MESENCHYME?EMBRYONIC STEM CELL INTERACTIONS

间充质？胚胎干细胞相互作用

• 批准号: 7958771
• 负责人: THADDEUS G GOLOS
• 金额: $ 4.68万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7958771
• 关键词: Adherent Culture; Cell Communication; Cell Line; Cells; Characteristics; Computer Retrieval of Information on Scientific Projects Database; Confocal Microscopy; Endometrium; Environment; Extracellular Matrix; Fluorescence; Fluorescent Dyes; Funding; Grant; Hormones; Human; Human Chorionic Gonadotropin; Immigration; Institution; Label; Laboratories; Mesenchymal; Mesenchyme; Methods; Modification; Monitor; Placental Hormones; Pregnancy; Primates; Quantum Dots; Research; Research Personnel; Resources; Source; Staining method; Stains; Surface; System; United States National Institutes of Health; Wisconsin; Work; embryonic stem cell; human embryonic stem cell; in vitro Model; matrigel; migration; nanoparticle; nonhuman primate; success; trophoblast; two-dimensional

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Objective: To define the influence of specific extracellular matrix components and placental mesenchymal cells on trophoblast differentiation from human embryonic stem cells. Our laboratory has established an in vitro model to study trophoblast differentiation. Human embryonic stem cells (hESC) will consistently differentiate to trophoblasts and secrete high levels of placental hormones when allowed to form embryoid bodies (EBs), and are transferred into a three-dimensional (3D) extracellular matrix (Matrigel) environment (ECM). Hormone secretion was enhanced in this 3D system in comparison with planar trophoblast outgrowths in standard adherent culture. The invasion of extracellular matrix and the uterine endometrium by extravillous trophoblasts is a crucial component of pregnancy success in human and nonhuman primates. To define the invasive characteristics of the hESC-derived extravillous trophoblasts, we worked to determine optimal systems for monitoring and quantifying trophoblast migration. We have determined that embryoid bodies derived from aggregates of approximately 500 hESC produce maximal levels of hCG, a marker of trophoblast differentiation, in comparison with larger embryoid bodies. Studies of migration with embryoid bodies stained with a lipophilic fluorescent dye proved unsatisfactory due to high background fluorescence in the migration chambers employed. Modifications have been introduced which provide better control over fluorescence, quantitation and migration. These changes include enrichment of embryoid body trophoblasts by selection following outgrowth on 2-dimensional Matrigel surfaces, and labeling with Quantum Dots fluorescent nanoparticles for quantification of migration by confocal microscopy. We conclude that the migration of hESC-derived trophoblasts, and its modulation by specific culture surfaces, can be quantitatively monitored with these methods. This work used federally approved hES cell lines.

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 目的：探讨特异性细胞外基质成分和胎盘间充质细胞对人胚胎干细胞向滋养层细胞分化的影响。 本实验室建立了滋养层细胞分化的体外模型。人类胚胎干细胞(HESC)在被允许形成类胚体(EBS)时会持续分化为滋养层细胞并分泌高水平的胎盘激素，并被转移到三维(3D)细胞外基质(Matrigel)环境中(ECM)。与标准贴壁培养中的平面滋养层细胞生长相比，该3D系统中的激素分泌增加。 绒毛外滋养层细胞对细胞外基质和子宫内膜的侵袭是人类和非人类灵长类动物妊娠成功的重要组成部分。为了确定hESC来源的绒毛外滋养层细胞的侵袭特性，我们致力于确定监测和量化滋养层细胞迁移的最佳系统。我们已经确定，与较大的类胚体相比，来自大约500个hESC聚集体的类胚体产生的hCG水平最高，hCG是滋养层细胞分化的标志。用亲脂荧光染料染色的类胚体迁移的研究被证明是不令人满意的，因为所使用的迁移小室中的背景荧光很高。已经引入了对荧光、定量和迁移提供更好控制的修饰。这些变化包括在二维Matrigel表面生长后通过选择富含类胚体滋养层细胞，以及用量子点荧光纳米颗粒标记通过共聚焦显微镜定量迁移。我们的结论是，这些方法可以定量监测hESC来源的滋养层细胞的迁移以及特定培养表面对其的调节。这项研究使用了联邦批准的HES细胞系。