A23187 and thapsigargin on serine incorporation into phosphatidylserine in intact and permeabilized glioma C6 cells at high and low Ca2+ concentrations. 813 13

The in vitro cell survival, localization and ultrastructural changes following irradiation were examined in 9L glioma cells sensitized with a new photosensitizer, lysyl chlorin p6 (LCP). In clonogenic assays, LCP was 10-100-fold more phototoxic than photofrin II on a microgram/mL basis. Lysyl chlorin p6 uptake was blocked when cells were incubated at 2 degrees C. In view of the chemical properties of LCP, this finding indicates that uptake probably occurred through the endocytic pathway. Fluorescence studies showed LCP localized in a region of the endocytic compartment similar in size, shape and distribution to that labeled by lucifer yellow CH (LY), as well as localizing diffusely throughout the perinuclear cytoplasm. Cells stained with both LY and LCP, however, had distinctly separate regions of staining. Lysyl chlorin p6 localization differed from that of fluorescent probes labeling the mitochondria, Golgi apparatus and endoplasmic reticulum. Ultrastructural changes at both 2 and 30 min after laser irradiation were similar. Mitochondria were often condensed or swollen and also had constrictions and cytoplasmic invaginations. The Golgi apparatus, perinuclear space and rough endoplasmic reticulum (RER) were dilated. These data demonstrate that LCP localizes in a portion of the endosomal compartment, but that morphologic damage initially occurs in the mitochondria, Golgi apparatus and RER.
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PMID:In vitro photodynamic effects of lysyl chlorin p6: cell survival, localization and ultrastructural changes. 828 20

Cholera toxin (CT) consists of a pentameric B subunit which binds to ganglioside GM1 on the cell surface and an A subunit which activates adenylylcyclase. The latter process involves the reduction of A to the A1 peptide which ADP-ribosylates the stimulatory G protein, Gs of adenylylcyclase. There is a distinct lag phase between toxin binding and activation of adenylylcyclase. Little is known about the events during this lag including where A1 is generated and how it gains access to Gs on the cytoplasmic side of the plasma membrane. We explored the effects of several inhibitors of intracellular trafficking on the response of human SK-N-MC neurotumor and Caco-2 intestinal tumor cells to CT. Whereas chloroquine or monensin had little or no effect on CT stimulation of cyclic AMP accumulation, brefeldin A (BFA) totally inhibited the response to CT in a time- and dose-dependent and reversible manner. BFA was effective when added at the same time as CT and had an IC50 of 30 ng/ml. BFA did not alter cell surface GM1 as cells treated with BFA for 30 min bound as much 125I-CT as control cells. Furthermore, BFA inhibited CT stimulation of GM1-treated rat glioma C6 cells. BFA treatment did not affect beta-adrenergic agonist stimulation of cyclic AMP. In addition, adenylylcyclase was activated by A1 peptide and NAD+ to the same extent in membranes from control and BFA-treated cells, or when BFA was added directly to the assay. Whereas control cells generated small amounts of A1 from bound CT with time, no A1 was detected in BFA-treated cells. BFA treatment did not prevent the internalization of CT but did inhibit its degradation. BFA is known to disrupt the organization of the Golgi complex, resulting in inhibition of protein transport from the endoplasmic reticulum and redistribution of Golgi enzymes to the endoplasmic reticulum. BFA also prevents the formation of non-clathrin-coated vesicles from Golgi membranes and thus vesicular transport between Golgi cisternae. We confirmed that BFA caused the morphological disruption of the Golgi apparatus in Caco-2 cells. The data support a role for a functional Golgi apparatus with its associated vesicular routing in CT action.
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PMID:Brefeldin A blocks the response of cultured cells to cholera toxin. Implications for intracellular trafficking in toxin action. 838 69

