CHARACTERIZATION OF RADIOIODINATED RECOMBINANT HUMAN TGF-BETA-1 BINDING TO BONE-MATRIX WITHIN RABBIT SKULL DEFECTS

Bone healing is regulated in part by the local production of TGF-beta1 and other growth factors produced by cells at the site of injury. The single application of recombinant human TGF-beta1 (rhTGF-beta1) to ca lvarial defects in rabbits induces an accelerated recruitment and prol iferation of osteoblasts within 3 days. This ultimately results in the formation of new bone and the complete closure of the defect within 2 8 days. The persistence and localization of [I-125]rhTGF-beta1 within an osseous defect was investigated after applying a single dose of [I- 125]rhTGF-beta1 formulated in a 3% methylcellulose vehicle. Normal bon e encompassing the defect site, the periosteum, and the gel film cover ing the dura were harvested at 0, 4, 8, and 24 h and 3, 7, and 16 days after [I-125]rhTGF-beta1 application. The defect site-associated radi oactivity was quantitated, visualized by autoradiography, and characte rized by TCA precipitation and SDS-PAGE. Radioactivity was observed in autoradiographs of gross specimens, histologic sections of the bone m atrix, and periosteal tissue surrounding the defect. There was a time- dependent decrease in TCA-precipitable radioactivity; however, radioac tivity was still associated with the bone matrix 16 days after applica tion of [I-125]rhTGF-beta1. SDS-PAGE and autoradiography of the radioa ctivity in homogenized bone and periosteal samples revealed a 25 kD ba nd, suggesting that the radioactivity remaining at the defect site rep resented intact [I-125]rhTGF-beta1. Results of this study indicate tha t rhTGF-beta1 may bind tightly to bone matrix in its active form, and this binding may be associated with the observed increase in osteoblas t number and bone matrix within the calvarial defects.