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L. K. Buie, M. Z. Karim, T. Borrás; Establishing an Elevated IOP Rat Model by Gene Transfer to the Trabecular Meshwork (TM). Invest. Ophthalmol. Vis. Sci. 2009;50(13):4841.
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To create an elevated IOP animal model that preserves the TM tissue and thus be useful to evaluate candidate TM drugs. Most elevated IOP models available are obtained by scarification and irreversible damaging of the TM. Our goal is to set up a gene transfer strategy to induce a controlled increase in TM resistance. We propose to overexpress the Bone Morphogenetic Protein 2 (BMP2) gene (an inducer of mineralization) to alter ECM configuration and raise IOP.
AdhBMP2 vector carries the human BMP2 cDNA driven by the CMV promoter (1). AdNull carries the same promoter without transgene. Brown Norway rats (n=35) received a single dose (intracameral injection of 5X109 viral particles) of AdhBMP2 (OS) or AdNull (OD). IOPs (Tonopen and calibrated Tonometer) were taken before injection (baseline) and once-twice a week thereafter. Animals were euthanized at 2 and 6 weeks, 2 and 4 months post injection. Differential pressure was evaluated by subtracting the mean integral pressure (defined as the cumulative IOP each eye received during the experiment) of the AdhBMP2-treated eyes vs that of the untreated. Anterior segments were fixed and processed for general morphology. RNA was extracted from the angle tissue, hybridized to BMP2 TaqMan ABI probe Hs00154192 and normalized to 18S.
AdhBMP2-injected eyes displayed a consistent increase in IOP up to 4 months post injection while those injected with AdNull maintained IOPs similar to baseline. Pressure elevation was evident at 10 days and the difference became significant at 24 days weeks post injection. In a representative tonopen experiment (n=10), at 2 weeks, the mean integral IOP of the treated eyes was 567.4 ± 4.9 and that of the untreated was 494.8 ± 12.5 mmHg-days. At 48 days, the mean integral IOP of the treated eyes was 1886.6 ± 58.3 mmHg-days and that of the untreated was 1403.2 ± 31.5 . The integral IOP differences were 72.6 (p ≤ 0.0001) at 16 days, and 483.4 (p ≤ 0.00002) at 48 days. Normalized TaqMan reactions of angle tissue RNA showed high expression of BMP2 in the treated eyes and no detection of the human transgene in the AdNull-injected eye. First morphological evaluation showed a normal TM. Further characterization is ongoing.
Gene transfer of BMP2 by injection of AdhBMP2 into the anterior chamber of the living rat induces elevated IOP while preserving the structure and cells of the TM. This gene transfer application strategy will be very useful for the evaluation of lowering IOP TM drugs. It will also open the door to the creation of large animal models tailored to the identification of specific drugs, and to the assay of other glaucoma pharmaceuticals targeting the TM.(1) Olmsted et al. JCB 82, 11 (2001)
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