| 1. |
- Lundqvist, C, et al.
(författare)
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Phosphatidylethanol affects inositol 1,4,5-trisphosphate levels in NG108-15 neuroblastoma x glioma hybrid cells
- 1993
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Ingår i: Journal of Neurochemistry. - : Wiley. - 1471-4159 .- 0022-3042. ; 60:2, s. 738-744
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Tidskriftsartikel (refereegranskat)abstract
- Phosphatidylethanol is formed by phospholipase D in animal cells exposed to ethanol. Previous reports have demonstrated that the degradation of phosphatidylethanol is slow, indicating that this lipid may be present in the cells after ethanol itself has disappeared. Accumulation of an abnormal alcohol metabolite may influence cellular functions. In the present study, cultivation of NG108-15 neuroblastoma x glioma hybrid cells in the presence of ethanol resulted in an accumulation of phosphatidylethanol and a simultaneous increase in basal inositol 1,4,5-trisphosphate levels. The direct effects of phosphatidylethanol on the phosphoinositide signal transduction system were examined through incorporation of exogenous phosphatidylethanol into membranes of ethanol-naive cells. An incorporation amounting to 2.8% of cellular phospholipids was achieved after a 5-h incubation with 30 microM phosphatidylethanol. Phosphatidylethanol was found to cause a time- and dose-dependent increase in the basal levels of inositol 1,4,5-trisphosphate. The effects on inositol 1,4,5-trisphosphate levels of exogenously added phosphatidylethanol and ethanol exposure for 2 days were not additive. No effect on bradykinin-stimulated inositol 1,4,5-trisphosphate production could be detected. However, the increase in basal inositol 1,4,5-trisphosphate levels indicates that phosphatidylethanol affects inositol 1,4,5-trisphosphate turnover and emphasizes the importance of considering phosphatidylethanol as a possible mediator of ethanol-induced effects on cellular processes.
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| 2. |
- Cardell, Monika, et al.
(författare)
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Time Course of the Translocation and Inhibition of Protein kinase C During Complete Cerebral Ischemia in the Rat
- 1993
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 61:4, s. 1308-1314
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Tidskriftsartikel (refereegranskat)abstract
- Abstract: The time course for the ischemia‐induced changes in the subcellular distribution of protein kinase C (PKC) (α), (β311). and (γ) and the activity of PKC were studied in the neocortex of rats subjected to 1, 2, 3, 5, 10, and 15 min of global cerebral ischemia. In the particulate fraction, a 14‐fold increase in PKC (γ) levels was seen at 3 min of ischemia, which further increased at 5–15 min of ischemia. At 15 min of ischemia, PKC (γ) and (βll) levels had increased two‐ and six‐fold, respectively. In the cytosolic fraction, a transient early 1.4‐fold increase in PKC (βll) and PKC (γ) levels was seen, whereas no change in the levels PKC (α) was noted. PKC (γ) levels then progressively declined, reaching 50% at 15 min of ischemia. At 5 min of ischemia, a 43% decrease in PKC activity was seen in the particulate fraction, reaching 50% at 15 min of ischemia concomitant with a 27% decrease in the cytosolic fraction. There was no change in the activator‐independent PKC activity. Pretreatment with the ganglioside AGF2 prevented the redistribution of PKC (γ) in the particulate fraction at 5 min. but not at 10 min of ischemia. The observed time course for the translocation of PKC (γ) parallels the ischemia‐induced release of neurotransmitters and increased levels of diacylglycerols, arachidonate, and intra‐cellular calcium and delineates this subspecies as especially ischemia‐sensitive. Ganglioside pretreatment delayed the translocation of PKC (γ), possibly by counteracting the effects of ischemia‐induced factors that favor PKC binding to cell membranes.
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| 3. |
- Hu, Bing Ren, et al.
(författare)
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Casein Kinase II Activity in the Postischemic Rat Brain Increases in Brain Regions Resistant to Ischemia and Decreases in Vulnerable Areas
- 1993
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 60:5, s. 1722-1728
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Tidskriftsartikel (refereegranskat)abstract
- Abstract: Casein kinase II (CKII) is a protein kinase acting in the intracellular cascade of reactions activated by growth factor receptors, and that has a profound influence on cell proliferation and survival. In this investigation, we studied the changes in the activity and levels of CKII in the rat brain exposed to 10. 15 and 20 min of transient forebrain ischemia followed by variable periods of reperfusion. The cytosolic CKII activity decreased during reperfusion by ∼ 30 and ∼ 50% in the selectively vulnerable areas, striatum and the CA1 region of the hippocampus, respectively. In the resistant CA3 region of hippocampus and neocortex, the activity increased by ∼ 20 and ∼ 60%, respectively. The postischemic changes in CKII activity were dependent on the duration of the ischemic insult. The levels of CKII did not change after ischemia, suggesting that the enzyme is modulated by covalent modification or is interacting with an endogenous inhibitor/activator. Treatment of the cytosolic fraction from cortex of rats exposed to ischemia and 1 h of reperfusion with agarose‐bound phosphatase decreased the activity of CKII to control levels, suggesting that CKII activation after ischemia involves a phosphorylation of the enzyme. The correlation between postischemic CKII activity and neuronal survival implies that preservation or activation of CKII activity may be important for neuronal survival after cerebral ischemia.
