Cellular commitment to mortality: Analyses of of mouse early embryo

细胞对死亡的承诺：小鼠早期胚胎的分析

• 批准号: 6431448
• 负责人: Minoru S Ko
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6431448
• 关键词: cell differentiation; cell senescence; complementary DNA; developmental genetics; early embryonic stage; embryogenesis; embryonic stem cell; female; genetic library; genome; germ cells; hematopoietic stem cells; kidney cell; laboratory mouse; male; mammalian embryology; ovary; urinary tract

The major goal is to understand the fundamental mechanisms for cellular commitment to mortality. We concentrate on the critical distinction between immortal early embryonic cells and mortal differentiating derivative cells. Our approach is to optimize the use of systematic genomic approaches for profiling gene expression patterns by large-scale cDNA sequencing and cDNA microarray technology with a current cohort of 15,257 genes collected from early mouse embryos. We are particularly focussing on four closely related experimental systems. (1) Differentiation of totipotent and immortal stem cells at E2.5 to mortal trophectoderm cells at E3.5. We have identified 16 novel genes that are dramatically up-regulated at the E3.5 blastocyst stage by comparing gene expression profiles between E2.5 morula stage and E3.5 blastocyst stage. To start to examine whether these genes might be involved in the immortal/mortal transition process, we are currently analyzing the nature of these genes in greater detail. (2) Differentiation and lineage commitment of pluripotent mouse embryonic stem (ES) cells. Expression profiling of mouse ES cells at six time points during the course of their differentiation has been done with the NIA mouse 15K cDNA microarrays. (3) Differentiation of the urogenital system: Germ line cells are often viewed as immortal, because they provide continuity from generation to generation and do not seem to age. To begin the analysis of gene expression regulation involved in germ line-somatic cell interactions, we conducted gene expression profiling by sequencing cell type-specific cDNA libraries that include E13.5 male and female primordial germ cells as well as two stages in the kidney-urogenital developmental axis, E12.5 female mesonephros and newborn ovary. We are currently analyzing genes differentially expressed in these cells, tissues, and organs more in detail. (4) Differentiation of hematopoietic stem cells: In an instance of particular interest, one of the genes (D7Wsu33e) that we previously isolated from E7.5 extraembryonic tissues turned out to be expressed essentially exclusively in hematopoietic cells. Accordingly the gene was renamed "Pan hematopoietic expression (Phemx)". The gene was strongly expressed in Lin- c-Kit+Sca-1+ hematopoietic stem cells, and expression was down-regulated after differentiation, but expression was still detected in all three hematopoietic lineages. We are currently analyzing the function of this gene.

主要目标是了解细胞死亡的基本机制。我们集中在不朽的早期胚胎细胞和致命的分化衍生细胞之间的关键区别。我们的方法是通过大规模cDNA测序和cDNA微阵列技术优化使用系统基因组方法来分析基因表达模式，目前从早期小鼠胚胎中收集了15，257个基因。我们特别关注四个密切相关的实验系统。(1)E2.5时全能和永生干细胞分化为E3.5时的致死滋养外胚层细胞。通过比较E2.5桑葚胚期和E3.5囊胚期的基因表达谱，我们发现了16个在E3.5囊胚期显着上调的新基因。为了开始研究这些基因是否可能参与不朽/凡人的转变过程，我们目前正在更详细地分析这些基因的性质。(2)多能小鼠胚胎干细胞的分化和谱系定型。用NIA小鼠15 KcDNA微阵列对小鼠ES细胞分化过程中6个时间点的表达谱进行了研究。(3)泌尿生殖系统的分化：生殖系细胞通常被认为是不朽的，因为它们提供了一代又一代的连续性，并且似乎不会老化。为了开始分析生殖系-体细胞相互作用中涉及的基因表达调控，我们通过对细胞类型特异性cDNA文库进行测序来进行基因表达谱分析，所述cDNA文库包括E13.5雄性和雌性原始生殖细胞以及肾脏-泌尿生殖器发育轴中的两个阶段，E12.5雌性中肾和新生儿卵巢。我们目前正在更详细地分析这些细胞、组织和器官中差异表达的基因。(4)造血干细胞的分化：在一个特别感兴趣的例子中，我们以前从E7.5胚胎外组织中分离的一个基因（D 7 Wsu 33 e）被证明基本上只在造血细胞中表达。因此，该基因被重新命名为“泛造血表达（Phemx）"。该基因在Lin-c-Kit +Sca-1+造血干细胞中强表达，分化后表达下调，但在所有三种造血谱系中仍检测到表达。目前，我们正在研究这个基因的功能。