Gene transfer to ex vivo stored corneas.
OBJECTIVE
We examined the efficiency and kinetics of recombinant adenovirus vector-mediated gene transfer to rat and rabbit cornea in culture ex vivo.


METHODS
A recombinant replication-defective adenovirus was used to transfer a lacZ marker gene to whole rat and rabbit corneas in culture. Histochemistry was used to localise transgene expression and a colorimetric assay to quantify recombinant protein expression.


RESULTS
After infection with recombinant virus and culture for 3 days, high-efficiency gene transfer was found, with expression in most endothelial cells of both species. Minimal expression was found in other corneal cell types. On histochemistry, longer duration of expression was found in rat than in rabbit endothelium. In both rat and rabbit cornea, highest levels of recombinant protein were found at days 3-7 after incubation with virus, decreasing to low or undetectable levels at 21 days.


CONCLUSIONS
Adenovirus vectors allow high-efficiency transgene expression in cornea, largely restricted to the endothelial cells of ex vivo cultured cornea. Kinetics of expression differ according to the species of cornea studied, a factor that must be considered if this vector is used in further studies.
Fehervari
Z
Z
Department of Immunology, Royal Postgraduate Medical School, Hammersmith Hospital, London, England.
Rayner
S A
SA
Oral
H B
HB
George
A J
AJ
Larkin
D F
DF
D000256:Adenoviridae / Q000235:genetics / Q000502:physiology*; D000818:Animals; D003315:Cornea / Q000378:metabolism / Q000821:virology*; D003673:Defective Viruses / Q000235:genetics; D004728:Endothelium, Corneal / Q000378:metabolism / Q000821:virology; D015967:Gene Expression Regulation, Viral; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006651:Histocytochemistry; D007763:Lac Operon / Q000235:genetics*; D009926:Organ Preservation; D011817:Rabbits; D051381:Rats; D011914:Rats, Inbred BN; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001616:beta-Galactosidase