Nanostructured Zirconia Surfaces Regulate Human Gingival Fibroblasts Behavior Through Differential Modulation of Macrophage Polarization.

Zirconia exhibits excellent biocompatibility and is widely used as dental implant materials in prosthodontics. Over the past years, research and development of dental implant biomaterials has focused on osseointegration, but few reports exist regarding the role of the immune environment on cellular responses to these materials. The present study investigates the effect of different nanostructured zirconia surface topographies on macrophage phenotypes and their influence on gingival fibroblast behavior. Three different nanostructured zirconia surfaces are characterized using scanning electron microscopy, atomic force microscopy, and water contact angle. Blank-machined zirconia (BMZ) surfaces were superior to RAW264.7 cell proliferation and adhesion. RAW264.7 seeded on all nanostructured zirconia surfaces polarized toward both inflammatory M1 and anti-inflammatory M2 macrophages with more M2 macrophage phenotype on BMZ surfaces. Meanwhile, conditioned media (CM) from RAW264.7 culture on three nanostructured zirconia surfaces inhibited cell apoptosis to human gingival fibroblasts (HGFs) but promoted HGF proliferation and secretion. Under modulation of RAW264.7 culture, HGFs cultured on BMZ surfaces significantly secreted more extracellular matrix with a higher expression of collagen-I (COL-I), vinculin (VCL), and fibronectin (FN) than those coated on self-glazed zirconia (CSGZ) and self-glazed zirconia (SGZ) surfaces. After being coated with a nano zirconia film, CSGZ surfaces showed certain changes in cell proliferation, adhesion, and protein production compared with SGZ surfaces. These findings will provide an overview of manipulating surface topography to modulate macrophage phenotypes in order to create an effective macrophage immune response and reinforce soft tissue integration.

Concentrated growth factor exudate enhances the proliferation of human periodontal ligament cells in the presence of TNF­α.

The purpose of this study was to evaluate the effects of concentrated growth factor exudate (CGFe) on human periodontal ligament cells (hPDLCs) stimulated by tumor necrosis factor (TNF)­α. From the peripheral blood of healthy donors, CGFe was prepared according to the Sacco protocol after 7 days of incubation. The hPDLCs were cultured by a tissue explant method and identified with anti­vimentin and anti­cytokeratin antibodies. Cells were subjected to four different treatments: i) Control; ii) TNF­α (10 ng/ml); iii) CGFe (concentration 50%); and iv) CGFe+TNF­α. The proliferation of hPDLCs was measured with Cell Counting Kit­8 assays. Osteogenic differentiation and mineralization were determined by Alizarin Red S staining, alkaline phosphatase activity, western blotting and reverse transcription­quantitative polymerase chain reaction. CGFe enhanced cell proliferation and upregulated ALP activity, the mineralization level, and osteogenic­associated osteocalcin, runt­related transcription factor 2 and Osterix gene expression in hPDLCs under inflammatory conditions induced by TNF­α. The present study demonstrated that CGFe enhanced hPDLC proliferation and osteogenic differentiation in the presence of TNF­α­induced inflammation. As CGFe can be obtained from the venous blood of patients, it generates no immune reaction. Thus, it is more economical and beneficial to use CGFe in clinical periodontal regeneration practice than synthetic growth factors.

Platelet­rich fibrin exudate promotes the proliferation and osteogenic differentiation of human periodontal ligament cells in vitro.

The purpose of the present study was to evaluate the effects of platelet­rich fibrin (PRF) exudate on the proliferation, osteogenic differentiation and mineralization of human periodontal ligament cells (hPDLCs) in vitro. In the present study PRF was obtained with permission, from the peripheral blood of healthy donors and PRF exudates were collected on the 7th day of incubation. hPDLCs were obtained from healthy premolars, cultured by a tissue explant method and identified with anti­vimentin and anti­cytokeratin antibodies. PRF exudates were added to hPDLCs in different concentrations to evaluate cell proliferation and osteogenic differentiation. The proliferation of hPDLCs was measured using a colorimetric assay. Osteogenic differentiation and mineralization were determined by Alizarin red staining, alkaline phosphatase activity (ALP), western blotting and reverse transcription­quantitative polymerase chain reaction. Cell proliferation was enhanced by addition of the PRF exudate, which also promoted the formation of mineralized matrix nodules and upregulated ALP activity and osteoblast­associated levels of osteocalcin, runt­related transcription factor and osterix gene expression. As these stimulatory effects occurred in a dose­dependent manner, it was concluded that high concentrations of the PRF exudate served an essential role in the proliferation, osteogenic differentiation and mineralization of hPDLCs in vitro. The present study demonstrated that PRF exudate enhanced hPDLC proliferation, induced the osteoblastic differentiation of hPDLCs into mineralized tissue­formation cells in vitro, and may therefore provide potential benefits for periodontal tissue engineering; contributing to the primary processes of periodontal tissue regeneration. From the perspective of both economics and biology, PRF has greater clinical benefits than analogous growth factors.