| 1. |
- Appelqvist, Hanna, et al.
(författare)
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Sensitivity to Lysosome-Dependent Cell Death is Directly Regulated by Lysosomal Cholesterol Content
- 2012
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Ingår i: PLOS ONE. - : Public Library of Science. - 1932-6203. ; 7:11
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Tidskriftsartikel (refereegranskat)abstract
- Alterations in lipid homeostasis are implicated in several neurodegenerative diseases, although the mechanisms responsible are poorly understood. We evaluated the impact of cholesterol accumulation, induced by U18666A, quinacrine or mutations in the cholesterol transporting Niemann-Pick disease type C1 (NPC1) protein, on lysosomal stability and sensitivity to lysosome-mediated cell death. We found that neurons with lysosomal cholesterol accumulation were protected from oxidative stress-induced apoptosis. In addition, human fibroblasts with cholesterol-loaded lysosomes showed higher lysosomal membrane stability than controls. Previous studies have shown that cholesterol accumulation is accompanied by the storage of lipids such as sphingomyelin, glycosphingolipids and sphingosine and an up regulation of lysosomal associated membrane protein-2 (LAMP-2), which may also influence lysosomal stability. However, in this study the use of myriocin and LAMP deficient fibroblasts excluded these factors as responsible for the rescuing effect and instead suggested that primarily lysosomal cholesterol content determined the cellular sensitivity to toxic insults. Further strengthening this concept, depletion of cholesterol using methyl-β-cyclodextrin or 25-hydroxycholesterol decreased the stability of lysosomes and cells became more prone to undergo apoptosis. In conclusion, cholesterol content regulated lysosomal membrane permeabilization and thereby influenced cell death sensitivity. Our data suggests that lysosomal cholesterol modulation might be used as a therapeutic strategy for conditions associated with accelerated or repressed apoptosis.
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| 2. |
- Fornander, Liselott, 1981-
(författare)
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How we talk : aspects of communication and team cognition of trauma resuscitation teams
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- A trauma team consists of professionals assembling in an ad hoc manner to resuscitate a patient with life-threatening injuries. In such a team, how the team manages to use non-technical skills, such as communication is important to achieve task management and decision making. Communication is practised in simulations, and the simulation environment has also been used in research to understand teamwork processes. This thesis is based on video observations of trauma teams working in real life (IRL) and in situ simulations of trauma resuscitations. The aim was to assess the creation of team cognition IRL and in simulation, to analyse verbal communication in the teams and the effects of real-time communication on team structure. In Study I, a grounded theory analysis was undertaken to understand how verbal and non-verbal interactions create team cognition. The analysis resulted in a theory pointing to “split vision” as a team’s ability to alter process modes between team positioning and sensitivity to the patient that was dependent on patient and situational values absorbed from team members’ implicit actions. In Study II, the verbal response modes taxonomy was applied to analyse both the grammatical and pragmatic meaning of verbal interactions that were compared between team roles. In the six most communicative team roles in the context of IRL trauma teamwork, pure mode communication dominated the way of delivering messages. In Study III, the structures of four IRL and four simulated trauma teams were analysed using a social network analysis of real-time communication. Overall, the teams were highly centralised, with the examining physician functioning as an information hub. In Study IV, communication from the same teams was categorised according to information and task management, as well as different coordination behaviours. We compared the IRL and simulation domains based on the proportions of utterances of each category/code, and found that “Give information after request” and closed-loop communication were more prevalent in simulation. Observing real-time communications using different methodologies gave a perspective on the conditions and possibilities for adaptation in terms of work prescriptions and team training, such as room for more communication and possible congestions of nodes in terms of occupancy and tacit communications and working modes thus far unnoticed by us and unreported. Such understanding complements established knowledge about coordination behaviours and interactive team cognition and should be taken into consideration in the practise and training of trauma teams.
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| 3. |
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| 4. |
- Andersson, Henrik, et al.
