A Developmental Craniofacial Atlas for the Zebrafish

斑马鱼颅面发育图谱

• 批准号: 7828074
• 负责人: CHARLES B KIMMEL
• 金额: $ 47.78万
• 依托单位: UNIVERSITY OF OREGON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-18 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7828074
• 关键词: 3-Dimensional; Address; Adolescent; Adult; Anatomy; Animal Model; Area; Atlases; Binding; Bone Development; Bone Matrix; Branchial arch structure; Cartilage; Cell Count; Cells; Chondrocytes; Cleft Palate; Color; Complex; Confocal Microscopy; Defect; Development; Diagnosis; Dimensions; Disease; Dissection; Dyes; Elements; Embryo; Fishes; Fluorescent Dyes; Genetic; Green Fluorescent Proteins; Growth; Head; Human; Image; Imagery; Information Networks; Internet; Jaw; Label; Larva; Learning; Life; Maintenance; Mesenchyme; Mesoderm; Methods; Minerals; Molecular; Morphogenesis; Mus; Nature; Neural Crest; Online Systems; Optics; Organism; Osteoblasts; Osteocalcin; Osteoclasts; Pattern; Pattern Formation; Phenotype; Physiologic pulse; Primordium; Proteins; Regulator Genes; Relative (related person); Resolution; Resource Informatics; Resources; Scientist; Series; Shapes; Skeleton; Specificity; Staging; Staining method; Stains; Structure; Technology; Transgenes; Transgenic Organisms; Zebrafish; base; bone; cell growth; cell type; craniofacial; cranium; fluorexon; genome-wide; innovation; interest; mutant; promoter; public health relevance; skeletal; spatiotemporal; tomography; virtual; zebrafish development

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06) Enabling Technologies, and specific Challenge Topic 06-DE-102 Structural and Molecular Atlases of Craniofacial Development. We propose to construct a developmental atlas for the craniofacial skeleton of zebrafish, a widely used model organism for understanding conserved features of development of the skull, and human disorders such as cleft palate. The atlas will provide a key resource for scientists wishing to discover how the complex anatomy of the zebrafish craniofacial skeleton develops. It will serve as a critical reference for learning the defects of the many zebrafish craniofacial mutants now available for study, as well as new mutants now being generated through genome-wide initiatives. It also will be useful for understanding development of the zebrafish skull compared with other organisms, such as mouse. The atlas will be web-based and freely available on FaceBase, a web resource rapidly becoming the standard for understanding craniofacial development, and on ZFIN, the specific informatics resource for zebrafish development and genetics. It will include a series of developmental stages beginning with the 1-day postfertilization embryo when the skeletal primordia are present, but bones and cartilages have not yet formed. It will continue with selected stages through the 3-week-old juvenile stage when nearly all of the elements of the skull are present, and shaped approximately as in the adult, but of miniature size. It will be 3-dimensional and interactive: The user will be able to virtually dissect the skull - calling up particular subsets of elements of interest, to rotate the image and observe a set of elements from any point of view, to section through the structure in any plane, and to examine how the structure changes at different developmental stages. A key innovation is that the atlas will be based entirely on images taken of the living, developing organism expressing transgenic fluorescent proteins. These markers will be driven by the promoters of key skeletal regulatory genes, including fli1, expressed in all craniofacial mesenchyme, and dlx5a, expressed in a subset of this mesenchyme that is restricted to the ventral elements of the skull, including the lower jaw. During stages of skeletal morphogenesis and growth, the cells making the skeleton will be revealed specifically by expression of markers including sox10 and foxp2a for cartilage, and osterix for bone. The fish expressing one of these markers also will be vitally counterstained with a fluorescent dye such as Alizarin red to reveal the mineralized bone. The atlas will feature, for every stage, high magnification, through-focus, 2-color image stacks made by confocal microscopy. Such imaging takes full advantage of the many currently available transgenic constructs, the optical clarity of the zebrafish, and of its relatively small size and the relatively small number of cells that build the skull. These attributes combine to provide cell-level resolution with unsurpassed specificity and detail throughout an extended period of development. The atlas will also include, especially for the later developmental stages, medium-resolution images at lower magnification made by optical projection tomography (OPT). The user will be able to interactively manipulate these OPT stacks in the same way as for the confocal stacks, and will allow better 3-d understanding of the more global features of skull organization and development. PUBLIC HEALTH RELEVANCE: The zebrafish craniofacial atlas that will come from this project will provide a resource for skeletal anatomy in an animal model that is valuable for understanding human craniofacial disorders such as cleft palate. The resource will provide for understanding normal morphological developmental in zebrafish as well as being a reference for diagnosing the nature of craniofacial mutant phenotypes.

