Adenovirus-mediated gene transfer to healing tendon--enhanced efficiency using a gelatin sponge.
Adenovirus-mediated gene transfer is a potential method for enhancing tendon healing. We investigated the transfection of Ad5CMVntLacZ, an adenovirus containing the reporter gene LacZ, in primary cultured human rotator cuff tendon cells and in a rat Achilles tendon healing model in vivo. Ad5CMVempty, the adenoviral vector containing no inserted gene, was used as a control for adenoviral transfection alone. Activity of beta-galactosidase,the protein expressed by LacZ gene, was measured using a beta-galactosidase assay and detected visually by X-gal staining. Cultured cells were successfully transfected without impairing cell viability. Maximal beta-galactosidase activity was detected when cells were transfected at the dose of 1000 PFU/cell. The duration of LacZ expression was six days with a peak value at 24 h post-transfection. A transfection rate of 100% was obtained at 5000 PFU/cell. Successful in vivo transfection by Ad5CMVntLacZ was obtained in healing rat Achilles tendon as confirmed by X-gal staining. 0.4% of tendon cells were transfected when Ad5CMVntLacZ was injected into the tendon at a dose of 10(6) PFU. The rate rose to 2% with 10(8) PFU and 3% with 10(9) PFU. The duration of LacZ expression in vivo was 17 days. Transfection efficiency was enhanced threefold and localization improved when a gelatin sponge was used to deliver the adenovirus. The results demonstrate that adenovirus can be used to deliver a gene of interest to cultured human rotator cuff tendon cells and healing tendon, with gelatin sponge implantation enhancing adenoviral transfection efficiency in vivo.
Dai
Quan
Q
The Orthopaedic Research Institute, University of New South Wales, St. George Hospital Campus, Sydney, NSW, Australia.
Manfield
Laura
L
Wang
Yao
Y
Murrell
George A C
GA
D000125:Achilles Tendon / Q000293:injuries / Q000502:physiology; D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002478:Cells, Cultured; D005780:Gelatin; D018014:Gene Transfer Techniques; D006801:Humans; D007763:Lac Operon; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D017006:Rotator Cuff / Q000166:cytology; D015503:Surgical Sponges; D013708:Tendon Injuries / Q000503:physiopathology / Q000628:therapy*; D014162:Transfection; D014945:Wound Healing
D005780:Gelatin