3-D Biomimetic Scaffolds for Bone Tissue Engineering

描述(由申请人提供)：口腔面部骨骼缺陷的重建是一项重大的临床挑战，每年进行超过100万次外科手术。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，从人和动物的骨髓抽提物中扩增的细胞，以及从骨碎片中获得的成骨细胞，都能够在体内形成新骨。然而，分化的速度、程度和新骨形成的程度是不可预测的，取决于细胞微环境中的因素，显然包括支持的生物材料。在这笔赠款的第一个周期中，我们开发了基于生物矿物在有机模板上三维自组装的仿生材料。我们证明，与聚合物对照组相比，通过控制有机模板上一层类骨矿物质的成核和生长，骨祖细胞的体外(细胞黏附、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)得到了重复性增强。通过改变矿物组成，我们能够合成一系列具有可控溶解性的矿物表面，初步数据表明，仿生层的溶解产物(即钙离子)本身增强了细胞功能，而不受直接底物介导的影响。总而言之，这些数据表明了可控地自组装纳米级矿物类似物的能力，提供了基于材料的细胞功能控制，以及细胞对生物材料扰动的反应是表面和溶液介导的(无机溶解因子)途径的叠加。综上所述，这些结果构成了在这次竞争性更新中检验全球假说的基础：生物材料提供的细胞外微环境通过溶液和底物介导的效应控制祖细胞增殖和分化为成骨细胞表型的能力，这些效应共同指导细胞再生矿化基质。进一步的假设是，较少的可溶性物质(表面介导的效应)促进祖细胞的增殖，而较多的可溶性物质(溶液介导的效应)促进细胞分化。这些假说是通过合成一系列仿生材料来验证的，其中包括表面自矿化为生物磷灰石的聚合物支架，根据热力学条件，这些材料的组成和溶解度受到控制，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开发，从而更好地控制祖细胞的骨形成。如果细胞对生物材料的反应是表面和可溶性因子介导的途径的叠加，这可能代表着理解细胞/生物材料相互作用的新范式，并导致基于直接呈现可溶性无机因子的治疗策略的发展，以一种侵入性较小的组织工程方法。 公共卫生相关性口腔面部骨骼缺陷的重建是一项重大的临床挑战，每年进行100多万次外科手术。由于现有技术的局限性，需要新的骨再生策略。从人骨髓抽提物中扩增的细胞能够在体内形成新骨。然而，新骨形成的程度是不可预测的，取决于细胞微环境中的因素，包括支持的生物材料。在这项提案中，我们将展示生物材料可以通过其材料的表面属性和从材料中释放的可溶离子物种来控制细胞功能和骨再生。这些研究的结果可能会导致生物材料的开发，从而更好地控制祖细胞的骨形成。如果细胞对生物材料的反应是表面和可溶性因子介导的途径的叠加，这可能代表着理解细胞/生物材料相互作用的新范式，并导致基于直接呈现可溶性无机因子的治疗策略的发展，以一种侵入性较小的组织工程方法。此外，表面和溶解因子效应之间的平衡也可以解释纳米结构材料优越的生物学性能。

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.