Biopsy of preimplantation mouse embryos: development of micromanipulated embryos and proliferation of single blastomeres in vitro.
Biopsy of preimplantation mouse embryos: development of micromanipulated embryos and proliferation of single blastomeres in vitro.
复制标题
植入前小鼠胚胎的活检：显微操作胚胎的发育和体外单个卵裂球的增殖。
作者：
L. Wilton;A. Trounson
我们已经开发出一种技术来采样植入前胚胎，这可能是，在未来，应用于遗传疾病的产前诊断。利用显微操作，我们从4细胞小鼠胚胎中吸出一个卵裂球。该程序对体外发育没有影响; 98%的对照和94%的活检胚胎在培养48小时后达到囊胚期。此外，在转移到假孕受体小鼠后，活检胚胎的胎儿发育率与对照胚胎无显著差异，尽管植入率显著降低（平均值+/- SD：活检53.1 +/- 4.0，对照81.8 +/- 8.4，p <0.001）。这是我们第一次从单个着床前卵裂球中培养出单层细胞。在不同的细胞外基质成分上体外培养单个活检的卵裂球。在用纤连蛋白（FN）、层粘连蛋白（LN）和层粘连蛋白和巢蛋白（LNC）的复合物包被的威尔斯孔中获得的细胞增殖显著高于在特异性较低的猪皮明胶（SSG）基质中获得的细胞增殖。培养6天后，在包被SSG、LN、FN和LNC的威尔斯孔中，每孔的平均（+/- SE）细胞核数分别为6.4 +/- 2.1、11.9 +/- 1.5、19.8 +/- 2.9和20.9 +/- 2.6。
We have developed a technique to sample the preimplantation embryo, which may, in the future, be applied to prenatal diagnosis of genetic disease. Using micromanipulation, we aspirated a single blastomere from 4-cell mouse embryos. This procedure had no effect on in vitro development; 98% of control and 94% of biopsied embryos reached the blastocyst stage after 48 h in culture. Furthermore, after transfer to pseudopregnant recipient mice, the rate of fetal development of biopsied embryos was not significantly different from control embryos, although implantation rate was significantly reduced (mean +/- SD: biopsied 53.1 +/- 4.0, control 81.8 +/- 8.4, p less than 0.001). For the first time we have produced monolayer cell cultures derived from single preimplantation blastomeres. Individual biopsied blastomeres were cultured in vitro on different extracellular matrix components. Significantly greater cell proliferation was obtained in wells coated with fibronectin (FN), laminin (LN), and a complex of laminin and nidogen (LNC) than in a less specific matrix of swine skin gelatin (SSG). Mean (+/- SE) cell nuclei number per well after 6 days in culture was 6.4 +/- 2.1, 11.9 +/- 1.5, 19.8 +/- 2.9, and 20.9 +/- 2.6 in wells coated with SSG, LN, FN, and LNC respectively.