Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2
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...
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doaj-7eb6da41a453437482ae355b09b7c5d42020-11-25T02:01:59ZengMDPI AGBiology2079-77372020-03-01935410.3390/biology9030054biology9030054Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2Azim Patar0Peter Dockery1Siobhan McMahon2Linda Howard3Discipline of Anatomy, College of Medicine Nursing and Health Sciences, National University of Ireland Galway, H91 YR71 Galway, IrelandDiscipline of Anatomy, College of Medicine Nursing and Health Sciences, National University of Ireland Galway, H91 YR71 Galway, IrelandDiscipline of Anatomy, College of Medicine Nursing and Health Sciences, National University of Ireland Galway, H91 YR71 Galway, IrelandRegenerative Medicine Institute (REMEDI), College of Medicine Nursing and Health Sciences, National University of Ireland Galway, H91 YR71 Galway, IrelandThe 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.https://www.mdpi.com/2079-7737/9/3/54ex vivo slice cultureorganotypic slice culturelentiviral vectorspinal cord injurytransection injuryneurotrophin-3short hairpin rnang2 proteoglycan |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Azim Patar Peter Dockery Siobhan McMahon Linda Howard |
spellingShingle |
Azim Patar Peter Dockery Siobhan McMahon Linda Howard 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 ex vivo slice culture organotypic slice culture lentiviral vector spinal cord injury transection injury neurotrophin-3 short hairpin rna ng2 proteoglycan |
author_facet |
Azim Patar Peter Dockery Siobhan McMahon Linda Howard |
author_sort |
Azim Patar |
title |
Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2 |
title_short |
Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2 |
title_full |
Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2 |
title_fullStr |
Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2 |
title_full_unstemmed |
Ex Vivo Rat Transected Spinal Cord Slices as a Model to Assess Lentiviral Vector Delivery of Neurotrophin-3 and Short Hairpin RNA against NG2 |
title_sort |
ex vivo rat transected spinal cord slices as a model to assess lentiviral vector delivery of neurotrophin-3 and short hairpin rna against ng2 |
publisher |
MDPI AG |
series |
Biology |
issn |
2079-7737 |
publishDate |
2020-03-01 |
description |
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. |
topic |
ex vivo slice culture organotypic slice culture lentiviral vector spinal cord injury transection injury neurotrophin-3 short hairpin rna ng2 proteoglycan |
url |
https://www.mdpi.com/2079-7737/9/3/54 |
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