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Recombinant adenovirus-mediated gene transfer into the adult rat retina
1Laboratory of Immunology, National Eye Institute, National Institute of Health, Bethesda, MD, USA.
Purpose:
The present study was designed to evaluate the feasibility of gene transfer into the retina of adult rats, using a recombinant replication-defective adenovirus vector expressing a reporter gene.
Methods:
Purified recombinant adenovirus expressing beta-galactosidase (lacZ) (Ad5.hCMV.lacZ) at doses ranging from 1.4 x 10(2) to 1.4 x 10(6) plaque-forming units (pfu) were injected into the subretinal space of adult Lewis rats. The presence of lacZ was determined by histochemical assay and reverse transcription and polymerase chain reaction analysis (RT PCR) of total RNA extracted from eyes injected with recombinant adenovirus expressing lacZ.
Results:
As assessed by biomicroscopy, the expression of lacZ was highest in the retinal pigment epithelium in a localized area corresponding to the site of injection. The level of lacZ expression was correlated with the amount of virus delivered to the subretinal space. Persistent but decreasing expression of lacZ was noted over time. RT PCR revealed the expression of messenger RNA for at least sixty days.
Conclusions:
The results of this study demonstrate that efficient and stable transfer of genetic material into the subretinal space of adult rats may be achieved using a recombinant adenoviral vector. The use of such vectors should prove useful in developing novel applications and approaches to the study of recombinant protein expression in vivo.