小鼠4-细胞期胚胎单卵裂球体外发育潜能的实验研究

目的:探讨小鼠4-细胞期胚胎的单卵裂球(4-cell stage mice embryo derived-blastomeres, 4c-MEDB)的全能性。方法:将小鼠4-细胞期胚胎分割产生单个卵裂球,在体外无透明带培养条件下,不加外源性细胞因子,观察其发育潜能及特性,并将得到的囊胚与正常囊胚进行形态学、免疫学鉴定比较,同时将4c-MEDB来源囊胚(individual blastomere derived-blastocysts,IBDB)移植到假孕母鼠子宫,观察其发育情况。结果:①体外无透明带培养条件下,4c-MEDB较正常胚胎发育滞后,囊胚获得率低;②4c-MEDB具有发育成为完整囊胚的潜能;③IBDB移植入假孕小鼠子宫后具有一定发育能力,能发育到原条期,但未获得足月妊娠。结论:在未添加外源性细胞因子的体外无透明带培养条件下,小鼠4-细胞期胚胎单卵裂球具有发育为完整囊胚的能力,所获囊胚移植入假孕小鼠子宫后可以发育到原条期。

In vitro Developmental Potential of Individual Blastomeres Derived from 4-Cell Stage Mice Embryo Cultured without Zona Pellucida

Objective： To study the in-vitro developmental potential of individual mouse blastomeres derived from 4-cell stage mice embryo（4c-MEDB）, and its future implications for human embryo stem cell （hESC） study. Methods： The 4-cell stage mice embryos were dissociated by enzymatic digestion and the resulting individual blastomeres（4c-MEDB） were cultured in in vitro conditions, free of zona pellucida and without any special growth factors, in order to observe their developmental potential（experimental group）. Untreated 4-cell embryos were cultured as the control （control group）. The resulted blastocysts （from both experimental and control groups） were compared with each other in the following items： morphology, diameter, cells number, incidence of blastocyst formation, and immunofluorescence staining （Oct-4）, on the other hand, the blastocysts were transplanted into pseudopregnant female mice uteri for further development. Results： （1） In vitro culture without zona pellucida, the 4c-MEDB were lagged behind the control, and the incidence of blastocyst formation from 4c-MEDB was less than that in the control （P〈0.05）. （2）4c-MEDB has the potential to develop to fully blastocyst（4c-MEDB）. （3） After transported into pseudopregnant female mice, IBDB can further develop to primitive streak stage, but it can＇t access to live birth in our study. Conclusion： In in vitro culture condition, free of zona pellucida and without any special growth factor, the 4c-MEDB has the potential to develop to fully blastocyst, the IBDB can further develop to primitive streak stage after transplanting into pseudopregnant female mice uteri.