Külli Jaako

Dehydroepiandrosterone sulphate prevents oxygen–glucose deprivation-induced injury in cerebellar granule cell culture

Decreased levels of dehydroepiandrosterone sulphate have been hypothesized to contribute to increased vulnerability of the ageing or stressed human brain to ischemia. To help to address the question of whether of dehydroepiandrosterone sulphate has a possible neuroprotective effect against ischemic neuronal injury, we tested its effect on the neurodegeneration induced by oxygen-glucose deprivation in rat cultured cerebellar granule cells. Dehydroepiandrosterone sulphate added to the medium after injury demonstrated a neuroprotective effect with a median inhibitory concentration of 0.5 microM. At 10 microM concentration almost full neuroprotection was observed. Even more pronounced neuroprotective effect was found when dehydroepiandrosterone sulphate was added for 48h before injury. Furthermore, partial neuroprotection of dehydroepiandrosterone sulphate was also found against 1-methyl-4-phenylpyridinium, colchicine, glutamate and N-methyl-D-aspartate-induced toxicity. Further analysis demonstrated that dehydroepiandrosterone sulphate eliminated the apoptotic features of the oxygen-glucose deprivation-induced neuronal death: DNA fragmentation and nuclear condensation/fragmentation.Thus, our data suggest that dehydroepiandrosterone sulphate may have therapeutic potential in the prevention and treatment of ischemic/hypoxic neuronal damage. The neuroprotective action of dehydroepiandrosterone sulphate was inhibited by both a GABA(A) receptor-linked chloride channel agonist and an antagonist, pentobarbital and picrotoxin, respectively. It seems that GABA(A) receptor-mediated neuronal inhibition as well as neuronal excitation can reduce the neuroprotective action of dehydroepiandrosterone sulphate.

Neurodegeneration and production of the new cells in the dentate gyrus of juvenile rat hippocampus after a single administration of ethanol

Administration of ethanol during brain development induces widespread neuronal loss in various structures of the brain. Here, we show that a single administration of ethanol given during the early postnatal period can induce not only neuronal death, but also an increase in proliferation of the progenitor cells in the dentate gyrus of hippocampal formation in rats. Ethanol (1.5 or 3 g/kg, i.p.) administered to 10-day-old rats induced massive neuronal degeneration as evidenced by TUNEL assay in the dentate gyrus. The neuronal death induced by a high dose of ethanol (3 g/kg) was accompanied by an enhanced proliferation of the progenitor cells labeled by bromodeoxyuridine (BrdU, 50 mg/kg, i.p.) in dentate gyrus. One and 3 weeks following ethanol or saline administration, ethanol-treated rats still had significantly more BrdU-labeled cells than control animals. In ethanol-treated rats, a higher proportion of newly born cells acquired the phenotype of immature postmitotic neurons whereas the final differentiation into calbindin-expressing granule cells remained unchanged. The proportion of astroglial cells was also increased in ethanol-treated rats. Thus, ethanol given in high doses not only induces neurodegeneration but also initiates the process of neuro- and gliogenesis, which might be responsible for the neuronal and glial reorganization of the dentate gyrus.

Effects of repeated citalopram treatment on kainic acid-induced neurogenesis in adult mouse hippocampus

Previous studies have demonstrated that systemic administration of kainic acid (KA) triggers a cascade of neuroplastic changes in the hippocampus. Intensive neurodegeneration accompanied by immune response and enhanced neurogenesis following local or systemic KA administration in rats and mice has been reported. KA-induced enhancement in proliferative activity of neuronal and glial precursors results in the appearance of immature hyperactive neurons which could be regarded as evidence of dysregulated neural plasticity. In this study we attempted to investigate whether administration of selective serotonin reuptake inhibitor (SSRI) citalopram could inhibit KA-induced reactive gliosis and dysregulated neurogenesis in mice. The results of our study demonstrate that repeated administration of citalopram counteracted KA-induced reactive gliosis and reduced aberrant proliferative activity in the dentate gyrus of the mouse brain. We found that the population of BrdU-positive cells expressing markers for young neurons was decreased following repeated citalopram administration compared to KA-treated animals. These results suggest that repeated citalopram administration could prevent activation of aberrant neuroplasticity in the damaged hippocampus.

Deficiency of prolyl oligopeptidase in mice disturbs synaptic plasticity and reduces anxiety-like behaviour, body weight, and brain volume.

