3-D Biomimetic Scaffolds for Bone Tissue Engineering

描述（由申请人提供）：口腔面部骨骼缺损的重建是一项重大的临床挑战，每年有超过100万例外科手术。由于现有技术的局限性，需要新的骨再生策略。我们和其他人已经证明，从人类和动物骨髓中提取的细胞，以及从骨碎片中收获的成骨细胞，能够在体内形成新的骨骼。然而，分化的速度和程度以及新骨形成的程度是不可预测的，并且取决于细胞微环境中的因素，其中显然包括支持生物材料。在这个资助的第一个周期中，我们开发了基于生物矿物在有机模板上的3D自组装的仿生材料。我们证明，与聚合物对照相比，通过控制一层骨样矿物质在有机模板上的成核和生长，骨祖细胞的体外（细胞粘附、增殖、细胞骨架组织、成骨分化）和体内功能（再生骨的体积分数）得到了可重复性的增强。通过改变矿物组成，我们能够合成一系列具有可控制溶解度的矿物表面，并且初步数据表明，仿生层的溶解产物（即Ca离子）可以独立于直接底物介导的作用，自行增强细胞功能。总的来说，这些数据证明了可控制自组装纳米级矿物类似物的能力，提供了对细胞功能的基于材料的控制，并且细胞对生物材料扰动的反应是表面和溶液介导（无机可溶性因子）途径的叠加。综上所述，这些结果构成了在这种竞争性更新中测试的整体假设的基础：生物材料提供的细胞外微环境通过溶液和底物介导的作用控制祖细胞增殖和向成骨细胞表型分化的能力，这两种作用共同指导细胞再生矿化基质。进一步假设，低可溶性物质（表面介导效应）促进祖细胞增殖，而高可溶性物质（溶液介导效应）促进细胞分化。这些假设是通过合成一系列仿生材料来验证的，这些仿生材料包括聚合物支架，其表面可以自矿化成生物磷灰石，根据热力学条件控制组成和溶解度，并评估人类祖细胞对这些仿生变体的体外和体内反应。这些研究的结果可能会导致生物材料的发展，更好地控制骨形成的祖细胞。如果细胞对生物材料的反应是表面和可溶性因子介导途径的叠加，这可能代表了理解细胞/生物材料相互作用的新范式，并且还导致基于可溶性无机因子的直接呈现的治疗策略的发展，以一种侵入性较小的方法进行组织工程。

DESCRIPTION (provided by applicant): Reconstruction of orofacial skeletal defects represents a major clinical challenge, with over 1 million surgical procedures performed each year. New strategies of regenerating bone are needed because of limitations with existing techniques. We and others have shown that cells expanded from human and animal bone marrow aspirates, as well as osteoblasts harvested from bone fragments, are capable of forming new bone in-vivo. However, the rate and extent of differentiation and degree of new bone formation are unpredictable, and dependent upon factors in the cellular microenvironment, which clearly include the supporting biomaterial. In the first cycle of this grant, we developed biomimetic materials based on the 3D self-assembly of biological minerals onto organic templates. We demonstrated that the in-vitro (cell adhesion, proliferation, cytoskeletal organization, osteogenic differentiation) and in-vivo function (volume fraction of regenerated bone) of osteoprogenitor cells was reproducibly enhanced, compared to polymer controls, by controlling the nucleation and growth of a layer of bone-like mineral onto an organic template. By altering mineral composition, we were able to synthesize a series of mineral surfaces which exhibited controllable solubility, and preliminary data suggests that the dissolution products of the biomimetic layer (i.e. Ca ions) enhance cell function by themselves, independent of direct substrate-mediated effects. Collectively, these data demonstrate the ability to controllably self-assemble nanoscale mineral analogues, providing material-based control over cell function, and that the response of cells to biomaterial perturbations is the superposition of surface and solution-mediated (inorganic soluble factor) pathways. Taken together, these results form the basis for the global hypothesis to be tested in this competing renewal: the extracellular microenvironment provided by a biomaterial controls the ability of progenitor cells to proliferate and differentiate toward an osteoblast phenotype through solution as well as substrate-mediated effects, which collectively can direct cells to regenerate a mineralized matrix. It is further hypothesized that less soluble materials (surface-mediated effects) promote progenitor cell proliferation, whereas more soluble materials (solution-mediated effects) promote cell differentiation. These hypotheses are tested by synthesizing a series of biomimetic materials that includes polymer scaffolds with surfaces that self-mineralize into a biological apatite of controlled composition and solubility, depending on thermodynamic conditions, and assessing the in-vitro and in-vivo response of human progenitor cells to these biomimetic variants. The results of these studies could lead to the development of biomaterials that better control bone formation by progenitor cells. If the response of cells to a biomaterial is the superposition of surface and soluble factor-mediated pathways, this could represent a new paradigm in understanding cell/biomaterial interactions, and also lead to the development of therapeutic strategies based on a direct presentation of soluble inorganic factors, in a less invasive approach to tissue engineering. PUBLIC HEALTH RELEVANCE Reconstruction of orofacial skeletal defects represents a major clinical challenge, with over 1 million surgical procedures performed each year. New strategies of regenerating bone are needed because of limitations with existing techniques. Cells expanded from human bone marrow aspirates are capable of forming new bone in-vivo. However, the degree of new bone formation is unpredictable, and dependent upon factors in the cellular microenvironment, including the supporting biomaterial. In this proposal, we will demonstrate that a biomaterial can control cell function and bone regeneration by both its surface properties of the material and soluble ionic species released from the material. Results of these studies could lead to the development of biomaterials that better control bone formation by progenitor cells. If the response of cells to a biomaterial is the superposition of surface and soluble factor-mediated pathways, this could represent a new paradigm in understanding cell/ biomaterial interactions, and also lead to the development of therapeutic strategies based on a direct presentation of soluble inorganic factors, in a less invasive approach to tissue engineering. Moreover, the balance between surface and soluble factor effects could also explain the superior biological performance of nanostructured materials.