Name: experimental strategies to bridge large tissue gaps in the injured spinal cord after acute and chronic lesion
Uploaded: 2016-04-05T16:00:00
Description: Watch this Scientific Journal Video about Experimental Strategies to Bridge Large Tissue Gaps in the Injured Spinal Cord after Acute and Chronic Lesion at JoVE.com

German Legal Casualty Insurance (DGUV), Research Commission of the Medical Faculty of the Heinrich-Heine-University

Institutional guidelines for animal safety and comfort were adhered to, and all surgical interventions and pre- and post-surgical animal care were provided in compliance with the German Animal Protection law (State Office, Environmental and Consumer Protection of North Rhine- Westphalia, LANUV NRW).
1. Complete Transection of the Thoracic Spinal Cord of Female Wistar Rats (220 - 250 g)
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	<li>Preparation of the Spinal Cord
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		<li>Use isoflurane inhalation anesthesia (2 - 3% of isoflurane...

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	<li>Ramer, L. M., Ramer, M. S., Bradbury, E. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Restoring+function+after+spinal+cord+injury:+towards+clinical+translation+of+experimental+strategies.">Restoring function after spinal cord injury: towards clinical translation of experimental strategies.</a> The Lancet. Neurology. 13 (12), 1241-1256 (2014).</li><li>McDonald, J. W., Howard, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Repairing+the+damaged+spinal+cord:+a+summary+of+our+early+success+with+embryonic+stem+cell+transplantation+and+remyelination.">Repairing the damaged spinal cord: a summary of our early success with embryonic stem cell transplantation and remyelination.</a> Prog. Brain Res. 137, 299-309 (2002).</li><li>Yoon, S. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complete+spinal+cord+injury+treatment+using+autologous+bone+marrow+cell+transplantation+and+bone+marrow+stimulation+with+granulocyte+macrophage-colony+stimulating+factor:+Phase+I/II+clinical+trial.">Complete spinal cord injury treatment using autologous bone marrow cell transplantation and bone marrow stimulation with granulocyte macrophage-colony stimulating factor: Phase I/II clinical trial.</a> Stem Cells. 25 (8), 2066-2073 (2007).</li><li>Brotchi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intrinsic+spinal+cord+tumor+resection.">Intrinsic spinal cord tumor resection.</a> Neurosurgery. 50 (5), 1059-1063 (2002).</li><li>Estrada, V., Tekinay, A., Muller, H. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+ECM+mimetics.">Neural ECM mimetics.</a> Prog. Brain Res. 214, 391-413 (2014).</li><li>Tetzlaff, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Systematic+Review+of+Cellular+Transplantation+Therapies+for+Spinal+Cord+Injury.">A Systematic Review of Cellular Transplantation Therapies for Spinal Cord Injury.</a> J.Neurotrauma. 28 (8), 1611-1682 (2010).</li><li>Brazda, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mechanical+microconnector+system+for+restoration+of+tissue+continuity+and+long-term+drug+application+into+the+injured+spinal+cord.">A mechanical microconnector system for restoration of tissue continuity and long-term drug application into the injured spinal cord.</a> Biomaterials. 34 (38), 10056-10064 (2013).</li><li>Estrada, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-lasting+significant+functional+improvement+in+chronic+severe+spinal+cord+injury+following+scar+resection+and+polyethylene+glycol+implantation.">Long-lasting significant functional improvement in chronic severe spinal cord injury following scar resection and polyethylene glycol implantation.</a> Neurobiol. Dis. 67, 165-179 (2014).</li><li>Rasouli, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Resection+of+glial+scar+following+spinal+cord+injury.">Resection of glial scar following spinal cord injury.</a> J.Orthop.Res. 27 (7), 931-936 (2009).</li><li>Schira, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Significant+clinical,+neuropathological+and+behavioural+recovery+from+acute+spinal+cord+trauma+by+transplantation+of+a+well-defined+somatic+stem+cell+from+human+umbilical+cord+blood.">Significant clinical, neuropathological and behavioural recovery from acute spinal cord trauma by transplantation of a well-defined somatic stem cell from human umbilical cord blood.</a> Brain. 135, 431-446 (2011).</li></ol>

Here two different surgical approaches are presented to bridge tissue gaps in the spinal cord after (1) acute complete transection and mMS implantation and (2) chronic spinal cord lesion and fibrous scar removal plus PEG matrix implantation. Both strategies lead to tissue preservation and axonal regeneration as well as to significant locomotor functional improvement of the treated animals. For mMS implantation an adequate fixation of the mMS within the spinal cord by the firm dura suture after surgery is a critical techn...

