The Study Of Tissue Engineered Cartilage Constructed By AdLacZ Gene Modified Goat Auricular Chondrocytes With F127 Pluronic

Objectives: AdLacZ gene transduced goat auricular chondrocytes combined with F127 pluronic scaffold were injected into nude mice subcutaneously. The in vivo effects were evaluated in order to explore preliminary parameters for future chondrogenic differentiation factor in gene therapy on cartilage reconstruction.Methods: Experiments were carried out in Shanghai Ninth People's Hospital affiliated to School of Medicine, Jiao Tong University, from Nov.2004 to Dec.2006. Animals including an adult male goat with average weight of 26-30kg as well as twenty-four 6w male nude mice with average weight of 20g were used in this experiment. Nude mice were randomly divided into four groups. Goat auricular chondrocytes were cultured and transduced with AdLacZ over-expression viruses. Gene transfer efficiency was tested in vitro. Those gene modified chondrocytes were then combined with biodegradable F127 pluronic scaffold and injected into either group of nude mice subcutaneously. Non-transduced chondrocytes combined with F127 scaffold or cells alone or scaffold along in vivo injections were carried out in the remaining groups under control. All samples were collected 2 and 4 weeks later at a time and evaluated via histology, alcian blue staining, Collagen II immunohistochemistry staining and X-gal staining.Results: Data were collected based on all the animals used. An MOI of 50 pfu/cell produced optimal effects in transfer efficiency over 80%. 2 weeks later, less mature catilageous like tissue formed at AdLacZ treated chondrocytes injection sites. 4 weeks later, comparatively more mature cartilage was observed in the sample, and II collagen positive staining was detected in a few cells. Finally, X-gal staining confirmed the expression of LacZ 4 weeks after injection. Non-transduced chondrocytes combined with F127 group showed similar HE and II collagen expression as AdLacZ transduced experimental group, but x-gal staining was negative while cells alone or F127 alone samples did not induce cartilage formation in this protocol.Conclusions: Adenovirus gene transduction achieved comparatively high transfer efficiency on primary cultured chondrocytes. F127 pluronic is a suitable scaffold for tissue engineered cartilage regeneration. This study provides valuable parameters for future research on cartilage regeneration in goat model with a regional ex vivo gene therapy method.