Bialorucki Callan, Subramanian Gayathri, Elsaadany Mostafa, Yildirim-Ayan Eda
Department of Bioengineering, College of Engineering, University of Toledo, Toledo, OH 43606, USA.
Department of Bioengineering, College of Engineering, University of Toledo, Toledo, OH 43606, USA; Department of Orthopaedic Surgery, University of Toledo Medical Center, Toledo, OH 43614, USA.
J Mech Behav Biomed Mater. 2014 Oct;38:143-53. doi: 10.1016/j.jmbbm.2014.06.018. Epub 2014 Jul 8.
The objective of this study was to understand the temporal relationship between in situ generated calcium content (mineralization) and the mechanical properties of an injectable orthobiologic bone-filler material. Murine derived osteoblast progenitor cells were differentiated using osteogenic factors and encapsulated within an injectable polycaprolactone nanofiber-collagen composite scaffold (PN-COL +osteo) to evaluate the effect of mineralization on the mechanical properties of the PN-COL scaffold. A comprehensive study was conducted using both an experimental and a predictive analytical mechanical analysis for mechanical property assessment as well as an extensive in vitro biological analysis for in situ mineralization. Cell proliferation was evaluated using a PicoGreen dsDNA quantification assay and in situ mineralization was analyzed using both an alkaline phosphatase (ALP) assay and an Alizarin Red stain-based assay. Mineralized matrix formation was further evaluated using energy dispersive x-ray spectroscopy (EDS) and visualized using SEM and histological analyses. Compressive mechanical properties of the PN-COL scaffolds were determined using a confined compression stress-relaxation protocol and the obtained data was fit to the standard linear solid viscoelastic material mathematical model to demonstrate a relationship between increased in situ mineralization and the mechanical properties of the PN-COL scaffold. Cell proliferation was constant over the 21 day period. ALP activity and calcium concentration significantly increased at day 14 and 21 as compared to PN-COL -osteo with undifferentiated osteoblast progenitor cells. Furthermore, at day 21 EDS, SEM and von Kossa histological staining confirmed mineralized matrix formation within the PN-COL scaffolds. After 21 days, compressive modulus, peak stress, and equilibrium stress demonstrate significant increases of 3.4-fold, 3.3-fold, and 4.0-fold respectively due to in situ mineralization. Viscoelastic parameters calculated through the standard linear solid mathematical model fit to the stress-relaxation data also indicate improved mechanical properties after in situ mineralization. This investigation clearly demonstrates that in situ mineralization can increase the mechanical properties of an injectable orthobiologic scaffold and can possibly guide the design of an effective osteoconductive injectable material.
本研究的目的是了解原位生成的钙含量（矿化）与可注射的骨生物填充材料的力学性能之间的时间关系。使用成骨因子分化源自小鼠的成骨细胞祖细胞，并将其封装在可注射的聚己内酯纳米纤维 - 胶原蛋白复合支架（PN - COL + osteo）中，以评估矿化对PN - COL支架力学性能的影响。采用实验性和预测性分析力学分析进行全面研究以评估力学性能，并进行广泛的体外生物学分析以研究原位矿化。使用PicoGreen双链DNA定量测定法评估细胞增殖，并使用碱性磷酸酶（ALP）测定法和基于茜素红染色的测定法分析原位矿化。使用能量色散X射线光谱法（EDS）进一步评估矿化基质的形成，并通过扫描电子显微镜（SEM）和组织学分析进行可视化。使用受限压缩应力松弛方案测定PN - COL支架的压缩力学性能，并将获得的数据拟合到标准线性固体粘弹性材料数学模型，以证明原位矿化增加与PN - COL支架力学性能之间的关系。在21天期间细胞增殖保持恒定。与含有未分化成骨细胞祖细胞的PN - COL - osteo相比，在第14天和第21天，ALP活性和钙浓度显著增加。此外，在第21天，EDS、SEM和冯·科萨组织学染色证实了PN - COL支架内矿化基质的形成。21天后，由于原位矿化，压缩模量、峰值应力和平衡应力分别显著增加3.4倍、3.3倍和4.0倍。通过拟合应力松弛数据的标准线性固体数学模型计算的粘弹性参数也表明原位矿化后力学性能得到改善。本研究清楚地表明，原位矿化可以提高可注射骨生物支架的力学性能，并可能指导有效的骨传导性可注射材料的设计。
