BMP-2控释涂层对加力状态下牙周膜干细胞成骨分化的影响

目的测试在加力状态下,层层自组装BMP-2控释涂层的缓释效果,并检测其对牙周膜干细胞(periodontal ligament stem cells, PDLSC)的成骨诱导作用。方法 通过层层自组装技术在Flexcell加力六孔板上负载[(PAH/BMP-2)/PSS][COL/ALG]20涂层,利用ELISA检测加力状态下缓释涂层21d内释放的BMP-2浓度；将4~6代PDLSCs种于预置BMP-2控释涂层的BioFlex专用六孔培养板,置于FX-4000T加力系统中,加载动态张应力(10%形变牵张力,20Hz),各组作用时间均为1h/次,3次/d,间隔4h加力,分别培养3、5、7d。对照组为非预置涂层添加BMP-2培养液(100ng/ml)的BioFlex板同条件下加力培养；使用用Real-Time PCR和Western印迹法分析PDLSCs在不同张应力作用前后,其表型标志Runx2, ALP和OCN在 mRNA和蛋白水平的表达变化情况。结果实验结果显示,与对照组相比,实验组加力3、5、7d后,Runx2、ALP和OCN标志物mRNA与蛋白表达差异有统计学意义(P＜0.05)。结论[(PAH/BMP-2)PSS]/[COL/ALG]20涂层对hPDLSCs在加力状态下成骨向分化能力具有显著增强作用。
Objective To assess the effect of self-assemble BMP-2 controlled-release nano-film on osteogenic differentiation of periodontal ligament Stem cells(PDLSCs) under mechanical force. Methods Bioflex Plates were coated with self-assembly [(PAH/BMP-2)/PSS][COL/ALG]20 multi-layers modified nano-films, and then the delivery concentration of BMP-2 was detected by ELISA. The PDLSCs were cultured into six-well Bioflex Plates pre-coated with the multi-layers modified nano-films, and then placed in the Flexcell FX-4000T Tension Plus System. PDLSC were subjected to the distinct mechanical forces(10% static tension force, 20Hz, 1h×3/d, for 3d, 5d or 7d). Control cells were cultured in BioFlex plates with BMP-2(100ng/ml) instead of BMP-2 nano-films. Then the expression of Runx2, ALP and OCN mRNA and protein was detected by Real-Time PCR and Western blot techniques, respectively. ResultsThe expressions of Runx2, ALP and OCN mRNA and protein in BMP-2 nano-biofilm group were significantly higher than those in control group(P＜0.05). Conclusion The BMP-2 controlled release nano-flim can significantly promote the osteoblast differentiation of hPDLSCs cultured under mechanical force