(författare)
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Orexin A Phosphorylates the gamma-Aminobutyric Acid Type A Receptor beta(2) Subunit on a Serine Residue and Changes the Surface Expression of the Receptor in SH-SY5Y Cells Exposed to Propofol
- 2015
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Ingår i: Journal of Neuroscience Research. - : WILEY-BLACKWELL. - 0360-4012 .- 1097-4547. ; 93:11, s. 1748-1755
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Tidskriftsartikel (refereegranskat)abstract
- Propofol activates the gamma-aminobutyric acid type A receptor (GABA(A)R) and causes a reversible neurite retraction, leaving a thin, thread-like structure behind; it also reverses the transport of vesicles in rat cortical neurons. The awakening peptide orexin A (OA) inhibits this retraction via phospholipase D (PLD) and protein kinase CE (PKCE). The human SH-SY5Y cells express both GABA(A)Rs and orexin 1 and 2 receptors. These cells are used to examine the interaction between OA and the GABAAR. The effects of OA are studied with flow cytometry and immunoblotting. This study shows that OA stimulates phosphorylation on the serine residues of the GABA(A)R beta(2) subunit and that the phosphorylation is caused by the activation of PLD and PKCE. OA administration followed by propofol reduces the cell surface expression of the GABA(A)R, whereas propofol stimulation before OA increases the surface expression. The GABA(A)R beta(2) subunit is important for receptor recirculation, and the effect of OA on propofol-stimulated cells may be due to a disturbed recirculation of the GABA(A)R. (C) 2015 Wiley Periodicals, Inc.
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| 5. |
- Björnström, Karin, 1971-, et al.
(författare)
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A tyrosine kinase regulates propofol-induced modulation of the beta-subunit of the GABA(A) receptor and release of intracellular calcium in cortical rat neurones
- 2002
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Ingår i: Acta Physiologica Scandinavica. - 0001-6772 .- 1365-201X. ; 175:3, s. 227-235
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Tidskriftsartikel (refereegranskat)abstract
- Propofol, an intravenous anaesthetic, has been shown to interact with the beta -subunit of the gamma -amino butyric acid(A) (GABA(A) ) receptor and also to cause changes in [Ca2+ ](i) . The GABA(A) receptor, a suggested target for anaesthetics, is known to be regulated by kinases. We have investigated if tyrosine kinase is involved in the intracellular signal system used by propofol to cause anaesthesia. We used primary cell cultured neurones from newborn rats, pre-incubated with or without a tyrosine kinase inhibitor before propofol stimulation. The effect of propofol on tyrosine phosphorylation and changes in [Ca2+ ](i) were investigated. Propofol (3 mu g mL(-1) , 16.8 mu M) increased intracellular calcium levels by 122 +/- 34% (mean +/- SEM) when applied to neurones in calcium free medium. This rise in [Ca2+ ](i) was lowered by 68% when the cells were pre-incubated with the tyrosine kinase inhibitor herbimycin A before exposure to propofol (P < 0.05). Propofol caused an increase (33 +/- 10%) in tyrosine phosphorylation, with maximum at 120 s, of the beta -subunit of the GABA(A) -receptor. This tyrosine phosphorylation was decreased after pre-treatment with herbimycin A (44 +/- 7%, P < 0.05), and was not affected by the absence of exogenous calcium in the medium. Tyrosine kinase participates in the propofol signalling system by inducing the release of calcium from intracellular stores and by modulating the beta -subunit of the GABA(A) -receptor.
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| 6. |
- Björnström, Karin, 1971-, et al.
(författare)
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Characterisation of the signal transduction cascade caused by propofol in rat neurons : From the GABAA receptor to the cytoskeleton
- 2008
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Ingår i: Journal of Physiology and Pharmacology. - 0867-5910 .- 1899-1505. ; 59:3, s. 617-632
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Tidskriftsartikel (refereegranskat)abstract
- The anaesthetic propofol interacts with the GABAA receptor, but its cellular signalling pathways are not fully understood. Propofol causes reorganisation of the actin cytoskeleton into ring structures in neurons. Is this reorganisation a specific effect of propofol as apposed to GABA, and which cellular pathways are involved? We used fluorescence-marked actin in cultured rat neurons to evaluate the percentage of actin rings caused by propofol or GABA in combination with rho, rho kinase (ROK), PI3-kinase or tyrosine kinase inhibitors, with or without the presence of extracellular calcium. Confocal microscopy was performed on propofol-stimulated cells and changes in actin between cellular compartments were studied with Western blot. Propofol (3 μg·ml-1), but not GABA (5 μM), caused transcellular actin ring formation, that was dependent on influx of extracellular calcium and blocked by rho, ROK, PI3-kinase or tyrosine kinase inhibitors. Propofol uses rho/ROK to translocate actin from the cytoskeleton to the membrane and its actin ring formation is dependent on an interaction site close to the GABA site on the GABAA receptor. GABA does not cause actin rings, implying that this is a specific effect of propofol.
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| 7. |
- Björnström, Karin, et al.
