Skeletal Phenotyping of Heterozygotes from IMPC Embryonic Lethal Lines

IMPC 胚胎致死系杂合子的骨骼表型

• 批准号: 10622475
• 负责人: PETER MAYE
• 金额: $ 51.9万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-02 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10622475
• 关键词: Adult; Affect; Animals; Architecture; Area; Big Data; Bioinformatics; Biological; Biology; Body Composition; Body Size; Bone Marrow; Breeding; Calibration; Cartilage; Cell Nucleus; Chromosome Mapping; Classification; Collagen; Communities; Complement; Critiques; Cryopreservation; Data; Developmental Biology; Diameter; Dimensions; Distal; Dyes; Embryo; Embryonic Development; Engineering; Enzyme-Linked Immunosorbent Assay; Epiphysial cartilage; Face; Fatty acid glycerol esters; Femur; Fiber; Formalin; Frequencies; Funding Agency; Genes; Genotype; Goals; Grant; Harvest; Health; Heterozygote; Histologic; Human; Image Analysis; Informatics; Injections; International; Investments; Knock-out; Knockout Mice; Knowledge; Life; Literature; Location; Maps; Marrow; Microscopy; Minerals; Modality; Molecular; Mus; Myelogenous; Osteoblasts; Osteoclasts; Osteogenesis; Outcome; Pathology; Pathway interactions; Pattern; Peptides; Phenotype; Physiologic calcification; Physiological; Population; Positioning Attribute; Pregnancy; Primary Cell Cultures; Process; Production; Program Development; Protocols documentation; Recommendation; Research; Research Personnel; Sampling; Serum; Shapes; Skeletal Development; Skeletal Muscle; Skeleton; Slide; Soleus Muscle; Source; Spectrum Analysis; Stains; Structure; Testing; Tissues; United States National Institutes of Health; Variant; Vertebral column; Work; animation; bone; bone mass; cortical bone; crosslink; design; expectation; experience; fetal; insight; knockout gene; lean body mass; mutant; osteoblast differentiation; osteoclastogenesis; outreach; phenotypic data; programs; repository; sample collection; screening; skeletal; skeletal abnormality; spine bone structure; substantia spongiosa; tibia; transcriptome sequencing; videoconference; web portal; young adult

Abstract The International Mouse Phenotyping Consortium (IMPC) has developed an invaluable repository of gene knockout (KO) mice and has performed a whole-animal assessment of each KO line. Approximately 1/3 of the KO lines are embryonic lethals that were characterized as heterozygous (HET) KO lines. Previously we performed high throughput µCT and histomorphometric analysis on 220 unselected viable homozygous (HOM) KO lines. This study found that ~12.5% have an unequivocal variance in total bone mass (trabecular bone volume and/or bone size), most of which were not detected by the IMPC screening. The results and interpretation of the KOs examined in the screen are available on a web portal (bonebase.org). Based on this experience, the phenotyping data on the IMPC webportal and the literature, we estimate that the HET population of KOs will be an equally rich source of genes that affect bone/body variance and skeletal health in the later adult years. This proposal will apply our skeletal phenotyping workflow to selected HET KO lines that are likely to have a significant bone/body mass phenotype. Both µCT and total body composition as determined by TD-NMR will be used as the screening modality. HET KO lines with significant variance in bone architecture will be processed for histomorphometry. Based on those results, certain lines will undergo studies of osteogenesis (Raman microscopy for the mineral/matrix composition, primary cell culture for osteoblast) and/or osteoclastogenesis (collagen crosslinks and primary cell culture for osteoclast). In HET KO lines with variance in body composition, histomorphometry of the skeletal muscle will be performed. The intent is to discriminate the impact of the HET KO as acting directly on the osteoblast or osteoclast, or whether the observed phenotype is a secondary effect of the HET KO on other tissues that can influence bone. In addition, HOM KO lethal lines that survive to 18.5 days of gestation will undergo a gross and histological skeletal examination to determine the impact of the gene on skeletal development. To complement these biological findings, a bioinformatics component is being added to map the KO genes that are affecting osteoblast or osteoclast directly to known or postulated molecular pathways. Does the gene participate in the function of the network or is it a product of a network? The final goal of the project is to place the biological and bioinformatic data into a classification structure that reveals differences and similarities between the various KO lines. We will engage internal and external experts in bone biology to review the data assembled for a specific HET KO line to critique and amplify our interpretation and ever-evolving classification schema. Recognizing that frequency and complexity of the genes affecting bone is a big-data challenge (estimated to be ~3,500 genes), our intent is to lay the ground work for how this information will be gathered, presented, interpreted, queried and eventually applied toward the goal of personalized skeletal human health.

