Biomedical subjects

Laura Korhonen

Publications and source records attributed to Laura Korhonen.

At least 19 recordsLinked to original sources

PubMed2006

Glucocorticoid hormones decrease proliferation of embryonic neural stem cells through ubiquitin-mediated degradation of cyclin D1.

Corticosteroids can influence brain function, and glucocorticoid hormone receptors (GRs) are present in brain tissue. We observed that GR and also mineralocorticoid receptor (MR) are expressed by embryonic rat neural stem cells (NSCs). NSCs in developing ventricular epithelium were positive for GR. Stimulation of cultured NSCs with the specific receptor ligands dexamethasone and corticosterone reduced cell proliferation, shown by 5'-bromo-2-deoxy-uridine labeling. The effect of the hormones was dose dependent and inhibited by the GR blocker mifepristone but not by spironolactone, blocking MR. Dexamethasone inhibited the cell cycle by decreasing the levels of cyclin D1 in NSCs. The hormone-induced decline was inhibited by MG132 (benzyloxycarbonyl-leucyl-leucyl-leucinal), showing an involvement of the ubiquitin proteasome system, In keeping with this, dexamethasone increased the ubiquitination of cyclin D1. In embryonic brain, dexamethasone inhibited cell proliferation of NSCs. This demonstrates that embryonic NSCs are critically influenced by glucocorticoids, which can have long-term effects in the brain.

Animals↗

PubMed2006

XIAP decreases caspase-12 cleavage and calpain activity in spinal cord of ALS transgenic mice.

Amyotrophic lateral sclerosis (ALS) is characterized by the selective degeneration of motor neurons. The cause for nerve cell demise is not clear but involves activation of the caspase family of cysteine proteases. We have shown that ER stress and caspase-12 activation occur in ALS transgenic mice carrying the mutant copper/zinc superoxide dismutase (SOD1) gene. In these mice, we found that the antiapoptotic proteins, X-linked Inhibitor of Apoptosis Protein (XIAP) and the related protein, MIAP2 were decreased. To study the role of this, we generated double transgenic mice expressing XIAP in ALS spinal cord neurons using the Thy1 promoter. Overexpression of XIAP inhibited caspase-12 cleavage and reduced calpain activity in the ALS mice. XIAP also reduced the breakdown of calpastatin that is an inhibitor of calpain. In the double transgenic mice, life span was increased by about 12%. These data support the view that XIAP has beneficial effects in ALS and extends survival. The neuroprotective effect of XIAP involves inhibition of caspases and the stabilization of the calpastatin/calpain system that is altered in the ALS mice.

Amyotrophic Lateral Sclerosis↗

PubMed2005

Downregulation of the HERG (KCNH2) K(+) channel by ceramide: evidence for ubiquitin-mediated lysosomal degradation.

The HERG (KCNH2) potassium channel underlies the rapid component of the delayed rectifier current (I(kr)), a current contributing to the repolarisation of the cardiac action potential. Mutations in HERG can cause the hereditary forms of the short-QT and long-QT syndromes, predisposing to ventricular arrhythmias and sudden cardiac death. HERG is expressed mainly in the cell membrane of cardiac myocytes, but has also been identified in cell membranes of a range of other cells, including smooth muscle and neurones. The mechanisms regulating the surface expression have however not yet been elucidated. Here we show, using stable HERG-expressing HEK 293 cells, that ceramide evokes a time-dependent decrease in HERG current which was not attributable to a change in gating properties of the channel. Surface expression of the HERG channel protein was reduced by ceramide as shown by biotinylation of surface proteins, western blotting and immunocytochemistry. The rapid decline in HERG protein after ceramide stimulation was due to protein ubiquitylation and its association with lysosomes. The results demonstrate that the surface expression of HERG is strictly regulated, and that ceramide modifies HERG currents and targets the protein for lysosomal degradation.

Cells, Cultured↗

PubMed2005

Bruce/apollon promotes hippocampal neuron survival and is downregulated by kainic acid.

Prolonged or excess stimulation of excitatory amino acid receptors leads to seizures and the induction of excitotoxic nerve cell injury. Kainic acid acting on glutamate receptors produces degeneration of vulnerable neurons in parts of the hippocampus and amygdala, but the exact mechanisms are not fully understood. We have here investigated whether the anti-apoptotic protein Bruce is involved in kainic acid-induced neurodegeneration. In the rat hippocampus and cortex, Bruce was exclusively expressed by neurons. The levels of Bruce were rapidly downregulated by kainic acid in hippocampal neurons as shown both in vivo and in cell culture. Caspase-3 was activated in neurons exhibiting low levels of Bruce causing cell death. Likewise, downregulation of Bruce using antisense oligonucleotides decreased viability and enhanced the effect of kainic acid in the hippocampal neurons. The results show that Bruce is involved in neurodegeneration caused by kainic acid and the downregulation of the protein promotes neuronal death.

