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Mechanisms and improvements of cell transplantation for nerve repair
Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy. Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Hand Surgery.
2011 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Trauma to the nervous system is a frequent clinical problem and new approaches to nerve repair are required. Autologous cell transplantation together with a suitable scaffold material could be used to create a bio-active artificial nerve graft to enhance regeneration. The work presented in this licentiate thesis attempts to improve both the biomaterial and cellular components of this repair strategy.

In the first study, by using common biodegradable polyesters, namely poly-ε-caprolactone (PCL) and poly -L,D- lactic acid (PLA), a thin film scaffold prototype was fabricated by using a solvent-evaporation method. These scaffolds, with thicknesses of approximately 10-20 µm, exhibited a heterogenous but continuous surface topography decorated with pore/pits of regulated sizes. The sizes of the pore/pits ranged from 0.5 to 30 µm2and could be modulated by varying the ratios of PLA and PCL. Biocompatibility of these scaffolds was demonstrated by using adipose derived stem cells (ADSC) differentiated into a Schwann cell-like phenotype (dADSC), which showed attachment and proliferation on the films, maintenance of glial cell markers expression and enhancement of neurite outgrowth in co-culture with dorsal root ganglia (DRG) neurons.

Transplantation of cells for nerve injuries remains sub-optimal due to their limited survival rates. In the second study, a chemical ischemia model (metabolically induced by sodium azide and 2-deoxyglucose) was established to investigate the differential effects of ischemia and serum deprivation on mesenchymal stem cells (MSCs). MSCs were more suseptible to combined than individual blockade of glycolysis and oxidative phosphorylation. Apoptotic and autophagy pathways were activated in the MSCs. Chemical ischemia or serum withdrawal alone induced a similar amount of cell death with significantly different intracellular ATP maintenance; but their effects were additive. The levels of various neurotrophin extracellular matrix and angiogenic factors expressed by the cells were shown to be differentially affected by ischemia but unaffected by changes in serum level. Stem cells isolated from both adipose tissue (ADSC) and bone marrow (BMSC) reacted similarly under these conditions. This chemical ischemia model will enable future screening of pharmacological agents to enhance the survival of MSCs under stress conditions.

The mechanism underlying the neurotrophic potential of MSCs is unknown. In the third study in this thesis it is hypothesised that MSCs, upon stimulation with different growth factors, could produce brain derived neurotrophic factor (BNDF) with a similar molecular mechanism to that described in the nervous system. Within 24 hours of stimulation, ADSC and BMSC showed high secretion levels of BDNF, and these cells were able to enhance axonal outgrowth in DRG neurons at levels similar to long-term differentiated MSCs. Both the neuronal activity dependent promoterBDNFexon IV, along with full length protein encodingBDNFexon IX, were up-regulated upon stimulation.BDNFgating transcription factor, cyclic cAMP responsive element binding (CREB) protein, was also found to be activated but blocking of CREB phosphorylation with the small molecule inhibitor H89 did not suppress expression of BDNF protein suggesting compensatory pathways are involved.

In summary, these studies indicate that MSCs are compatible with polyester based microporous scaffolds but the cells are highly susceptible to the stress conditions mimicking the hostile milieu at a nerve injury site. Preliminary studies hint at the molecular mechanism regulation BDNF expression in MSC and imply the interactions between MSCs and axons may play a role in the neurotrophic activity of the stem cells.

Place, publisher, year, edition, pages
Umeå: Umeå universitet , 2011. , 56 p.
Keyword [en]
Mesenchymal stem cells, microporous scaffold, biomaterials, ischemia, cell death, brain derived neurotrophic factor
National Category
Surgery
Identifiers
URN: urn:nbn:se:umu:diva-56298OAI: oai:DiVA.org:umu-56298DiVA: diva2:533224
Available from: 2012-06-13 Created: 2012-06-13 Last updated: 2012-06-14Bibliographically approved
List of papers
1. In vitro evaluation of polyester-based scaffolds seeded with adipose derived stem cells for peripheral nerve regeneration
Open this publication in new window or tab >>In vitro evaluation of polyester-based scaffolds seeded with adipose derived stem cells for peripheral nerve regeneration
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2010 (English)In: Journal of Biomedical Materials Research. Part A, ISSN 1549-3296, E-ISSN 1552-4965, Vol. 95, no 3, 701-708 p.Article in journal (Refereed) Published
Abstract [en]

