PAPER 20: CELL-BASED GENE TRANSFER AND TISSUE ENGINEERING IN FRACTURE REPAIR

Abstract

Aim of the study: To evaluate the use of a gelfoam sponge as a scaffold material in delivering osteoblast cells transfected with the VEGF gene for fracture repair.

Methods: In vitro: Osteoblasts were cultured from periosteum of rabbit bone and labeled with the visible CMTMR. Commercially available gelfoam with 12 pieces (each 3 × 3 × 3 mm3) was impregnated and cultured with the labelled cells (1×106) in a 12 wells plate for 1, 3 and 7 days. We embedded the gelfoam with labeled cells in an OCT compound enface, and the sections were then examined under a fluorescent microscope. In vivo: Osteoblasts were transfected with VEGF by use of SuperFect (Qiagen Inc) and cultured for 24 hours. The gelfoam pieces were impregnated with the transfected cells (5×106) saline solution for 30 minutes and placed into a segmental bone defect created in the rabbit tibia for 7 (n=3) and 14 (n=3) days. The specimens including the new bone were cut through each site of the segmental defect and embedded in paraffin. The sections were dewaxed and immunostained with mouse anti-human VEGF.

Results: In vitro: CMTMR-labeled cells survived and were detected within gelfoam at different time intervals (days 1, 3 and 7). In vivo: Immunostained VEGF proteins were visualized in the tissues surrounding the residual gel-foam at the fracture site at days 7 and 14 post surgery.

Conclusion: Our results indicate that the labeled/transfected cells are capable of growth in a gelfoam sponge both in vitro and in vivo.