Collas P, Robl J M
Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst 01003.
Biol Reprod. 1991 Sep;45(3):455-65. doi: 10.1095/biolreprod45.3.455.
The present study characterized the profile of nuclear remodeling in nuclear transplant rabbit embryos and investigated the relationship between chromatin behavior after transfer and embryo development. The developmental potential and pattern of remodeling of donor nuclei from cleavage-, morula-, and blastocyst- (inner cell mass ICM, and trophectoderm, TE) stage donors were evaluated. In addition, we determined whether a modification in the synchrony between blastomere fusion and oocyte activation altered the profile of nuclear remodeling and affected development of reconstituted embryos. Development to blastocysts was similar with 8- and 32-cell-stage donor nuclei (42% and 33%, respectively, p greater than 0.1). However, it was reduced with ICM transplants (17%, p less than 0.05), and development of TE transplants did not progress beyond the 8-cell stage. Upon blastomere fusion into nonactivated oocyte cytoplasm, nuclear remodeling was characterized by premature chromosome condensation (PCC), followed by pronuclear (PN) formation and swelling. PCC occurred synchronously within 1.2-1.5 h post-fusion with all stages of donor nuclei (p greater than 0.1). PN formation in 8- and 32-cell transplants occurred approximately 4 h after fusion, and was synchronous to that of female pronuclei in activated oocytes; however, it was delayed in ICM and TE transplants (p less than 0.01). With all stages of donor nuclei, final nuclear diameter was similar to, or larger than, that of female pronuclei. Fusion to activated oocyte cytoplasm, as opposed to nonactivated cytoplasm, prevented PCC and extensive nuclear swelling (16.0 +/- 0.7 vs. 30 +/- 0.7 microns, respectively, p less than 0.01). Nuclear diameter in early embryos was smaller (p less than 0.01), and development to blastocysts was reduced (p less than 0.05). The results indicate that remodeling of the donor nucleus is not essential for development to blastocysts; however, it is beneficial. Furthermore, complete reprogramming seems possible only after remodeling of the donor nucleus, i.e., PCC in nonactivated cytoplasm, followed by nuclear swelling upon activation of the oocyte.
本研究对核移植兔胚胎中的核重塑情况进行了表征，并研究了移植后染色质行为与胚胎发育之间的关系。评估了来自卵裂期、桑葚胚期和囊胚期（内细胞团ICM和滋养外胚层TE）供体的供体细胞核的发育潜力和重塑模式。此外，我们确定了卵裂球融合与卵母细胞激活之间同步性的改变是否会改变核重塑情况并影响重构胚胎的发育。8细胞期和32细胞期供体细胞核发育至囊胚的比例相似（分别为42%和33%，p大于0.1）。然而，ICM移植后的发育比例降低（17%，p小于0.05），TE移植的发育未超过8细胞期。当卵裂球融合到未激活的卵母细胞细胞质中时，核重塑的特征是过早染色体凝聚（PCC），随后是原核（PN）形成和肿胀。PCC在融合后1.2 - 1.5小时内与所有阶段的供体细胞核同步发生（p大于0.1）。8细胞期和32细胞期移植中的PN形成在融合后约4小时发生，并且与激活卵母细胞中的雌原核同步；然而，在ICM和TE移植中延迟（p小于0.01）。对于所有阶段的供体细胞核，最终核直径与雌原核相似或更大。与未激活的细胞质融合相比，与激活的卵母细胞细胞质融合可防止PCC和广泛的核肿胀（分别为16.0±0.7微米和30±0.7微米，p小于0.01）。早期胚胎中的核直径较小（p小于0.01），发育至囊胚期的比例降低（p小于0.05）。结果表明，供体细胞核的重塑对于发育至囊胚不是必需的；然而，它是有益的。此外，似乎只有在供体细胞核重塑后，即未激活细胞质中的PCC，随后卵母细胞激活时的核肿胀，才可能进行完全重编程。
