| Autor: |
Marius Ader, G. Jane Farrar, Naomi Chadderton, Marian M. Humphries, Mary O'Reilly, Anna-Sophia Kiang, Peter Humphries, Arpad Palfi, Paul F. Kenna, Sophia Millington-Ward, Niamh McNally, Gearoid Tuohy |
| Rok vydání: |
2006 |
| Předmět: |
|
| Zdroj: |
Molecular Therapy. 13 |
| ISSN: |
1525-0016 |
| DOI: |
10.1016/j.ymthe.2006.08.1116 |
| Popis: |
Top of pageAbstract Retintis pigmentosa (RP) is a term used to describe a group of inherited retinopathies primarily involving photoreceptor cell loss. The clinical and genetic heterogeneity inherent in RP has been highlighted with over 35 disease genes now characterised (http://www.sph.uth.tmc.edu/retnet/home.htm). One of the first genes to be identified as causative of autosomal dominantly inherited RP (adRP) was the rhodopsin gene (RHO) which encodes the light sensitive pigment present in rod photoreceptor cells. Over 100 mutations have been characterised in this gene, highlighting the mutational heterogeneity present in adRP. Gene therapies targeting a disorder such as RHO-linked adRP will likely require suppression of the mutant allele causative of the disease pathology. For this reason, RNA interference (RNAi) has been explored as a means to provide potent suppression of the target RHO gene both in cell culture and organotypic culture, that is, in retinal explant tissue. A number of synthesised siRNAs have been found to elicit potent suppression of expression of RHO in HELA cells as assessed at the mRNA and protein levels. In addition, retinal explants from transgenic mice expressing a RHO gene, were electroporated with short hairpin RNA constructs (shRNAs) at post-natal day zero. Suppression of RHO expression was evaluated by quantitative real-time rt PCR 14 days subsequent to electroporation of shRNAs and notably potent suppression of the target gene was obtained in these retinal explants. Results from the study provide evidence that RNAi may be used as a means of suppressing expression of RHO in cell culture and in the target cell type for adRP, that is, photoreceptors. A therapeutic strategy for adRP, in addition to suppression of the mutant and wild-type RHO genes, will require a replacement element whereby a RHO gene capable of avoiding suppression is supplied. A transgenic mouse has been generated which expresses a wild-type human RHO protein from a cDNA modified at an siRNA target site, exploiting the degeneracy of the genetic code, to avoid suppression by RNAi. Retinal function and thickness as analysed using ERG and histology suggests that the modified human replacement RHO gene rescues the disease pathology in a knockout rho mouse model. This work demonstrates that the replacement element of the strategy works well in a transgenic animal. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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