Gene Transfer into Corneal Keratocytes using a Hybrid EBV/retroviral Vector
Journal of the Korean Ophthalmological Society
;
: 571-577, 2002.
Article
in Korean
| WPRIM
| ID: wpr-97862
ABSTRACT
PURPOSE:
We tried to determine the feasibility and efficiency of foreign gene transfer into corneal keratocytes using a hybrid EBV/retroviral vector as an investigative trial for gene therapy in corneal diseases.METHODS:
LZRSpBMN-Z, alac Z-transducing hybrid EBV/retroviral vector, was transfected into Phoenix(T M) amphotropic packaging cells based on a 293T cell line and then collected without/with puromycin selection (puro (-)/puro (+) vector respectively). Cultured human and rabbit keratocytes were transduced with lac-Z gene using the puro (-) or puro (+) vector solutions, then stained with 5-bromo-4-chloro-3-indolyl galactopyranoside (X-gal). FACS-Gal analysis of transduced corneal keratocytes was also performed for calculating gene transfer efficiency. In addition, as an in vivo trial, we tried to transduce rabbit keratocytes by topical application of the vector supernatants following PRK or lamellar dissection of rabbit corneas.RESULTS:
In vitro, both cultred human and rabbit keratocytes were transduced successfully with lac - Z gene. Transduction efficiency was 22% and 16% for human and rabbit keratocytes respectively with puro (-) vector, and slightly increased to 24% and 22% with puro (+) vector. In vivo corneas, however, no keratocytes were stained with X-gal.CONCLUSIONS:
A hybrid EBV/retroviral vector, LZRSpBMN-Z, successfully transduced corneal keratocytes in in vitro conditions but not in vivo corneas.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Puromycin
/
Genetic Therapy
/
Cell Line
/
Product Packaging
/
Cornea
/
Corneal Diseases
/
Corneal Keratocytes
/
Galactose
Limits:
Humans
Language:
Korean
Journal:
Journal of the Korean Ophthalmological Society
Year:
2002
Type:
Article
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