Retroviral vector-mediated gene tranfer into keratocyte in vitro and in vivo Original Reearch Article

PURPOE: To determine the potential of omatic gene tranfer a a technique for modulating corneal wound healing after uperficial keratectomy. METHOD: The tranduction of human and rabbit keratocyte with β-galactoidae and herpe implex viru thymidine kinae gene wa performed. In vitro, human and rabbit keratocyte were tranduced with retroviral vector bearing β-galactoidae or Htk (herpe implex viru thymidine kinae) gene. In vivo, rabbit keratocyte were tranduced by topical application of vector upernatant after a uperficial keratectomy. In vitro and in vivo, expreion of the β-galactoidae gene wa examined with hitochemical taining. In vitro, ganciclovir cytotoxicity in Htk gene-tranduced keratocyte and bytander effect in co-culture of Htk(+) and Htk(−) keratocyte were meaured by determining the degree of confluency of cell in 6-well plate after 10 day of incubation. Corneal haze in rabbit wa meaured after tranduction with Htk and ubequent treatment with topical ganciclovir. REULT: In vitro, both human and rabbit keratocyte were tranduced uccefully with both β-galactoidae and Htk gene. Tranduction efficiency wa greater with human (22%) than with rabbit (16%) cell, and both Htk-tranduced cell line howed doe-dependent ganciclovir cytotoxicity and a ignificant bytander effect. In vivo, expreion of β-galactoidae within vimentin-poitive corneal tromal cell confirmed tranduction of keratocyte in the rabbit after uperficial tromal keratectomy with an efficiency of 25% to 40%. Potoperative application of topical ganciclovir reduced corneal tromal haze in rabbit. CONCLUION: The ability to genetically tranduce tromal keratocyte provide a new trategy for undertanding the important cellular and molecular event that influence corneal wound healing, thu offering a potential approach to decreae or prevent corneal haze and carring after uperficial keratectomy.