(författare)
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Rho and Rho Kinase are involved in the signal transduction cascade caused by propofol
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Background: Propofol is known to interact with the γ-aminobutyric acidA (GABAA) receptor, however, activating the receptor alone is not sufficient for producing anaesthesia. Propofol tyresine phosphorylates the GABAA receptor and reorganises the actin cytoskeleton, eausing ring structures and rnembrane ruffles. Propofol, but not GABA, the endogenous tigand for the GABAA receptor, tyresine phosphorylates actin, both in the membrane and cytoskeletal fractions of the neuron.Aim: How does propofol cause the actin reorganisation and is this a specific effect of propofol? Is the small membrane associated G-protein rho involved in the signal cascade towards the actin reorganisation?Methods: Westem blotting (WB) was used to visualize tyresine phosphorylated immunoprecipitated proteins and changes in actin between the different cellularcompartments after inhibition with rho (C3 exotoxin) and rho kinase (ROK) (HA-1077) inhibitors. Fluoreseenee mireoscopy after rhodamine-phalloidin labelling of actin was used to calculate the number of actin ring structures caused by propofol or GABA, in same experiments combined with pre-incubation with C3 exotoxin, HA- 1077 or the tyrosine kinase inhibitor Herbimycin A. Propofol-stimulated cells were studied with confocal microscopy.Results: Propofol eaused an increased tyresine phosphorylation, that was reduced by C3 exotoxin, of a 160 kDa protein after two minutes stimulation. The 160 kDa protein is still unidentified. The actin ring structures caused by propofol was shown with confocal microscopy to go almost through the entire cell. The amount of rings were reduced by C3 exatoxin as well as HA-1077. Furthermore, w hen a tyrosine kinase bioeker was used no ring structures were formed. However, GABA did not produce any ring structures. When the actin content of the cellular campartments were analysed, C3 exatoxin treated cells showed an increased amount of actin in the cytoskeletal fraction, simultaneausly with a decrease in both the membrane and the cytosol fractions. The ROK bioeker on ly eaused a reduction of actin in the cytosol/membrane fractions, but no increase was observed in the cytoskeleton.Conclusion: Propofol, but not GABA, eauses actin ring structures in neurons. Propofol uses the rho and rho kinase pathway to reorganize the actin cytoskeleton into ring structures, which is also dependent on a tyresine klnase. Propofol also eauses an unidentified rho dependent 160 kDa protein to be tyresine phosphorylated. The activation eaused by propofol of rho and rho kinase causes actin to be moved from the cytoskeleton to the cell membrane and cytosol. This reorganisation of actin might influence the GABAA receptor by keeping it open, thus allowing the cell to be hyperpolarized for longer time, and consequently maintain anaesthesia.
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| 8. |
- Björnström, Karin, 1971-, et al.
(författare)
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The difference between sleep and anaesthesia is in the intracellular signal : propofol and GABA use different subtypes of the GABAA receptor β subunit and vary in their interaction with actin
- 2003
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Ingår i: Acta Anaesthesiologica Scandinavica. - : Wiley. - 0001-5172 .- 1399-6576. ; 47:2, s. 157-164
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Tidskriftsartikel (refereegranskat)abstract
- Background: Propofol is known to interact with the γ-aminobutyric acidA (GABAA) receptor, however, activating the receptor alone is not sufficient for producing anaesthesia.Methods: To compare propofol and GABA, their interaction with the GABAA receptor β subunit and actin were studied in three cellular fractions of cultured rat neurons using Western blot technique.Results: Propofol tyrosine phosphorylated the GABAA receptor β2 (MW 54 and 56 kDa) and β3 (MW 57 kDa) subtypes. The increase was shown in both the cytoskeleton (β2(54) and β2(56) subtypes) and the cell membrane (β2(54) and β3 subtypes). Concurrently the 56 kDa β2 subtype was reduced in the cytosol. Propofol, but not GABA, also tyrosine phosphorylated actin in the cell membrane and cytoskeletal fraction. Without extracellular calcium available, the amount of actin decreased in the cytoskeleton, but tyrosine phosphorylation was unchanged. GABA caused increased tyrosine phosphorylation of β2(56) and β3 subtypes in the membrane and both β2 subtypes in the cytoskeleton but no cytosolic tyrosine phosphorylation.Conclusion: The difference between propofol and GABA at the GABAA receptor was shown to take place in the membrane, where the β2(54) was increased by propofol and instead the β2(56) subtype was increased by GABA. Only propofol also tyrosine phosphorylated actin in the cell membrane and cytoskeletal fraction. This interaction between the GABAA receptor and actin might explain the difference between anaesthesia and physiological neuronal inhibition.
