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(Investigative Ophthalmology and Visual Science. 2001;42:3294-3300.)
© 2001 by The Association for Research in Vision and Ophthalmology, Inc.

Gene Transfer Mediated by Recombinant Baculovirus into Mouse Eye

Françoise Haeseleer1, Yoshikazu Imanishi1, David A. Saperstein1 and Krzysztof Palczewski1,2,3

1 From the Departments of Ophthalmology, 2 Pharmacology, and 3 Chemistry, University of Washington, Seattle.

PURPOSE. To determine the efficiency of baculoviruses (BVs) to transfer recombinant genes in vivo into murine ocular tissues.

METHODS. Recombinant (r)BVs carrying fluorescent protein (FP) cDNA under the control of cytomegalovirus (CMV) immediate early promoter were constructed. Initially, cultured HEK293 and ARPE19 cells were infected with these rBVs and analyzed for efficiency and stability of transgene expression. The rBV-CMV green (G)FP was also injected into the intravitreal and subretinal space of mouse eye. Mice were periodically analyzed to determine the efficiency and stability of expression by histologic examination under fluorescence microscopy. The effect of rBV-CMV-GFP on the physiology of the retina was analyzed by electroretinography.

RESULTS. cDNAs encoding fluorescent proteins were efficiently transduced in HEK293 and ARPE19 cells in vitro. GFP expression in vivo was observed exclusively in retinal pigment epithelial (RPE) cells after subretinal injections. Intravitreal injections of rBV resulted in GFP expression in the corneal endothelium, lens, RPE, and retina. GFP expression was observed for up to 14 days after injection. The infiltration of macrophages, observed 2 days after injection in the area of GFP transduction, had dissipated by day 8 after injection. No alteration in ERG responses was observed 6 weeks after injection of rBV-CMV-GFP.

CONCLUSIONS. BV efficiently transduces cultured RPE cells and many cell types in vivo in the eye, including endothelial, epithelial, and neuronal cells. BV may be a useful vector for transferring genes in cultured cells and in vivo into ocular tissue.




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Copyright © 2001 by the Association for Research in Vision and Ophthalmology