Recombinant adeno-associated virus serotype 2 (rAAV2)-An efficient vector for gene delivery in condylar cartilage, glenoid fossa and TMJ disc in an experimental study in vivo.
OBJECTIVE
To elaborate whether rAAV2 can be used for future TMJ gene therapy, we examined the infection efficiencies of rAAV2 in vitro, and the transgene expression pattern mediated by rAAV2 in glenoid fossa, TMJ disc and condylar cartilage in vivo.


METHODS
Different dosages of rAAV2-eGFP (MOI: 5 x 10(4), 1 x 10(4), 5 x 10(3)) were applied to primary cultured condylar chondrocytes of rats. Infection efficiencies were analysed by FACSCalitur at different time points. Vastatin, a molecule not naturally expressed in TMJ, was used as a reporter for detection of rAAV2 mediated transgene expression in vivo. Thirty SD rats were injected with either rAAV2-sec-Vastatin (experimental group) or rAAV2-eGFP (control group) into both sides of TMJ. They were sacrificed at the indicated time (7, 14, 21, 30 and 60 days of injection) and the TMJ samples were collected for RT-PCR and immunostaining analysis.


RESULTS
High dosage (MOI 5 x 10(4)) of rAAV2-eGFP can achieve desirable transduction efficiencies in vitro after 5 days. Transgene expression of rAAV-sec-Vastatin persisted for about 21 days in glenoid fossa, around 7 days in TMJ disc and at least 60 days in condylar cartilage in vivo. In condylar cartilage, transgene expression was found in the proliferative layer and chondroblast layer (day 7), chondrocyte layer (day 14), pre-hypertrophic and hypertrophic layer (day 21), hypertrophic layer and deep hypertrophic layer (day 30 and 60).


CONCLUSIONS
Recombinant AAV2 could be considered as a promising vector for gene therapy in TMJ which can mediate therapeutic gene expression in glenoid fossa, articular disc and condylar cartilage in vivo.
Li
Qianfeng
Q
Faculty of Dentistry, The University of Hong Kong, PPDH, Sai ying pun, HKSAR, China. qianfengll@yahoo.com.cn
Dai
Juan
J
Rabie
A Bakr M
AB
D000818:Animals; D002358:Cartilage, Articular / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D024102:Collagen Type VIII / Q000096:biosynthesis*; D000229:Dependovirus / Q000235:genetics*; D005260:Female; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D008335:Mandibular Condyle / Q000378:metabolism*; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D013701:Temporal Bone / Q000378:metabolism; D019224:Temporomandibular Joint Disc / Q000378:metabolism*; D019076:Transgenes
D024102:Collagen Type VIII; C441971:vastatin; D049452:Green Fluorescent Proteins