包封负载EGCG的介孔聚多巴胺纳米粒子的壳聚糖/F127 复合水凝胶的构建及骨再生研究

Construction and bone regeneration study of chitosan/F127 composite hydrogels encapsulating mesoporous polydopamine nanoparticles loaded with EGCG

  • 摘要: 背景 骨缺损修复是临床骨科的常见难题,传统修复材料存在生物活性低、药物缓释效果差等缺陷。目的 结合表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)与介孔聚多巴胺(mesoporous polydopamine,MPDA)的特性,并利用壳聚糖/F127 水凝胶良好的生物相容性及机械性能,构建可以促进骨缺损修复的复合水凝胶。方法 以普通培养基处理的细胞作为对照(control,Ctrl)组;制备负载EGCG的MPDA(MPDA@EGCG)纳米粒子,并通过扫描电子显微镜、纳米粒径电位仪对其粒径、电位、形态学进行观察;将上述纳米粒子与壳聚糖/F127 水凝胶相结合制备复合水凝胶(MPDA@EGCGChitosan/F127,MPDA@EGCG-CF),并通过扫描电子显微镜、流变仪、CCK-8 试剂盒、细胞活/死染色、溶血实验、碱性磷酸酶染色及定量分析、茜素红染色及定量分析、免疫印迹试验对该复合水凝胶的形态学、流变特性、体外释放特性、安全性以及体外促成骨能力进行检测。结果 成功构建MPDA@EGCG-CF水凝胶。体外释放结果显示,该复合水凝胶可在局部持续释放EGCG长达一周以上。形态学观察结果显示,该水凝胶有着良好的形态学特征,为多孔结构。细胞实验及溶血实验结果显示,该复合材料有着良好的安全性;碱性磷酸酶染色、茜素红染色及免疫印迹试验结果表明,相较于Ctrl 组以及包封非载药MPDA水凝胶组,其具备明显的体外促成骨分化能力(P<0.05)。结论 所构建具有骨诱导性的包封负载EGCG的MPDA的壳聚糖/F127 复合水凝胶可实现EGCG的局部长期释放,在体外可明显促进成骨分化,展现出良好的骨再生应用潜力。

     

    Abstract: Background Bone defect repair is a common challenge in clinical orthopedics, and traditional repair materials are limited by drawbacks such as low biological activity and poor drug controlled-release performance. Objective To construct a composite hydrogel capable of promoting bone defect repair by combining the properties of epigallocatechin gallate (EGCG) and mesoporous polydopamine (MPDA), while leveraging the excellent biocompatibility and mechanical properties of chitosan/F127 hydrogel. Methods Cells treated with regular culture medium were used as the control (control group, Ctrl group). EGCG-loaded MPDA (MPDA@EGCG) nanoparticles were prepared, and their particle size, Zeta potential, and morphology were characterized using scanning electron microscopy (SEM) and a nanoparticle size and Zeta potential analyzer. Subsequently, the aforementioned nanoparticles were incorporated into chitosan/F127 hydrogel to fabricate the composite hydrogel (MPDA@EGCG-CF). The morphology, rheological properties, in vitro release behavior, biosafety, and in vitro osteogenic capacity of the composite hydrogel were evaluated via SEM, a rheometer, the CCK-8 assay, live/dead cell staining, hemolysis test, alkaline phosphatase staining with quantitative analysis, alizarin red staining with quantitative analysis, and Western blot analysis. Results The MPDA@EGCG-CF hydrogel was successfully constructed. In vitro release studies demonstrated that the composite hydrogel could achieve sustained local release of EGCG for more than one week. Morphological observations via SEM revealed that the hydrogel exhibited favorable morphological characteristics with a porous structure. Cell experiments and hemolysis test confirmed the good biosafety of the composite material. Results from ALP staining, alizarin red staining, and Western blot analysis indicated that, compared with the control (Ctrl) group and the hydrogel group encapsulating non-drug-loaded MPDA, the MPDA@EGCG-CF hydrogel displayed significantly enhanced in vitro osteogenic differentiation capacity (P<0.05). Conclusion The constructed osteoconductive chitosan/F127 composite hydrogel encapsulating EGCG-loaded MPDA can realize long-term local release of EGCG and remarkably promote osteogenic differentiation in vitro, exhibiting promising application potential for bone regeneration.

     

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