The subcellular distribution of annexin V, a calcium-dependent phospholipid- and membrane-binding protein, in a human-derived cell line, GL15, was investigated by immunocytochemistry at light and electron microscope levels. Annexin V was found diffusely in the cytoplasm and associated with plasma membranes, membranes delimiting cytoplasmic vacuoles, membranes of the endoplasmic reticulum, and filamentous structures the identity of which remains to be established. By immunocytochemistry at the light microscope level and immunochemistry, the expression of annexin V in these cells was found to depend on cellular growth stage, being maximal soon after plating and progressively declining thereafter. However, re-expression of annexin V was observed whenever cell proliferation slowed down or arrested. These findings suggest that annexin V in glioma cells is mostly expressed in connection with cell differentiation. Also, the present ultrastructural data suggest that plasma membranes, membranes of the endoplasmic reticulum and the cytoskeleton are prominent sites of action of annexin V in vivo, thus lending support to the possibility that this protein might have a role in the regulation of cytoskeleton elements and/or of the structural organization of membranes.
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PMID:Immunocytochemical analyses of annexin V (CaBP33) in a human-derived glioma cell line. Expression of annexin V depends on cellular growth state. 849 46

GRP78, a molecular chaperone expressed in the endoplasmic reticulum, is a "glucose-regulated protein" induced by stress responses that deplete glucose or intracisternal calcium or otherwise disrupt glycoprotein trafficking. Previously we showed that chronic ethanol exposure increases the expression of GRP78. To further understand the mechanism underlying ethanol regulation of GRP78 expression, we studied the interaction between ethanol and classical modulators of GRP78 expression in NG108-15 neuroblastoma x glioma cells. We found that, in addition to increasing basal levels of GRP78 mRNA ("induction"), ethanol produced greater than additive increases in the induction of GRP78 mRNA by the "classical" GRP inducers A23187, brefeldin A, and thapsigargin ("potentiation"). Both the ethanol induction and potentiation responses modulated grp78 gene transcription as determined by stable transfection analyses with the rat grp78 promoter. Ethanol potentiated the action of all classical inducers of grp78 transcription that were studied. In contrast, co-treatment with the classical GRP inducers thapsigargin and tunicamycin produced only simple additive increases in grp78 promoter activity. Transient transfection studies with deletion mutants of the rat grp78 promoter showed that cis-acting promoter sequences required for ethanol induction differ from those mediating responses to classical GRP inducers. Furthermore, linker-scanning mutations of the grp78 promoter suggested that the ethanol potentiation response required a cis-acting promoter element different from those involved in induction by ethanol or classical inducing agents. While the ethanol induction response required 16-24 h to be detectable, ethanol potentiation of thapsigargin occurred within 6 h. The potentiation response also decayed rapidly after ethanol removal. In addition, the protein kinase A inhibitor Rp-cAMPS and protein phosphatase inhibitor okadaic acid both increased ethanol potentiation of thapsigargin while Sp-cAMPS, an activator of protein kinase A, decreased ethanol potentiation. Taken together, our findings suggest two mechanisms by which ethanol regulates grp78 transcription, both differing from the action of classical GRP inducers such as thapsigargin. One mechanism (potentiation) involves a protein phosphorylation cascade and potentiates the action of classical GRP inducers. In contrast, GRP78 induction by ethanol involves promoter sequences and a mechanistic pathway separate from that of the ethanol potentiation response or classical GRP78 inducers. These studies show that ethanol produces a novel and complex regulation of grp78 transcription which could be of particular importance during neuronal exposure to GRP-inducing stressors as might occur with central nervous system injury.
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PMID:Interaction of ethanol with inducers of glucose-regulated stress proteins. Ethanol potentiates inducers of grp78 transcription. 857 45

