Lentiviral vector-mediated knockdown of the neuroglycan 2 proteoglycan or expression of neurotrophin-3 promotes neurite outgrowth in a cell culture model of the glial scar.

Eleanor Donnelly, Padraig Strappe, Lisa McGinley, Nicholas Madigan, Elizabeth Guerts, Gemma Rooney, Anthony Windebank, John Fraher, Peter Dockery, Timothy O'Brien, Siobhan S. McMahon

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

BackgroundFollowing spinal cord injury, a highly inhibitory environment for axonal regeneration develops. One of the main sources of this inhibition is the glial scar that is formed after injury by reactive astrocytes. The inhibitory environment is mainly a result of chondroitin sulphate proteoglycans (CSPGs). Neuroglycan 2 (NG2), one of the main inhibitory CSPGs, is up-regulated following spinal cord injury.MethodsSmall interfering RNA (siRNA) was designed to target NG2 and this short hairpin RNA (shRNA) was cloned into a lentiviral vector (LV). The neurotrophic factor neurotrophin-3 (NT-3) promotes the growth and survival of developing neurites and has also been shown to aid regeneration. NT-3 was also cloned into a LV. In vitro assessment of these vectors using a coculture system of dorsal root ganglia (DRG) neurones and Neu7 astrocytes was carried out. The Neu7 cell line is a rat astrocyte cell line that overexpresses NG2, thereby mimicking the inhibitory environment following spinal cord injury.Results and DiscussionThese experiments show that both the knockdown of NG2 via shRNA and over-expression of NT-3 can significantly increase neurite growth, although a combination of both vectors did not confer any additional benefit over the vectors used individually. These LVs show promising potential for growth and survival of neurites in injured central nervous system tissue (CNS).
Original languageEnglish
Pages (from-to)863-872
Number of pages10
JournalJournal of Gene Medicine
Volume12
Issue number11
DOIs
Publication statusPublished - Nov 2010

Fingerprint

Neurotrophin 3
Neurites
Proteoglycans
Spinal Cord Injuries
Neuroglia
Astrocytes
Small Interfering RNA
Cicatrix
Chondroitin Sulfate Proteoglycans
Cell Culture Techniques
Regeneration
Growth
Nerve Tissue
Cell Line
Nerve Growth Factors
Spinal Ganglia
Coculture Techniques
Central Nervous System
RNA
Neurons

Cite this

Donnelly, Eleanor ; Strappe, Padraig ; McGinley, Lisa ; Madigan, Nicholas ; Guerts, Elizabeth ; Rooney, Gemma ; Windebank, Anthony ; Fraher, John ; Dockery, Peter ; O'Brien, Timothy ; McMahon, Siobhan S. / Lentiviral vector-mediated knockdown of the neuroglycan 2 proteoglycan or expression of neurotrophin-3 promotes neurite outgrowth in a cell culture model of the glial scar. In: Journal of Gene Medicine. 2010 ; Vol. 12, No. 11. pp. 863-872.
@article{087921874b244277b60bf34a63b75349,
title = "Lentiviral vector-mediated knockdown of the neuroglycan 2 proteoglycan or expression of neurotrophin-3 promotes neurite outgrowth in a cell culture model of the glial scar.",
abstract = "BackgroundFollowing spinal cord injury, a highly inhibitory environment for axonal regeneration develops. One of the main sources of this inhibition is the glial scar that is formed after injury by reactive astrocytes. The inhibitory environment is mainly a result of chondroitin sulphate proteoglycans (CSPGs). Neuroglycan 2 (NG2), one of the main inhibitory CSPGs, is up-regulated following spinal cord injury.MethodsSmall interfering RNA (siRNA) was designed to target NG2 and this short hairpin RNA (shRNA) was cloned into a lentiviral vector (LV). The neurotrophic factor neurotrophin-3 (NT-3) promotes the growth and survival of developing neurites and has also been shown to aid regeneration. NT-3 was also cloned into a LV. In vitro assessment of these vectors using a coculture system of dorsal root ganglia (DRG) neurones and Neu7 astrocytes was carried out. The Neu7 cell line is a rat astrocyte cell line that overexpresses NG2, thereby mimicking the inhibitory environment following spinal cord injury.Results and DiscussionThese experiments show that both the knockdown of NG2 via shRNA and over-expression of NT-3 can significantly increase neurite growth, although a combination of both vectors did not confer any additional benefit over the vectors used individually. These LVs show promising potential for growth and survival of neurites in injured central nervous system tissue (CNS).",
author = "Eleanor Donnelly and Padraig Strappe and Lisa McGinley and Nicholas Madigan and Elizabeth Guerts and Gemma Rooney and Anthony Windebank and John Fraher and Peter Dockery and Timothy O'Brien and McMahon, {Siobhan S.}",
note = "Imported on 12 Apr 2017 - DigiTool details were: month (773h) = Nov 2010; Journal title (773t) = Journal of Gene Medicine. ISSNs: 1099-498X;",
year = "2010",
month = "11",
doi = "10.1002/jgm.1509",
language = "English",
volume = "12",
pages = "863--872",
journal = "Journal of Gene Medicine",
issn = "1099-498X",
publisher = "John Wiley & Sons",
number = "11",

