Osteoblast proliferation and differentiation on dentin slices are modulated by pretreatment of the surface with tetracycline or osteoclasts

Background: implant surface roughness and chemical composition, as well as other factors, affect the ability of osteogenic cells to form bone adjacent to an implant. The same principles may also apply to the tooth root and so me reports have shown that surface modification of the root may lead to imp roved restoration of the periodontal apparatus. The most common of these su rface modification techniques involves demineralization with citric acid or treatment with tetracycline to expose collagen fibrils, in addition, durin g normal bone remodeling, osteoclasts demineralize the extracellular matrix , leaving resorption pits and exposed collagen fibrils. in this study, the effect of different dentin surface-preparation techniques on osteoblasts we re compared. Methods: Slices of sperm whale dentin were mechanically polished and surfac es were treated with tetracycline-HCl (TCN) or were cultured with mouse bon e marrow cells to create a surface with osteoclast (OC) resorption pits or left untreated. Profilometry, x-ray photoelectron spectroscopy (XPS), and s canning electron microscopy (SEM) were used to evaluate the 3 different den tin surfaces. MG63 osteoblast-like cells were cultured on the 3 different s urfaces and the effect of dentin surface preparation technique on MG63 cell proliferation (cell number), differentiation (alkaline phosphatase specifi c activity of isolated cells and cell layer lysates; osteocalcin production ), and local factor production (transforming growth factor (TGF)-beta1 and prostaglandin E-2 (PGE(2)) compared. Results: Profilometry showed the polished and TCN surfaces were smooth with comparable R-a values, whereas the OC surfaces were slightly rougher due t o resorption pits which covered 3.7% of the surface. XPS measurements showe d that TCN treatment reduced the Ca and P content of the surface, indicatin g that it had dissolved the mineral. Osteoclast-resorption also reduced the Ca and P content, but to a lesser extent. MG63 cell proliferation on polis hed dentin and tissue culture polystyrene was equivalent. In contrast, cell s grown on the TCN- and OC-treated surfaces exhibited increased proliferati on. No effect of surface treatment on cell alkaline phosphatase activity wa s observed, but activity in the cell layer lysates was increased on the TCN - and OC-treated surfaces. Osteocalcin production was reduced on all dentin surfaces, but the greatest reduction was found on the TCN-treated surface. Production of both TGF-beta1 and PGE(2) was increased on the treated surfa ces. All effects were greatest in cultures grown on the TCN-treated dentin, Conclusions: These data indicate that demineralization of the dentin surfac e promotes proliferation of osteoblasts and early differentiation events li ke production of alkaline phosphatase and autocrine mediators such as PGE(2 ) and TGF-beta1. However, later differentiation events like osteocalcin pro duction are decreased. Osteoclast-mediated bone resorption elicits similar responses; less than 4% of the dentin surface resulted in approximately 75% of the response caused by TCN treatment. These observations suggest that g reater attention should be paid to the effects of osteoclastic resorption i n designing methods for enhancing bone and cementum formation adjacent to r oot surfaces.