负载褪黑素复合屏障膜的制备及成骨性能研究

Preparation and Osteogenic Properties of Melatonin-loaded Composite Barrier Membrane

目的:探讨负载不同浓度褪黑激素(melatonin,MT)纳米屏障膜的理化性能及其对小鼠胚胎成骨细胞前体细胞MC3T3-E1增殖和成骨分化的影响。方法:利用静电纺丝技术制备载不同浓度MT(0、0.01、0.05、0.1 mol/L)的丝素蛋白(SF)/聚L-丙交酯-己内酯(PLCL)/MT(SP/MT)生物膜。使用扫描电镜、水接触角、傅里叶变换红外分析进行样品表征及模拟体内环境测定屏障膜的降解和缓释行为。利用DPPH法测定抗氧化能力、CCK-8与细胞粘附实验评估样品的生物相容性。使用碱性磷酸酶(alkaline phosphatase,ALP)染色分析成骨分化关键酶,RT-qPCR检测ALP、OCN、OPN、Runx2成骨相关mRNA表达水平。结果:MT成功负载于SF/PLCL中并持续缓释27 d,SP/MT的抗氧化活性与MT浓度呈正相关。此外,结果显示SP/MT-0.1 mol/L成骨分化关键酶及成骨相关mRNA表达水平明显高于对照组(P<0.05)。结论:SP/MT-0.1 mol/L屏障膜具有优异的理化生物性能,并可以促进MC3T3-E1成骨分化潜能。

Objective:To explore the physical and chemical properties of nano-barrier membranes loaded with different concentrations of melatonin(MT)and explore their effect on the proliferation and osteogenic differentiation of mouse embryonic osteoblast precursor cells(MC3T3-E1).Methods:SF/PLCL/MT biofilms loaded with different concentrations of MT(0,0.01,0.05,and 0.1 mol/L)were prepared by electrospinning technology.SEM,water contact angle,and FTIR were used to characterize the samples,and the degradation and sustained-release behaviors of barrier membranes were measured in an in vitro environment.The antioxidant capacity of the samples was determined by dpph assay.In addition,the biocompatibility of the samples was evaluated by CCK-8 and cell adhesion assay.ALP activity was detected to analyze the key enzymes of osteogenic differentiation,and RT-qPCR was used to test the mRNA expression levels of ALP,OCN,OPN,and Runx2.Results:MT was successfully loaded in SP and sustained for 27 days.The oxidation resistance of SP/MT was positively associated with the concentration of MT.In addition,the expression levels of key enzymes and mRNA related to osteogenic differentiation of SP/MT-0.1 mol/L were significantly higher than those of control group(P<0.05).Conclusion:SP/MT-0.1 mol/L barrier membrane has excellent physicochemical and biological properties and can promote the osteogenic differentiation potential of MC3T3-E1.