ADENOVIRUS-MEDIATED HEME OXYGENASE-1 GENE-TRANSFER INTO RABBIT OCULAR-TISSUES

Purpose. Heme oxygenase-1 (HO-1) is a stress protein induced up to 100 -fold within a few hours after exposure to oxidative stress, and it ha s been shown to counteract oxidative injury induced by ultraviolet lig ht or free radicals. The current study was undertaken to determine whe ther the HO-1 gene can be introduced into adult rabbit ocular tissues by microinjection of a recombinant replication-deficient adenovirus hu man HO-1 cDNA (Adv-HHO). Methods. Human HO-1 gene was used for transfe ction studies to differentiate endogenous from transfected HO. The pur ified Adv-HHO construct (10(8) pfu/ml) was mixed with lipofectamine an d microinjected into the anterior chamber, vitreous cavity, and subret inal space of New Zealand rabbit eyes. After 2 weeks, total RNA was ex tracted from different ocular tissues, reverse transcription-polymeras e chain reaction was performed using specific human HO-1 primers, and amplification products were subjected to Southern hybridization. Resul ts. Transfection with the Adv-HHO construct into rabbit corneal epithe lial cells in culture resulted in a functional expression of the human HO-1 gene; the human HO-1 mRNA was detected, and enzyme activity incr eased threefold. Human HO-1 mRNA was detected in the retina after micr oinjection of the Adv-HHO construct into the subretinal space. Microin jection into the vitreous resulted in HO-1 mRNA expression in the corn eal endothelium, iris, lens, and retina; after intracameral injection of the Adv-HHO construct, human HO-1 mRNA was detected in corneal epit helium and endothelium, ciliary body, lens, and iris. Regardless of th e injection site, transfected human HO-1 mRNA was undetectable in tiss ues outside the eye, that is, brain, liver, and kidney. Conclusions. T hese results demonstrated a tissue-selective functional transfer of th e human HO-1 gene into rabbit ocular tissues in vivo. This technique m ay be a promising means for delivering HO-1 gene in vivo as a protecti ve mechanism against oxidative stress that contributes to the pathogen esis of ocular diseases such as cataract, light-induced injury, age-re lated macular degeneration, and diabetic retinopathy.