Novel Bioceramic Nano-Composite Prepones Osteoblast Differentiation

Objectives: To substantiate the hypothesis that hydroxyapatite(HAP)/GEMOSIL (gelatin modified silane) temporally accelerates transcription factors of mouse osteoblasts (MC3T3).

Methods: MC3T3 were cultured on bioceramic material plates and analyzed for cell proliferation and differentiation and compared against control plates without material coating. Bioceramic (HAP/GEMOSIL) was synthesized using hydroxyapatite-gelatin nanocomposites and aminosilane. Cells were seeded on 35-mm dishes at a density of 100000 cells per milliliter of media. Cells were pre-cultured for four days to reach confluence and evaluated 0, 3, and 7 days post-differentiation. To test our hypothesis, runt-related transcription factor 2 (RUNX2) and Alkaline Phosphatase (ALP) activity were quantified by real-time polymerase chain reaction (RT-PCR) method and colorimetric analysis, respectively. In addition, 5-bromo-2-deoxyuridine (BrdU) incorporation was used to confirm cell proliferation after pre-culturing for 24 hours. Two-way ANOVA was performed to compare experiment to control, in which the two independent factors were time and material. Visual indication for calcification was achieved by Alizarin Red staining.

Results: Real-time PCR results confirmed that RUNX2 on material coated plates were significantly higher relative to control plates on Day 0, similar values on Day 3, and lower values on Day 7. This indicates HAP/GEMOSIL may cause differentiation antecedently. Other translational factors (e.g., collagen type Iα1 and bone sialoprotein) quantified by RT-PCR demonstrated that material plates did not increase its expression. Post-translational factor such as ALP did not reveal any perceptible differences. BrdU assay showed positive stain (10% of total cells) on material with a cell density of nearly 8 times more than control. This suggests that HAP/GEMOSIL may also accelerate cell proliferation.

Conclusion: HAP/GEMOSIL material, if applicable to clinic as an alloplastic graft, may promote initial bone healing. This is the first experiment to exhibit that HAP/GEMOSIL encourages in vitro osteoblastic activity.(Supported in part by AAOF, UNC Dental School)