In utero mouse embryo phenotyping with high-frequency ultrasound

项目摘要/摘要 这项建议的目标是通过分割选定的器官来表现妊娠早期到中期的小鼠胚胎。 使用高频超声(HFU)在宫内采集3D数据集的系统。国际老鼠Phe- 包括NIH基因敲除(KO)小鼠表型鉴定计划(KOMP2)在内的诺型鉴定联盟(IMPC)将 在接下来的十年里产生20,000个小鼠品系，包括许多重要的人类结构出生模型 缺陷和先天疾病。提供有效fi流水线的表型方法的发展 对KO小鼠品系胚胎生长缺陷的分析是这项工作的高度优先事项。宫内3D 成像方法，能够对各种器官系统进行体积和纵向分析 妊娠早期到中期的小鼠胚胎，将提供额外的有益fit和关键的额外体内数据。 目前不可用。商业高频超声系统在许多研究中心广泛使用，这在很大程度上要归功于 美国国立卫生研究院资助的小动物成像研究项目和共享仪器项目。高频超声是 因此，在子宫内提供可定量分析的3D图像数据是一种很好的候选模式 并存档以支持KOMP2/IMPC胚胎致命表型管道和未来的表型工作。 我们建议开发和验证宫内3D高频超声图像采集协议和图像处理方法。 允许对胚胎发育进行非侵入性的纵向研究，特别是检测和 KO表型的特征。将在子宫内收集分期小鼠胚胎的体积高频超声数据 在E9.5至15.5之间，以建立正常发育数据库。算法将被开发为 分割3D区域并提取量化胚胎阶段和识别区域变化的参数 正常胚胎和KO胚胎。我们将使用定制的环形阵列系统和Visualsonics Vevo获取数据 2100年。fiNe分辨率环形阵列数据将用于初步开发图像处理算法和 然后，算法将适用于Vevo 2100数据。我们将比较得出的定量参数 从两台扫描仪获得的分割结果中确保Vevo 2100能够提供 等量突变检测和定量fi阳离子。将使用野生型和EN1和Gli2进行初步测试 有已知缺陷的突变体。最后，将采集和处理协议应用于3D Vevo 2100来自5-10个KOMP2 KO小鼠品系的数据，这些小鼠在各种器官系统中存在胚胎缺陷，以验证 用高频超声检测和鉴定这些突变胚胎的表型。

Project Summary/Abstract The goal of this proposal is to phenotype early- to mid-gestational mouse embryos by segmenting select organ systems in 3D data sets acquired in utero with high-frequency ultrasound (HFU). The International Mouse Phe- notyping Consortium (IMPC), which includes the NIH Knockout (KO) Mouse Phenotyping Program (KOMP2), will generate 20,000 mouse strains in the next decade, including many important models of human structural birth defects and congenital diseases. The development of phenotyping methods that provide for efficient pipeline analyses of defects in embryonic growth in the KO mouse strains is a high priority for this effort. An in utero 3D imaging approach, enabling volumetric and longitudinal analyses of a variety of organ systems over a range of early- to mid-gestational stage mouse embryos, would provide added benefit and critical additional in vivo data not currently available. Commercial HFU systems are widely available in many research centers largely thanks to the NIH-funded Small Animal Imaging Research Programs and Shared Instrumentation Programs. HFU is therefore an excellent candidate modality to provide in utero 3D image data that can be quantitatively analyzed and archived to support the KOMP2/IMPC embryonic lethal phenotyping pipeline and future phenotyping efforts. We propose to develop and validate in utero 3D HFU image-acquisition protocols and image-processing meth- ods that permit noninvasive, longitudinal studies of embryonic development and, in particular, the detection and characterization of KO phenotypes. Volumetric HFU data will be collected in utero from mouse embryos staged between E9.5 to 15.5 in order to establish a database of normal development. Algorithms will be developed to segment 3D regions and extract parameters that quantify embryonic stage and identify regional changes between normal and KO embryos. We will acquire data with a custom, annular-array system and with a VisualSonics Vevo 2100. The fine-resolution annular-array data will be used to initially develop the image-processing algorithms and then the algorithms will be adapted for Vevo 2100 data. We will compare the quantitative parameters derived from the segmentation results obtained from the two scanners to ensure that the Vevo 2100 is able to provide equivalent mutant detection and quantification. Initial testing will be undertaken using wild-type and En1 and Gli2 mutants that have known defects. Finally, the acquisition and processing protocols will be applied to 3D Vevo 2100 data from 5-10 KOMP2 KO mouse lines with embryonic defects in a variety of organ systems to validate the HFU methods for detecting and characterizing phenotypes in these mutant embryos.