Engineering biomimetic knee menisci with zonal and anisotropic variations

项目总结 该方案旨在组织工程一个各向异性的新半月板，它也捕捉到区域变化 存在于天然组织中。随后，同种异体半月板结构将被植入Leporine。 模型实现半月板修复和替换。假设：1)区域变化， 通过优化细胞培养和无支架培养，可以设计出各向异性的半月板状结构 条件；2)多水平刺激的战略性时间应用(在细胞、分子和结构- 级别)将允许跨不同行动级别的协同作用，以增强 成熟的新半月板；以及3)同种异体结构可以成功地植入Leporine模型。这些 假设将通过以下三个具体目标进行检验：1)创建具有以下特性的各向异性新半月面 模仿原生组织的区域变异；2)增强近地天体的功能和组织特性 半月板通过多层次外生刺激与时间协调协同；3)发展 手术固定技术，植入兔新半月板。以前，自然的半月板是 在形态和生物力学上都是高度各向异性和区域变异的，这促使我们的 目前的组织工程学方法可以模拟这些特征。同种异体Leporine细胞将被用于 在目标1中形成组织上和区域上模仿的新半月板结构；这一目标将实现 通过使用新颖的空间和时间变化播种技术。各向异性和组织性 工程新半月板的性能将通过操纵分子、细胞和 具体地说，转化生长因子-β1和静水压力将作用于细胞水平以 增加基质产量；溶血磷脂酸和软骨素酶-ABC将用于对齐和紧凑 在分子水平上的基质；以及特定于半月板的机械刺激将在 构造标高。在这项建议中，为了避免使用原代细胞，我们将调查传代细胞的使用 同种异体细胞在体内修复和替换半月板(目标3)。在成功演示后 在Leporine模型中，我们将确定同样扩展绵羊和人类的方法 用于未来研究的细胞。这种方法寻求解决组织稀缺的问题，并旨在提供一种 解决了半月板修复和更换这一复杂问题。

PROJECT SUMMARY This proposal aims to tissue engineer an anisotropic neo-meniscus that also captures the regional variations present in the native tissue. Subsequently, allogeneic neo-meniscal constructs will be implanted in a leporine model to achieve both meniscus repair and replacement. It is hypothesized that: 1) regionally variant, anisotropic, meniscus-shaped constructs can be engineered by optimizing cell culture and scaffold-free culture conditions; 2) the strategic temporal application of multi-level stimuli (at cellular-, molecular-, and construct- levels) will allow for synergisms across the different levels of action to enhance the functional properties of the maturing neo-menisci; and 3) allogeneic constructs can be successfully implanted in a leporine model. These hypotheses will be tested via the following three specific aims: 1) To create an anisotropic neo-meniscus with regional variations mimicking native tissue; 2) to enhance functional and organizational properties of the neo- meniscus via multi-level exogenous stimulation synergized by temporal coordination; and 3) to develop surgical fixation techniques and implant the neo-menisci in the rabbit. Previously, the native meniscus was found to be highly anisotropic and regionally variant both morphologically and biomechanically, motivating our current tissue engineering approach to mimic these characteristics. Allogeneic leporine cells will be used to form organizationally and regionally mimetic neo-meniscal constructs in Aim 1; this goal will be accomplished via the use of novel spatial and temporally variant seeding techniques. The anisotropic and organizational properties of the engineered neo-meniscus will then be enhanced by manipulating molecular-, cellular-, and construct-level targets in Aim 2. Specifically, TGF-β1 and hydrostatic pressure will act on the cellular level to increase matrix production; lysophosphatidic acid and chondroitinase-ABC will be used to align and compact the matrix at the molecular level; and meniscus-specific mechanical stimulation will direct anisotropy at the construct level. In this proposal, to avoid the use of primary cells, we will investigate the use of passaged allogeneic cells toward in vivo repair and replacement of the meniscus (Aim 3). Upon successful demonstration of repair/replacement in the leporine model, we will determine methods to likewise expand sheep and human cells for future studies. This approach seeks to address the issue of tissue scarcity and aims to provide a solution to the complex problem of meniscus repair and replacement.