Three-dimensional nano-architected scaffolds with tunable stiffness for efficient bone tissue growth.
Three-dimensional nano-architected scaffolds with tunable stiffness for efficient bone tissue growth.
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DOI：
10.1016/j.actbio.2017.09.007
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发表时间：
2017-11
影响因子：
9.7
通讯作者：
Greer JR
中科院分区：
文献类型：
--
作者：
Maggi A;Li H;Greer JR
导致骨科植入物失败的确切机制尚不清楚；认为骨-种植体界面处的微力学环境调节着种植体的结构稳定性。在这项工作中，我们试图了解种植体的3D机械环境如何影响早期骨整合期间的骨形成。我们采用双光子光刻（TPL）直接激光写入技术制备了具有四面十面体周期几何结构的三维刚性聚合物支架（以下简称纳米晶格），其支撑尺寸与成骨细胞的黏附大小相同（~2μm），孔径大小与细胞大小相同（~ 10μm）。这些纳米晶格中的一些随后被涂上薄的Ti或W保形层，最后在所有样品上沉积18nm厚的TiO2外层，以确保生物相容性。对各类型纳米晶格的纳米力学实验表明，其刚度范围为0.7 ~ 100 MPa。将成骨细胞样细胞（SAOS-2）播种在每个纳米格上，并在矿化培养基中培养2、8和12天后，通过跟踪矿物质分泌物和细胞内f-肌动蛋白和血管素浓度来探索它们的机械敏感反应。实验显示，与较硬的纳米晶格相比，最柔性化的纳米晶格细胞内的f-肌动蛋白多出约20%，Ca和P分泌量多出约40%，而在较硬的纳米晶格中，这种细胞反应几乎无法区分。我们构建了一个简单的现象学模型，似乎捕捉到观察到的支架刚度和f-肌动蛋白浓度之间的关系。该模型预测了最大f-肌动蛋白浓度的最佳支架刚度范围，这似乎与成骨细胞驱动的矿物沉积直接相关。该研究表明，当钛涂层三维支架的刚度与软骨相似（~ 0.5-3MPa）时，可以为细胞生长提供最佳的微环境。这些发现有助于更好地理解成骨细胞的机械敏感性，并可能对开发更有效、更安全的骨假体具有深远的意义。二十多年来，由于缺乏对三维（3D）环境中细胞行为的全面了解，制造导致最佳骨重塑的假体一直是一个挑战。文献表明，2D基质刚度在决定细胞行为方面起着重要作用，然而，制造技术的局限性和表征细胞-支架相互作用的困难限制了我们对3D支架刚度如何影响细胞反应的理解。本研究表明支架结构刚度影响成骨细胞的细胞反应。具体来说，本研究表明，与所有其他纳米晶格相比，在刚度为0.7MPa的最柔顺纳米晶格上生长的细胞表达的细胞内f-actin浓度高出约20%，分泌的Ca和P含量高出约40%。这表明，具有接近软骨刚度的骨支架可以作为新型合成骨移植材料的最佳3D支架。
The precise mechanisms that lead to orthopedic implant failure are not well understood; it is believed that the micromechanical environment at the bone-implant interface regulates structural stability of an implant. In this work, we seek to understand how the 3D mechanical environment of an implant affects bone formation during early osteointegration. We employed two-photon lithography (TPL) direct laser writing to fabricate 3-dimensional rigid polymer scaffolds with tetrakaidecahedral periodic geometry, herewith referred to as nanolattices, whose strut dimensions were on the same order as osteoblasts’ focal adhesions (~2μm) and pore sizes on the order of cell size, ~10μm. Some of these nanolattices were subsequently coated with thin conformal layers of Ti or W, and a final outer layer of 18nm-thick TiO2 was deposited on all samples to ensure biocompatibility. Nanomechanical experiments on each type of nanolattice revealed the range of stiffnesses of 0.7–100 MPa. Osteoblast-like cells (SAOS-2) were seeded on each nanolattice, and their mechanosensitve response was explored by tracking mineral secretions and intracellular f-actin and vinculin concentrations after 2, 8 and 12 days of cell culture in mineralization media. Experiments revealed that the most compliant nanolattices had ~20% more intracellular f-actin and ~40% more Ca and P secreted onto them than the stiffer nanolattices, where such cellular response was virtually indistinguishable. We constructed a simple phenomenological model that appears to capture the observed relation between scaffold stiffness and f-actin concentration. This model predicts a range of optimal scaffold stiffnesses for maximum f-actin concentration, which appears to be directly correlated with osteoblast-driven mineral deposition. This work suggests that three-dimensional scaffolds with titania-coated surfaces may provide an optimal microenvironment for cell growth when their stiffness is similar to that of cartilage (~0.5–3MPa). These findings help provide a greater understanding of osteoblast mechanosensitivity and may have profound implications in developing more effective and safer bone prostheses. Creating prostheses that lead to optimal bone remodeling has been a challenge for more than two decades because of a lack of thorough knowledge of cell behavior in three-dimensional (3D) environments. Literature has shown that 2D substrate stiffness plays a significant role in determining cell behavior, however, limitations in fabrication techniques and difficulties in characterizing cell-scaffold interactions have limited our understanding of how 3D scaffolds’ stiffness affects cell response. The present study shows that scaffold structural stiffness affects osteoblasts cellular response. Specifically this work shows that the cells grown on the most compliant nanolattices with a stiffness of 0.7MPa expressed ~20% higher concentration of intracellular f-actin and secreted ~40% more Ca and P compared with all other nanolattices. This suggests that bone scaffolds with a stiffness close to that of cartilage may serve as optimal 3D scaffolds for new synthetic bone graft materials.
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