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Wulf Paschen

Publications and source records attributed to Wulf Paschen.

17 recordsLinked to original sources
PubMed2005

Cellular abnormalities linked to endoplasmic reticulum dysfunction in cerebrovascular disease--therapeutic potential.

Unfolded proteins accumulate in the lumen of the endoplasmic reticulum (ER) as part of the cellular response to cerebral hypoxia/ischemia and also to the overexpression of the mutant genes responsible for familial forms of degenerative diseases such as Alzheimer's disease, Parkinson's disease, Huntington's disease, amyothrophic lateral sclerosis, and Huntington's disease, as well as other disorders that are caused by an expanded CAG repeat. This accumulation arises from an imbalance between the load of proteins that need to be folded and processed in the ER lumen and the ER folding/processing capacity. To withstand such potentially lethal conditions, stress responses are activated that includes the shutdown of translation to reduce the ER work load and the activation of the expression of genes coding for proteins involved in the folding and processing reactions, to increase folding/processing capacity. In transient cerebral ischemia, ER stress-induced suppression of protein synthesis is believed to be too severe to permit sufficient activation of the genetic arm of the ER stress response. Mutations associated with Alzheimer's disease down-regulate the ER stress response and make cells more vulnerable to conditions associated with ER stress. When the functioning of the ER is severely impaired and affected cells can no longer withstand these stressful conditions, programmed cell death is induced, including a mitochondria-driven apoptotic pathway. Raising the resistance of cells to conditions that interfere with ER functions and activating the degradation and refolding of unfolded proteins accumulated in the ER lumen are possible strategies for blocking the pathological process leading to cell death at an early stage.

Animals↗
PubMed2005

Spinal cord trauma activates processing of xbp1 mRNA indicative of endoplasmic reticulum dysfunction.

The contribution of the various subcellular compartments in the induction of cell injury triggered by spinal cord trauma has not been clearly elucidated yet. In the present study, we investigated changes in mRNA levels of processed xbp1, ho-1, and hsp70 induced in mice by hemisection or contusion of the spinal cord. The expression of these genes is upregulated under conditions associated with endoplasmic reticulum (ER; xbp1, ho-1) dysfunction or impairment of cytoplasmic function (hsp70) respectively. When the functioning of the ER or the cytoplasm is impaired, unfolded proteins accumulate in these compartments. This is the warning signal for activation of the unfolded protein response (ER) and heat-shock response (cytoplasm) respectively. Spinal cord trauma activated the expression of these genes starting at 3 h and peaking at 6 h of recovery (processed xbp1, 12-fold and fivefold increase; ho-1, fourfold and eightfold increase; hsp70, fourfold, no increase, after contusion and hemisection, respectively). After 6 h of recovery, the rise in hsp70 mRNA levels was confined to the traumatized segment (fourfold), whereas a significant increase in processed xbp1 and ho-1 mRNA levels was also observed in the adjacent segments. This suggests a spread of the pathological process from the site of the primary impact into the surrounding tissue. After induction of spinal cord trauma processed xbp1 mRNA levels rose in a delayed fashion. This implies that the pathological process that causes impairment of ER functioning, starts with a delay of a few hours after induction of trauma and may therefore be amenable to therapeutic intervention.

Animals↗
PubMed2004

Spreading depression activates unfolded protein response.

Preconditioning is a process where a preceding non-lethal form of stress activates a stress response that protects cells against an otherwise lethal form of stress. Preconditioning can be induced in various ways including short-term ischemia or spreading depression. Here we investigated the effect of 1 h repetitive spreading depression on the unfolded protein response (UPR), a stress response activated under conditions associated with endoplasmic reticulum (ER) dysfunction. Spreading depression induced processing of xbp1 mRNA, indicative of an activation of UPR. Processing of xbp1 was paralleled by a rise in grp78 mRNA levels resulting from an activation of a signal transduction pathway that depends on protein synthesis. Preconditioning-induced activation of UPR may preserve ER functioning under pathological conditions interfering with ER functions.