Prolyl oligopeptidase (PREP) has been implicated in neurodegeneration and neuroinflammation and has been considered a drug target to enhance memory in dementia. However, the true physiological role of PREP is not yet understood. In this paper, we report the phenotyping of a mouse line where the PREP gene has been knocked out. This work indicates that the lack of PREP in mice causes reduced anxiety but also hyperactivity. The cortical volumes of PREP knockout mice were smaller than those of wild type littermates. Additionally, we found increased expression of diazepam binding inhibitor protein in the cortex and of the somatostatin receptor-2 in the hippocampus of PREP knockout mice. Furthermore, immunohistochemistry and tail suspension test revealed lack of response of PREP knockout mice to lipopolysaccharide insult. Further analysis revealed significantly increased levels of polysialylated-neural cell adhesion molecule in PREP deficient mice. These findings might be explained as possible alteration in brain plasticity caused by PREP deficiency, which in turn affect behaviour and brain development.

Neuroprotective and memory enhancing properties of a dual agonist of the FGF receptor and NCAM.

The fibroblast growth factor receptor (FGFR) plays a vital role in the development of the nervous system regulating a multitude of cellular processes. One of the interaction partners of the FGFR is the neural cell adhesion molecule (NCAM), which is known to play an important role in neuronal development, regeneration and synaptic plasticity. Thus, simultaneous activation of FGFR- and NCAM-mediated signaling pathways may be expected to affect processes underlying neurodegenerative diseases. We here report the identification of a peptide compound, Enreptin, capable of interacting with both FGFR and NCAM. We demonstrate that this dual specificity agonist induces phosphorylation of FGFR and differentiation and survival of primary neurons in vitro, and that these effects are inhibited by abrogation of both NCAM and FGFR signaling pathways. Furthermore, Enreptin crosses the blood-brain barrier after subcutaneous administration, enhances long-term memory in normal mice and ameliorates memory deficit in mice with induced brain inflammation. Moreover, Enreptin reduces cognitive impairment and neuronal death induced by Aβ25-35 in a rat model of Alzheimers disease, and reduces the mortality rate and clinical signs of experimental autoimmune encephalomyelitis in rats. Thus, Enreptin is an attractive candidate for the treatment of neurological diseases.

Pharmacological approach for targeting dysfunctional brain plasticity: Focus on neural cell adhesion molecule (NCAM)

Brain plasticity refers to the ability of the brain to undergo functionally relevant adaptations in response to external and internal stimuli. Alterations in brain plasticity have been associated with several neuropsychiatric disorders, and current theories suggest that dysfunctions in neuronal circuits and synaptogenesis have a major impact in the development of these diseases. Among the molecules that regulate brain plasticity, neural cell adhesion molecule (NCAM) and its polysialylated form PSA-NCAM have been of particular interest for years because alterations in NCAM and PSA-NCAM levels have been associated with memory impairment, depression, autistic spectrum disorders and schizophrenia. In this review, we discuss the roles of NCAM and PSA-NCAM in the regulation of brain plasticity and, in particular, their roles in the mechanisms of depression. We also demonstrate that the NCAM-mimetic peptides FGL and Enreptin are able to restore disrupted neuronal plasticity. FGL peptide has also been demonstrated to ameliorate the symptoms of depressive-like behavior in NCAM-deficient mice and therefore, may be considered a new drug candidate for the treatment of depression as well as other neuropsychiatric disorders with disrupted neuroplasticity.

PSA modification of NCAM supports the survival of injured retinal ganglion cells in adulthood.

Neural cell adhesion molecule (NCAM) is known as the cell surface glycoprotein, and it belongs to the immunoglobulin superfamily of adhesion molecules. Polysialic acid (PSA) is a carbohydrate attached to NCAM via either of two specific sialyltransferases: ST8SiaII and ST8SiaIV. Polysialylated neural cell adhesion molecule (PSA-NCAM) mediates cell interactions, plays a role in axon growth, migration, synaptic plasticity during development and cell regeneration. Some evidence has shown that PSA-NCAM supports the survival of neurons. It was demonstrated that PSA-NCAM is present in abundance in the retina during development and in adulthood. The aim of this study was to investigate whether PSA-NCAM promotes retinal ganglion cell (RGC) survival in transgenic mice with deficiencies in sialyltransferases or NCAM or after the administration of endoneuraminidase (Endo-N). RGC injury was induced by intravitreal administration of kainic acid (KA). These studies showed that injection of Endo-N after 14 days enhances the toxicity of KA to RGCs in wild-type (WT) mice by 18%. In contrast, in knockout mice (ST8SiaII-/-, ST8SiaIV-/-, NCAM-/-), survival of RGCs after KA injury did not change. Deficiencies of either ST8SiaII or ST8SiaIV did not influence the level of PSA-NCAM in the adult retina, however, in neonatal animals, decreased levels of PSA-NCAM were observed. In knockout ST8SiaII-/- adults, a reduced number of RGCs was detected, whereas in contrast, increased numbers of RGCs were noted in NCAM-/- mice. In conclusion, these data demonstrate that PSA-NCAM supports the survival of injured RGCs in adulthood. However, the role of PSA-NCAM in the adult retina requires further clarification.

