Utilizing tissue clearing based 3-D imaging to quantitatively study neural regulation of craniofacial mesenchymal stem cells

利用基于组织透明化的3D成像定量研究颅面间充质干细胞的神经调节

• 批准号: 9762081
• 负责人: JIAN Q. FENG
• 金额: $ 18.56万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-10 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9762081
• 关键词: 3-Dimensional; Address; Adult; Aftercare; Antibodies; Blood Vessels; Bone Injury; Bone Marrow; Bone Regeneration; Bone remodeling; Brain; Caliber; Calvaria; Cells; Chemicals; Communities; Denervation; Development; Distant; Dura Mater; Femur; Fluorescence; Harvest; Image; Image Analysis; Imagery; Imaging Techniques; Injury; Location; Manuscripts; Mesenchymal Stem Cells; Methods; Microscope; Minerals; Mus; Names; Nerve; Nerve Fibers; Nervous system structure; Neurons; Neurosciences; Organ; Pattern; Periodontal Ligament; Periosteum; Peripheral Nervous System; Regulation; Research; Research Personnel; Sampling; Signal Transduction; Site; Solvents; Spatial Distribution; Spinal Cord; Stains; Stem cells; Surgical sutures; System; Techniques; Testing; Three-Dimensional Image; Three-Dimensional Imaging; Three-dimensional analysis; Tissues; Tooth structure; base; bone; cell motility; computerized data processing; connectome; craniofacial; craniofacial bone; craniofacial complex; craniofacial tissue; cranium; density; design; experimental study; improved; injury and repair; migration; neuroregulation; neurotransmission; neurovascular; preservation; relating to nervous system; skeletal; soft tissue; tibia; tool

Utilizing tissue clearing based 3-D imaging to quantitatively study neural regulation of craniofacial mesenchymal stem cells Recent discoveries of bone regulation by the nervous system has led to the identification of new regulatory axes between the nervous system and bone1-3. Various studies indicate that signal from the central or peripheral nervous system directly or indirectly regulates the bone remodeling process1, 2. One technical challenge on studying the nerve-bone interactions is to visualize nerve fibers on or within bones because of their fine size (~1µm diameter) and the distant location of the neuron cell bodies. In addition, 2-D information obtained from sections made it difficult or impossible to quantify spatial interactions between complex craniofacial tissues. Development of 3-D imaging based tissue clearing technique provides solutions to these two issues. Tissue clearing technique is a recent breakthrough in the neuroscience field. By rendering tissue transparent, 3-D images of the whole organ can be achieved directly with a confocal microscope without sectioning4. We recently developed a new tissue clearing technique named PEG Associated Solvent System (PEGASOS). The PEGASOS method efficiently clears both hard and soft tissue to high transparency and protects endogenous fluorescence with no loss. The PEGASOS method was initially developed to study the neuron connectome within the brain (manuscript in revision). Pilot experiments suggested that it can also be applied on the craniofacial hard tissue. Skull bone, teeth and femur harvested from adult mice can be cleared to nearly invisible after treatment without losing GFP fluorescence. We were able to acquire the 3-D images of vascular network within the tooth, suture, dura and calvarial bone. We were also able to acquire the 3-D images of nerves within the femur periosteum, tibia bone marrow, suture and periodontal ligamental space. Quantitative analysis of these 3-D images indicated that vasculatures and nerves within craniofacial hard tissue present highly heterogenous distribution pattern. In contrast to previous study, our preliminary results suggested that Gli1+ cells within the suture are closely associated with the vasculatures. In the current proposal, we propose to improve the PEGASOS method for craniofacial hard tissue research. With the spatial information provided by PEGASOS based 3-D imaging, we would like to quantitatively test the hypothesis that craniofacial MSCs are associated with the neurovascular bundle (NVB) and nerves are essential for stem cell migration towards the injury site

利用基于组织清晰度的三维成像进行定量研究 颅面间充质干细胞的神经调控 最近神经系统对骨骼调节的发现导致了新的 神经系统和骨骼之间的调节轴1-3。各种研究表明，信号 从中枢或外周神经系统直接或间接调节骨重塑 进程1、进程2。 研究神经-骨骼相互作用的一个技术挑战是将神经纤维显示在或 在骨骼内，因为它们的细小(直径约1微米)和神经细胞的遥远位置 身体。此外，从各部分获得的2-D信息使得很难或不可能 量化复杂的头面部组织之间的空间相互作用。三维成像技术的发展 基于组织的透明技术为这两个问题提供了解决方案。组织清除技术 是神经科学领域最近的一项突破。通过渲染组织透明的3-D图像 用共聚焦显微镜可以不经切片直接实现对整个器官的观察。 我们最近开发了一种新的组织清除技术，名为聚乙二醇联用溶剂 系统(PEGASOS)。PEGASOS方法有效地清除了硬组织和软组织到高 透明，保护内源荧光，不会损失。PEGASOS方法是 最初开发是为了研究大脑中的神经元连接体(手稿正在修改中)。引航员 实验表明，它也可以应用于颅面硬组织。头盖骨， 成年小鼠的牙齿和股骨在治疗后可以被清除到几乎看不见的位置 而不会失去GFP荧光。我们能够获得血管网络的三维图像 在牙齿、缝线、硬脑膜和头盖骨内。我们还获得了 股骨骨膜、胫骨骨髓、骨缝和牙周膜间隙内的神经。 对这些三维图像的定量分析表明，血管和神经 颅面硬组织呈高度不均匀分布。与之前的对比 研究表明，我们的初步结果表明，缝线内的Gli1+细胞密切相关 与血管有关。 在目前的方案中，我们建议改进PEGASOS方法治疗颅面部硬化症 组织研究。利用基于PEGASOS的3-D成像提供的空间信息， 我们想要定量地检验颅面间充质干细胞与 神经血管束(NVB)和神经是干细胞迁移所必需的 向受伤地点进发

项目成果

Author Correction: Tissue clearing of both hard and soft tissue organs with the PEGASOS method.

作者更正：使用 PEGASOS 方法对硬组织和软组织器官进行组织清除。

• DOI: 10.1038/s41422-019-0180-5
• 发表时间: 2019
• 期刊: Cell research
• 影响因子: 44.1
• 作者: Jing,Dian;Zhang,Shiwen;Luo,Wenjing;Gao,Xiaofei;Men,Yi;Ma,Chi;Liu,Xiaohua;Yi,Yating;Bugde,Abhijit;Zhou,BoO;Zhao,Zhihe;Yuan,Quan;Feng,JianQ;Gao,Liang;Ge,Woo-Ping;Zhao,Hu
• 通讯作者: Zhao,Hu