Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2
Azim Patar
1
Discipline of Anatomy, College of Medicine Nursing and Health Sciences, National University of Ireland Galway, H91 YR71 Galway, Ireland
2
Department of Neuroscience, School of Medical Sciences, Universiti Sains Malaysia, Gelugor 11800, Malaysia
3
Regenerative Medicine Institute (REMEDI), College of Medicine Nursing and Health Sciences, National University of Ireland Galway, H91 YR71 Galway, Ireland
*
Authors to whom correspondence should be addressed.
â€
Linda Howard and Siobhan McMahon should be considered joint senior authors.
Biology 2020, 9(3), 54; https://doi.org/10.3390/biology9030054
Received: 20 January 2020 / Revised: 11 March 2020 / Accepted: 13 March 2020 / Published: 15 March 2020
(This article belongs to the Special Issue Models of CNS Regeneration: Mind the Gap!)
The failure of the spinal cord to regenerate can be attributed both to a lack of trophic support for regenerating axons and to upregulation of inhibitory factors such as chondroitin sulphate proteoglycans including NG2 following injury. Lentiviral vector-mediated gene therapy is a possible strategy for treating spinal cord injury (SCI). This study investigated the effect of lentiviral vectors expressing Neurotrophin-3 (NT-3) and short-hairpin RNA against NG2 (NG2 sh) to enhance neurite outgrowth in in vitro and ex vivo transection injury models. Conditioned medium from cells transduced with NT-3 or shNG2 lentiviruses caused a significant increase in neurite length of primary dorsal root ganglia neurons compared to the control group in vitro. In an ex vivo organotypic slice culture (OSC) transduction with Lenti-NT-3 promoted axonal growth. Transducing OSCs with a combination of Lenti-NT-3/NG2 sh lead to a further increase in axonal growth but only in injured slices and only within the region adjacent to the site of injury. These findings suggest that the combination of lentiviral NT-3 and NG2 sh reduced NG2 levels and provided a more favourable microenvironment for neuronal regeneration after SCI. This study also shows that OSCs may be a useful platform for studying glial scarring and potential SCI treatments.
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Keywords:
ex vivo slice culture; organotypic slice culture; lentiviral vector; spinal cord injury; transection injury; neurotrophin-3; short hairpin RNA; NG2 proteoglycan
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MDPI and ACS Style
Patar, A.; Dockery, P.; McMahon, S.; Howard, L. Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2. Biology 2020, 9, 54.
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