Long-term superfusion with bradykinin causes oscillations of cytosolic Ca2+ activity ([Ca2+]i) in Fura-2 loaded rat glioma cells. The [Ca2+]i rise is associated with synchronous plasma membrane hyperpolarization oscillating with a frequency of 0.8-1.8 per min. The initial large transient [Ca2+]i rise, induced immediately with bradykinin admission results from InsP3-mediated Ca2+ release, whereas the subsequent oscillations depend mainly on Ca2+ influx, as demonstrated: (i) by blockade of [Ca2+]i oscillations by reduction of [Ca2+]ex' or addition of Ca(2+)-channel blockers; and (ii) evidence from Mn2+ quench experiments. Suppression of [Ca2+]i oscillations with high K+ depolarization and with block of Ca(2+)-dependent K+ channels proves that membrane hyperpolarization is required for Ca2+ influx during the oscillation. Ca2+ release from intracellular stores by inhibitors of endoplasmic reticulum Ca(2+)-ATPase attenuates or blocks the [Ca2+]i oscillations. This suggests that bradykinin-induced Ca2+ influx is controlled by the filling state of the stores. The [Ca2+]i oscillations are suppressed by hypertonic medium and enhanced by hypotonic medium. Cell swelling enhances Ca2+ influx. We propose the following model for generation of the oscillations in the glial cell line: InsP3-induced Ca2+ release from internal stores periodically evokes Ca2+ influx through Ca(2+)-permeable cation channels. Hyperpolarization of the plasma membrane due to the activation of Ca(2+)-dependent K+ channels enhances the Ca2+ influx. The concomitant K+ efflux could lead to cell shrinkage which suppresses Ca2+ influx. Cell volume and membrane potential probably serve as feedback regulators during the [Ca2+]i oscillations.
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PMID:[Ca2+]i oscillations induced by bradykinin in rat glioma cells associated with Ca2+ store-dependent Ca2+ influx are controlled by cell volume and by membrane potential. 868 72

The effects of 2,5-di-tert-butylhydroquinone (DBHQ) and thimerosal on phosphatidylserine synthesis by the base exchange reaction and on calcium mobilization in intact glioma C6 cells were compared with that of thapsigargin, a selective inhibitor of the endoplasmic reticulum Ca(2+)-ATPase. It has been found that all these agents inhibit phosphatidylserine synthesis by 70%, but their effectiveness are different. The data show that this inhibition is caused by Ca2+ depletion of the endoplasmic reticulum, indicating that phosphatidylserine synthesis requires high concentration of Ca2+ within this structure. On this basis and on literature data, a new model for the localization of the serine base exchange enzyme in the endoplasmic reticulum membrane is proposed.
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PMID:Phosphatidylserine synthesis in glioma C6 cells is inhibited by Ca2+ depletion from the endoplasmic reticulum: effects of 2,5-di-tert-butylhydroquinone and thimerosal. 871 2

The effect of sphingosine on intracellular calcium signalling in glioma C6 cells was studied with Fura-2 video imaging technique. Sphingosine had a direct effect on changes in cytosolic Ca2+ concentration only when applied at high concentration of 100 microM, causing the cytosolic Ca2+ level to rise. However, at a much lower concentration of 15 microM sphingosine diminished calcium responses triggered by thapsigargin (a specific inhibitor of calcium pump in the endoplasmic reticulum) and ionomycin (calcium ionophore). Since responses to thapsigargin and ionomycin were blocked in Ca(2+)-free medium, we postulate that sphingosine is acting on the intracellular calcium stores. Additionally, sphingosine (at 15 microM and 100 microM) markedly decreases thapsigargin-induced sustained elevation in cytosolic Ca2+ concentration, indicating its inhibitory effect on thapsigargin-evoked Ca2+ influx. Sphingosine is a known inhibitor of protein kinase C and the involvement of this enzyme is postulated in the modulatory effects of sphingosine on intracellular calcium dynamics.
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PMID:Sphingosine stimulates calcium mobilization and modulates calcium signals evoked by thapsigargin in glioma C6 cells. 876

