title: Targeted decorin gene therapy delivered with adeno-associated virus effectively retards corneal neovascularization in vivo.
abstract: Decorin, small leucine-rich proteoglycan, has been shown to modulate angiogenesis in nonocular tissues. This study tested a hypothesis that tissue-selective targeted decorin gene therapy delivered to the rabbit stroma with adeno-associated virus serotype 5 (AAV5) impedes corneal neovascularization (CNV) in vivo without significant side effects. An established rabbit CNV model was used. Targeted decorin gene therapy in the rabbit stroma was delivered with a single topical AAV5 titer (100 µl; 5×10(12) vg/ml) application onto the stroma for two minutes after removing corneal epithelium. The levels of CNV were examined with stereomicroscopy, H&E staining, lectin, collagen type IV, CD31 immunocytochemistry and CD31 immunoblotting. Real-time PCR quantified mRNA expression of pro- and anti-angiogenic genes. Corneal health in live animals was monitored with clinical, slit-lamp and optical coherence tomography biomicroscopic examinations. Selective decorin delivery into stroma showed significant 52% (p<0.05), 66% (p<0.001), and 63% (p<0.01) reduction at early (day 5), mid (day 10), and late (day 14) stages of CNV in decorin-delivered rabbit corneas compared to control (no decorin delivered) corneas in morphometric analysis. The H&E staining, lectin, collagen type IV, CD31 immunostaining (57-65, p<0.5), and CD31 immunoblotting (62-67%, p<0.05) supported morphometric findings. Quantitative PCR studies demonstrated decorin gene therapy down-regulated expression of VEGF, MCP1 and angiopoietin (pro-angiogenic) and up-regulated PEDF (anti-angiogenic) genes. The clinical, biomicroscopy and transmission electron microscopy studies revealed that AAV5-mediated decorin gene therapy is safe for the cornea. Tissue-targeted AAV5-mediated decorin gene therapy decreases CNV with no major side effects, and could potentially be used for treating patients.
authors: Harry S. Truman Memorial Veterans' Hospital, Columbia, Missouri, United States of America. mohanr@health.missouri.edu
authors: Rajiv R
authors: RR
authors: Mohan
authors: Jonathan C K
authors: JC
authors: Tovey
authors: Ajay
authors: A
authors: Sharma
authors: Gregory S
authors: GS
authors: Schultz
authors: John W
authors: JW
authors: Cowden
authors: Ashish
authors: A
authors: Tandon
mesh_terms: D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D003315:Cornea / Q000098:blood supply*; D017931:DNA Primers; D058575:Decorin / Q000235:genetics / Q000627:therapeutic use*; D000229:Dependovirus / Q000235:genetics*; D005260:Female; D018390:Gene Targeting; D015316:Genetic Therapy; D007150:Immunohistochemistry; D009389:Neovascularization, Pathologic / Q000628:therapy*; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D060888:Real-Time Polymerase Chain Reaction; D041623:Tomography, Optical Coherence
chemical_list: D017931:DNA Primers; D058575:Decorin; D012333:RNA, Messenger