Analysis of development and differentiation suppression mechanism in the tetraploid mouse embryos

哺乳动物发育早期禁止有丝分裂或细胞融合产生四倍体胚胎。然而，这些胚胎不能发育到妊娠足月，也不能生出活的幼崽。四倍体细胞能在体外存活并维持，但不能形成活的胎儿。它们发育的失败被认为取决于细胞大小的增加，从而导致代谢不足和细胞周期的延迟。本研究在细胞查素B诱导的四倍体小鼠胚胎中，直接证实了四倍体细胞周期的延迟，尽管桑葚胚和囊胚的形成与正常胚胎一样按时进行。有丝分裂禁止的持续产生了一个细胞中2到4个巨大的细胞核和多核。对于多倍体的分析，我们采用直接r带FISH方法对细胞周期间期进行分析，而不是在有丝分裂期进行核型分析。通过该方法，从MHC H类区获得的BAC的信号为2个单点，四倍体为4个单点。结果表明，直接r带FISH方法是鉴别染色体间基重复的有效方法。利用绿色荧光转基因小鼠制备了嵌合小鼠，以获得胚胎发生过程中单倍体细胞和四倍体细胞的痕迹。两种细胞系在紫外光下很容易被识别，在GFP-Tg小鼠衍生细胞中显示绿色荧光。通过对四倍体和部署体胚胎嵌合体的分析，发现四倍体来源的细胞主要构成胚胎外组织，如胎盘和胚胎膜，而部署体来源的细胞则主要构成胚胎。有丝分裂抑制法得到的四倍体与电细胞融合法得到的四倍体结果不同。有丝分裂抑制与细胞融合对四倍体的差异有待进一步研究。少

Mitosis prohibition or cell fusion at early developmental stage produces the tetraploid embryos in mammals. However, these embryos can not develop for full term of gestation and never deliver living young. The tetoploid cells survive and are maintained in vitro, but the cells can not form living fetus. The failure of their develop ment are considered to depend on increment of the cell size which results in insufficiency of metabolism and delay of cell cycles. In the present study of the tetraploid mouse embryo induced by cytochalsin B, the delay of tetra ploid cell cycles were confirmed directly in spite of the morula and blastcyst formatidh is performed on time as normal deploid embryos. And continuation of mitosis prohibition produced enormously large nucleus and multi nuclei from 2 to 4 in a cell. For the analysis of the polyploidy, we used the direct R-banding FISH method for the interphase of cell cycle, unlike karyotype analysis at the mitotic phase. By this method the signal of … More BAC clone from MHC class H area was found as 2 single dots in deploid and 4 single dots in tetraploid. It shows that the direct R-banding FISH method is a useful techniques for distinction of chromosomes duplication in interp base. For the trace of the deploid and tetraploid cell during embryogenesis, the chimeric mouse were made using green fluorescent transgenic mouse. Both cell lineages were readily identified under the uv lihgt, as green fluorescent in the GFP-Tg mouse derived cell. As a result of the analysis of chimra between deploid and tetra ploid embryos, tetraploid derived cells composed mainly the extraembryonic tissue, such as placenta and embryonic membrane, in contarast to the deploid derived cells contributed to the embryos. Above mentioned results from the tetraploidy by mitosis inhibition were differ from the tetraploidy by the electric cell fusion. 'The further study should be performed on the difference of tetraploidy between by mitosis inhibition and cell fusion in detail. Less