Chondroitinase ABC-I and -II linked chitosan microbeads increase neurtire length in CSPG-enriched astrocyte culture

Poster Presentation - Theme 5: NeuroscienceAfter spinal cord injury, axonal regrowth is often restricted due to upregulation of chondroitin sulfate proteoglycans (CSPGs) at the lesion site. Chondroitinase ABC I and II (recombinant ChABC-I & -II) cleave CS moieties of the PGs enhancing prospects of axonal regrowth through the lesion. We attempted to use glutaraldehyde to immobilize ChABC-I or-II separately on chitosan beads. Immobilized ChABC-I demonstrated CS-cleaving activity both in biochemical assay and in ...postprin

The Mechanisms of Chondroitin Sulphate Lyases Treatment in Promotion of Axonal Growth

Poster PresentationIn Injured nerves, chondroitin sulphate (CS) is upregulated forming barriers with astrocytes/fibroblasts and other extracellular matrix molecules, and thereby hampering nerve regeneration. Cleavage of CS using chondroitin sulphate lyases (Proteus vulgaris) promises axon regrowth through the barrier but the enzymatic efficacy remains to be improved. Two subtypes, endolyase and exolyase, have been found coexisting in the original host but only the former has been exploited for treatment of injured nerve tracts. We hypothesise that the two subtypes are necessary for enzymatic efficacy on CSs. We therefore prepared recombinant enzymes of both subtypes. Enzyme kinetics study revealed feedback inhibition by limit digestion products: that of the endolyase by tetrasaccharides and that of the exolyase by the disaccharides. When the two subtypes were used in combination, the digestion efficiency increased. We then used TGF beta-1 to induce CS production by astrocytes in culture, mimicking reactive glia in injured nerves. In co-cultures of such astrocytes with cortical neurons, treatment with combinations of the two subtypes resulted in increased neurite lengths as compared to co-cultures treated with one of the subtypes. The limit digestion products of CS were further tested for their effects on neurite extension on astrocytes that had been treated with TGF beta-1. The CS disaccharides, both 4- and 6-sulphated but not the tetrasaccharides, promoted neurite extension significantly. Taken together, the combinatorial use of the ChABC subtypes not only improved efficacy of enzyme activity on the axon-restrictive CS moiety, but also increased the yield of CS disaccharides which contributed to axonal growth.published_or_final_versio

Heparanase 1 and Heparanase 2 expression in Hippocampal Neurons

Poster presentation - Theme 5: NeuroscienceSelective modification (weakening or strengthening) of synaptic connections between neurons within the hippocampal circuit contribute to the learning and memory processes in the brain. Enzymatic heparanase-1 (hpa1) was found expressed in hippocampus by both immunohistochemistry and in-situ hybridization. Western blot analysis of neuronal secretions however revealed the pro-form, i.e. proheparanase, which do not have enzymatic activity is secreted by hippocampal neurons. Proheparanase could trigger AMPA receptor internalization upon binding to the ...postprin

Molecular and cellular aspects of plasticity after neural injury

This review focuses on our effort to address plasticity of the nervous system after neural injury. We have used different animal models to examine cellular mechanisms of plasticity underlying the pathological and repair processes. After severance of sensory input from one inner ear, topographic representations of spacecentered coordinates in the brain undergo plastic changes. During vestibular compensation, tissue plasticity constitutes an important component for functional recovery of neuronal network. In Parkinsonian animals, modulation of signaling via glutamatergic synapses, neurotrophins and neurokinins contributes to the protection of basal ganglion neurons from degeneration, thereby delaying deterioration of motor functions. With the use of animal models of neural injury, we further overcome the molecular restriction at the glial scar to enhance neural regrowth and remyelination, pointing to the possibility of developing new therapeutic strategies to stimulate neural plasticity and repair in the adult nervous system.published_or_final_versio

Semaphorin3A, associated with perineuronal nets, regulates the development of the maturation of the central vestibular circuitry

