Delivery of transforming growth factor-β2-perturbing antibody in a collagen vehicle inhibits cranial suture fusion in calvarial organ culture

Objective: To determine whether antibody perturbation of Tgf-β, delivered in a collagen gel, could inhibit cranial suture fusion. Design: Attachment and proliferation of osteoblasts cultured on a collagen gel with or without anti-Tgf-β2 antibody were determined by AlamarBlue dye assay and cell morphology by toluidine-blue staining. In rat calvarial organ culture, collagen gel with and without anti-Tgf-β2 antibody was injected subperiosteally over the posterior frontal suture of postnatal day 15 rat calvariae. A quantitative analysis of suture fusion was used to measure suture bridging in histological serial sections at various time points. Results: Attachment and proliferation for cells cultured on collagen gel with anti-Tgf-β2 antibody were similar to collagen gel controls. Although proliferation was lower than on tissue culture plastic, cells treated with anti-Tgf-β2 antibody maintained an osteoblastic morphology. After 7, 10, and 15 days in organ culture, anti-Tgf-β2 antibody treatment caused a reduction in the percent bridging of posterior frontal sutures, compared with controls. Sutures exposed to anti-Tgf-β2 antibody and fibroblast growth factor-2 concurrently did not show an inhibition of bony bridging. Conclusions: These results support previous reports suggesting a role for Tgf-β2 in cranial suture fusion. In cell culture the collagen gel, both with and without anti-Tgf-β2 antibody, promoted similar osteoblast attachment, proliferation, and osteoblastic morphology. In organ culture anti-Tgf-β2 antibody was delivered in a bioactive state via a collagen gel to inhibit cranial suture fusion. Also, the results suggest that the inductive effect of fibroblast growth factor-2 is not dependent on Tgf-β2 activity. Together, these results provide further support for the role of Tgf-β2 in cranial suture fusion.