Animals↗

PubMed2005

OX1 orexin receptors activate extracellular signal-regulated kinase in Chinese hamster ovary cells via multiple mechanisms: the role of Ca2+ influx in OX1 receptor signaling.

Activation of OX1 orexin receptors heterologously expressed in Chinese hamster ovary (CHO) cells led to a rapid, strong, and long-lasting increase in ERK phosphorylation (activation). Dissection of the signal pathways to ERK using multiple inhibitors and dominant-negative constructs indicated involvement of Ras, protein kinase C, phosphoinositide-3-kinase, and Src. Most interestingly, Ca2+ influx appeared central for the ERK response in CHO cells, and the same was indicated in recombinant neuro-2a cells and cultured rat striatal neurons. Detailed investigations in CHO cells showed that inhibition of the receptor- and store-operated Ca2+ influx pathways could fully attenuate the response, whereas inhibition of the store-operated Ca2+ influx pathway alone or the Ca2+ release was ineffective. If the receptor-operated pathway was blocked, an exogenously activated store-operated pathway could take its place and restore the coupling of OX1 receptors to ERK. Further experiments suggested that Ca2+ influx, as such, may not be required for ERK phosphorylation, but that Ca2+, elevated via influx, acts as a switch enabling OX1 receptors to couple to cascades leading to ERK phosphorylation, cAMP elevation, and phospholipase C activation. In conclusion, the data suggest that the primary coupling of orexin receptors to Ca2+ influx allows them to couple to other signal pathways; in the absence of coupling to Ca2+ influx, orexin receptors can act as signal integrators by taking advantage of other Ca2+ influx pathways.

Animals↗

PubMed2005

Hippocalcin protects against caspase-12-induced and age-dependent neuronal degeneration.

Hippocalcin is a neuronal calcium binding protein, but its physiological function in brain is unknown. We show here that hippocampal neurons from hippocalcin-deficient mice are more vulnerable to degeneration, particularly using thapsigargin, elevating intracellular calcium. Caspase-12 was activated in neurons lacking hippocalcin, while calpain was unchanged. Neuronal viability was accompanied by endoplasmic reticulum (ER) stress and a change in the relative induction of the ER chaperone, BiP/GRP78. Neuronal apoptosis inhibitor protein (NAIP), known to interact with hippocalcin, was not altered, but hippocampal neurons from gene-deleted mice were more sensitive to excitotoxicity caused by kainic acid. In addition, an age-dependent increase in neurodegeneration occurred in the gene-deleted mice, showing that hippocalcin contributes to neuronal viability during aging.

Aging↗

PubMed2005

19-Nortestosterone influences neural stem cell proliferation and neurogenesis in the rat brain.

Abuse of androgenic anabolic steroids can affect brain function leading to behavioural changes. In this study, the effects of the testosterone analogue, 19-nortestosterone, on rat neural stem cells was examined. The androgen receptor is expressed by cultured embryonic and adult neural stem cells, and is also present in the ventricular epithelium during development and in the adult brain in, among others, dentate gyrus. In neural stem cells stimulated with epidermal growth factor, nandrolone reduced cell proliferation, especially in adult ones. The decrease was abolished by flutamide, a receptor antagonist. Nandrolone also decreased the BrdU labelling of neural stem cells in the dentate gyrus, demonstrating an effect of the hormone on cell proliferation in vivo. The effect of nandrolone was observed with both female and male rats but it was more pronounced in pregnant rats, indicating an involvement of oestrogen in nandrolone action. Nandrolone also decreased the number of newly born neuronal cells in the dentate gyrus of male rats. The results show that nandrolone has important effects on the proliferation and differentiation of neural stem cells expressing the cognate androgen receptor. The data show that the use of nandrolone may severely affect the formation of neural stem cells and could therefore have long-term negative consequences in the brain.

