In vitro transduction of human osteobalstic cells using retroviral vectors

Objectives:The involvement of cytokines in degeneration and inflammation of human tissue is well established. Interleukin-1 (IL-1) is a major agent in the pathophysiology of periarticular bone resorption in rheumatoid arthrit is and in osteoporosis. Because the use of recombinant cytokines and growth factors is Limited due to their short half lives, techniques are needed to get a permanent release of these therapeutic proteins. The rational of thi s study was to show that retroviral transduction of human osteoblastic cell s is possible in vitro using the marker gene LacZ and the potentially thera peutic gene encoding for human interleukin-1 receptor antagonist protein (I L-1Ra). Different transduction techniques were combined to improve the rate of transduction in vitro. Methods: Osteoblastic cells were isolated from human spongious bone and cul tured in vitro. The beta-galactosidase (LacZ) gene and the cDNA of IL-1Ra w ere introduced into the isolated cells by retrovirus mediated gene transfer . LacZ activity was determined by Xgal staining, IL-1Ra was measured quanti tatively by ELISA. Results: The transfer of retroviral IL-1Ra led to IL-1Ra expression of 8614 to 10089 pg IRAP/50 000 cells/48 h. By combining different techniques to i mprove transduction, the X-gal staining established a rate of transduction of 60%. Conclusion: Our results demonstrate that retroviral transduction of human o steobalstic cells is possible in vitro, and leads to high levels of the syn thesized transgene product. The rate of retroviral transduction can be acce lerated in vitro.