Enhanced short-term plasmid transfection of filtration surgery tissues

PURPOSE. To quantify and localize plasmid transfection of filtration surger y tissues using two delivery techniques. METHODs. Full-thickness filtering procedures were performed on eyes of New Zealand albino rabbits. in 10 eyes, naked plasmid DNA in saline was either injected beneath Tenon's capsule at the filtration site or absorbed into a collagen shield that was then placed external to the sclerostomy and under the Tenon's capsule. Forty-eight hours after surgery, levels of the reporte r gene, chloramphenicol acetyltransferase (CAT) were measured in samples of ocular tissues, in two additional eyes, the beta -galactosidase (beta -Gal ) reporter gene expression was localized histologically. RESULTS. Injection of plasmid DNA in saline vehicle into the filtration ble b produced readily detectable CAT activity in bleb tissue (conjunctiva, Ten on's capsule, and sclera) whereas CAT activity was nearly undetectable in s amples of the cornea, iris-ciliary body, and tissues located opposite the b leb site. Delivery of the plasmid DNA into the bleb through a collagen shie ld increased CAT activity 30-fold over injection of plasmid in saline (2711 +/- 567 mU/mg versus 92 +/- 38 mU/mg). beta -Gal activity was imaged only in the region of the bleb, and microscopic examination showed beta -Gal act ivity localized to Tenon's capsule fibroblasts, with minimal beta -Gal acti vity observed in inflammatory cells or scleral fibroblasts, CONCLUSIONS. Transfection of filtration tissues is enhanced by absorption o f naked DNA into a collagen shield. Furthermore, transfection is localized to the fibroblasts and inflammatory cells of the filtration bleb site. Gene therapy using naked plasmid DNA and a simple collagen shield delivery vehi cle may be useful for regulating wound healing after glaucoma surgery.