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Neural Crest Stem Cells protect Spinal Cord Slice Cultures from Excitotoxic Neuronal Damage and Inhibit Glial Activation
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences. (The Spineab)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Neural Crest Stem Cells (NCSC) possess anti-inflammatory properties and they could therefore have protective effects on neurons after spinal cord injury (SCI). We investigated if NCSCs reduce neuronal loss and glial activation after excitotoxic injury in spinal cord slice cultures (SCSCs). SCSCs subjected to N-Methyl-D-Aspartate (NMDA)-induced injury were either treated by transplantation of NCSC or with interleukin-1 receptor antagonist (IL1RA). Immunohistochemistry for Neuronal Nuclei (NeuN), glial fibrillary acidic protein (GFAP) and MacII were used to study neuronal and glial populations, and TUNEL staining to visualize apoptotic cells. Migration and differentiation of NCSCs on SCSCs, laminin or a hyaluronic acid hydrogel were investigated by immunohistochemistry. NCSCs counteracted the neuronal loss observed after NMDA-induced excitotoxicity and reduced the proportion of apoptotic cells. Transplantation of NCSCs also reduced the number of activated microglial cells in the white matter and the number of activated astrocytes in the grey matter of NMDA-injured SCSCs. The majority of NCSCs migrated superficially across the surface of SCSCs with some of them penetrating into the tissue. NCSCs did not show signs of neuronal or glial differentiation, however, the markers SOX2 and Krox20 were expressed which indicated an undifferentiated state of NCSCs. In conclusion, NCSCs have neuroprotective, anti-apoptotic and anti-inflammatory effects on SCSCs subjected to excitotoxicity, similar to the effects of IL1RA. Since the majority of NCSCs did not migrate through SCSCs they are likely to exert their actions through the secretion of soluble factors.

Keyword [en]
Neuroprotection, Suppressed glial activation, excitotoxicity, apoptosis
National Category
Medical and Health Sciences
Research subject
Medical Science
Identifiers
URN: urn:nbn:se:uu:diva-251476OAI: oai:DiVA.org:uu-251476DiVA: diva2:806231
Available from: 2015-04-19 Created: 2015-04-19 Last updated: 2015-07-07
In thesis
1. Neuroprotection in the Injured Spinal Cord: Novel Strategies using Immunomodulation, Stem cell Transplantation and Hyaluronic acid Hydrogel carriers
Open this publication in new window or tab >>Neuroprotection in the Injured Spinal Cord: Novel Strategies using Immunomodulation, Stem cell Transplantation and Hyaluronic acid Hydrogel carriers
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The overall aim of this thesis was to establish strategies to minimize secondary damage to the injured spinal cord. Secondary damage that follows spinal cord injury (SCI) involves inflammatory and excitotoxic pathways. Regulation of these pathways using immunomodulatory and neuroprotective substances potentially protects the injured spinal cord from further damage. We also developed and studied resorbable biomaterials to be used as carriers for potential neuroprotectants to the injured spinal cord.

We used transversal spinal cord slice cultures (SCSCs) derived from postnatal mice as a model. SCSCs were maintained on different biomaterials and were studied after treatment with immunomodulatory and/or neurotrophic factors. They were further excitotoxically injured and subsequently treated with interleukin-1 receptor antagonist (IL1RA) or by neural crest stem cell (NCSC)-transplantation.

The results show that biocompatible and resorbable hydrogels based on hyaluronic acid (HA) preserved neurons in SCSCs to a much higher extent than a conventional collagen-based biomaterial or standard polyethylene terephthalate (PET) membrane inserts. Glial activation was limited in the cultures maintained on HA-based hydrogel. The anti-inflammatory factor IL1RA protected SCSCs from degenerative mechanisms that occur during in vitro incubation, and IL1RA also protected SCSCs from excitotoxic injury induced by N-Methyl-d-Aspartate (NMDA). IL1RA specifically protected neurons that resided in the ventral horn, while other neuronal populations such as dorsal horn neurons and Renshaw cells did not respond to treatment. Finally, transplantation of NCSCs onto excitotoxically injured SCSCs protected from neuronal loss, apoptosis and glial activation, while NCSCs remained undifferentiated.

The results presented in this thesis indicate that carriers based on HA seem to be more suitable than conventional collagen-based biomaterials since they enhance neuronal survival per se. The observed neuroprotection is likely due to biomechanical properties of HA. IL1RA protects SCSCs from spontaneous degeneration and from NMDA-induced injury, suggesting that excitotoxic mechanisms can be modulated through anti-inflammatory pathways. Different neuronal populations are affected by IL1RA to various degrees, suggesting that a combination of different neuroprotectants should be used in treatment strategies after SCI. Finally, NCSCs seem to protect SCSCs from excitotoxic injury through paracrine actions, since they remain undifferentiated and do not migrate into the tissue during in vitro incubation.

It seems that combinations of neuroprotectants and carrier substances should be considered rather than one single strategy when designing future treatments for SCI. Incorporation of neuroprotectants such as IL1RA combined with stem cells in injectable biocompatible carriers based on HA is the final goal of our group in the treatment of SCI.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2015. 59 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1106
Keyword
Hyaluronic Acid-based hydrogel, motorneurons, microglial cells, Interleukin-1 Receptor Antagonist, Renshaw cells, excitotoxicity, neuroinflammation, Neural Crest Stem Cells
National Category
Medical and Health Sciences
Research subject
Medical Science
Identifiers
urn:nbn:se:uu:diva-251477 (URN)978-91-554-9255-7 (ISBN)
Public defence
2015-06-12, Grönwallsalen, Akademiska sjukhuset, Akademiska sjukhuset ing.70, Uppsala, 09:00 (English)
Opponent
Supervisors
Available from: 2015-06-04 Created: 2015-04-19 Last updated: 2016-04-21

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Schizas, Nikos

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