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| 4. |
- Hu, Bing Ren, et al.
(författare)
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Depression of Neuronal Protein Synthesis Initiation by Protein Tyrosine Kinase Inhibitors
- 1993
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 61:5, s. 1789-1794
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Tidskriftsartikel (refereegranskat)abstract
- Abstract— Growth factors stimulate cellular protein synthesis, but the intracellular signaling mechanisms that regulate initiation of mRNA translation in neurons have not been clarified. A rate‐limiting step in the initiation of protein synthesis is the formation of the ternary complex among GTP, eukaryotic initiation factor 2 (elF‐2), and the initiator tRNA. Here we report that genistein, a specific tyrosine kinase inhibitor, decreases tyrosine kinase activity and the content of phosphotyrosine proteins in cultured primary cortical neurons. Genistein inhibits protein synthesis by >80% in a dose‐dependent manner (10–80 μg/ml) and concurrently decreases ternary complex formation by 60%. At the doses investigated, genistein depresses tyrosine kinase activity and concomitantly stimulates PKC activity. We propose that a protein tyrosine kinase participates in the initiation of protein synthesis in neurons, by affecting the activity of elF‐2 directly or through a protein kinase cascade.
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| 5. |
- Karlsson, J E, et al.
(författare)
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Glial and neuronal marker proteins in the silicone chamber model for nerve regeneration
- 1993
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 60:3, s. 1098-1104
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Tidskriftsartikel (refereegranskat)abstract
- In the present study, neuronal and Schwann cell marker proteins were used to biochemically characterize the spatiotemporal progress of degeneration/regeneration in the silicone chamber model for nerve regeneration. Rat sciatic nerves were transected and the proximal and distal stumps were inserted into a bridging silicone chamber with a 10-mm interstump gap. Using dot immunobinding assays, S-100 protein and neuronal intermediate filament polypeptides were measured in different parts of the nerve 0-30 days after transection. In the most proximal nerve segment, all the measured proteins were transiently increased. In the proximal and distal stumps adjacent to the transection, the studied proteins were decreased indicating degeneration of the nerve. Within the silicone chamber, the regenerating nerve expressed the Schwann cell S-100 protein already at 7 days, whereas the neurofilament polypeptides appeared later. These observations are corroborated by previous morphological studies. The biochemical method described provides a new and fast approach to the study of nerve regeneration.
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| 6. |
- Kurihara, Junichi, et al.
(författare)
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Changes in the Tyrosine Phosphorylation of Mitogen‐Activated Protein Kinase in the Rat Hippocampus During and Following Severe Hypoglycemia
- 1994
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 63:6, s. 2346-2348
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Tidskriftsartikel (refereegranskat)abstract
- Abstract: The changes in the levels of tyrosine‐phosphorylated proteins in the cytosolic fraction of the rat hippocampus subjected to severe hypoglycemia were analyzed. A marked increase in tyrosine phosphorylation of a 43‐kDa protein was observed at 30 min of isoelectric EEG and 30 min and 1 h of recovery. Immunostaining of the same blot with antibody against mitogen‐activated protein (MAP) kinase demonstrated a double band of ∼42 and 43 kDa. The increased tyrosine phosphorylation of MAP kinase during hypoglycemic coma and the early recovery period suggests that MAP kinase may be involved in neuronal degeneration and repair.
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| 7. |
- Larsson, Christer, et al.
(författare)
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Activation of protein kinase C in permeabilized human neuroblastoma SH-SY5Y cells
- 1992
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Ingår i: Journal of Neurochemistry. - : Wiley. - 1471-4159 .- 0022-3042. ; 59:2, s. 644-651
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Tidskriftsartikel (refereegranskat)abstract
- The activation of protein kinase C was investigated in digitonin-permeabilized human neuroblastoma SH-SY5Y cells by measuring the phosphorylation of the specific protein kinase C substrate myelin basic protein4-14. The phosphorylation was inhibited by the protein kinase C inhibitory peptide PKC19-36 and was associated to a translocation of the enzyme to the membrane fractions of the SH-SY5Y cells. 1,2-Dioctanoyl-sn-glycerol had no effect on protein kinase C activity unless the calcium concentration was raised to concentrations found in stimulated cells (above 100 nM). Calcium in the absence of other activators did not stimulate protein kinase C. Phorbol 12-myristate 13-acetate was not dependent on calcium for the activation or the translocation of protein kinase C. The induced activation was sustained for 10 min, and thereafter only a small net phosphorylation of the substrate could be detected. Calcium or dioctanoylglycerol, when applied alone, only caused a minor translocation, whereas in combination a marked translocation was observed. Arachidonic acid (10 microM) enhanced protein kinase C activity in the presence of submaximal concentrations of calcium and dioctanoylglycerol. Quinacrine and p-bromophenacyl bromide did not inhibit calcium- and dioctanoylglycerol-induced protein kinase C activity at concentrations which are considered to be sufficient for phospholipase A2 inhibition.
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| 8. |
- Remgård, Pär, et al.