Prolyl endopeptidase is involved in the degradation of neural cell adhesion molecules in vitro

ABSTRACT Membrane-associated glycoprotein neural cell adhesion molecule (NCAM) and its polysialylated form (PSA-NCAM) play an important role in brain plasticity by regulating cell–cell interactions. Here, we demonstrate that the cytosolic serine protease prolyl endopeptidase (PREP) is able to regulate NCAM and PSA-NCAM. Using a SH-SY5Y neuroblastoma cell line with stable overexpression of PREP, we found a remarkable loss of PSA-NCAM, reduced levels of NCAM180 and NCAM140 protein species, and a significant increase in the NCAM immunoreactive band migrating at an apparent molecular weight of 120 kDa in PREP-overexpressing cells. Moreover, increased levels of NCAM fragments were found in the concentrated medium derived from PREP-overexpressing cells. PREP overexpression selectively induced an activation of matrix metalloproteinase-9 (MMP-9), which could be involved in the observed degradation of NCAM, as MMP-9 neutralization reduced the levels of NCAM fragments in cell culture medium. We propose that increased PREP levels promote epidermal growth factor receptor (EGFR) signaling, which in turn activates MMP-9. In conclusion, our findings provide evidence for newly-discovered roles for PREP in mechanisms regulating cellular plasticity through NCAM and PSA-NCAM. Summary: NCAM and its polysialylated form (PSA-NCAM) are important regulators of brain plasticity. We demonstrate that prolyl endopeptidase (PREP) is involved in the regulation of NCAM and PSA-NCAM.

Extracellular prolyl oligopeptidase derived from activated microglia is a potential neuroprotection target

Prolyl oligopeptidase (PREP) is an abundant peptidase in the brain and periphery, but its physiological functions are still largely unknown. Recent findings point to a role for PREP in inflammatory processes. This study assessed the cellular and extracellular PREP activities in cultures of mouse primary cortical neurons, microglial cells and astrocytes, and immortalized microglial BV‐2 cells under neuroinflammatory conditions induced by lipopolysaccharide (LPS) and interferon gamma (IFNγ). Furthermore, we evaluated the neuroprotective effect of a specific PREP inhibitor, KYP‐2047, in a neuroinflammation model based on a coculture of primary cortical neurons and activated BV‐2 cells. The inflammatory insult reduced intracellular and increased extracellular PREP activity specifically in microglial cells, suggesting that activated microglia excretes active PREP. A targeted proteomics approach revealed up‐regulation in PREP protein levels in BV‐2 cell growth medium but down‐regulation in crude membrane‐bound PREP after LPS+IFNγ. In the coculture of BV‐2 cells and primary neurons, an increase in extracellular PREP activity was also detected after inflammation. KYP‐2047 (10 μmol/L) significantly protected neurons against microglial toxicity and reduced the levels of the pro‐inflammatory cytokine tumour necrosis factor alpha. In conclusion, these data point to an extracellular role for microglial PREP in the inflammatory process. Inhibition of PREP during neuroinflammation is a potential target for neuroprotection. Thus, PREP inhibitors may offer a novel therapeutic approach for the treatment of neurodegenerative disorders with an inflammatory component including Parkinsons and Alzheimers diseases.

Survival of retinal ganglion cells in transgenic mice with deficiencies in sialyltransferases or neural cell adhesion molecule (NCAM) or after the administration of neuraminidase

Neural cell adhesion molecule (NCAM) plays important roles in the regulation of the brain plasticity during its development and in adulthood. NCAM functions may be regulated by the addition of long linear homopolymers of alpha 2-8-linked sialic acid (PSA). PSA is attached to NCAM via either of two specific sialyltransferases: ST8SiaII and ST8SiaIV. PSA-NCAM is expressed abundantly in the retina and optic nerve during development and adulthood. In the retina PSA-NCAM is expressed in the glial cells in close proximity to retinal ganglion cell (RGC). The functions of the PSA-NCAM in the retina remain unknown. The aim of this study was to investigate the roles of PSA-NCAM in the survival of RGCs after administration of the exitotoxin kainic acid (KA). Intraocular administration of KA induced reduction in the density of RGCs approximately by 60%. Administration of endoneuraminidase (Endo-N) an enzyme, which removes PSA residues from the surface of NCAM, enhanced the toxic effect of KA on RGC. In knockout mice with constitutive deficiency of either ST8SiaII or ST8SiaIV genes, the levels of PSA-NCAM did not differ from those in wild type mice. The toxicity of KA on RGC in these animals also did not differ from control. It should be noted, however, that in knockout ST8SiaII-/- adult mice a reduced number of RGCs was found despite the presence of high levels of PSA-NCAM. These data suggest that during development ST8SiaII ensures high levels of PSA-NCAM, which necessary for the developmental survival of RGCs. The PSA-NCAM in the adult retina ensures the resistance of RGCs to injury.