Animals↗
PubMed2004

Brain trauma induces X-box protein 1 processing indicative of activation of the endoplasmic reticulum unfolded protein response.

Brain trauma was induced in mice using a closed head injury (CHI) model. At 1, 6 or 24 h after trauma, brains were dissected into the cortex, striatum and hippocampus. Changes in levels of processed X-box protein 1 (xbp1), glucose-regulated protein 78 (grp78), growth arrest and DNA damage-inducible gene 153 (gadd153) and heat-shock protein 70 (hsp70) mRNA, indicating impaired endoplasmic reticulum (ER) and cytoplasmic functioning, were evaluated by quantitative PCR. In the cortex, processed xbp1 mRNA levels rose to 2000% of control 1 h after CHI, and stayed high throughout the experiments. In the hippocampus and striatum, processed xbp1 mRNA levels rose in a delayed fashion, peaking at 6 h (1000% of control) and 24 h after CHI (1500% of control) respectively. Levels of grp78 mRNA were only slightly increased in the cortex 24 h after CHI (150% of control), and were unchanged or transiently decreased in the hippocampus and striatum. Levels of gadd153 mRNA did not change significantly after trauma. A transient rise in hsp70 mRNA levels was observed only in the cortex, peaking at 1 h after CHI (600% of control). Processing of xbp1 mRNA is a sign of activation of the unfolded protein response indicative of ER dysfunction. The results suggest that brain trauma induces ER dysfunction, which spreads from the ipsilateral cortex to the hippocampus and striatum. These observations may have clinical implications and should therefore be considered for future investigations on therapeutic intervention of brain injury caused by contusion-induced neurotrauma.

Animals↗
PubMed2004

GADD34 protein levels increase after transient ischemia in the cortex but not in the CA1 subfield: implications for post-ischemic recovery of protein synthesis in ischemia-resistant cells.

Transient cerebral ischemia is a pathological process whereby an irreversible suppression of protein synthesis is believed to contribute to the extent of cell death in vulnerable neurons. Endoplasmic reticulum (ER) dysfunction has been identified as being responsible for ischemia-induced shut-down of translation. Recovery from ER dysfunction is facilitated by GADD34, a protein that dephosphorylates eukaryotic initiation factor (eIF)2alpha-P and thus reactivates protein synthesis. We investigated ischemia-induced changes in GADD34 levels in wild-type and Cu2+/Zn2+ SOD (SOD1) over-expressing rats. Transient global cerebral ischemia was induced by common carotid artery occlusion. Tissue samples were taken from the vulnerable hippocampal CA1 subfield and the resistant cerebral cortex of the right and left hemispheres for evaluation of changes in gadd34 mRNA and GADD34 protein levels. In wild-type animals, we found significantly lower GADD34 levels than in SOD1 transgenes but no differences in gadd34 mRNA levels, implying that superoxides regulate gadd34 translation. After ischemia, GADD34 protein levels were significantly increased in the cortex but not in the CA1 subfield, and these changes occurred earlier in SOD1 transgenic than in wild-type animals. The rise in gadd34 mRNA levels did not differ in the cortex and CA1 subfield, implying that gadd34 expression is regulated at the translational level.

Animals↗
PubMed2004

Endoplasmic reticulum dysfunction in brain pathology: critical role of protein synthesis.

The endoplasmic reticulum (ER) is a subcellular compartment playing a central role in folding and processing membrane and secretory proteins. The importance of these reactions for normal cellular function is indicated by the fact that blocking of these processes is potentially lethal for cells. Under conditions associated with ER dysfunction, unfolded proteins accumulate in the ER lumen. This is the warning signal of two stress responses: the unfolded protein response (UPR) required for inducing the new synthesis of chaperons to refold the unfolded proteins, and the ER-associated degradation (ERAD) to degrade unfolded proteins at the proteasome. Cells in which UPR and ERAD cannot be activated to such an extent that ER function is restored die by apoptosis. In acute pathological states of the brain, including stroke, neurotrauma and epileptic seizures, and in degenerative diseases ER function is impaired in multiple ways. These include oxidative stress, nitric oxide-induced inactivation of the ER calcium pump resulting in disturbances of ER calcium homeostasis and impairment of UPR and ERAD. Furthermore, proteasomal function is impaired which causes secondary ER dysfunction. The only way to escape this potentially lethal cycle is to induce UPR and thus to activate new synthesis of ER chaperon GRP78 to levels sufficient to refold unfolded proteins. ER dysfunction may induce a state of tolerance, impair cellular functions, or induce apoptosis, depending on the severity and duration and the cell type affected. This review focuses on the possible role of ER dysfunction in the pathological process induced by transient cerebral ischemia.