The intracellular localization of soluble and membrane-bound isoforms of rat and human catechol O-methyltransferase (COMT) was studied by expressing the recombinant COMT proteins either separately or together in mammalian cell lines (HeLa and COS-7 cells) and in rat primary neurons. The distribution of soluble and membrane-bound COMT enzyme was visualized by immunocytochemistry. For comparison, the localization of native COMT was studied in rat C6 glioma cells by immunoelectron microscopy. Staining of cells expressing membrane-bound COMT with a COMT-specific antiserum revealed an immunofluorescence signal in intracellular reticular structures and in the nuclear membrane. Double-staining of the cells with antisera against proteins specific for the rough endoplasmic reticulum indicated that they colocalized with membrane-bound COMT, suggesting that it resided in the endoplasmic reticulum. Notably, no COMT-specific fluorescence of plasma membranes was detected. The signal in the endoplasmic reticulum was also evident in the cells expressing both recombinant COMT forms. Intracellular native COMT reaction was detected by immunoelectron microscopy in rat C6 glioma cells and an intense cytoplasmic signal was seen in the primary neurons infected with the recombinant Semliki Forest virus. The cells expressing recombinant soluble COMT revealed intense nuclear staining together with diffuse cytoplasmic immunoreactivity, suggesting that a part of soluble COMT is transported to nuclei. Western blotting from rat liver and brain revealed soluble COMT in the nuclei. Enzyme activity measurements from liver cytoplasmic and nuclear fractions suggested that about 5% of the soluble COMT resided in nuclei. The intracellular localization of both COMT forms implies that COMT acts in the cytoplasm and possibly also in the nuclear compartment, and that the physiological substrates of COMT enzymes may have to be internalized before their methylation by COMT.
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PMID:Expression and intracellular localization of catechol O-methyltransferase in transfected mammalian cells. 903 Jul 72

The aim of the present study was to examine the possible role of protein kinase C (PKC) in thrombin-induced Ca2+ signalling. As shown before, continuous superfusion of rat glioma cells with thrombin caused sustained [Ca2+]i oscillations through activation of cell surface receptors [Czubayko U, Reiser G (1995) Neuroreport 6: 1249]. These oscillations were inhibited by protease nexin-1. Addition of PKC inhibitors, i. e. staurosporine (0.2-20 microM), bisindolylmaleimide (1 microM) or chelerythrine (1 microM), irreversibly suppressed thrombin-induced [Ca2+]i oscillations. Thereafter application of 2,5-di(tert-butyl)-1,4-benzohydroquinone (t-BuBHQ, 20 microM) or thapsigargin (1 microM) (inhibitors of sarco/endoplasmic reticulum Ca2+-ATPase) caused no [Ca2+]i response, indicating that intracellular Ca2+ stores were completely empty. We tested whether PKC affects the refilling of internal Ca2+ stores in thrombin-stimulated cells, by monitoring the amount of Ca2+ release caused by t-BuBHQ in the presence or absence of PKC inhibitors or activators. The amount of Ca2+ released by t-BuBHQ, which was normalized by comparison with the thrombin-induced Ca2+ response, was decreased by simultaneous incubation with staurosporine or chelerythrine, but enhanced with the PKC activator oleoyl acetyl glycerol. Furthermore, the capacitative Ca2+ entry was reduced by inhibition or downregulation, and increased by activation, of PKC. Capacitative Ca2+ entry was induced in these experiments by depletion of Ca2+ stores by the addition of thapsigargin or t-BuBHQ. In contrast, the inhibition of PKC during thrombin-induced depletion of intracellular stores did not influence the Ca2+ entry but nearly completely abolished the refilling of the internal stores. Thus we conclude that during thrombin receptor stimulation activation of PKC is required to maintain the refilling of intracellular Ca2+ stores for sustained [Ca2+]i oscillations. Thus, the control by PKC of the capacitative Ca2+ entry is apparently different depending on whether it is induced by sarco/endoplasmic reticulum Ca2+-ATPase inhibition or by activation of the thrombin receptor.
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PMID:Activity of protein kinase C is necessary for sustained thrombin-induced [Ca2+]i oscillations in rat glioma cells. 906 47


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