}

Donnelly, E, Strappe, P, McGinley, L, Madigan, N, Guerts, E, Rooney, G, Windebank, A, Fraher, J, Dockery, P, O'Brien, T & McMahon, SS 2010, 'Lentiviral vector-mediated knockdown of the neuroglycan 2 proteoglycan or expression of neurotrophin-3 promotes neurite outgrowth in a cell culture model of the glial scar.', Journal of Gene Medicine, vol. 12, no. 11, pp. 863-872. https://doi.org/10.1002/jgm.1509

Lentiviral vector-mediated knockdown of the neuroglycan 2 proteoglycan or expression of neurotrophin-3 promotes neurite outgrowth in a cell culture model of the glial scar. / Donnelly, Eleanor; Strappe, Padraig; McGinley, Lisa; Madigan, Nicholas; Guerts, Elizabeth; Rooney, Gemma; Windebank, Anthony; Fraher, John; Dockery, Peter; O'Brien, Timothy; McMahon, Siobhan S.

In: Journal of Gene Medicine, Vol. 12, No. 11, 11.2010, p. 863-872.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Lentiviral vector-mediated knockdown of the neuroglycan 2 proteoglycan or expression of neurotrophin-3 promotes neurite outgrowth in a cell culture model of the glial scar.

AU - Donnelly, Eleanor

AU - Strappe, Padraig

AU - McGinley, Lisa

AU - Madigan, Nicholas

AU - Guerts, Elizabeth

AU - Rooney, Gemma

AU - Windebank, Anthony

AU - Fraher, John

AU - Dockery, Peter

AU - O'Brien, Timothy

AU - McMahon, Siobhan S.

N1 - Imported on 12 Apr 2017 - DigiTool details were: month (773h) = Nov 2010; Journal title (773t) = Journal of Gene Medicine. ISSNs: 1099-498X;

PY - 2010/11

Y1 - 2010/11

N2 - BackgroundFollowing spinal cord injury, a highly inhibitory environment for axonal regeneration develops. One of the main sources of this inhibition is the glial scar that is formed after injury by reactive astrocytes. The inhibitory environment is mainly a result of chondroitin sulphate proteoglycans (CSPGs). Neuroglycan 2 (NG2), one of the main inhibitory CSPGs, is up-regulated following spinal cord injury.MethodsSmall interfering RNA (siRNA) was designed to target NG2 and this short hairpin RNA (shRNA) was cloned into a lentiviral vector (LV). The neurotrophic factor neurotrophin-3 (NT-3) promotes the growth and survival of developing neurites and has also been shown to aid regeneration. NT-3 was also cloned into a LV. In vitro assessment of these vectors using a coculture system of dorsal root ganglia (DRG) neurones and Neu7 astrocytes was carried out. The Neu7 cell line is a rat astrocyte cell line that overexpresses NG2, thereby mimicking the inhibitory environment following spinal cord injury.Results and DiscussionThese experiments show that both the knockdown of NG2 via shRNA and over-expression of NT-3 can significantly increase neurite growth, although a combination of both vectors did not confer any additional benefit over the vectors used individually. These LVs show promising potential for growth and survival of neurites in injured central nervous system tissue (CNS).

AB - BackgroundFollowing spinal cord injury, a highly inhibitory environment for axonal regeneration develops. One of the main sources of this inhibition is the glial scar that is formed after injury by reactive astrocytes. The inhibitory environment is mainly a result of chondroitin sulphate proteoglycans (CSPGs). Neuroglycan 2 (NG2), one of the main inhibitory CSPGs, is up-regulated following spinal cord injury.MethodsSmall interfering RNA (siRNA) was designed to target NG2 and this short hairpin RNA (shRNA) was cloned into a lentiviral vector (LV). The neurotrophic factor neurotrophin-3 (NT-3) promotes the growth and survival of developing neurites and has also been shown to aid regeneration. NT-3 was also cloned into a LV. In vitro assessment of these vectors using a coculture system of dorsal root ganglia (DRG) neurones and Neu7 astrocytes was carried out. The Neu7 cell line is a rat astrocyte cell line that overexpresses NG2, thereby mimicking the inhibitory environment following spinal cord injury.Results and DiscussionThese experiments show that both the knockdown of NG2 via shRNA and over-expression of NT-3 can significantly increase neurite growth, although a combination of both vectors did not confer any additional benefit over the vectors used individually. These LVs show promising potential for growth and survival of neurites in injured central nervous system tissue (CNS).

U2 - 10.1002/jgm.1509

DO - 10.1002/jgm.1509

M3 - Article

VL - 12

SP - 863

EP - 872

JO - Journal of Gene Medicine

JF - Journal of Gene Medicine

SN - 1099-498X

IS - 11

ER -