Animals↗
PubMed2003

Loading neurons with BAPTA-AM activates xbp1 processing indicative of induction of endoplasmic reticulum stress.

Loading cells with the calcium chelator BAPTA-AM is an analytical tool which has been used to suppress a rise in cytoplasmic calcium activity under various experimental conditions and thus, to evaluate the role of elevated cytoplasmic calcium levels in the process under investigation. BAPTA-AM may, however, not only have an isolated effect on cytoplasmic processes but also on functions of other subcellular compartments such as the endoplasmic reticulum (ER). Under conditions associated with ER dysfunction, the unfolded protein response is activated which is characterized by suppression of translation and processing of xbp1 mRNA, resulting in activation of the expression of genes coding for ER stress proteins. To investigate whether BAPTA-AM causes ER stress, primary neuronal cell cultures were loaded with varying amounts of BAPTA-AM. Exposure of cells to BAPTA-AM induced a marked rise in processed xbp1 mRNA levels, correlating with exposure times and BAPTA-AM concentrations in the medium used for loading. The increase in processed xbp1 mRNA was associated with suppression of protein synthesis and induction of cell injury. The results of this study indicate that loading primary neuronal cell cultures with BAPTA-AM activates xbp1 processing, implying that this calcium chelator does not have an isolated effect on cytoplasmic calcium activity but also an affect on ER function.

Animals↗
PubMed2003

Conditions associated with ER dysfunction activate homer 1a expression.

Homer proteins physically link metabotropic glutamate receptors with IP3 receptors located at the endoplasmic reticulum (ER) and thereby modulate receptor-activated calcium signaling. Homer 1a, the short form of constitutively expressed homer 1 proteins, exerts dominant negative activity with respect to homer 1 proteins by interfering with the formation of multiprotein complexes. Homer 1a is an immediate early gene, the expression of which is activated by various stimuli including glutamate receptor activation. The mechanisms underlying activation of homer 1a expression are however, not fully understood. Here, we show that homer 1a expression is induced in neuronal cell cultures under experimental conditions associated with ER dysfunction. Increased homer 1a mRNA levels were found in 2 sets of cultures: in those exposed to thapsigargin, a specific inhibitor of ER Ca2+-ATPase, after a transient depletion of ER calcium stores through exposure to calcium-free medium supplemented with EGTA, and in those exposed to a proteasome inhibitor known to induce ER dysfunction. Thus, homer 1a expression may be activated by impairment of ER functioning just as it is by glutamate receptor activation.

Animals↗
PubMed2003

Transient cerebral ischemia activates processing of xbp1 messenger RNA indicative of endoplasmic reticulum stress.

Cells respond to conditions associated with endoplasmic reticulum (ER) dysfunction with activation of the unfolded protein response, characterized by a shutdown of translation and induction of the expression of genes coding for ER stress proteins. The genetic response is based on IRE1-induced processing of xbp1 messenger RNA (mRNA), resulting in synthesis of new XBP1proc protein that functions as a potent transcription factor for ER stress genes. xbp1 processing in models of transient global and focal cerebral ischemia was studied. A marked increase in processed xbp1 mRNA levels during reperfusion was observed, most pronounced (about 35-fold) after 1-h occlusion of the right middle cerebral artery. The rise in processed xbp1 mRNA was not paralleled by a similar increase in XBP1proc protein levels because transient ischemia induces severe suppression of translation. As a result, mRNA levels of genes coding for ER stress proteins were only slightly increased, whereas mRNA levels of heat-shock protein 70 rose about 550-fold. Under conditions associated with ER dysfunction, cells require activation of the entire ER stress-induced signal transduction pathway, to cope with this severe form of stress. After transient cerebral ischemia, however, the block of translation may prevent synthesis of new XBP1proc protein and thus hinder recovery from ischemia-induced ER dysfunction.

