Objectives:
We have previously reported preliminary data showing inhibition of bone formation by gingival fibroblasts in vitro (PEF IADR, Dublin 2006). Here we extend these observations; specifically the aim of this study was to test the effects of conditioned medium (CM) from gingival fibroblasts on osteoblast differentiation, proliferation and migration.
Methods:
Rat gingival fibroblasts (RGF) and periodontal ligament cells (PDL) were cultured from tissue explants. The rat osteoblastic cell line ROS 17/2.8 was used to assess osteoblast differentiation. Cultures of ROS cells were stimulated with CM from RGF and PDL cultures and differentiation was assessed by measuring alkaline phosphatase (ALP) activity and osteocalcin levels. The effects of CM on the osteotropic action of 10-8M dexamethasone (Dx) and 100ng/ml BMP-2 were also tested. In addition, the effects of CM on chemotactic migration to PDGF were tested using a micro-Boyden chamber.
Results:
Treatment with of ROS cells increased ALP activity by 4 fold with Dx and 7 fold with BMP-2. RGF CM inhibited constitutive ALP expression by 55% and reduced osteocalcin levels by 50%. RGF CM also largely blocked the effects of Dx and BMP-2 on ALP activity (eg. BMP 1.14 ± 0.07 ALP Units, BMP + RGF CM 0.36±0.12).
In contrast to the effects of RGF CM, CM from PDL directly stimulated ALP expression in ROS cells, and acted synergistically to enhance the action of Dx and BMP-2. Neither CM tested had any effects on osteoblast proliferation or chemotactic responses to PDGF.
Conclusions:
Gingival fibroblasts release factors which inhibit osteoblast differentiation and can block the osteogenic effects of Dx or BMP-2. In contrast PDL cells release factors that appear to promote osteoblast differentiation. The results support the hypothesis that gingival connective tissues may directly inhibit bone regeneration during periodontal and implant-associated surgical procedures.