Rescue of photoreceptors by BDNF gene transfer using in vivo electroporation in the RCS rat of retinitis pigmentosa.
OBJECTIVE
To investigate the feasibility of introducing brain-derived neurotrophic factor (BDNF) gene into retinal pigment epithelial cells in vivo by electroporation and whether this method can rescue photoreceptors of retinitis pigmentosa in Royal College Surgeons (RCS) rats.


METHODS
The BDNF-GFP fusion eukaryotic-expressing plasmid was constructed and subretinally or intravitreously injected into the eyes of RCS rats followed by in vivo electroporation. The expression of BDNF mRNA and protein was detected by RT-PCR and Western immunoblot analysis. The number of surviving photoreceptors was counted, and the TdT-dUTP terminal nick-end labeling (TUNEL) method was used to detect the apoptotic retinal cells at different timepoints after introduction of BDNF plasmid.


RESULTS
Treated eyes showed a significantly higher rescue ratio and a lower number of TUNEL-positive photoreceptors than did the control eyes at various timepoints.


CONCLUSIONS
These findings provide evidence that electroporation is an effective method for gene transfer into retinal pigment epithelial cells, and the rescue of photoreceptors can be achieved by BDNF gene transfection with electroporation.
Department of Ophthalmology, Eye and ENT Hospital, Fudan University, Shanghai, China.
Meng
M
Zhang
Xiaofen
X
Mo
Yuan
Y
Fang
Wenyi
W
Guo
Jihong
J
Wu
Shenghai
S
Zhang
Qian
Q
Huang
D000818:Animals; D015153:Blotting, Western; D019208:Brain-Derived Neurotrophic Factor / Q000235:genetics*; D002452:Cell Count; D018274:Electroporation / Q000379:methods*; D015870:Gene Expression / Q000502:physiology; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D020287:In Situ Nick-End Labeling; D008856:Microscopy, Fluorescence; D020419:Photoreceptor Cells, Vertebrate / Q000473:pathology / Q000502:physiology*; D010957:Plasmids; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011922:Rats, Mutant Strains; D055213:Retinal Pigment Epithelium / Q000378:metabolism*; D012174:Retinitis Pigmentosa / Q000503:physiopathology / Q000628:therapy*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D019208:Brain-Derived Neurotrophic Factor; D012333:RNA, Messenger; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins