Recombinant AAV-transduced iris pigment epithelial cell transplantation may transfer vector to native RPE but suppress systemic dissemination.
OBJECTIVE
To determine whether adenoassociated virus (AAV) vectors transduced into iris pigment epithelial (IPE) cells and transplanted into the subretinal space of rats will transfer the AAV genome to the host cells and whether the vectors are disseminated systemically.


METHODS
Recombinant (r)AAV was transduced into rat IPE cells and transplanted into the subretinal space of rats. For the control, rAAVs alone were injected subretinally. The transplanted IPE cells were detected by LacZ staining. Immunohistochemistry, electron microscopy, electroretinography, and fluorescein-dextran angiography were performed. DNA was extracted from various organs and blood and examined for the AAV genome by polymerase chain reaction.


RESULTS
No toxicity from rAAV transduction was observed in vitro. LacZ was expressed in the transplanted cells 1 and 2 weeks after transplantation. At 4 and 12 weeks, fewer transplanted cells were detected than at 1 week, and LacZ expression was occasionally detected at the level of host retinal pigment epithelial (RPE) cells. Expression was also detected in ciliary body epithelial cells. The electroretinograms and fluorescein-dextran angiography were only mildly altered. Significantly lower levels of AAV genome were detected in the organs and blood of rats receiving rAAV-IPE cell transplants than with direct intravenous injection of AAV vectors.


CONCLUSIONS
AAV-mediated LacZ was expressed in the transplanted cells after subretinal transplantation, and the transplanted IPE cells may transfer the rAAV to host tissues, such as RPE cells, long after the transplantation. This method of gene delivery did not lead to systemic dissemination of the vectors.
Yoshioka
Yuki
Y
Department of Ophthalmology and Visual Science, Tohoku University Graduate School of Medicine, Sendai, Japan.
Abe
Toshiaki
T
Wakusawa
Ryosuke
R
Moriya
Takuya
T
Mochizuki
Shizue
S
Saigo
Yoko
Y
Saito
Takae
T
Murata
Hiromi
H
Tokita
Yumi
Y
Iseya
Takashi
T
Sugano
Eriko
E
Tomita
Hiroshi
H
Sagara
Yoshiko
Y
Tamai
Makoto
M
D000818:Animals; D002470:Cell Survival; D017690:Cell Transplantation; D002478:Cells, Cultured; D004279:DNA, Viral / Q000032:analysis; D000229:Dependovirus / Q000235:genetics*; D003911:Dextrans; D004596:Electroretinography; D005110:Extracellular Space; D005452:Fluoresceins; D005822:Genetic Vectors; D016679:Genome, Viral; D007150:Immunohistochemistry; D007498:Iris / Q000166:cytology*; D007763:Lac Operon / Q000502:physiology; D008856:Microscopy, Fluorescence; D010857:Pigment Epithelium of Eye / Q000637:transplantation* / Q000821:virology*; D016133:Polymerase Chain Reaction; D051381:Rats; D020318:Rats, Long-Evans; D012160:Retina / Q000601:surgery* / Q000821:virology; D014161:Transduction, Genetic; D001616:beta-Galactosidase / Q000378:metabolism
D004279:DNA, Viral; D003911:Dextrans; D005452:Fluoresceins; C052526:fluorescein-dextran; D001616:beta-Galactosidase