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Dose Optimization for Long-term rAAV-mediated RNA Interference in the Nigrostriatal Projection Neurons.

Ulusoy, Ayse LU ; Sahin, Gurdal LU ; Björklund, Tomas LU ; Aebischer, Patrick and Kirik, Deniz LU (2009) In Molecular Therapy 17. p.1574-1584
Abstract
Short-hairpin RNA (shRNA)-mediated gene knockdown is a powerful tool for targeted gene silencing and an emerging novel therapeutic strategy. Recent publications, however, reported unexpected toxicity after utilizing viral-mediated shRNA knockdown in vivo. Thus, it is currently unclear whether shRNA-mediated knockdown strategy can be used as a safe and efficient tool for gene silencing. In this study, we have generated rAAV vectors expressing shRNAs targeting the rat tyrosine hydroxylase (TH) mRNA (shTH) for testing the efficacy of in vivo TH knockdown in the nigral dopaminergic neurons. At high titers, not only the shTH vectors but also the scrambled and green fluorescence protein (GFP)-only controls caused cell death. In a dose-response... (More)
Short-hairpin RNA (shRNA)-mediated gene knockdown is a powerful tool for targeted gene silencing and an emerging novel therapeutic strategy. Recent publications, however, reported unexpected toxicity after utilizing viral-mediated shRNA knockdown in vivo. Thus, it is currently unclear whether shRNA-mediated knockdown strategy can be used as a safe and efficient tool for gene silencing. In this study, we have generated rAAV vectors expressing shRNAs targeting the rat tyrosine hydroxylase (TH) mRNA (shTH) for testing the efficacy of in vivo TH knockdown in the nigral dopaminergic neurons. At high titers, not only the shTH vectors but also the scrambled and green fluorescence protein (GFP)-only controls caused cell death. In a dose-response study, we identified a dose window leading to >60% decrease in TH(+) neurons without any change in vesicular monoamine transporter-2 (VMAT2) expression. Moreover, using the safe and efficient dose, we showed that dopamine (DA) synthesis rate was significantly reduced and this lead to emergence of motor deficits in the shTH-expressing rats. Interestingly, these animals showed very robust and long-lasting recovery after a single systemic L-3,4-dihydroxyphenylalanine (L-DOPA) administration beyond what can be achieved in 6-hydroxydopamine (6-OHDA)-lesioned rats. Our results have implications for both mechanistic and therapeutic studies utilizing long-term shRNA-mediated gene silencing in the nigrostriatal projection system.Molecular Therapy (2009); doi:10.1038/mt.2009.142. (Less)
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author
organization
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type
Contribution to journal
publication status
published
subject
in
Molecular Therapy
volume
17
pages
1574 - 1584
publisher
Nature Publishing Group
external identifiers
  • wos:000269534600012
  • pmid:19584816
  • scopus:69949144286
ISSN
1525-0024
DOI
10.1038/mt.2009.142
language
English
LU publication?
yes
id
8142ccbf-8001-4621-8b57-e36a503fe896 (old id 1453299)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/19584816?dopt=Abstract
date added to LUP
2009-08-04 10:51:07
date last changed
2017-10-22 03:48:47
@article{8142ccbf-8001-4621-8b57-e36a503fe896,
  abstract     = {Short-hairpin RNA (shRNA)-mediated gene knockdown is a powerful tool for targeted gene silencing and an emerging novel therapeutic strategy. Recent publications, however, reported unexpected toxicity after utilizing viral-mediated shRNA knockdown in vivo. Thus, it is currently unclear whether shRNA-mediated knockdown strategy can be used as a safe and efficient tool for gene silencing. In this study, we have generated rAAV vectors expressing shRNAs targeting the rat tyrosine hydroxylase (TH) mRNA (shTH) for testing the efficacy of in vivo TH knockdown in the nigral dopaminergic neurons. At high titers, not only the shTH vectors but also the scrambled and green fluorescence protein (GFP)-only controls caused cell death. In a dose-response study, we identified a dose window leading to >60% decrease in TH(+) neurons without any change in vesicular monoamine transporter-2 (VMAT2) expression. Moreover, using the safe and efficient dose, we showed that dopamine (DA) synthesis rate was significantly reduced and this lead to emergence of motor deficits in the shTH-expressing rats. Interestingly, these animals showed very robust and long-lasting recovery after a single systemic L-3,4-dihydroxyphenylalanine (L-DOPA) administration beyond what can be achieved in 6-hydroxydopamine (6-OHDA)-lesioned rats. Our results have implications for both mechanistic and therapeutic studies utilizing long-term shRNA-mediated gene silencing in the nigrostriatal projection system.Molecular Therapy (2009); doi:10.1038/mt.2009.142.},
  author       = {Ulusoy, Ayse and Sahin, Gurdal and Björklund, Tomas and Aebischer, Patrick and Kirik, Deniz},
  issn         = {1525-0024},
  language     = {eng},
  pages        = {1574--1584},
  publisher    = {Nature Publishing Group},
  series       = {Molecular Therapy},
  title        = {Dose Optimization for Long-term rAAV-mediated RNA Interference in the Nigrostriatal Projection Neurons.},
  url          = {http://dx.doi.org/10.1038/mt.2009.142},
  volume       = {17},
  year         = {2009},
}