仿珍珠结构的多级套层骨仿生微支架构建及其原位骨再生机理研究

Collagen is one of the commonly used biomaterial in bone tissue engineering to repair bone defecs. However, the mechanical strength and osteoconductivity of collagen scaffolds are still unsatisfactory. Our previous study showed that the incorporation of graphene oxide into collagen scaffolds could enhanced the mechanical strength, and the coating of bonelike apatite on the scaffolds could promoted the osteoconductivity. The fabricated graphene oxide-collagen-bonelike apatite biomimetic scaffolds could regenerated in situ bone defect to some extent. While the depth of bonelike apatite in the scaffolds were not enough to develop an uniformly mineralized and large-size (800μmφ<φ<2500μmφ) microscaffolds. Therefore, we propose the development of pearl-like struture and multi-layered biomimetic bone microscaffolds for in situ bone regeneration. We use the microchips and layer-by-layer methods to develop 1500μmφ-1200μmφ-800μmφ micro-scaffolds which could exhibit an uniform distribution of the bonelike apatite in the scaffolds. After assembling the microscaffolds into large-scale scaffolds and putting them in critical bone defect of rat, the newly formed bone tissues will be analyzed to evaluate the bone regeneration ability of the scaffolds. And the tissue-induced orthotopic bone regeneration mechanism of homing endogenous cells and growth factors will be revealed. Therefore providing new ideas for constructing bone tissue engineering scaffolds.

胶原蛋白是骨组织工程中修复骨缺损常用的生物材料，但胶原支架存在力学性能不强、骨传导性欠佳等问题。申请人前期研究发现，胶原支架中复合氧化石墨烯可以增加其力学性能，涂层骨样磷灰石可以提高其骨传导性，结合两者特点制备的氧化石墨烯-胶原蛋白-骨样磷灰石骨仿生支架具有一定的原位骨再生能力。但是我们发现骨样磷灰石浸润的深度不够，难以构建内部均匀矿化的大尺寸微支架（800μmφ<φ<2500μmφ）。为此，申请人提出仿珍珠结构的多级套层骨仿生微支架构建及其骨再生策略，通过微阵列芯片法，以层层套叠的方式，构建1500μmφ-1200μmφ-800μmφ多级套层骨仿生微支架，达到内部均匀矿化的效果，通过大鼠临界骨缺损模型，对组装后的仿珍珠结构微支架的原位骨再生效果进行评估，并揭示其诱导宿主来源的细胞、生长因子的募集达到组织诱导型原位骨再生机理，为骨组织工程支架的选择提供新的思路。

胶原蛋白（Collagen, Col）是骨组织工程中修复骨缺损常用的生物材料，但胶原支架存在力学性能不强、骨传导性欠佳等问题。前期研究发现，胶原支架中复合氧化石墨烯（Graphene Oxide, GO）可以增加其力学性能，胶原支架表面涂层骨样磷灰石（Apatite）可以提高其骨传导性，结合两者特点制备的氧化石墨烯-胶原蛋白-骨样磷灰石骨仿生支架具有一定的原位骨再生能力。但是我们发现骨样磷灰石浸润的深度不够，难以构建内部均匀矿化的大尺寸微支架（800μmφ<φ<2500μmφ）。基于此，我们提出仿珍珠结构的多级套层骨仿生微支架构建及其骨再生策略，即通过微阵列芯片法，以层层套叠的方式，构建1500μmφ-1200μmφ-800μmφ多级套层骨仿生微支架，达到内部均匀矿化的效果。通过扫描电镜、红外光谱、弹性模量等表征微支架的理化性质；通过细胞活死染色、细胞扫描电镜、MTT实验、成骨分化等评估微支架的生物相容性、成骨活性等；通过大鼠临界骨缺损模型，对组装后的仿珍珠结构微支架的原位骨再生效果进行评估，并揭示其诱导宿主来源的细胞、生长因子的募集达到组织诱导型原位骨再生机理。结果表明，微支架具有疏松多孔结构，表面均匀分布羟基磷灰石；同时，微支架为间充质干细胞的增殖、成骨分化提供了合适的微环境，使成骨相关基因和蛋白上调；最后，骨缺损原位修复效果评估提示微支架能够募集宿主来源的细胞、蛋白来促进骨组织的新生。本研究提出了一种仿生手段构建内部均匀矿化的微支架，为骨组织工程支架的选择提供新的思路。