Primate-specific miRNAs in Embryonic and Placental Development

胚胎和胎盘发育中灵长类动物特异性 miRNA

• 批准号: 10405364
• 负责人: Jenna Ann Schmidt
• 金额: $ 4.55万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2022-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10405364
• 关键词: Adult; Blood; Cell Line; Child Health; Chromosome 19; Chromosome Mapping; Chromosome abnormality; Communities; Competence; Development; Diagnostic; Embryo; Embryonic Development; Environment; Ethics; Experimental Animal Model; Fetal Growth Retardation; Fetus; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Genes; Goals; Health; Human; In Vitro; Individual; Investigational Therapies; Knock-out; Knowledge; Lead; Longevity; Macaca; Macaca mulatta; Maps; Mentors; Methods; MicroRNAs; Modeling; Modification; Outcome; Outcomes Research; Pathology; Pathway interactions; Personal Satisfaction; Phase; Placenta; Placentation; Pre-Eclampsia; Pre-implantation Embryo Development; Pregnancy; Pregnancy Complications; Primates; RNA; Research; Resources; Rhesus; Role; Scientist; Therapeutic Agents; Transcend; Universities; Wisconsin; base; cardiovascular disorder risk; career development; cell type; experience; experimental study; fetal; fetal programming; gene therapy; genome editing; implantation; improved; in utero; in vivo; innovation; mRNA Expression; member; migration; nonhuman primate; overexpression; precision medicine; primate development; progenitor; research and development; stem; stem cell differentiation; tool; trophoblast; trophoblast stem cell

PROJECT ABSTRACT Placental pathologies stem from poor early placental development characterized by shallow invasion of trophoblasts, and are also associated with aberrant expression of miRNAs belonging to the primate-specific chromosome 19 miRNA cluster (C19MC). C19MC miRNAs are thought to have roles in trophoblast invasion and migration, however, their role(s) in primate embryonic and early placental development is not well-defined. I hypothesize that C19MC miRNAs will have primate-specific roles in trophectoderm lineage specification and early development of the primate placenta. The overall objective of my proposal is to determine the role of these miRNAs in primate trophoblast lineage specification and placental development in a rhesus macaque model, and reveal gene networks regulated by miRNAs in primate placentation. Towards this objective, I propose three Specific Aims. Specific Aim 1 (K99-phase). To define the expression of miRNAs in the primate embryo and trophoblast stem cells (TSC). This aim will establish the miRNA signature during embryo development through specification of the trophectoderm lineage and determine whether these miRNAs are expressed in a stage- or cell-type-specific manner. Specific Aim 2 (K99-phase). To determine the functional role(s) of C19MC members in TSC and differentiated trophoblast function. This experiment will directly overexpress C19MC miRNA members in TSC to identify genes and pathways regulated by these miRNAs, assessing their functional roles. Specific Aim 3 (R00-phase). To use genome editing strategies to perturb C19MC expression to evaluate the impact of aberrant C19MC miRNA expression on embryo development and primate placentation. This Aim will 1) repress and overexpress C19MC cluster and individual C19MC miRNAs expression in TSCs, 2) apply C19MC genome editing to embryos to evaluate the role of cluster expression on primate preimplantation embryo development, and 3) determine the impact of embryonic genome editing on trophectoderm function and differentiation in an in vitro implantation culture model. I have recently developed macaque TSCs with methods described by Okae et al. (2018), and have experience with rhesus IVF to derive embryos for genome editing and in vitro implantation experiments. TSC and embryo resources will be used to define miRNA and mRNA expression during embryo development and trophoblast lineage specification as well as identify miRNA-regulated gene networks in early placentation. Overall, the proposed research will establish a basic understanding of miRNA expression and miRNA target gene regulation in the embryo to placenta transition. Ultimately, we envision that the nonhuman primate model will allow us to extend these approaches to transfer of edited embryos to recipient dams, and to develop in vivo strategies to directly target the placenta for modification of miRNA expression for experimental and therapeutic purposes.

项目摘要 胎盘病理源于早期胎盘发育不良，其特征是浅层侵入 滋养层细胞，并且还与属于灵长类特异性的 miRNA 的异常表达有关 19 号染色体 miRNA 簇 (C19MC)。 C19MC miRNAs被认为在滋养层侵袭和 然而，它们在灵长类胚胎和早期胎盘发育中的作用尚不明确。我 假设 C19MC miRNA 在滋养外胚层谱系规范中具有灵长类特异性作用 以及灵长类动物胎盘的早期发育。我提案的总体目标是确定角色 这些 miRNA 在灵长类滋养层谱系规范和恒河猴胎盘发育中的作用 模型，并揭示灵长类动物胎盘形成中受 miRNA 调控的基因网络。为了这个目标，我建议 三个具体目标。 具体目标 1（K99 阶段）。确定 miRNA 在灵长类动物胚胎和滋养层干中的表达 细胞（TSC）。该目标将通过规范 miRNA 来建立胚胎发育过程中的 miRNA 特征。 滋养外胚层谱系并确定这些 miRNA 是否以特定阶段或细胞类型表达 方式。 具体目标 2（K99 阶段）。确定 C19MC 成员在 TSC 和分化中的功能角色 滋养层功能。本实验将直接在TSC中过表达C19MC miRNA成员来鉴定基因 以及这些 miRNA 调节的途径，评估它们的功能作用。 具体目标 3（R00 阶段）。使用基因组编辑策略扰乱 C19MC 表达来评估 异常 C19MC miRNA 表达对胚胎发育和灵长类动物胎盘的影响。这个目标 1) 抑制和过表达 TSC 中的 C19MC 簇和单个 C19MC miRNA 表达，2) 将 C19MC 基因组编辑应用于胚胎，以评估簇表达对灵长类动物植入前的作用 胚胎发育，3) 确定胚胎基因组编辑对滋养外胚层功能的影响和 体外植入培养模型中的分化。 我最近用 Okae 等人描述的方法开发了猕猴 TSC。 (2018)，并有经验 与恒河猴体外受精获得胚胎用于基因组编辑和体外植入实验。 TSC 和胚胎 资源将用于定义胚胎发育和滋养层期间的 miRNA 和 mRNA 表达 谱系规范以及识别早期胎盘形成中 miRNA 调节的基因网络。总体而言， 拟议的研究将建立对 miRNA 表达和 miRNA 靶基因调控的基本了解 在胚胎到胎盘的过渡过程中。最终，我们设想非人类灵长类动物模型将使我们能够 将这些方法扩展到将编辑的胚胎转移到受体母鼠中，并开发体内策略 直接靶向胎盘以修饰 miRNA 表达，用于实验和治疗目的。