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

利用基于组织透明的 3D 成像进行定量研究 颅面间充质干细胞的神经调控 神经系统对骨调节的最新发现导致了新的发现 各种研究表明神经系统和骨骼之间的调节轴1-3。 来自中枢或周围神经系统的直接或间接调节骨重塑 流程1、2。 研究神经与骨骼相互作用的一项技术挑战是可视化上或下的神经纤维 由于其尺寸细小（直径约 1 µm）且神经元细胞位置较远，因此位于骨骼内 此外，从剖面获得的二维信息使得很难或不可能。 量化复杂颅面组织之间的空间相互作用。 3D 成像的发展。 基于组织清除技术为这两个问题提供了解决方案。 是神经科学领域的一项最新突破，通过渲染组织透明的 3D 图像。 整个器官的观察可以直接用共焦显微镜实现，无需切片4。 我们最近开发了一种新的组织透明技术，名为 PEG 联合溶剂 系统（PEGASOS）。PEGASOS 方法可有效清除硬组织和软组织。 PEGASOS 方法具有透明性并保护内源荧光。 最初是为了研究大脑内的神经元连接组而开发的（试验手稿正在修订中）。 实验表明它也可以应用于颅面硬组织、 从成年小鼠身上摘取的牙齿和股骨在治疗后可以被清除到几乎看不见 在不丢失 GFP 荧光的情况下，我们能够获取血管网络的 3D 图像。 我们还能够获取牙齿、缝线、硬脑膜和颅骨内的 3D 图像。 股骨骨膜、胫骨骨髓、缝线和牙周膜间隙内的神经。 这些 3D 图像的定量分析表明，血管内的脉管系统和神经 与以前相比，颅面部硬组织呈现出高度异质的分布模式。 研究中，我们的初步结果表明缝合线内的 Gli1+ 细胞密切相关 与脉管系统。 在当前的提案中，我们建议改进颅面硬骨的PEGASOS方法 利用基于 PEGASOS 的 3D 成像提供的空间信息， 我们想定量检验颅面部 MSC 相关的假设 神经血管束 (NVB) 和神经对于干细胞迁移至关重要 朝向受伤部位

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