The Effect Of Brushite Coating Doped With Strontium On Proliferation And Differentiation Of ME3T3-E1Cells

Objectives To study the physicochemical properties of brushite coating deposited ontitanium surface, and to compare the effect of cytotoxicity, proliferation, differentiationand the expression of gene of different concentration of strontium in titanium-brushitelayer for MC3T3-E1cells. To explore the feasibility of titanium-brushite compositebecoming new implant materials, and to discuss whether strontium can help brushite toplay a role in osteogenesis and improve the biological performance. We hope this studycan provide some reference data for in vivo study and clinical application of toothimplant.Methods1Ca-P coating was deposited on titanium surface through electrochemicaldeposition. Analysed the coating composition through EDS and XRD, and observed themicrostructure of material surface through SEM.2Cultivated MC3T3-E1cells, andobserved the cell growth pattern through Giemsa staining.3detected the toxicity andappreciation of preosteoblasts among the titanium group, and the strontium-brushitegroup of different concentrations.4Investigated its biocompatibility through cultivatedMC3T3-E1cells in vitro on the composite materials, and observed the adhesion,morphology, proliferation of cells.5Detected the content of ALP for quantitative.6Cellscalcified nodules were stained with Alizarin red.7Real-Time PCR was used toquantitative analyse the effect of brushite coating materials with different quantity ofstrontium on preosteoblast about the expression of BMP-2and COL-I.Results1We used electrochemical deposition to prepare the coating which Ca, P ratiowas1:1, the main components was brushite; SEM observation showed that the coatingsurface was uneven, the crystal was dense, like a cauliflower, more rough than thetitanium surface.2MC3T3-E1cell line belongs to the adherent cell. Cell morphologywas spindle shaped, irregular triangle or polygon in the microscope, each protrusionextending connection each other, the cell nucleus was round, cell aggregation was tooclose to overlapping growth.3The CCK-8cell toxicity test showed that the experimentalgroup toxicity were lower than grade2, the absorbance value of0.5%,1%and5%grouphad a significant difference compared with the other groups (P＜0.05).4MC3T3-E1cellwhich adhere and spreading on the coating material was good, no cell shrinkage, no changed round, and no morphology changing, SEM showed that cells grew into the poresof material. With the extension of time, the cell confluent overlaied coating surface.5The ALP expression of cells was higher level on the strontium containing coating. Theactivity of ALP of cells on brushite was higher than the pure titanium at the fourteenthdays (P＜0.05).6The osteoblast mineralized nodule staining showed that the amount ofcontaining strontium group nodule formation increased, the time was shortening.7Theexpression levels of BMP-2in the experimental group were significantly higher than thatof control group(P＜0.05). The expression of differentiation marker gene COL-Ⅰinstrontium group was higher than the control group.Conclusions1The brushite coating and strontium have no toxicity on osteoblasts, thebrushite coating can promote the proliferation of preosteoblast. They have a goodcompatibility and bioactivity of biological.2Strontium-brushite not only improves theproliferation of osteoblast on the titanium-brushite composite, but also promotes the cellcalcification, as well as the high expression of ALP and BMP-2. The addition ofstrontium promotes the differentiation of preosteoblast, and can assist the brushite toproduce osteogenic effects.