Kerkis Alexandre, Fonseca Simone A S, Serafim Rui C, Lavagnolli Thais M C, Abdelmassih Soraya, Abdelmassih Roger, Kerkis Irina
Clínica e Centro de Pesquisa em Reprodução Humana Roger Abdelmassih, São Paulo, SP, Brasil.
Cloning Stem Cells. 2007 Winter;9(4):535-48. doi: 10.1089/clo.2007.0031.
Pioneer work in male mouse embryonic stem (ES) cells differentiation into germ cells (GC) showed generations of male or female gametes in separate experiments, using genetically manipulated or preselected ES cells. In an attempt to produce both types of gametes from male mouse ES cells without any genetic manipulation or preselection, we induce the differentiation by retinoic acid (RA) within nonadherent embryoid bodies (EB). It seems that gamete-like cell formation occurs in the correct manner based on the expression of early and late GC-specific genes such as Oct-4, Mvh, Stella, Dazl, Piwil 2, Pdrd 1, Rex 14, Rnf 17, Bmp8b, Acrosin, Stra-8, Haprin, LH-R, Gdf9, Zp3, Zp2, Sycp1, and Sycp3. Immunofluorescence analysis of morphologically well-formed GC and presumptive gametes showed positive labeling for SSEA1, Oct-4, EMA-1, FE-J1, Dazl, Fragilis, Mvh, Acrosin, and acetylated alpha-tubulin. Conventional cytogenetic and FISH analysis indicated a chromosome reduction in ES-derived GC. Our data suggest that ES cells with XY chromosomes can produce under the same experimental conditions both types of presumptive gametes, and this production depends on their positional and temporal information within the EB context.
在雄性小鼠胚胎干细胞（ES细胞）分化为生殖细胞（GC）方面的开创性工作表明，在单独的实验中，利用基因操作或预先选择的ES细胞可产生雄性或雌性配子。为了在不进行任何基因操作或预先选择的情况下从雄性小鼠ES细胞中产生两种类型的配子，我们通过视黄酸（RA）在非贴壁胚状体（EB）中诱导分化。基于早期和晚期GC特异性基因如Oct-4、Mvh、Stella、Dazl、Piwil 2、Pdrd 1、Rex 14、Rnf 17、Bmp8b、顶体蛋白酶、Stra-8、Haprin、促黄体激素受体、生长分化因子9、透明带蛋白3、透明带蛋白2、联会复合体蛋白1和联会复合体蛋白3的表达，似乎配子样细胞以正确的方式形成。对形态良好的GC和推定配子进行免疫荧光分析显示，阶段特异性胚胎抗原1、Oct-4、EMA-1、FE-J1、Dazl、脆性蛋白、Mvh、顶体蛋白酶和乙酰化α微管蛋白呈阳性标记。传统的细胞遗传学和荧光原位杂交分析表明，ES细胞来源的GC中染色体数目减少。我们的数据表明，具有XY染色体的ES细胞在相同的实验条件下可以产生两种类型的推定配子，并且这种产生取决于它们在EB环境中的位置和时间信息。
