Functional full-term placentas formed from parthenogenetic embryos using serial nuclear transfer

Abstract

SUMMARYMammalian parthenogenetic embryos invariably die in mid-gestation from imprinted gene defects and placental hypoplasia. Based on chimera experiments, trophoblastic proliferation is supposed to be inhibited in the absence of a male genome. Here, we show that parthenogenetic mouse embryonic cell nuclei can be reprogrammed by serial rounds of nuclear transfer without using any genetic modification. The durations of survival in uteri of cloned foetuses derived from green fluorescent protein (GFP)-labelled parthenogenetic cell nuclei were extended with repeated nuclear transfers. After five repeats, live cloned foetuses were obtained up to day 14.5 of gestation; however, they did not survive longer even when we repeated nuclear transfer up to nine times. All foetuses showed intestinal herniation and possessed well-expanded large placentas. When embryonic stem (ES) cells derived from fertilised embryos were aggregated with the cloned embryos, full-term offspring with large placentas were obtained from the chimeric embryos. Those placentas were derived from parthenogenetic cell nuclei, judging from GFP expression. The patterns of imprinted gene expression and methylation status were similar to their parthenogenetic origin, except for Peg10, which showed the same level as in the normal placenta. These results suggest that there is a limitation for foetal development in the ability to reprogramme imprinted genes by repeated rounds of nuclear transfer. However, the placentas of parthenogenetic embryos can escape epigenetic regulation when developed using nuclear transfer techniques and can support foetal development to full gestation.

Downregulation ofH19Improves the Differentiation Potential of Mouse Parthenogenetic Embryonic Stem Cells

“…In vitro culture of B-ESCs and P-ESCs can lead to alterations in DNA methylation and covalent modifications of histones such as histone acetylation and methylation [19,33,[45][46][47][48]. Moreover, in P-ESCs the expression of several imprinted genes is altered due to absence of a paternal chromosomal complement [19,46,47].…”

“…The efficiency of full-term development significantly increases when the H19/Igf2 DMR together with the DMR found within a second imprinted gene cluster Dlk1/Dio3 on chromosome 12 are deleted [23,32]. Most recently Hikichi et al showed that the development of PG embryos can be extended following serial nuclear transfer (NT) and is associated with a decrease in H19 expression [33]. Altogether, these results suggest that the correct expression of H19 is important for normal embryonic development.…”

Ascorbic acid improves pluripotency of human parthenogenetic embryonic stem cells through modifying imprinted gene expression in the Dlk1-Dio3 region

“…Hikichi and co-workers established a new type of pESCs by transferring traditional ESC nuclei into oocytes and deriving these pESCs again at the blastocyst stage (NT-pES), leading to two to five times enhancement of differentiation ability, compared with the original pESCs [ 29 ]. The same group reported development of parthenogenetic embryos to term using the serial nuclear transfer method [ 30 ]. Epigenetic modification was evidently altered during reprogramming mediated by nuclear transfer, and aberrant methylation observed in cloned embryos and animals.…”

Derivation of androgenetic embryonic stem cells from m-carboxycinnamic acid bishydroxamide (CBHA) treated androgenetic embryos

“…Most of the chimeras from aES cells had skeletal abnormality and other defects [10,23], and no chimeras were generated from two mature sperms derived aES cells assisted with SCNT previously. Some labs tried to erase the strict imprinting of uniparental ES cells by serial nuclear transfer method [24] or long-term cell culture [25]. But these methods were either complicated or time-consuming.…”