To overcome the disadvantages of autografts for peripheral nerve repair, different methods such as artificial nerve conduits have been investigated for an alternative approach. This study demonstrated that solvent casting is a simple but efficient method to create thin polyester-based scaffolds for stem cell delivery. Using poly (ε-caprolactone) and poly (D,L-lactic acid), we produced scaffold films containing heterogenous depressions (pits) on the air surface with a size ranging from 0.5 to 30 μm(2). These scaffolds were moderately hydrophobic; however, they supported the differentiation of adipose derived stem cells (ADSC) into a Schwann cell-like phenotype. The differentiated ADSC (dADSC) expressed S100 protein and glial fibrillary acidic protein and readily adhered to the films and proliferated at a similar rate to those cultured on tissue culture polystyrene. Cells were also positive for proliferating cell nuclear antigen. Furthermore, dADSC retained functional activity and significantly enhanced neurite outgrowth from dorsal root ganglia neurons. This study suggests polymer scaffolds combined with dADSCs could be a promising therapy for peripheral nerve injuries.

Keyword
stromal cells; blends; repair; tissue; transplantation; morphology; therapies; conduit; lineage; films
National Category
Biomaterials Science
Identifiers
urn:nbn:se:umu:diva-42249 (URN)10.1002/jbm.a.32889 (DOI)000283959800005 ()20725987 (PubMedID)
Available from: 2011-04-06 Created: 2011-04-06 Last updated: 2017-12-11Bibliographically approved
2. The reactions of adipose tissue and bone marrow-derived stem cells in an ischaemia-like microenvironment
Open this publication in new window or tab >>The reactions of adipose tissue and bone marrow-derived stem cells in an ischaemia-like microenvironment
2012 (English)In: Journal of tissue engineering and regenerative medicine, ISSN 1932-7005, Vol. 6, no 6, 473-485 p.Article in journal (Refereed) Published
Abstract [en]

Mesenchymal stem cells (MSCs) from adipose tissue and bone marrow are promising cell sources for autologous cell therapy of nerve injuries, as demonstrated by their intrinsic neurotrophic potential. However, extensive death of transplanted cells limits their full benefits. This study investigated the effects of ischaemia (metabolically induced by sodium azide and 2-deoxyglucose) and serum-derived mitogens on the viability and functional profile of MSCs in vitro. MSCs were more susceptible to combined, rather than individual, blockade of glycolysis and oxidative phosphorylation. Apoptosis and autophagy were involved in ischaemia-induced cell death. Chemical ischaemia alone and serum withdrawal alone induced a similar amount of cell death, with significantly different intracellular ATP maintenance. Combined ischaemia and serum deprivation had additive effects on cell death. Expression of the extracellular matrix (ECM) molecules laminin and fibronectin was attenuated under ischaemia and independent of serum level; however, BDNF and NGF levels remained relatively constant. Strong upregulation of VEGF and to a lesser extent angiopoietin-1 was observed under ischaemia but not in serum withdrawal conditions. Importantly, this study demonstrated similar reactions of MSCs derived from adipose and bone marrow tissue, in ischaemia-like and mitogen-deprived microenvironments in terms of viability, cellular energetics, cell death mechanisms and expression levels of various growth-promoting molecules. Also, the results suggest that ischaemia has a larger impact on the ability of MSCs to survive transplantation than withdrawal of mitogens. Copyright © 2011 John Wiley & Sons, Ltd.

Place, publisher, year, edition, pages
John Wiley & Sons, 2012
Keyword
apoptosis, autophagy, cell death, ischaemia, mesenchymal stem cell, trauma
National Category
Neurosciences
Identifiers
urn:nbn:se:umu:diva-53241 (URN)10.1002/term.452 (DOI)21751425 (PubMedID)
Available from: 2012-03-19 Created: 2012-03-19 Last updated: 2012-06-13Bibliographically approved
3. Intrinsic mechanisms underlying the de novo BDNF expression of mesenchymal stem cells
Open this publication in new window or tab >>Intrinsic mechanisms underlying the de novo BDNF expression of mesenchymal stem cells
(English)Manuscript (preprint) (Other academic)
National Category
Surgery
Identifiers
urn:nbn:se:umu:diva-56296 (URN)
Available from: 2012-06-13 Created: 2012-06-13 Last updated: 2012-06-13Bibliographically approved

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