摘要 国际小鼠表型鉴定协会（IMPC）已经开发了一个非常宝贵的基因库， 敲除（KO）小鼠，并对每个KO系进行了全动物评估。的约1/3 KO系是被表征为杂合（HET）KO系的胚胎致死。以前我们 对220例可存活的纯合子（HOM）进行了高通量µCT和组织形态计量学分析 KO线。这项研究发现，约12.5%的人的总骨量（骨小梁）存在明确的差异 体积和/或骨大小），其中大部分未被IMPC筛查检测到。的结果和 对屏幕中检查的科斯的解释可在门户网站（bonebase.org）上获得。基于此 经验，IMPC门户网站和文献上的表型数据，我们估计HET 科斯的人口将是一个同样丰富的基因来源，影响骨/身体的变化和骨骼健康， 成年后的岁月该提案将我们的骨骼表型工作流程应用于选定的HET KO系， 可能具有显著的骨/体重表型。µCT和全身成分， 通过TD-NMR测定的浓度将用作筛选模式。在骨中具有显著差异的HET KO系 将处理架构以进行组织形态测定。根据这些结果，某些线路将进行研究 骨生成（矿物/基质成分的拉曼显微镜检查，成骨细胞的原代细胞培养） 和/或破骨细胞生成（胶原交联和破骨细胞的原代细胞培养）。在HET KO线路中， 身体组成的变化，将进行骨骼肌的组织形态测定。目的是 区分HET KO直接作用于成骨细胞或破骨细胞的影响，或者 观察到的表型是HET KO对其他组织的继发效应，其可影响骨。此外，本发明还提供了一种方法， 存活至妊娠18.5天的HOM KO致死系将经历大体和组织学骨骼检查。 检查以确定基因对骨骼发育的影响。为了补充这些生物 研究发现，生物信息学成分正在被添加到映射KO基因，影响成骨细胞或 破骨细胞直接与已知或假定的分子途径。基因是否参与了 网络，还是网络的产物？该项目的最终目标是将生物和生物信息学 数据转化为一个分类结构，揭示了各种KO系之间的差异和相似性。我们 将聘请骨生物学方面的内部和外部专家，以审查为特定HET KO收集的数据 线批判和放大我们的解释和不断发展的分类模式。认识到 影响骨骼的基因的频率和复杂性是一个大数据挑战（估计约有3，500个基因）， 我们的目的是为如何收集、呈现、解释和查询这些信息奠定基础 并最终应用于个性化骨骼人类健康的目标。

项目成果

Predicting the targets of IRF8 and NFATc1 during osteoclast differentiation using the machine learning method framework cTAP.

• DOI: 10.1186/s12864-021-08159-z
• 发表时间: 2022-01-07
• 期刊: BMC genomics
• 影响因子: 4.4
• 作者: Wang H;Joshi P;Hong SH;Maye PF;Rowe DW;Shin DG
• 通讯作者: Shin DG

Mesenchyme-specific loss of Dot1L histone methyltransferase leads to skeletal dysplasia phenotype in mice.

DOT1L组蛋白甲基转移酶的间充质特异性损失导致小鼠的骨骼发育异常表型。

• DOI: 10.1016/j.bone.2020.115677
• 发表时间: 2021-01
• 期刊: Bone
• 影响因子: 4.1
• 作者: Sutter PA;Karki S;Crawley I;Singh V;Bernt KM;Rowe DW;Crocker SJ;Bayarsaihan D;Guzzo RM
• 通讯作者: Guzzo RM

Intrafibrillar Mineralized Collagen-Hydroxyapatite-Based Scaffolds for Bone Regeneration.

• DOI: 10.1021/acsami.0c00275
• 发表时间: 2020-03
• 期刊: ACS applied materials & interfaces
• 影响因子: 9.5
• 作者: Le Yu;D. Rowe;I. Perera;Jiyao Zhang;S. Suib;X. Xin;M. Wei