Animals↗
PubMed2003

Shutdown of translation: lethal or protective? Unfolded protein response versus apoptosis.

Shutdown of translation is a highly conserved response of cells to a severe form of metabolic, thermal, or physical stress. After the metabolic stress induced by transient cerebral ischemia, translational recovery is observed only in cells that withstand the transient interruption of blood supply, implying that restoration of translation critically determines the final outcome. On the other hand, apoptosis is believed to play a role in ischemia-induced cell death. Apoptosis is an active process that is blocked by agents known to suppress protein synthesis. Thus, the question arises whether stress-induced suppression of protein synthesis is protective or toxic for the affected cells. Accepting the notion that endoplasmic reticulum (ER) dysfunction is the mechanism underlying shutdown of translation after transient cerebral ischemia, an attempt may be made to try to solve the protein synthesis paradox by understanding the role of protein synthesis suppression in conditions associated with ER dysfunction. Endoplasmic reticulum dysfunction-induced accumulation of unfolded proteins in the ER lumen is the trigger of two signal transduction pathways: PKR-like ER kinase-induced shutdown of translation to suppress new synthesis of proteins that cannot be correctly folded, and IRE1-induced expression of ER stress genes, a protein synthesis-dependent pathway needed to restore ER functions. Together these comprise the unfolded protein response. They are also induced after transient ischemia, implying a dual effect of protein synthesis suppression, a protective and a pathologic effect during early and prolonged reperfusion.

Animals↗
PubMed2002

Down-regulation of parkin protein in transient focal cerebral ischemia: A link between stroke and degenerative disease?

Ubiquitylated protein aggregates are characteristic features of neurodegenerative disorders that are also found in acute pathological states of the brain such as stroke. Many of the proteins connected to neurodegenerative diseases play a role in the ubiquitin-proteasomal pathway. Mutation of one of these proteins, the E3 ubiquitin ligase parkin, is the cause of autosomal recessive juvenile Parkinson's disease. Here we show that transient focal cerebral ischemia of 1-h duration induces marked depletion of parkin protein levels, to 60%, 36%, 33%, and 25% of controls after 1, 3, 6, and 24 h of reperfusion, but that ischemia does not cause lower protein levels of E2 ubiquitin-conjugating enzymes Ubc6, Ubc7, or Ubc9. After 3 h of reperfusion, when parkin protein levels were already reduced to <40% of control, ATP levels were almost completely recovered from ischemia and we did not observe DNA fragmentation, suggesting that parkin depletion preceded development of neuronal cell death. Up-regulation of the expression of parkin has been shown to protect cells from injury induced by endoplasmic reticulum (ER) dysfunction, and this form of cellular stress is also triggered by transient cerebral ischemia. However, in contrast to observations in neuroblastoma cells, we saw no up-regulation of parkin expression in primary neuronal cell cultures after induction of ER dysfunction. Our data thus suggest that ischemia-induced depletion of parkin protein may contribute to the pathological process resulting in cell injury by increasing the sensitivity of neurons to ER dysfunction and the aggregation of ubiquitylated proteins during the reperfusion period.

Amino Acid Sequence↗
PubMed2002

Genes associated with pro-apoptotic and protective mechanisms are affected differently on exposure of neuronal cell cultures to arsenite. No indication for endoplasmic reticulum stress despite activation of grp78 and gadd153 expression.