A traumatic injury to the spinal cord not only leads to the loss of axons but it further results in tissue defects which hinder any regenerative responses (for review see 1,2). Spinal cord tissue is often lost through secondary degeneration leading to cyst formation or holes in and around the lesion area. Most experimental therapeutic interventions focus on incomplete spinal cord damages like partial transection, crush or contusion injuries with a remaining rim of healthy tissue. For complete injuries like total transections resulting from traumatic accidents or surgical interventions, like tumor resections, only very limited treatment options are available today3,4. After complete transection, mechanic tension of the tissue results in spinal stump retraction, leaving a small gap in the spinal cord. Most strategies focus on filling this gap with tissue, cells or matrices5,6.
Here, a different strategy is presented, namely re-adaptation of the separated stumps using a novel microconnector device7. In order to readapt the two stumps, mechanical force has to be applied as a slight negative pressure to accomplish this (Figure 1). The mechanical microconnector system (mMS) is a multi-channel system of polymethylmethacrylate (PMMA) with honeycomb-shaped holes (Figure 1A) and provided with an outlet tubing system. It is implanted into the tissue gap resulting from complete spinal cord transection in the rat (Figure 1C). One tube can be connected to a vacuum pump to apply negative pressure to the mMS (Figure 1D). The pressure pulls the disconnected spinal cord stumps into the honeycomb-shaped holes of the mMS, which have microstructured walls to hold the tissue in place when pressure is released (Figure 1B). The tubing can be left intact after surgery and attached to an osmotic minipump in order to infuse substances into the lesion core (Figure 1E-F).
Besides an acute transection of the spinal cord another type of complete lesion results from surgical removal of a spinal tumor or a solid chronic lesion scar leading to large tissue gaps of several millimeters, which cannot be overcome by the mMS so far. The majority of patients with spinal cord trauma suffer from chronic injuries. In these patients, a fully developed scar occupies the lesion core. The surgical removal of the lesion scar is a concept for treatment which is currently investigated after experimental SCI8,9. While the resection procedure itself can be performed without causing considerable additional damage, the resulting tissue gap needs to be bridged with a suitable matrix which allows and promotes the regeneration of tissue and, in the specific case of spinal cord injuries, the regeneration of nerve fibers to maintain and promote locomotor functions. It was found that low-molecular weight polyethylene glycol (PEG 600) is a very suitable material for this purpose. Its lack of immunogenicity and the very low viscosity allow smooth integration into the surrounding tissue. Insertion of the biopolymer alone promotes the invasion of beneficial cells, including endothelial cells, peripheral Schwann cells, and astrocytes, and - very importantly - the regeneration and elongation of axons of descending and ascending fiber tracts as well as their ensheathment by compact myelin8. These regenerative responses were found to be accompanied by long-lasting functional improvements. The combination of resection of scar tissue and subsequent implantation of PEG 600 presents a safe and simple, yet very efficient means of bridging substantial spinal cord tissue defects.

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experimental strategies to bridge large tissue gaps in the injured spinal cord after acute and chronic lesion

After a spinal cord injury (SCI) a scar forms in the lesion core which hinders axonal regeneration. Bridging the site of injury after an insult to the spinal cord, tumor resections, or tissue defects resulting from traumatic accidents can aid in facilitating general tissue repair as well as regenerative growth of nerve fibers into and beyond the affected area. Two experimental treatment strategies are presented: (1) implantation of a novel microconnector device into an acutely and completely transected thoracic rat spinal cord to readapt severed spinal cord tissue stumps, and (2) polyethylene glycol filling of the SCI site in chronically lesioned rats after scar resection. The chronic spinal cord lesion in this model is a complete spinal cord transection which was inflicted 5 weeks before treatment. Both methods have recently achieved very promising outcomes and promoted axonal regrowth, beneficial cellular invasion and functional improvements in rodent models of spinal cord injury.
The mechanical microconnector system (mMS) is a multi-channel system composed of polymethylmethacrylate (PMMA) with an outlet tubing system to apply negative pressure to the mMS lumen thus pulling the spinal cord stumps into the honeycomb-structured holes. After its implantation into the 1 mm tissue gap the tissue is sucked into the device. Furthermore, the inner walls of the mMS are microstructured for better tissue adhesion.
In the case of the chronic spinal cord injury approach, spinal cord tissue - including the scar-filled lesion area - is resected over an area of 4 mm in length. After the microsurgical scar resection the resulting cavity is filled with polyethylene glycol (PEG 600) which was found to provide an excellent substratum for cellular invasion, revascularization, axonal regeneration and even compact remyelination in vivo.

Tissue Preservation, Axonal Regrowth and Functional Benefit of mMS Implantation after Acute Complete Transection of the Spinal Cord 
It is demonstrated that the acute implantation of the mMS stabilized the completely transected spinal cord stumps and decreased shrinkage of the tissue (Figure 2A versus B). As visualized by Trichrome staining in sagittal sections, the green connective tissue staining of the fibrotic scar in th...

Watch this Scientific Journal Video about Experimental Strategies to Bridge Large Tissue Gaps in the Injured Spinal Cord after Acute and Chronic Lesion at JoVE.com

Experimental Strategies to Bridge Large Tissue Gaps in the Injured Spinal Cord after Acute and Chronic Lesion

Severe spinal cord injuries often result in tissue defects. Two possibilities are described to successfully bridge such gaps to promote tissue adaptation, regenerative responses and functional improvement in rats via implantation of a mechanical microconnector system after acute injury and five weeks after complete spinal cord transection.

After a spinal cord injury (SCI) a scar forms in the lesion core which hinders axonal regeneration. Bridging the site of injury after an insult to the ...

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