Biophysical signals, biomaterial surface characteristics and hMSC differentiation

描述(申请人提供)：随着我们人口的老龄化，骨移植修复骨缺损的需求正在迅速增加。自体骨移植和同种异体骨移植已成功地治疗了骨缺损。然而，这些方法并不理想，因为自体移植的可用性有限，可能导致供体部位的发病率，而同种异体移植可能被免疫排斥，并有可能传播疾病。假定人骨髓间充质干细胞(HMSC)与生物相容性支架相结合，在治疗骨缺损方面具有巨大的潜力，否则将采用自体或同种异体骨移植。遗憾的是，hMSC的体外扩增可以增加hMSC的治疗潜力，但会降低其成骨能力。在体外扩增人骨髓间充质干细胞的同时保持甚至增强其成骨能力，可以极大地提高其治疗潜力。因此，该项目的目标是确定特定的生物材料表面特征和生物物理信号，它们在优化hMSC向成骨细胞分化的过程中协同作用。我们的总体假设是，生物材料表面的特性，特别是纳米级的形貌，使细胞对流体流动敏感，从而增加流体流动对hMSC扩张的影响，同时也增强hMSC的成骨能力。利用独特的流体流动方案、新的生物材料表面特性、原子力显微镜、基因工程和转基因动物模型，我们将确定一个优化hMSC向成骨细胞分化的环境以及参与这一机制的信号转导途径。然后，我们将检查这些经过预处理的hMSC在体内是否比未经预处理的hMSC更具成骨能力。我们将通过在5年的时间内完成4个目标来实现这一点：目的1，在存在和不存在特定信号通路抑制剂的情况下，检测表面形貌对hMSC黏附、增殖和分化的影响；目标2，确定hMSC在不同纳米级形貌表面的硬度和机械敏感性；目标3，在有和没有PLC/钙调神经磷酸酶和PLC/ERK信号通路抑制剂的情况下，检测流体流动对hMSC增殖和分化的影响；以及目标4，检测种植在HA/TCP支架上的hMSC的体内成骨作用。通过完成这些目标，我们不仅将为骨组织工程开发新的策略，而且还将为调控hMSC的增殖和分化提供机制方面的见解。公共卫生相关性：随着老年人口的增加，对肌肉骨骼病理学新的治疗方法的需求也将增加。利用成人干细胞进行组织工程就是这样一种方法。该项目将开发新的肌肉骨骼组织工程方案，结合纳米技术、成人干细胞和生物物理力，将导致替代因疾病、损伤和衰老而造成的骨丢失的策略。

DESCRIPTION (provided by applicant): The need for bone grafts to repair bone defects is rapidly accelerating as our population ages. Bone defects have been successfully treated using autografts and allografts. However, these are less than ideal approaches since autograft availability is limited and can result in donor-site morbidity while allografts can be immunologically rejected and have the potential to transmit disease. The use of putative human mesenchymal stem cells (hMSC) combined with biocompatible scaffolds has great potential for treating bone defects, which would otherwise be treated with autografts or allografts. Unfortunately, the expansion of hMSC in vitro, which could increase the therapeutic potential of hMSC, reduces their osteogenic potential. The ability to expand hMSC in vitro while maintaining, or even enhancing, their osteogenic potential, could greatly enhance their therapeutic potential. Therefore, the goal of this project is to identify specific biomaterial surface characteristics and biophysical signals that interact synergistically in optimizing hMSC differentiation toward the osteoblastic lineage. Our overall hypothesis is that biomaterial surface characteristics, specifically nanoscale topography, sensitize cells to fluid flow thus increasing the effect of fluid flow on expansion of hMSC while also enhancing the osteogenic potential of hMSC. Using unique fluid flow protocols, novel biomaterial surface characteristics, atomic force microscopy, genetic engineering and transgenic animal models we will identify an environment that optimizes differentiation of hMSC towards the osteoblastic lineage and the signal transduction pathways involved in this mechanism. We will then examine whether these pre-treated hMSC are more osteogenic in vivo than non pre-treated hMSC. We will accomplish this through the completion, over a 5 year period, of 4 aims: Aim 1, Examine the effect of surface topography on adhesion, proliferation and differentiation of hMSC, in the presence and absence of inhibitors of specific signaling pathways; Aim 2, Determine stiffness and mechanosensitivity of hMSC on surfaces with varying nanoscale topographies; Aim 3, Examine the effect of fluid flow, in the presence and absence of inhibitors of the PLC/calcineurin and PLC/ERK signaling pathways, on hMSC proliferation and differentiation; and Aim 4, Examine in vivo osteogenesis of hMSC seeded onto HA/TCP scaffolds. By completing these aims we will not only develop novel strategies for bone tissue engineering but also provide mechanistic insight into the regulation of hMSC proliferation and differentiation. PUBLIC HEALTH RELEVANCE: As the aged population increases the need for novel therapeutic approaches to musculoskeletal pathology will also increase. Tissue engineering exploiting adult stem cells is one such approach. This project will develop novel musculoskeletal tissue engineering protocols combining nanotechnology, adult stem cells and biophysical forces that will lead to strategies to replace bone loss to disease, injury and aging.