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| 9. |
- Björnström, Karin, et al.
(författare)
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Volatile anesthetics cause changes in intracellular calcium, tyrosine phosphorylation and actin morphology
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Background: The cellular effects of anesthetics is poorly known. The GABAA receptor has been suggested as the main target for most anesthetics. In previous studies we have shown that propofol tyresine phosphorylates the GABAA receptor ß subunit, increases intracellular calcium and changes the actin morphology of neurons.Aim: To investigate the effects of the volatile anesthetics sevoflurane, isoflurane and nitmus oxide on changes in [Ca2+]i tyrosine phosphorylation and actin morphology in cultured rat neurons.Methods: Western blotting (WB) was used to visualize tyrosine phosphorylated proteins. Fluorescence microscopy after rhodamine-phalloidin labelling of actin was used to calculate the number of actin ring structures eaused by sevoflurane. Intracellular calcium was measured with the calcium-binding probe Fura-2 on single cells.Results: A protein of approx. 60 kDa increased dose-dependently in tyresine phosphorylation by sevoflurane in the membrane and cytoskeletal fractions, and was simultaneausly reduced in the cytosol. Isoflurane instead increased the tyresine phosphorylation of the same protein in the cytosol with only a slight increase in the membrane and no changes in the cytoskeletal fraction. Nitrous oxide did not cause any changes campared to air in the cytosol and was not detectable in the membrane. However, in the cytoskeletal fraction, the increase in tyrosine phosphorylation was high compared to air. Sevoflurane but not nitrous oxide or air increased the [Ca2+]i· Sevoflurane also eaused actin ring structures with a maximum after 20 minutes.Conclusion: Sevoflurane, isoflurane and nitrous oxide all have different signal pathways. The 60 kDa protein is probably the GABAA receptor ß subunit. According to the changes in tyrosine phosphorylation, changes in actin morphology and intracellular calcium, sevoflurane behaves most like the intravenous anesthetic propofol.
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| 10. |
- Björnström Karlsson, Karin, 1971-
(författare)
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Cellular mechanisms of anaesthetic agents
- 2003
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Anaesthesia is given to approximate 5% of the Swedish population annually, with the great advantage of painless surgery, but it also has side effects such as depression of blood pressure that might give a heart infarction. Exactly how anaesthetic agents cause anaesthesia is poorly known. Most anaesthetics have been shown to interact with the GABAA receptor, whose endogenous ligand GABA causes down-regulation of the brain and sleep. To further explore the cellular signal system used by anaesthetics this study was performed.First, two different malignant cell lines, PC-12 and SH-SY5Y, were tested, to evaluate if they could replace animal cells; however, they did not respond with increased intracellular calcium [Ca2+]i upon stimulation with propofol, as the normal rat neurons do. This is probably due to differences in the intracellular signaling systems in these malignant cells. Therefore, the studies in this thesis were performed on rat neurons.Propofol, an intravenous anaesthetic, was shown to cause a bicucullin insensitive increase in [Ca2+]i, where the release from intracellular stores was dependent on a tyrosine kinase. Sevoflurane, a volatile anaesthetic, also caused an ilrunediate increase in [Ca2+]i, but not nitrous oxide. Increased [Ca2+], is supposed to augment the influx of chloride ions through the GABAA receptor, hence hyperpolarising the neuron, and thereby make it anaesthetised.Tyrosine phosphorylation of the GABAA receptor is necessary for its function. Propofol tyrosine phosphorylates another ß2 subunit in the membrane then GABA. Propofol, but not GABA, also caused a tyrosine phosphorylation of actin in both the cytoskeletal and cell membrane fraction. Together these changes might explain the difference between sleep and anaesthesia. Isoflurane, sevoflurane and nitrous oxide all tyrosine phosphmylate a protein, suggested to be the GABAA receptor ß subunit, in different cellular compartments. This might explain their different clinical effects.Propofol and sevoflurane, but not GABA, causes actin rings to be formed in the cell, and for propofol the signal goes via rhoA and rho kinase, that also are involved in the translocation of actin to the cellular membrane. An unl~own 160 kDa protein is tyrosine phosphorylated by propofol, is part of the rho signalling pathway and is regulated by rho, This unknown protein might be involved in the actin reorganisation.
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