Age Factors↗

PubMed2004

Caspase-12 cleavage and increased oxidative stress during motoneuron degeneration in transgenic mouse model of ALS.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by loss of motoneurons in the spinal cord and brain stem. We have characterized motoneuron death in transgenic mice carrying the mutant human copper/zinc superoxide dismutase, as a model for familial ALS. Previous studies have shown the involvement of mitochondria in nerve cell demise in these animals. We report here an early cleavage of caspase-12, residing in the endoplasmic reticulum (ER), in the spinal cord during the course of the disease. Apart from caspase-12, caspase-9, and caspase-3 were activated in the transgenic ALS mice. Staining with an antibody for nitrotyrosine, as a marker for oxidative stress, showed a large increase in the ALS mice. The results indicate that oxidative and ER induced stress causing caspase-12 activation are involved in neuronal death and disease progression in ALS. Caspase-12 and the ER pathway for cell death may constitute potential novel targets for the treatment of ALS.

Adaptation, Physiological↗

PubMed2004

Mitochondrial proteins in neuronal degeneration.

In this review, we highlight recent findings about the role of some mitochondrial proteins in neurological diseases. Studies in mice gene-deleted for Omi/HtrA2 and AIF showed the involvement of these mitochondrial proteins in selective cell degeneration in the spinal cord and brain. In humans, mutations in the mitochondrial protein, Paraplegin, cause an autosomal form of hereditary spastic paraplegia with an enhanced sensitivity to oxidative stress. Reactive oxygen species and decreased respiratory chain activity in mitochondria also contribute to common neurological diseases. The mitochondrial uncoupling protein, Ucp-2, was found to be neuroprotective in experimental stroke and brain trauma. Recent proteomic and profiling studies have revealed the existence of additional mitochondrial proteins with unknown functions. The elucidation of the physiological functions of mitochondrial proteins may lead to new insights into the role of these organelles in cell degeneration and to identification of novel drug targets for the prevention and treatment of different diseases.

ATPases Associated with Diverse Cellular Activitie↗

PubMed2004

Differential regulation of X-chromosome-linked inhibitor of apoptosis protein (XIAP) and caspase-3 by NMDA in developing hippocampal neurons; involvement of the mitochondrial pathway in NMDA-mediated neuronal survival.

N-methyl-d-aspartate (NMDA) subtype of glutamate receptors plays an important role during brain development and receptor activation has been shown to promote neuronal survival. The mechanisms by which NMDA increases neuronal survival are not fully understood. Here, we show that treatment with low concentrations of NMDA upregulates the neuroprotective molecule, X-chromosome-linked inhibitor of apoptosis protein (XIAP) in cultured hippocampal neurons at the post-transcriptional level. In contrast, NMDA treatment decreased mRNA and protein levels of caspase-3 in these neurons. The activation of the caspase-3 was also inhibited by NMDA, and the neurons were more resistant towards death caused by high concentrations of glutamate and staurosporine. Data on cytochrome c release in hippocampal neurons showed that NMDA pretreatment inhibits the mitochondrial pathway of cell death. The results demonstrate that the activation of NMDA receptors induces an anti-apoptotic program in hippocampal neurons that involves mitochondria and alterations of the levels of caspase-3 and XIAP. These findings are of importance for understanding the function of NMDA receptors in the control of neuronal survival during development and in other conditions.

Animals↗

PubMed2004

The ubiquitin proteasome system in synaptic and axonal degeneration: a new twist to an old cycle.

The ubiquitin proteasome system (UPS) contributes to the pathophysiology of neurodegenerative diseases, and it is also a major determinant of synaptic protein degradation and activity. Recent studies in rodents and in the fruit fly Drosophila have shown that the activity of the UPS is involved in axonal degeneration. Increased knowledge of the UPS in synaptic and axonal reactions may provide novel drug targets for treatments of neuronal injuries and neurodegenerative disorders.

Animals↗

PubMed2004

X-linked inhibitor of apoptosis (XIAP) protein protects against caspase activation and tissue loss after neonatal hypoxia-ischemia.

Nine-day-old transgenic XIAP overexpressing (TG-XIAP) and wild-type mice were subjected to left carotid artery ligation and 10% O(2) for 60 min, leading to widespread infarctions in the ipsilateral hemisphere during reperfusion. The activation of caspase-3 and -9 seen in wild-type animals was virtually abolished in TG-XIAP mice. Tissue loss was significantly reduced from 54.4 +/- 4.1 mm(3) (mean +/- SEM) in wild-type mice to 33.1 +/- 2.1 mm(3) in the TG-XIAP mice. Injured neurons displayed stronger XIAP staining during reperfusion, particularly in the nuclei. XIAP was colocalized with XAF-1, Smac, and HtrA2 in injured neurons after hypoxia-ischemia (HI). XIAP was cleaved after HI, and Smac immunoprecipitation co-precipitated a 25-kDa C-terminal fragment of XIAP, indicating that Smac preferentially bound to cleaved XIAP. These findings provide the first evidence that increased XIAP levels protect the neonatal brain against HI.