The effect of arsenite exposure on cell viability, protein synthesis, energy metabolism and the expression of genes coding for cytoplasmic (hsp70) and endoplasmic reticulum (ER; gadd153, grp78, grp94) stress proteins was investigated in primary neuronal cell cultures. Furthermore, signs of ER stress were evaluated by investigating xbp1 mRNA processing. Arsenite levels of 30 and 100 microM induced severe cell injury. Protein synthesis was reduced to below 20% of control in cultures exposed to 30 and 100 microM arsenite for 1 h, and it remained markedly suppressed until 24 h of exposure. Arsenite induced a transient inhibition of energy metabolism after 1 h of exposure, but energy state recovered completely after 3 h. Arsenite exposure affected the expression and translation of genes coding for HSP70 and GRP78, GRP94, GADD153 to different extents. While hsp70 mRNA levels rose drastically, approximally 550-fold after 6 h exposure, HSP70 protein levels did not change over the first 6 h. On the other hand, gadd153 mRNA levels rose only approximately 14-fold after 6 h exposure, while GADD153 protein levels were markedly increased after 3 and 6 h exposure. HSP70 protein levels were markedly increased and GADD153 protein levels decreased to almost control levels in cultures left in arsenite solution for 24 h, i.e. when only a small fraction of cells had escaped arsenite toxicity. Arsenite exposure of neurons thus induced an imbalance between pro-apoptotic and survival-activating pathways. Despite the marked increase in gadd153 mRNA levels, we did not observe signs of xbp1 processing in arsenite exposed cultures, indicating that arsenite did not produce ER stress.

Animals↗
PubMed2002

Mechanisms underlying suppression of protein synthesis induced by transient focal cerebral ischemia in mouse brain.

Transient global cerebral ischemia triggers suppression of the initiation step of protein synthesis, a process which is controlled by endoplasmic reticulum (ER) function. ER function has been shown to be disturbed after transient cerebral ischemia, as indicated by an activation of the ER-resident eIF2alpha kinase PERK. In this study, we investigated ischemia-induced changes in protein levels and phosphorylation states of the initiation factors eIF2alpha, eIF2B epsilon, and eIF4G1 and of p70 S6 kinase, proteins playing a central role in the control of the initiation of translation. Transient focal cerebral ischemia was induced in mice by occlusion of the left middle cerebral artery. Transient ischemia caused a long-lasting suppression of global protein synthesis. eIF2alpha was transiently phosphorylated after ischemia, peaking at 1-3 h of recovery. eIF2B epsilon and p70 S6 kinase were completely dephosphorylated during ischemia and phosphorylation did not recover completely following reperfusion. In addition, eIF2B epsilon, eIF4G1, and p70 S6 kinase protein levels decreased progressively with increasing recirculation time. Thus, several different processes contributed to ischemia-induced suppression of the initiation of protein synthesis: a long-lasting dephosphorylation of eIF2B epsilon and p70 S6K starting during ischemia, a transient phosphorylation of eIF2alpha during early reperfusion, and a marked decrease of eIF2B epsilon, eIF4G1, and p70 S6K protein levels starting during vascular occlusion (eIF4G1). Study of the mechanisms underlying ischemia-induced suppression of the initiation step of translation will help to elucidate the role of protein synthesis inhibition in the development of neuronal cell injury triggered by transient cerebral ischemia.

Animals↗
PubMed2002

Phosphorylation state, solubility, and activity of calcium/calmodulin-dependent protein kinase II alpha in transient focal ischemia in mouse brain.

During and after middle cerebral artery occlusion in mice, CaMKII alpha protein was irreversibly translocated from the soluble to the Triton X-100-nonsoluble fraction. This decrease in solubility had a strong effect on activity: CaMKII alpha was almost completely inactivated after being translocated. Results from solubilization experiments suggest that different mechanisms underlie the conversion of CaMKII alpha protein from a soluble to a detergent nonsoluble form in ischemic as opposite to nonischemic tissue. Analysis of the phosphorylation state of CaMKII alpha revealed that in the total homogenate and the Triton X-100-nonsoluble fraction, CaMKII alpha phosphorylated at only one site was the dominant phosphorylated form, whereas in the soluble fraction CaMKII phosphorylated at two sites was the predominant phosphorylated species. Investigation of the mechanisms underlying ischemia-induced changes in the solubility of CaMKII alpha could help to elucidate processes triggered by transient focal cerebral ischemia that lead to neuronal cell injury.

