Tendon healing in vitro: adeno-associated virus-2 effectively transduces intrasynovial tenocytes with persistent expression of the transgene, but other serotypes do not.
BACKGROUND
Transfer of exogenous growth factor genes to injured tendons offers a promising method for strengthening tendon repairs. Adeno-associated virus vectors have advantages of being both nonpathogenic and nontoxic. The authors explored the efficiency of transduction of intrasynovial tenocytes with different serotypes of adeno-associated virus (AAV) and the persistency of its expression of a growth factor transgene.


METHODS
Tenocytes were obtained from cultures of rat intrasynovial tendons and distributed to 82 wells in eight culture plates and to 30 culture dishes. The tenocytes in the wells were treated with AAV1, AAV2, AAV3, AAV4, AAV5, AAV7, and AAV8 vectors containing the lacZ gene, and plasmid vectors (pCMVbeta-lacZ). The tenocytes were stained with in situ beta-galactosidase 5 days later. The basic fibroblast growth factor (bFGF) gene was cloned to the AAV2 vector to construct the AAV2-bFGF vector, which transduced tenocytes in culture dishes. Expression of the transgene was measured over 3 weeks and analyzed statistically.


RESULTS
AAV2 effectively delivered exogenous genes to proliferating intrasynovial tenocytes. In contrast, other tested adeno-associated viruses transduced tenocytes minimally or not at all. The efficiency of gene transfer by AAV2, indicated by the percentage of cells with positive beta-galactosidase staining, was significantly greater than that by a plasmid vector (p = 0.001). Expression of the bFGF gene in tenocytes transduced with the AAV2-bFGF was significantly higher than that in the control over the 3-week period (p < 0.01).


CONCLUSIONS
Gene transfer to tenocytes by AAV2 is more efficient than that by a plasmid vector. However, other adeno-associated virus serotypes cannot effectively transduce tenocytes. The bFGF gene can be delivered to intrasynovial tenocytes by the AAV2 vector effectively, and the gene transfer significantly increases expression of bFGF gene over 3 weeks.
Wang
Xiao Tian
XT
Providence, R.I.; and Tochigi, Yokohama, and Tokyo, Japan From the Department of Surgery, Roger Williams Medical Center, Boston University School of Medicine; Jichi Medical School; Yokohama City University; and Graduate School of Medicine, University of Tokyo.
Liu
Paul Y
PY
Tang
Jin Bo
JB
Mizukami
Hiroaki
H
Xin
Ke-Qin
KQ
Ozawa
Keiya
K
Ushijima
Hiroshi
H
D000256:Adenoviridae / Q000145:classification; D000818:Animals; D002478:Cells, Cultured; D016222:Fibroblast Growth Factor 2 / Q000235:genetics*; D015870:Gene Expression / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013710:Tendons / Q000166:cytology*; D014161:Transduction, Genetic; D019076:Transgenes / Q000235:genetics*; D014945:Wound Healing
D016222:Fibroblast Growth Factor 2