Animals↗

PubMed2003

Cystatin-B is expressed by neural stem cells and by differentiated neurons and astrocytes.

Mutation in the gene encoding cystatin-B (CSTB) has been shown to cause progressive myoclonus epilepsy. Mice with a gene deletion of CSTB exhibit increased apoptosis of specific neurons but the physiological role of CSTB in brain cells is not fully understood. In the present study, we have examined the expression of CSTB in neural stem cells (NSC) and in differentiating mature brain cells. The results show that CSTB is present in embryonic and adult NSC and in the neuroepithelium. CSTB was expressed by both neurons and glial cells differentiated from NSC and in hippocampal cultures. CSTB localized mainly to the nucleus in NSC and in neurons, whilst in astrocytes CSTB was also in the cytoplasm. Double labeling showed that CSTB was present in the lysosomes in glial cells. The results demonstrate a nuclear expression of CSTB in NSC and in neurons, suggesting novel function for this molecule.

Animals↗

PubMed2003

Regulation of sympathetic neuron and neuroblastoma cell death by XIAP and its association with proteasomes in neural cells.

XIAP (X chromosome-linked inhibitor of apoptosis protein) has been shown to inhibit cell death in a variety of cells. XIAP binds to active caspases, but XIAP also has a carboxy-terminal RING domain that can regulate cell death via protein degradation. Here we have studied the function of full-length and RING-deleted XIAP in mouse sympathetic neurons microinjected with expression plasmids and in neuroblastoma cells stably overexpressing these proteins. Both full-length and RING-deleted XIAP-protected sympathetic neurons against death induced by nerve growth factor (NGF) withdrawal to about the same extent. However, the two proteins were differentially localized in transfected neurons, with RING-deleted XIAP present in the cytoplasm and full-length XIAP found mostly in cytoplasmic protein aggregates, as revealed by transmission electron microscopy. The occurrence of these aggregates was blocked by lactacystin, a proteasome inhibitor. In neuroblastoma cells, RING-deleted XIAP protected against death induced by staurosporine, thapsigargin, or serum withdrawal, whereas full-length XIAP was without effect. Full-length, but not RING-deleted, XIAP was degraded and ubiquitinated in the neuroblastoma cells. The results show that the presence of the RING domain differentially affected the neuroprotective ability of XIAP in sensory neurons and neuroblastoma cells. The RING domain was essentially required for the proteasomal association of XIAP and for its ubiquitination.

Animals↗

PubMed2003

Transgenic mice overexpressing XIAP in neurons show better outcome after transient cerebral ischemia.

X-chromosome linked inhibitor of apoptosis protein (XIAP) is a member of the inhibitor of apoptosis protein (IAP) family and known to inhibit death of various cells under different experimental conditions. Although present in brain tissue, little is known about the physiology of the IAPs in nerve cells. Here we report on the establishment of transgenic mice with overexpression of human XIAP in brain neurons. The mice developed normally, and were more resistant to brain injury caused by transient forebrain ischemia after occlusion of the middle cerebral artery compared to control mice. The XIAP transgenic animals exhibited significantly smaller brain damage, as shown by TUNEL labelling, less reduction in brain protein synthesis, and less active caspase-3 after ischemia compared with controls. Upregulation of RhoB, which is an early indicator of neurological damage, was markedly reduced in the XIAP-overexpressing mice, which had also a better neurological outcome than control animals. This together with the increase in XIAP in normal mouse brain in regions surviving the infarct demonstrates that XIAP is an important factor promoting neuronal survival after ischemia. The results suggest that interference with the levels and the activity of XIAP in neurons may provide targets for the development of drugs limiting neuronal death after ischemia, and possibly in other brain injuries.

Animals↗

PubMed2003

Increase in Bcl-2 phosphorylation and reduced levels of BH3-only Bcl-2 family proteins in kainic acid-mediated neuronal death in the rat brain.

