Development of cloned embryos from adult rabbit fibroblasts:: Effect of activation treatment and donor cell preparation
Development of cloned embryos from adult rabbit fibroblasts:: Effect of activation treatment and donor cell preparation
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DOI：
10.1095/biolreprod64.1.257
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发表时间：
2001-01-01
影响因子：
3.6
通讯作者：
Yang, XZ
中科院分区：
文献类型：
--
作者：
Dinnyés, A;Dal, YP;Yang, XZ
本研究旨在研究不同激活方案下成年兔成纤维细胞克隆胚胎的体外和体内发育。研究了血清饥饿和供体细胞传代数对克隆效率的影响。在实验一中，卵母细胞被电脉冲激活，或者被电脉冲激活，然后用6-二甲氨基嘌呤（DMAP）培养。实验二采用实验一的最佳激活方案，对培养0 ~ 15代的成年兔成纤维细胞进行克隆。实验三考察供体细胞血清饥饿对克隆的影响。最后，实验四进行胚胎移植。这些实验表明，电脉冲和DMAP联合处理导致了更好的孤雌生殖囊胚发育（高达29%），并且在支持核移植胚胎的体外发育（16%-18%囊胚）方面，融合前和融合后细胞质的激活并没有什么不同，来自非传代细胞的成纤维细胞发育成囊胚的能力低于传代细胞（6%对17%）。供体细胞的血清饥饿改善了卵裂（高达71%），但没有改善囊胚发育（13%），并且没有获得后代，无论如何处理，成年兔成纤维细胞的细胞周期分析表明，传代6和12细胞比传代0细胞更可能处于G(0)/G(1)，这与传代细胞组中胚胎发育的改善一致。
This research was to study the in vitro and in vivo development of cloned embryos derived from adult rabbit fibroblasts following various activation protocols. Effects of serum starvation and passage number of donor cells on the efficiency of cloning were also examined. In experiment I, oocytes were activated either by electric pulses or by electric pulses followed by culture with 6-dimethylaminopurin (DMAP). For experiment II, the best activation protocol from experiment I was employed for cloning using adult rabbit fibroblasts that were cultured for 0-15 passages. In experiment III, the effect of serum starvation of the donor cells on cloning was examined. Finally, in experiment IV, embryo transfers were conducted. These experiments showed that combined electrical pulse and DMAP treatment resulted in superior parthenogenetic blastocyst development (up to 29%), and that activation of the cytoplast before versus after fusion was not different in supporting the in vitro development of nuclear transferred embryos (16%-18% blastocysts), Adult fibroblasts derived from nonpassaged cells were less capable of developing into blastocysts than passaged cells (6% vs. 17%), Serum starvation of donor cells improved cleavage (up to 71%) but did not improve blastocyst development (13%), and no progeny was obtained, irrespective of the treatment, Cell-cycle analysis of adult rabbit fibroblast cells showed that passage 6 and 12 cells were more likely to be in G(0)/G(1) than passage 0 cells, which agrees with the improved embryo development in the passaged-cell groups.