4-D Imaging Cell/Scaffold Interplays During In Vivo Bone Repair Process

4-D 成像细胞/支架在体内骨修复过程中的相互作用

• 批准号: 8114748
• 负责人: David W. Rowe
• 金额: $ 17.98万
• 依托单位: UNIVERSITY OF CONNECTICUT STORRS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-15 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8114748
• 关键词: Algorithms; Animals; Apatites; Architecture; Biocompatible Materials; Biomimetics; Bone Development; Bone Regeneration; Bone Tissue; Calvaria; Cell Communication; Cells; Cellular Structures; Collagen; Communities; Complex; Defect; Development; Developmental Biology; Dyes; Elements; Event; Failure; Fluorescence; Green Fluorescent Proteins; Growth Factor; Histology; Human; Image; Image Analysis; Knowledge; Lasers; Learning; Life; Luciferases; Microscope; Microscopy; Modeling; Modification; Mus; Osteogenesis; Outcome; Participant; Photons; Population; Preclinical Testing; Procedures; Process; Proliferating; Property; Protocols documentation; Relative (related person); Reporter; Research; Research Personnel; Resolution; Screening procedure; Series; Signal Transduction; Site; Source; Specimen; Staging; Staining method; Stains; Stem cells; System; Testing; Three-Dimensional Imaging; Time; Tissue Engineering; Tissues; Transgenic Mice; Wound Healing; base; bone; cell preparation; cellular imaging; design; embryonic stem cell; improved; in vivo; induced pluripotent stem cell; knowledge base; medical schools; mineralization; molecular marker; novel; osteogenic; osteoprogenitor cell; progenitor; repaired; response; scaffold; skeletal; skeletal regeneration; skeletal tissue; spatial relationship; success; two-photon

DESCRIPTION (provided by applicant): Skeletal tissue engineering is rapidly approaching the stage for human application despite the fact that rigorous preclinical testing to understand the cellular and environmental determinants of success or failure has not been developed. As traditional histological and molecular markers of tissue formation do not indicate the degree of bone formation, the knowledge of the proliferative properties of the progenitor cells that invest in the scaffold and the relative host/donor contributions to the repair has not been achieved. To overcome this problem, we have developed or acquired a series of GFP reporter transgenic mice that mark the source and level of differentiation of the major cellular components in bone repair. Murine models have been developed to assess these events utilizing 2D cryo-histology that preserves the activity of the reporters in mineralized tissues. However, details of temporal and spatial cell/scaffold interactions during the dynamic bone repair process are still not understood. Gaining this knowledge is vital for improving existing, or developing new research strategies and therapies. Thus, the primary objective of this application is to demonstrate to the tissue engineering community that it is possible to develop an in vivo 4D time-lapse imaging platform to visualize cell/scaffold/new bone interplay in a mouse calvarial defect using 2-photon microscopy. This will be the first time that a 3D cell- scaffold repair system is visualized in real-time with the emission of cell-specific GFP signals during the dynamic bone regeneration. We will then use this newly established platform to view different stages of bone development in two novel scaffolds created recently within our research group. The results will be used in an attempt to interpret prior observations, that lamellar scaffolds have superior bone forming ability to that of the cellular structure, irrespective of the type of progenitor cells seeded. We hypothesize that the way the osteoprogenitor cells initially distribute, proliferate and progress toward osteogenic differentiation in vivo determines the ultimate outcome of bone formation in a scaffold. Successful implementation of the 4D imaging platform will be transformative to the field because it will provide the essential information as to why a particular strategy succeeds or fails. The platform will become the primary imaging base of understanding the spatial relationships, for appreciating temporal events between the cellular elements the cellular elements and cell/scaffold interactions during bone formation. It will allow investigators to begin to understand and ultimately optimize scaffold design and cellular participants as well as growth factor selection and release profile design for early stages of skeletal regeneration. This knowledge base will be crucial for the bone tissue engineering community. PUBLIC HEALTH RELEVANCE: More than 1.3 million bone-repair procedures are conducted per year in the USA, which created a huge demand in bone re-generating materials. However, very little is known to date about the temporal and spatial interactions between scaffold and cells during the dynamic process of bone repair. In this study, we propose to establish a 4D in vivo imaging platform to visualize in real-time cellular activities and bone development in scaffolds, and thereby guide us to design better scaffold/cell complex for bone repair.

描述（由申请人提供）：尽管尚未开发出严格的临床前测试来了解成功或失败的细胞和环境决定因素，但骨骼组织工程仍在迅速接近人类应用阶段。由于组织形成的传统组织学和分子标记并未表明骨形成程度，因此尚未实现投资支架上投资的祖细胞的增殖特性以及对修复的相对宿主/供体贡献的知识。为了克服这个问题，我们开发了或获得了一系列GFP报告基因转基因小鼠，这些小鼠标记了骨修复中主要细胞成分的分化的来源和水平。已经开发了使用2D冷冻历史的鼠模型来评估这些事件，从而保留了记者在矿化组织中的活性。但是，在动态骨修复过程中的时间和空间细胞/支架相互作用的细节仍然尚不清楚。获得这些知识对于改善现有或制定新的研究策略和疗法至关重要。 因此，该应用的主要目的是向组织工程界证明可以使用2光子显微镜在小鼠钙纳学缺陷中可视化小鼠/支架/新骨相互作用的体内4D延时成像平台。这将是第一次，在动态骨再生过程中，通过细胞特异性GFP信号的发射实时可视化3D细胞修复系统。然后，我们将使用这个新建立的平台在最近在我们的研究小组中创建的两个新颖的脚手架中查看骨骼发育的不同阶段。结果将用于解释先前的观察结果，即层状支架具有优质的骨形成能力，而骨骼结构的骨骼能力不论所播种的祖细胞的类型如何。我们假设骨基源细胞最初在体内分布，增殖和进步的方式决定了脚手架中骨形成的最终结果。 成功实施4D成像平台将对该领域进行变革，因为它将提供有关特定策略为何成功或失败的基本信息。该平台将成为理解空间关系的主要成像基础，以欣赏细胞元素之间的时间事件，细胞元素与骨形成过程中细胞/脚手架相互作用。它将允许研究人员开始理解并最终优化脚手架设计和细胞参与者，以及对于骨骼再生的早期阶段的生长因子选择和释放曲线设计。这个知识库对于骨组织工程社区至关重要。 公共卫生相关性：在美国，每年进行超过130万次的骨修复程序，这对骨重生材料产生了巨大的需求。然而，关于骨修复动态过程中支架与细胞之间的时间和空间相互作用的迄今为止，鲜为人知。在这项研究中，我们建议建立一个4D体内成像平台，以在脚手架中的实时细胞活动和骨骼发育中可视化，从而指导我们设计更好的脚手架/细胞复合体以进行骨骼修复。