Animals↗
PubMedDate not supplied

Endoplasmic reticulum stress response and neurodegeneration.

The endoplasmic reticulum (ER) is a subcellular compartment playing a central role in calcium storage and signaling. Disturbances of ER calcium homeostasis constitute a severe form of stress interfering with central functions of this structure including the folding and processing of newly synthesized membrane and secretory proteins. Blocking the folding and processing reactions results in the accumulation of unfolded proteins forming potentially toxic aggregates. To restore ER functioning, specific stress responses are activated one of which is the unfolded protein response (UPR). UPR is characterized by a shutdown of global protein synthesis and activation of expression of genes coding for ER-resident proteins that are involved in the folding and processing reactions. ER calcium homeostasis is therefore inevitably associated with major cellular functions, including gene transcription and translation. ER calcium homeostasis und ER functions are believed to be impaired in various degenerative diseases of the brain including Alzheimer's, Parkinson's and Huntington's disease, and amyotrophic lateral sclerosis. ER functioning has also been shown to be disturbed in acute pathological states of the brain such as ischemia and trauma, which have been identified as risk factors for the development of degenerative diseases. This implies that there are common underlying pathomechanisms. This review will summarize new observations suggesting that impairment of ER functioning may be a common denominator of pathological processes resulting in neuronal cell injury in acute disorders and degenerative diseases of the brain.

Animals↗
PubMedDate not supplied

Endoplasmic reticulum: a primary target in various acute disorders and degenerative diseases of the brain.

Changes in neuronal calcium activity in the various subcellular compartments have divergent effects on affected cells. In the cytoplasm and mitochondria, where calcium activity is normally low, a prolonged excessive rise in free calcium levels is believed to be toxic, in the endoplasmic reticulum (ER), in contrast, calcium activity is relatively high and severe stress is caused by a depletion of ER calcium stores. Besides its role in cellular calcium signaling, the ER is the site where membrane and secretory proteins are folded and processed. These calcium-dependent processes are fundamental to normal cell functioning. Under conditions of ER dysfunction unfolded proteins accumulate in the ER lumen, a signal responsible for activation of the unfolded protein response (UPR) and the ER-associated degradation (ERAD). UPR is characterized by activation of two ER-resident kinases, PKR-like ER kinase (PERK) and IRE1. PERK induces phosphorylation of the eukaryotic initiation factor (eIF2alpha), resulting in a shut-down of translation at the initiation step. This stress response is needed to block new synthesis of proteins that cannot be correctly folded, and thus to protect cells from the effect of unfolded proteins which tend to form toxic aggregates. IRE1, on the other hand, is turned after activation into an endonuclease that cuts out a sequence of 26 bases from the coding region of xbp1 mRNA. Processed xbp1 mRNA is translated into the respective protein, an active transcription factor specific for ER stress genes such as grp78. In acute disorders and degenerative diseases, the ER calcium pool is a primary target of toxic metabolites or intermediates, such as oxygen free radicals, produced during the pathological process. Affected neurons need to activate the entire UPR to cope with the severe form of stress induced by ER dysfunction. This stress response is however hindered under conditions where protein synthesis is suppressed to such an extent that processed xbp1 mRNA is not translated into the processed XBP1 protein (XBP1(proc)). Furthermore, activation of ERAD is important for the degradation of unfolded proteins through the ubiquitin/proteasomal pathway, which is impaired in acute disorders and degenerative diseases, resulting in further ER stress. ER functioning is thus impaired in two different ways: first by the direct action of toxic intermediates, produced in the course of the pathological process, hindering vital ER reactions, and second by the inability of cells to fully activate UPR and ERAD, leaving them unable to withstand the severe form of stress induced by ER dysfunction.

Acute Disease↗