兰州大学机构库 >口腔医学院
负载ALN的3D打印nHA/SA/PCL复合支架的制备及性能
Alternative TitleSynthesis and properties of 3D printing nHA/SA/PCL composite scaffold loaded with ALN
周红丽
Subtype硕士
Thesis Advisor何祥一
2023-05-30
Degree Grantor兰州大学
Place of Conferral兰州
Degree Name医学硕士
Degree Discipline口腔医学
Keyword纳米羟基磷灰石 Nano-hydroxyapatite, sodium alginate 海藻酸钠 polycaprolactone 聚己内酯 alendronate sodium 阿仑膦酸钠 3D printing 3D打印 scaffold 支架
Abstract

目的:对于由肿瘤、外伤以及炎症等原因导致的骨缺损,机体的自我修复能力受限,而目前尚无理想的骨替代材料能够促进缺损的骨组织再生。本研究采用挤压式三维(Three-dimension,3D)打印技术制备负载阿仑膦酸钠(Alendronate,ALN)的多孔纳米羟基磷灰石(Nano-hydroxyapatite,nHA)/海藻酸钠(Sodium alginate,SA)/聚己内酯(Polycaprolactone,PCL)复合支架,以模仿骨组织的构造和力学性能;采用各种表征手段探究该支架的材料属性、药物缓释能力以及体外生物学行为,以初步探索其作为人工植入物的应用前景。

方法:(1)将不同含量的nHA(分别为50wt%、60wt%、70wt%)与SA共混获得均匀的打印墨水,通过测试各组墨水的流变性来评估其打印的可行性;(2)采用挤压式3D打印机按照预定的设计进行支架的打印,并分别命名为nHA50、nHA60和nHA70支架。随后采用浸渍法将PCL组装到复合支架表面,得到的支架分别命名为nHA50P、nHA60P和nHA70P支架;(3)采用扫描电子显微镜(Scanning electron microscope,SEM)、傅里叶变换红外光谱仪(Fourier transform infrared spectroscopy,FTIR)、X射线衍射(X-ray diffraction,XRD)对支架的形貌、结构及组分进行表征,并进行孔隙率检测;(4)检测所有支架的机械性能、亲水性能和溶胀性能,以评估是否达到组织工程支架的应用要求;(5)将ALN载入性能更为优异的nHA60P支架和作为对照组的nHA60支架中,获得nHA60P-ALN和nHA60-ALN支架并进行FTIR检测。同时在14天中定期收集支架的浸提液进行吸光度(Optical density,OD)的检测,分析PCL涂层对支架中ALN的控缓释作用;(6)采用CCK-8法对nHA60、nHA60P和nHA60P-ALN支架的细胞毒性及促增值能力进行检测。

结果:(1)不同nHA含量的打印墨水均具有剪切变稀的能力,且支撑性良好,符合挤压式3D打印的要求;(2)nHA50、nHA60和nHA70支架均按照预定的形状打印成型,nHA50P、nHA60P和nHA70P支架的涂层均匀涂覆于表面,所有支架的孔隙规则且相互连通,具有良好的成形性;(3)SEM结果显示,所有支架表面呈现凹凸不平的粗糙表面,内部则为疏松的多孔结构,有利于细胞的生长活动。FTIR和XRD结果显示,复合支架中存在nHA、SA和PCL成分且性质没有发生明显的改变。六组支架的孔隙率在86-91%之间,提供了较大的表面积便于细胞的粘附生长;(4)力学实验显示,随着nHA比例的提高,nHA50、nHA60和nHA70无涂层支架与nHA50P、nHA60P和nHA70P涂层支架的压缩强度和模量均呈现先增加后减少的趋势,且PCL涂层支架均高于无涂层支架,因此nHA60P支架的压缩强度最高,达到了16 MPa。亲水性实验显示,nHA50、nHA60和nHA70支架的接触角依次降低(P<0.05),而nHA50P、nHA60P和nHA70P支架的接触角因其表面涂覆PCL有所增加,但均小于90°,依然可定义为亲水性材料。溶胀实验表明所有复合支架均在12 h后达到溶胀平衡,其中以nHA60P支架的溶胀率最低,这有助于支架在植入体内后保持良好的结构稳定性;(5)对nHA60-ALN和nHA60P-ALN进行FTIR检测,显示ALN被成功的载入且其性质未发生明显改变。药物缓释实验表明,与nHA60-ALN支架相比,nHA60P-ALN支架的药物释放曲线更为平缓,因此PCL涂层可以延长ALN的释放时间,起到很好的控缓释作用;(6)细胞毒性实验显示成骨细胞在nHA60、nHA60P和nHA60P-ALN支架浸提液中的生存率均高于80%,且在支架中载入0.5wt%的ALN并不会影响成骨细胞的活动。随后将各组支架与成骨细胞共培养,各组支架材料均可以促进成骨细胞的增殖,且以nHA60P-ALN支架的促增殖效果最佳。

结论:将nHA与SA结合,再辅以性能优良的PCL和促进成骨细胞增殖的ALN,最终制备出来的载药复合支架具有疏松多孔的微观结构、优良的力学性能、较好的亲水性、适宜的溶胀性、良好的药物缓释作用和生物相容性等特性,为临床修复骨缺损提供了一种新的候选材料,并为ALN在组织工程的应用打下了坚实的基础。

Other Abstract

Objective: For bone defects caused by tumor, trauma, inflammation and other causes, the body's self-repair ability is limited, and currently there is no ideal bone replacement material that can promote the regeneration of bone tissue defects. In this study, the porous nano-hydroxyapatite (nHA)/ sodium alginate (SA)/ polycaprolactone (PCL) composite scaffolds loaded with Alendronate (ALN) were prepared by extrusion three-dimension (3D) printing technology to simulate the structure and mechanical properties of bone tissue; The material properties, drug sustained-release ability and in vitro biological behavior of the scaffolds were investigated by various characterization methods to initially explore its application prospects as an artificial implant.

