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J. Sassani, P.J. McLaughlin, K.J. Malefyt, I.S. Zagon; Gene Expression of the Opioid Growth Factor Receptor and the Regulation of Corneal Re–Epithelialization . Invest. Ophthalmol. Vis. Sci. 2006;47(13):5025.
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© ARVO (1962-2015); The Authors (2016-present)
The opioid growth factor (OGF) interacts with the OGF receptor (OGFr) to regulate cell proliferation by a tonic inhibitory action. Disruption of the OGF–OGFr axis by the opioid antagonist naltrexone (NTX) results in acceleration of corneal re–epithelialization in rat, rabbit, and human. To examine the molecular nature of growth regulation by the OGF–OGFr axis, DNA constructs designed to overexpress (Sense) or underexpress (Antisense) OGFr were delivered to the corneal epithelium of adult rats by particle–mediated gene transfer using a gene gun.
The plasmid pcDNA3.1–OGFr, carrying the rat OGFr cDNA in both sense (Sense rats) and antisense (AS rats) orientation, under the control of the CMV immediate early promoter, was delivered twice at 300 psi into one eye of each rat; empty vectors were transfected into other animals (EV rats). This regimen resulted in >90% transfection of the central and peripheral corneal epithelium without penetration into the stroma. After a 24–hr latency, a corneal abrasion was created using a 3 mm trephine. Corneal wounds were tracked photographically with fluorescein dye, and areas quantitated using Optimas software. Overexpression of OGFr would be expected to result in delayed wound healing because of the enhanced ability of receptor – inhibitory peptide interaction.
At 16 hr, animals transfected with OGFr–Sense constructs had residual wounds that were 22.3% larger than those in rats transfected with empty vectors. At 24 and 28 hr, the residual wounds in Sense group were 49.1% and 46.7% larger, respectively, than there empty vector counterparts. Disruption of the OGF–OGFr axis would be predicted to accelerate corneal wound healing if this peptide and receptor complex played a direct role in re–epithelialization. At 16 hr after wounding, the residual defect was 40% smaller for AS rats than for EV rats. After 24 hr, the defect in rats transfected with antisense was 98% smaller than for EV animals. Rats in all groups were injected with BrdU to determine DNA synthesis rates. AS rats had significantly increased levels of BrdU labeled cells in the basal (>3–fold) and suprabasal (up to 8–fold) layers of cornea, limbus, and conjunctiva relative to both EV and Sense animals.
These data reveal that the OGF–OGFr axis plays a direct role in re–epithelialization processes in the cornea. Moreover, these gene gun studies support the use of gene therapy for ocular diseases.
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