(författare)
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A Fast Axonally Transported Protein of the Frog Sciatic Sensory Axons Undergoes Similar Qualitative Changes During Regeneration In Vitro and In Vivo
- 1991
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 57:6, s. 1907-1912
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Tidskriftsartikel (refereegranskat)abstract
- The adult frog sciatic sensory neurons have been shown to regenerate in vitro. If a crush injury is made at the beginning of culture, regeneration starts after 3.4 days and proceeds at a rate of ∼0.8 mm/day for several days. Two‐dimensional gel electrophoresis was used to study the patterns of radiolabeled, fast axonally transported proteins during the first 7 days of regeneration. Interest was focused on one protein, referred to as rrp31 (regeneration‐related protein 31), which changed in apparent pl from 4.9 to 5.3 when the outgrowth of new fibers started. The change was noticeable 3 days after injury and became prominent during day 5 of culturing. By day 7 the pl changed again, this time toward the original value. The in vitro results were supported by experiments in vivo. In this case the change occurred earlier, with a peak only 3 days after injury, after which the pl decreased. If adenosine at 1 mM was included in the culturing medium, the outgrowth of sensory axons was inhibited in a nontoxic way, and the pl changes of rrp31 were prevented. The temporal nature of the pl changes suggests a role for rrp31 in the initiation of the regeneration process.
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| 9. |
- Simonsson, Per, et al.
(författare)
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G proteins coupled to phospholipase C: molecular targets of long-term ethanol exposure
- 1991
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Ingår i: Journal of Neurochemistry. - : Wiley. - 1471-4159 .- 0022-3042. ; 56:6, s. 2018-2026
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Tidskriftsartikel (refereegranskat)abstract
- Long-term ethanol exposure is known to inhibit bradykinin-stimulated phosphoinositide hydrolysis in cultures of neuroblastoma x glioma 108-15 cells. In the present study, [3H]bradykinin binding, GTP-binding protein function, and phospholipase C activity were assayed in cells grown for 4 days in 100 mM ethanol with the aim of elucidating the molecular target of ethanol on signal transduction coupled to inositol trisphosphate and diacylglycerol formation. Ethanol exposure reduced guanosine 5'-O-(3-thiotriphosphate) [GTP(S)]- and, to a lesser extent, NaF/AlCl3-stimulated phosphoinositide hydrolysis, whereas it had no effect on the enzymatic activity of a phosphatidylinositol 4,5-bisphosphate-specific phospholipase C. [3H]Bradykinin binding in the absence of GTP(S) was not influenced by ethanol exposure. However, the reduction in [3H]bradykinin binding seen in control cells after addition of GTP analogue was inhibited in cells grown in ethanol-containing medium. The results indicate that long-term ethanol exposure exerts its effects on receptor-stimulated phosphoinositide hydrolysis primarily at the level of the GTP-binding protein.
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| 10. |
- Wieloch, T., et al.
(författare)
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Changes in the Activity of Protein Kinase C and the Differential Subcellular Redistribution of Its Isozymes in the Rat Striatum During and Following Transient Forebrain Ischemia
- 1991
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 56:4, s. 1227-1235
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Tidskriftsartikel (refereegranskat)abstract
- Abstract: The changes in the levels of protein kinase C [PKC(α, βII, γ)] were studied in cytosolic and particulate fractions of striatal homogenates from rats subjected to 15 min of cerebral ischemia induced by bilateral occlusion of the common carotid arteries and following 1 h, 6 h, and 48 h of reperfusion. During ischemia the levels of PKC(βII) and ‐(γ) increased in the particulate fraction to 390% and 590% of control levels, respectively, concomitant with a decrease in the cytosolic fraction to 36% and 20% of control, respectively, suggesting that PKC is redistributed from the cytosol to cell membranes. During reperfusion the PKC(βII) levels in the particulate fraction remained elevated at 1 h postischemia and decreased to below control levels after 48 h reperfusion, whereas PKC(γ) rapidly decreased to subnormal levels. In the cytosol PKC(βII) and ‐(γ) decreased to 25% and 15% of control levels at 48 h, respectively. The distribution of PKC(α) did not change significantly during ischemia and early reperfusion. The PKC activity in the particulate fraction measured in vitro by histone IIIS phosphorylation in the presence of calcium, 4β‐phorbol 13‐myristate 12‐acetate, and phosphatidylserine (PS) significantly decreased by 52% during ischemia, and remained depressed over the 48‐h reperfusion period. In the cytosolic fraction PKC activity was unchanged at the end of ischemia, and decreased by 47% after 6 h of reperfusion. The appearance of a stable cytosolic 50‐kDa PKC‐immunoreactive peptide or an increase in the calcium‐and PS‐independent histone IIIS phosphorylation was not observed. Consequently, during ischemia PKC, preferentially PKC(γ) and PKC(βII), is translocated from the cytosol and inserted into cell membranes, concomitant with a decrease in PKC activity. In the reperfusion phase the depression of PKC activity persists and the enzyme is degraded. The observed translocation and downregulation of PKC during ischemia and reperfusion may be of significance for the development of ischemic neuronal damage.
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