Methods: (1) Different contents of nHA (they are 50wt%, 60wt% and 70wt% respectively) were mixed with SA to obtain uniform printing inks. Rheological properties of each set of inks were tested to evaluate their printing feasibility; (2) The extrusion 3D printer was used to print the scaffolds according to the predetermined design, and named nHA50, nHA60 and nHA70 scaffold respectively. The PCL was then assembled on the surface of the composite scaffold by dipping method. The obtained scaffolds were named nHA50P, nHA60P and nHA70P scaffold, respectively; (3) Scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) were used to characterize the morphology, structure and component of the scaffolds , and to detect the porosity; (4) The mechanical properties, hydrophilicity and swelling properties of all scaffolds were tested to evaluate whether they met the application requirements of tissue engineering scaffolds; (5) The ALN was loaded into the nHA60P scaffold with better performance and the nHA60 scaffold as the control group, and the FTIR detection was performed after the nHA60P-ALN and nHA60-ALN scaffold were obtained. Optical density (OD) values of the scaffolds were collected periodically for 14 days to analyze the slow-release effect of PCL coating on ALN in the scaffolds; (6) The cytotoxicity and growth promoting ability of nHA60、nHA60P and nHA60P-ALN scaffolds were detected by CCK-8 method.

Results: (1) Printing inks with different nHA contents have the ability to shear thinning and have good support, which meets the requirements of extrusion 3D printing; (2) nHA50, nHA60 and nHA70 scaffolds were printed and formed according to the predetermined shape, and the PCL coatings of nHA50P, nHA60P and nHA70P scaffolds were uniformly coated on the surface. The pores of all scaffolds were regular and interconnected, showing good formability; (3) SEM results showed that the surface of all scaffolds presented uneven rough surface, and the interior was loose porous structure, which was conducive to the growth of cells. The results of FTIR and XRD showed that nHA, SA and PCL components existed in the composite scaffold and their properties did not change significantly. The porosity of the six scaffolds ranged from 86-91%, providing a large surface area for cell adhesion and growth; (4) Mechanical experiments showed that with the increase of nHA ratio, the compressive strength and modulus of nHA50, nHA60 and nHA70 uncoated scaffolds and nHA50P, nHA60P and nHA70P coated scaffolds increased first and then decreased, while the PCL coated scaffolds were higher than the uncoated scaffolds. Therefore, the nHA60P scaffold had the highest compressive strength of 16 MPa. Hydrophilic experiments showed that the contact angles of nHA50, nHA60 and nHA70 scaffolds decreased successively (P < 0.05), and the contact angles of nHA50P, nHA60P and nHA70P scaffolds were increased due to PCL coating, but they were all less than 90°, which can still be defined as hydrophilic materials. Swelling experiments showed that all the composite scaffolds reached swelling equilibrium after 12 hours, and nHA60P scaffold had the lowest swelling rate, which was conducive to maintaining good structural stability after implantation; (5) FTIR detection of nHA60-ALN and nHA60P-ALN showed that ALN was loaded successfully and no obvious change in its properties was found. Drug release experiments showed that the drug release curve of nHA60P-ALN scaffold was smoother than that of nHA60-ALN scaffold, so the PCL coating could prolong the release time of ALN and play a good role in controlling slow release; (6) Cytotoxicity tests showed that the survival rates of osteoblast in the extracting liquid of nHA60, nHA60P and nHA60P-AlN scaffold were all higher than 80%, and loading 0.5wt% of ALN in the scaffolds did not affect the activity of the cells. Subsequently, each group of scaffolds were then co-cultured with osteoblast. All three groups of scaffold materials could promote osteoblast proliferation, and nHA60P-ALN scaffolds had the best effect on cell proliferation.

Conclusion: By combining nHA with SA, supplemented by excellent PCL and ALN, which promote bone cell proliferation, finally the prepared drug-carrying composite scaffold has the properties of loose and porous microstructure, excellent mechanical properties, good hydrophilicity, suitable swelling, good drug slow release and biocompatibility, which provides a new candidate material for clinical repair of bone defects and lays a solid foundation for the application of ALN in tissue engineering.

Subject Area生物医用材料
MOST Discipline Catalogue医学 - 口腔医学
URL查看原文
Language中文
Other Code262010_220200927350
Document Type学位论文
Identifierhttps://ir.lzu.edu.cn/handle/262010/537600
Collection口腔医学院
Affiliation
兰州大学口腔医学院
Recommended Citation
GB/T 7714
周红丽. 负载ALN的3D打印nHA/SA/PCL复合支架的制备及性能[D]. 兰州. 兰州大学,2023.
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