Oligonucleotide-polyethylenimine complexes targeting retinal cells: structural analysis and application to anti-TGFbeta-2 therapy.
OBJECTIVE
The aim of this study was to characterize oligonucleotide-polyethylenimine (ODN/PEI) complex preparation for potential transfection of retinal cells in vitro and in vivo.


METHODS
The effect of medium preparation [HEPES-buffered saline (HBS), water] on particle size and morphology was evaluated. Cultured Lewis rat retinal Müller glial (RMG) cells were transfected using fluorescein isothiocyanate (FITC)-ODN/PEI complexes specifically directed at transforming growth factor beta (TGFbeta)-2. Efficacy of transfection was evaluated using confocal microscopy, and regulation of gene expression was assayed using quantitative real-time RT-PCR and ELISA assay. One, 24, and 72 h after injection of FITC-ODN/PEI complexes into the vitreous of rat eyes, their distribution was analyzed on eye sections.


RESULTS
Complexes prepared in HBS were smaller than complexes prepared in pure water and presented a core-shell structure. These particles showed a high cellular internalization efficacy, along with a significant and specific down-regulation of TGFbeta-2 expression and production in RMG cells, correlating with specific inhibition of cell growth at 72 h. In vivo, complexes efficiently transfect retinal cells and follow a transretinal migration at 24 h. After 72 h, ODN seems to preferentially target RMG cells without inducing any detectable toxicity.


CONCLUSIONS
Specific down-regulation of TGFbeta-2 expression using ODN/PEI complexes may have potential interest for the treatment of retinal diseases associated with glial proliferation.
Gomes dos Santos
Ana L
AL
INSERM U 598, 15 rue de l'Ecole de Médecine, 75270, Paris, Cedex 06, France.
Bochot
Amélie
A
Tsapis
Nicolas
N
Artzner
Franck
F
Bejjani
Riad Antoine
RA
Thillaye-Goldenberg
Brigitte
B
de Kozak
Yvonne
Y
Fattal
Elias
E
Behar-Cohen
Francine
F
D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D015536:Down-Regulation; D016503:Drug Delivery Systems; D004563:Electrochemistry; D004797:Enzyme-Linked Immunosorbent Assay; D005614:Freeze Fracturing; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D009457:Neuroglia / Q000378:metabolism; D009841:Oligonucleotides / Q000008:administration & dosage / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics*; D011094:Polyethyleneimine / Q000737:chemistry*; D051381:Rats; D011917:Rats, Inbred Lew; D012160:Retina / Q000166:cytology / Q000187:drug effects*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics*; D053781:Transforming Growth Factor beta2
D009841:Oligonucleotides; D016212:Transforming Growth Factor beta; D053781:Transforming Growth Factor beta2; D011094:Polyethyleneimine