Roscovitine同期化供核细胞对猴—猪异种核移植胚胎发育的影响

目的]探讨Roscovitine同期化供核细胞对食蟹猴—猪异种体细胞核移植胚胎体外发育的影响,为提高灵长类的核移植效率奠定基础.方法]活体采集4岁雄性食蟹猴的耳组织,经组织块培养获得纯化的食蟹猴耳成纤维细胞,细胞经固定、染色后用流式细胞仪分析细胞周期分布.以不同同期化方法处理的食蟹猴耳纤维细胞为供体细胞,以体外成熟、去核的猪卵母细胞为受体细胞,利用电融合法构建食蟹猴—猪异种核移植胚胎,观察异种核移植重构胚胎的体外发育情况.结果]以15 μmol/L Roscovitine处理食蟹猴耳成纤维细胞24、48、72 h获得的G0/G1期细胞比率分别为76.51％、89.69％和90.49％,其中处理48和72h的G0/G1期细胞比率显著高于对照组（P＜0.05）.血清饥饿、接触抑制72 h同期化获得的G0/G1期细胞比率分别为91.12％和90.46％.Roscovitine同期化处理食蟹猴耳成纤维细胞48 h可有效提高食蟹猴—猪异种核移植重构胚胎的囊胚形成率（15.05％）,显著高于血清饥饿同期化处理和接触抑制同期化处理的效果（P＜0.05）.结论]Roscovitine可有效同期化食蟹猴耳成纤维细胞在G0/G1期,最终提高食蟹猴—猪异种核移植重构胚胎的囊胚形成率.

Effects of Roscovitine-treated donor cells on synchronization and development of monkey-porcine inter-species nuclear transplantation embryos

Objective]The present study evaluated the effects of Roscovitine on long-tailed macaque ear-derived fi- broblast cell cycle synchronization and embryo development of monkey-porcine interspecies nuclear transfer to provide ref- erences for improving nuclear transfer of primates. Method ] Cells were obtained from the ear of a 4-year-old male mon- key. Cell cycle was distributed by fluorescence-activated cell sorting when ceils were fixed and stained. As donor ceils, long-tailed macaque ear fibroblast cells were introduced into pig enucleated oocytes to investigate development of inter- species cloned embryos. Result]Cell rate of G0/G1 phase under treatment tot 48 and 72 hours by 15 Ixmol/L Roscovitine （89.69%,90.49%） was significantly higher than the control group （P〈0.05）. The cell rate of G0/G1 phase was 91.12% and 90.46% when cells were treated by serum starvation and contact inhibition for 72 hours. Blastocyst formation rate was vast- ly improved under Roscovitine treatment for 48 hours, which was signifcantly higher than that treated by serum starvation and contact inhibition （P〈0.05）. Conclusion ]In conclusion, we suggested that roscovitine treatment of long-tailed macaque ear-derived fibroblastel cells can be used for synchronization of donor cells at the G1 or GO phase which improves the de- velopment of cloned monkey-porcine embryo.