描述（由申请人提供）：本申请涉及广泛的挑战领域 (06) 支持技术，以及具体的挑战主题 06-DE-102 颅面发育的结构和分子图谱。我们建议为斑马鱼的颅面骨骼构建发育图谱，斑马鱼是一种广泛使用的模型生物，用于了解头骨发育的保守特征以及腭裂等人类疾病。该图集将为希望了解斑马鱼颅面骨骼的复杂解剖结构如何发育的科学家提供关键资源。它将作为了解目前可用于研究的许多斑马鱼颅面突变体的缺陷以及目前通过全基因组计划产生的新突变体的重要参考。它还有助于了解斑马鱼头骨与其他生物（例如小鼠）的发育情况。该图集将基于网络并在 FaceBase（一种迅速成为了解颅面发育标准的网络资源）和 ZFIN（斑马鱼发育和遗传学的特定信息学资源）上免费提供。它将包括从受精后 1 天的胚胎开始的一系列发育阶段，此时骨骼原基已经存在，但骨骼和软骨尚未形成。它将继续进行选定的阶段，直到 3 周大的幼年阶段，此时几乎所有头骨元素都已存在，并且形状与成人相似，但尺寸较小。它将是 3 维和交互式的：用户将能够虚拟地解剖头骨 - 调用感兴趣元素的特定子集，旋转图像并从任何角度观察一组元素，以剖析结构在任何平面上，并检查结构在不同发育阶段如何变化。一个关键的创新是，该图谱将完全基于表达转基因荧光蛋白的活体、发育中的生物体拍摄的图像。这些标记将由关键骨骼调节基因的启动子驱动，包括在所有颅面间质中表达的 fli1 和在该间质的一个子集中表达的 dlx5a，该间质仅限于颅骨的腹侧元件，包括下颌。在骨骼形态发生和生长阶段，构成骨骼的细胞将通过标记物的表达来具体揭示，这些标记物包括软骨的 sox10 和 Foxp2a 以及骨的 osterix。表达这些标记之一的鱼也将用茜素红等荧光染料进行重要复染，以显示矿化骨骼。该图谱将针对每个阶段提供由共焦显微镜制作的高放大倍率、离焦、2 色图像堆栈。这种成像充分利用了许多目前可用的转基因结构、斑马鱼的光学清晰度、其相对较小的尺寸和相对较少数量的构成头骨的细胞。这些属性相结合，可在整个发育过程中提供具有无与伦比的特异性和细节的细胞级分辨率。该图集还将包括由光学投影断层扫描 (OPT) 制作的较低放大倍率的中等分辨率图像，特别是针对后期的发展阶段。用户将能够以与共焦堆栈相同的方式交互式操作这些 OPT 堆栈，并且可以更好地 3D 理解头骨组织和发育的更全局特征。 公共健康相关性：该项目产生的斑马鱼颅面图谱将为动物模型中的骨骼解剖学提供资源，这对于了解人类颅面疾病（例如腭裂）非常有价值。该资源将为了解斑马鱼的正常形态发育提供依据，并为诊断颅面突变表型的性质提供参考。