Poster Presentation - Theme 5: NeuroscienceDuring the formative period of neural circuits, perineuronal nets (PN) are established to restrict plasticity of the circuit. The role of PN in vestibular plasticity can be tested by studying the emergence of negative geotaxis with postnatal maturation of the vestibular circuitry for gravity detection. Using rats as model, we observed that negative geotaxis was mature by postnatal day (P) 9, in correlation with consolidation of PN around GABAergic neurons …postprin

Perineuronal chondroitin sulfates and semaphorin 3A regulate postnatal maturation of the vestibular circuit for gravity detection

Conference Theme: The Extracellular Matrix NichePoster Presentation: abstract no. P30Perineuronal chondroitin sulfate proteoglycan networks are implicated in restricting plasticity of the mature CNS. Less is known of the impact of ligands associated with the perineuronal network (PN) in the formative period of neural circuits. The emergence of negative geotaxis with postnatal maturation of the vestibular circuitry for gravity detection offers a behavioral readout in tests for roles of the PN and associated ligands in vestibular plasticity. Using postnatal rats as model, we found that negative geotaxis was mature by postnatal day (P) 9, in correlation with ...postprin

Chitosan nanofiber composed nerve conduit for directing axonal growth

Poster presentation - Theme 5: NeuroscienceSchwann cell-seeded guidance channels have been exploited to bridge and guide axonal re-growth across gaps in lesioned nerves. By orienting the Schwann cell growth on aligned nanofibers, we hypothesized that axonal growth can be guided along the designated direction towards the target. Chitosan as the choice scaffold material given its biocompatibility and the tunable susceptibility to biodegradation. Chitosan was dissolved in trifluoroacetic acid/methylene chloride solution and for electrospinning onto a high speed rotating collector ...postprin

Expression of chondroitin sulfotransferase in cranial motor neurons for cell migration in rat embryonic hindbrain

Poster presentation - Theme 5: NeuroscienceNeuronal migration allows the proper positioning of neurons for establishing functional connectivity of defined neural circuits. We and others reported the restrictive role of Chondroitin sulfate (CS) moieties in axonal fasciculation. CS moieties of proteoglycans are therefore hypothesized to control the timely orchestration of cranial motor neuron migration during hindbrain development by the varying sulfation patterns of the chondroitins between the migrating and ready-to-migrate neurons. Hindbrain explants of E11.5 Sprague Dawley rats were ...postprin

Regulatory role of proheparanase with peri-synaptic heparan sulfate proteoglycan and AMPA-type glutamate receptor in synaptic plasticity

Poster Presentation: P59AMPA-type glutamate receptors (AMPAR) govern excitatory synaptic transmission. Perineuronal heparan sulfates (HS) have been implicated in controlling the open-state of AMPAR. Our finding of neuronal heparanase expression in adult rats led us to test (1) if neuronal heparanase is secreted and (2) if the secreted form acts on perineuronal HS to modulate synaptic plasticity. Neuronal secretion of heparanase was triggered by phorbol ester of rat hippocampal neurons in culture. Western blot analysis of the secreted product revealed enzymatically inactive proheparanase, but not the enzymatically active heparanase. Synaptosomes prepared from phorbol ester-treated rat cortexslices showed enrichment in proheparanase; co-immunoprecipitation studies further showed association of AMPAR subunits (GluA1 and GluA2/3) with both syndecan-3 (a transmembrane HS-proteoglycan) and proheparanase, suggesting their partnership in the peri-synaptic environment. Treatment of hippocampal neurons in culture with recombinant proheparanase triggered internalization of proheparanase, perineuronal HS-proteoglycans and AMPARs, suggesting their clustering as a functional complex. Heparitinase pre-treatment of hippocampal neuron cultures reduced proheparanase-induced internalization of AMPARs, suggesting that the HS moiety is critical for effecting the partnership. Treatment of hippocampal slices with recombinant proheparanase resulted in down-regulation of both basal synaptic strength and LTP at Schaffer collateral synapses. These results reveal a novel role of neuronal proheparanase in resetting AMPAR and perineuronal HS levels at the synapse and thus the modulation of synaptic plasticity.postprin

Human induced pluripotent stem cell-derived sensory neurons for fate commitment of bone marrow-derived Schwann cells: Implications for re-myelination therapy

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