Involvement of the Wnt-ß-catenin signalling antagonists, sclerostin and dickkopf-related protein 1, in chronic periodontitis.

AIM: The regulation of Wnt-ß-catenin signalling, which is crucial for osteoblast differentiation and for bone resorption, is driven by critical inhibitors such as sclerostin (SOST) and dickkopf-related protein 1 (DKK1). As such, the aim of this study was to evaluate the involvement of SOST and DKK1 in human chronic periodontitis. MATERIAL AND METHODS: Gingival biopsies and serum were sampled from systemically healthy non-periodontitis (n = 15) and chronic periodontitis subjects (n = 15). The mRNA and protein levels of SOST, DKK1 and TNF-α in periodontal tissues were measured by qPCR and by enzyme-linked immunosorbent assay (ELISA) respectively. Serum levels of SOST, DKK1 and TNF-α were assessed by ELISA. RESULTS: The mRNA and protein levels of SOST, DKK1 and TNF-α were significantly increased in the gingival tissues of the chronic periodontitis when compared to the non-periodontitis group (p < 0.05). In addition, circulating levels of SOST and TNF-α, but not DKK1, were higher in the periodontitis group than in the non-periodontitis group (p < 0.05). CONCLUSION: SOST and DKK1 were upregulated in the periodontal tissues of chronic periodontitis subjects, suggesting a possible role of these molecules on periodontal tissues.

In-vitro analysis of rhBMP-2 effects in human osteogenic cells.

This study evaluated the in vitro expression of bone-related proteins by osteoblasts in the presence of different concentrations of human recombinant bone morphogenetic protein-2 (rhBMP-2). Immortalized human fetal osteoblastic cell line 1.19 (hFOB) were exposed to different concentrations of rhBMP-2 (10, 50, or 100 ng/mL) for 72 h. Cell proliferation and viability (MTT assay), as well as the expression of fibronectin, osteonectin, and osteopontin were assessed by indirect immunofluorescence and Western blot. Neither of the 3 concentrations of rhBMP-2 caused statistically significant alterations in cell proliferation and viability, although the concentration of 100 ng/mL showed lower values for both assays after both 48 and 72 h of exposure. There was no alteration in the expression of noncollagenous proteins, as analyzed by immunofluorescence, when compared with the control group. Furthermore, in the Western blot assay we observed a statistically significant decrease in fibronectin and osteonectin at 100 ng rhBMP-2/mL (p < 0.05) by comparison with the medium alone. The expression of osteopontin decreased slightly in all 3 concentrations of rhBMP-2 tested; however, the change was not statistically significant (p > 0.05). In this in-vitro study, the tested concentrations of rhBMP-2 appeared to decrease the expression of important bone-related molecules in pre-osteoblast cells.

Low doses of 15d-PGJ2 induce osteoblast activity in a PPAR-gamma independent manner.

Peroxisome proliferator-activated receptor-gamma (PPARγ) regulates both glucose metabolism and bone mass. Evidence suggests that the therapeutic modulation of PPARγ with synthetic agonists activity may elicit undesirable effects on bone. However, there is no information regarding its natural agonist 15d-PGJ2, besides its excellent anti-inflammatory action. In the present study the effects of 15d-PGJ2 on osteoblastic cells were determined. Osteoblastic cells (MC3T3) were cultured in an osteogenic medium in the presence of 1, 3 or 10 µM of 15d-PGJ2 during 21 days and alizarin and Von Kossa staining were employed. The protein expression (type-I collagen, osteonectin, osteopontin, RANKL, osteoprotegerin, HDAC-9c and PPAR-γ) was evaluated after 3 days in the presence of 15d-PGJ2 by western blotting and indirect immunofluorescence methods. The production of mineralized extracellular matrix was observed by transmission electron microscopy. After 72 h of culture, the mRNA was extracted for RT-qPCR analysis of RUNX expression. In the presence of all 3 tested 15d-PGJ2 doses, alizarin red and Von kossa staining were positive demonstrating the ability to the osteoblast differentiation. Type-I collagen and osteonectin proteins expression were up-regulated (p < 0.05) after 72 h in the presence of the smaller doses of 15d-PGJ2. In contrast, osteopontin, RANKL and OPG expression did not significantly alter. In the presence of 15d-PGJ2 it was possible to visualize mineralized nodules in the extracellular matrix confirmed with the increased RUNX mRNA expression. 15d-PGJ2 at small doses increased the osteoblast activity and the bone-related proteins expression.