Modeling human trophoblast stem cells using iPS cells derived from molar placenta

使用源自臼齿胎盘的 iPS 细胞模拟人类滋养层干细胞

• 批准号: 8511232
• 负责人: Mana M Parast
• 金额: $ 19.38万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-15 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8511232
• 关键词: Activities of Daily Living; Animal Model; BMP4; Biological Assay; Blood Vessels; Blood flow; CDKN1C gene; CDX2 gene; Cell Culture System; Cell Culture Techniques; Cell Line; Cell physiology; Cell surface; Cells; Chorionic villi; Chronic; Commit; Complete Hydatidiform Moles; Conceptus; DNA; Data; Derivation procedure; Development; Diagnosis; Disease; Dose; Embryo; Epithelial; Erythroblasts; Evaluation; Fertilization; Fetal Growth Retardation; Fetal Tissues; Fibroblasts; First Pregnancy Trimester; Gases; Gene Expression; Gene Expression Profiling; Genes; Genome; Growth Factor; HLA G antigen; Hospitals; Human; Human Chorionic Gonadotropin; Hydatidiform Mole; Hypoxia; In Vitro; Lead; Life; Maintenance; Mediating; Mesoderm; Modeling; Molecular; Molecular Profiling; Morbidity - disease rate; Morphology; Mus; Nutrient; Organ; Ovum; Partial Hydatidiform Mole; Phenotype; Placenta; Placentation; Pluripotent Stem Cells; Population; Pre-Eclampsia; Pregnancy; Pregnancy Complications; Protocols documentation; Sampling; Sendai virus; Side; Stem cells; Subfamily lentivirinae; Surface; Syncytiotrophoblast; System; Teratoma; Time; Tissues; Triploidy; Tumor Suppressor Proteins; Villous; Villus; Woman; base; blastocyst; cell behavior; cytotrophoblast; embryo tissue; fetal; fetal blood; human embryonic stem cell; human embryonic stem cell line; imprint; in vivo; induced pluripotent stem cell; mortality; mouse model; paternal imprint; pluripotency; pregnancy disorder; progenitor; public health relevance; sperm cell; success; transcription factor; trophoblast

DESCRIPTION (provided by applicant): The placenta is a fetal-derived organ, whose proper development and function are pivotal in pregnancy success. Trophoblast cells comprise the epithelial compartment of the placenta, and mediate nutrient/gas exchange functions and establishment of maternal blood flow into the feto-placental unit. Abnormal differentiation of trophoblast during early embryonic life is thought to lead to chronic feto-placental hypoxia and result in pregnancy complications associated with both maternal and fetal morbidity and mortality, such as preeclampsia and fetal growth restriction. One major limitation in studying these diseases is the lack of a useful human trophoblast stem cell culture system, where differentiation and lineage specification can be examined at the molecular level. Current human trophoblast cell lines (i.e. HTR8, BeWo, JEG3) are limited in their differentiation potential and functional resemblance to trophoblast in vivo. Also, despite the ability to derive trophoblast stem cells from blastocysts, mouse models are of limited use, due to major differences in placental morphology, trophoblast subtypes, and lineage-specific functional abilities. Human trophoblast stem cells have yet to be derived. Recently, several groups have shown trophoblast differentiation following BMP4 treatment of human embryonic stem cells (hESCs); however, it has been difficult to identify trophoblast stem cells in this system, partly due to the mixture of other resulting lineages, in particular mesoderm, in early cultures following BMP4 addition. We have recently shown that induced pluripotent stem cells (hiPSCs), similar to hESCs, are capable of differentiating into trophoblast following BMP4 treatment. We therefore propose to take advantage of complete molar gestations, which, being exclusively of paternal origin, are unable to give rise to any embryonic tissue and result in a subtype of gestational trophoblastic disease characterized by abnormal placental tissue with hydropic chorionic villi and trophoblastic proliferation. We will isolate fibroblast from these molar placentas and reprogram them into iPS cells using lenti- or Sendai virus expressing Klf4, Sox2, Oct4, and c-Myc. We will then differentiate these iPSCs into trophoblast using BMP4 and characterize the resulting cells at early timepoints following treatment. We hypothesize that these molar iPSC-derived trophoblast will provide a more uniform cell culture system for defining the human "trophoblast stem" cell signature. Characterization of these cells will further provide us with surface markers and lineage-specific transcription factors by which to define this cell population and will help to establish culture conditions for their maintenance in vitro.

描述（由申请人提供）：胎盘是胎儿衍生的器官，其适当的发育和功能在妊娠成功中是关键的。滋养细胞包括胎盘的上皮区室，并介导营养/气体交换功能以及将母体血流建立到feto-plactartental单元中。胚胎早期生活期间滋养细胞的异常分化被认为会导致慢性胎儿缺氧，并导致与母体和胎儿的发病率和死亡率相关的妊娠并发症，例如先兆子痫和胎儿生长限制。研究这些疾病的一个主要局限性是缺乏有用的人滋养细胞细胞培养系统，在该系统中可以在分子水平上检查分化和谱系规范。当前的人类滋养细胞细胞系（即HTR8，BEWO，JEG3）的分化潜力和功能相似之处与体内的滋养细胞相似。另外，尽管能够得出滋养细胞茎 由于胎盘形态的主要差异，滋养细胞亚型和谱系特异性功能能力，小鼠模型的使用有限，因此小鼠模型的使用有限。人滋养细胞干细胞尚未得出。最近，在BMP4治疗人类胚胎干细胞（HESC）后，几组显示了滋养细胞的分化。然而，很难在该系统中识别该系统中的滋养细胞干细胞，部分原因是在添加BMP4后，早期培养物中其他结果谱系的混合物，尤其是中胚层的混合物。我们最近表明，类似于hESC的诱导多能干细胞（HIPSC）能够在BMP4处理后能够区分成滋养细胞。因此，我们建议利用完整的摩尔妊娠，这些妊娠仅是父亲起源的，无法引起任何胚胎组织，并导致妊娠滋养细胞疾病的亚型，其特征在于胎盘异常的胎盘组织，其水力发电粘性绒毛和滋养性蛋白质和滋养性扩散。我们将从这些摩尔胎盘中分离成纤维细胞，并使用表达KLF4，SOX2，OCT4和C-MYC的扁豆或仙台病毒将其重新编程为IPS细胞。然后，我们将使用BMP4将这些IPSC区分为滋养层细胞，并在处理后的早期时间点表征所得的细胞。我们假设这些摩尔IPSC衍生的滋养层细胞将提供更均匀的细胞培养系统，用于定义人类“滋养细胞干燥的干细胞”细胞特征。这些细胞的表征将进一步为我们提供表面标记和谱系特异性的转录因子，通过这些因素来定义该细胞群体，并有助于建立体外维持的培养条件。