3D打印PLA-HA复合材料与骨髓基质细胞的相容性研究

目的 探讨骨髓基质细胞与3D打印聚乳酸-羟基磷灰石(Three-dimensional printed polylactic acid-hydroxyapatite,3D printed PLA-HA)复合支架材料的相容性,为骨组织工程选择合适的支架材料提供理论依据。方法将标记绿色荧光蛋白(Green fluorescent protein,GFP)的骨髓基质细胞分别与3D打印PLA-HA复合材料和β-磷酸三钙(betatricalcium phosphate,β-TCP)材料体外复合培养,采用荧光显微镜观察细胞在支架材料上的生长黏附情况,并通过扫描电镜观察细胞在支架中的形态变化;CCK8(Cell Counting Kit8)法检测细胞于不同时间点在材料上的黏附率以及增殖情况;碱性磷酸酶(Alkaline phosphatase,ALP)活性测定法检测3 d、7 d、14 d的碱性磷酸酶活性变化。结果 骨髓基质细胞能够在3D打印PLA-HA复合材料和β-TCP材料上黏附生长;β-TCP组细胞黏附率高于3D打印PLA-HA组(P<0.05),但3D打印PLA-HA组在12 h细胞黏附率可达到60%以上;细胞增殖实验显示,3D打印PLA-HA组在体外培养第4天和第7天的细胞增殖量(OD值)均高于β-TCP组与对照组(P<0.05);3D打印PLA-HA组以及β-TCP组在第3天、第7天和第14天的ALP含量均高于对照组,且3D打印PLA-HA组第7天ALP含量较β-TCP组增多(P<0.05)。结论 3D打印PLA-HA复合材料具有良好的细胞相容性,可作为骨组织工程的支架材料。

Biocompatibility Research of Three-dimensional Printed Polylactic acid/Hydroxyapatite Composite with Bone Marrow Stromal Cells in vitro

Objective To explore the biocompatibility of three-dimensional printed polylactic acid-hydroxyapatite（3D printed PLA-HA） as the composite scaffolds with bone marrow stromal cells for selecting the suitable scaffold in bone tissue engineering. Methods GPF positive bone marrow stromal cells were seeded on three-dimensional printed polylactic acidhydroxyapatite composite scaffolds and beta-tricalcium phosphate（β-TCP） and then cultured in vitro. The cell adhesion and proliferation were observed by fluorescence microscope, and cell morphology was scrutinized by scanning electron microscopy. Cell Counting Kit 8（CCK8） were used to test the adhesion rate and proliferation of cells on two kinds of scaffolds at different time points; Alkaline phosphatase（ALP） activity were detected on the third, seventh and fourteenth day.Results Bone marrow stromal cells exhibited the adhesion and proliferation ability well in both three-dimensional printed polylactic acid-hydroxyapatite composite scaffolds and beta-tricalcium phosphate; The adhesion rate in β-TCP group was higher than in PLA-HA group（P〈0.05）, while the adhesion rate in PLA-HA group was still more than 60% after cultured for12 hours; The amount of cell proliferation（the value of OD） 4 and 7 days after culture in PLA-HA group were both higher than in β-TCP and control group respectively（P〈0.05） in vitro. The secretion of ALP 7 and 14 days after culture in groupPLA-HA as well as β-TCP were both higher than in control group 3（P〈0.05）. Meanwhile, the ALP secretion in PLA-HA group was higher on the third day compared with β-TCP group（P〈0.05）. Conclusion Three-dimensional printed polylactic acid-hydroxyapatite composite scaffolds present good biocompatibility in vitro and it can be selected as biomaterials in bone tissue engineering.