Kainic acid induces excitotoxicity and nerve cell degeneration in vulnerable regions of rat brain, most markedly in hippocampus and amygdala. Part of the cell death following kainic acid is apoptotic as shown by caspase 3 activation and chromatin condensation. Here we have studied the regulation of pro- and anti-apoptotic proteins belonging to the Bcl-2 family in rat hippocampus and amygdala by kainic acid in relationship to ensuing neuronal death. The pro-apoptotic protein Bax was up-regulated in hippocampus 6 h after kainic acid administration. The increase in Bax was followed by the appearance of TdT-mediated dUTP nick end labelling-positive cells which were prominent at 24 h. Immunohistochemistry for active Bax revealed a punctuated labelling of neurons in the CA3 and hilar regions of hippocampus as well as in amygdala. Double staining for NeuN, a marker for nerve cells, and TdT-mediated dUTP nick end labelling showed that mainly neurons undergo degeneration after kainic acid treatment. In contrast to Bax, the pro-apoptotic BH3-only Bcl-2 proteins Bim and Harakiri/DP5 were down-regulated by kainic acid. This was also observed for the anti-apoptotic proteins Bcl-x and Bcl-w. Immunoreactive Bcl-2 was up-regulated in hippocampus after kainic acid together with an increase in the phosphorylation of serine-87 in Bcl-2, suggesting a post-transcriptional modification of the protein. This was confirmed using immunoprecipitation of total Bcl-2 from hippocampus and amygdala which revealed an increase in serine-87 phospho-Bcl-2 after kainic acid. Inhibition of the c-jun N-terminal protein kinase pathway reduced both serine-87 phosphorylation and cell death after kainic acid. This indicates an important role of Bcl-2 phosphorylation in controlling neuronal death after kainic acid. In contrast to the situation in trophic factor-deprived neurons, no up-regulation of Bim or Harakiri/DP5 proteins occurred after kainic acid, suggesting alternative pathways for regulation of cell death in excitotoxicity. The results indicate that not only the relative levels of Bcl-2 family proteins but also conformation changes and post-translational modifications contribute to neuronal death following kainic acid.

Animals↗

PubMed2002

Nerve growth factor in serum of children with systemic lupus erythematosus is correlated with disease activity.

Nerve growth factor (NGF) is a neurotrophic factor, which is expressed both in the nervous system and in peripheral organs. NGF is also present in mast cells, and in B- and T-lymphocytes, and may play a role in the immune cell development and differentiation. Various cytokines have been shown to affect NGF expression, and NGF is elevated in inflammation and in some autoimmune diseases. Here we have studied NGF concentrations in serum of pediatric patients with systemic lupus erythematosus (SLE) using a two-site enzyme-linked immunosorbent assay (ELISA). We have further correlated the levels of NGF to the inflammatory state of the disease. The mean value of serum NGF in SLE patients was significantly increased compared with controls (3346 vs 627pg/ml). There was a correlation between the activity of SLE and the levels of NGF. The results show that NGF is elevated in childhood SLE and that the levels are correlated with disease activity. The present results suggest that NGF may play a role in the pathogenesis of SLE and may have a prognostic value in evaluating the course of the disease and in outlining the medication.

Adolescent↗

PubMed2002

Neuronal apoptosis inhibitory protein: Structural requirements for hippocalcin binding and effects on survival of NGF-dependent sympathetic neurons.

Neuronal apoptosis inhibitory protein (NAIP) has been linked to the inherited disease, spinal muscular atrophy (SMA), which occurs in children with degeneration of the motorneurons. In the nervous system, NAIP is expressed by specific classes of neurons including spinal motorneurons. Recently, NAIP was shown to interact with hippocalcin, which belongs to the neuronal calcium sensor (NCS) protein family. Here we have studied this interaction in more detail, using deletions and a mutagenesis of the third baculovirus inhibitory repeat (BIR) motif in NAIP, and functional assays for neuronal death. The results showed that specific amino acids and the zinc finger domain in BIR3 are needed for efficient interaction of NAIP with hippocalcin. Cotransfections of NAIP-BIR3 and hippocalcin resulted in translocation and colocalisation of the two proteins in neuroblastoma cells. This was accompanied by an enhanced resistance towards cell death induced by high levels of calcium. In contrast, expression of NAIP-BIR3 and hippocalcin in sympathetic neurons did not protect against death induced by nerve growth factor (NGF) withdrawal. The results demonstrate a functional interaction of hippocalcin with NAIP-BIR3, which in neuroblastoma cells leads to rescue of cells after high intracellular calcium, but which in sympathetic neurons had no significant effect. The results indicate that NAIP in conjunction with hippocalcin can affect the survival of some, but not all neural cells, and this interaction may play a role in the neurodegenerative processes in SMA, and possible other human disorders.

Animals↗