Somatic cell nuclear transfer alters peri-implantation trophoblast differentiation in bovine embryos

Abstract

Abnormal placental development limits success in ruminant pregnancies derived from somatic cell nuclear transfer (SCNT), due to reduction in placentome number and consequently, maternal/fetal exchange. In the primary stages of an epithelial-chorial association, the maternal/fetal interface is characterized by progressive endometrial invasion by specialized trophoblast binucleate/giant cells (TGC). We hypothesized that dysfunctional placentation in SCNT pregnancies results from aberration in expression of genes known to be necessary for trophoblast proliferation (Mash2), differentiation (Hand1), and function (IFN-t and PAG-9). We, therefore, compared the expression of these factors in trophoblast from bovine embryos derived from artificial insemination (AI), in vitro fertilization (IVF), and SCNT prior to (day 17) and following (day 40 of gestation) implantation, as well as TGC densities and function. In preimplantation embryos, Mash2 mRNA was more abundant in SCNT embryos compared to AI, while Hand1 was highest in AI and IVF relative to SCNT embryos. IFN-t mRNA abundance did not differ among groups. PAG-9 mRNA was undetectable in SCNT embryos, present in IVF embryos and highest in AI embryos. In postimplantation pregnancies, SCNT fetal cotyledons displayed higher Mash2 and Hand1 than AI and IVF tissues. Allelic expression of Mash2 was not different among the groups, which suggests that elevated mRNA expression was not due to altered imprinting status of Mash2. The day 40 SCNT cotyledons had the fewest number of TGC compared to IVF and AI controls. Thus, expression of genes critical to normal placental development is altered in SCNT bovine embryos, and this is expected to cause abnormal trophoblast differentiation and contribute to pregnancy loss.

High developmental potential in vitro and in vivo of cattle embryos cloned without micromanipulators

“…1). The overall recovery was 11/18 (61.1%) which compares favourably to data reported by others [27,28] thus confirming the developmental potential of the adult cell line derived from Wagyu cattle used in our experiments.…”

Review Paper: A Review of the Pathology of Abnormal Placentae of Somatic Cell Nuclear Transfer Clone Pregnancies in Cattle, Sheep, and Mice

“…1, 2) were associated with relative immaturity of vessels of the placental villi, as determined by reduced or no immunohistochemical expression of von Willebrand factor and calponin (Figs. [3][4][5][6], 79 a marker of late differentiation in vascular smooth muscle cells. 56 Placental vasculogenesis was significantly reduced as indicated by lack of CD34 expression by individual foetal blood vessels, 79 similar to the abnormal placental vasculogenesis observed with spontaneous abortions in humans.…”

“…19 Thus, although the 2 gene products exert somewhat opposing effects, their simultaneous overexpression leads to failure to stimulate binucleate cell differentiation by trophoblastic cells and reduced binucleate cell numbers. 3 In placentae of nonviable bovine SCNT embryos at 60 days of gestation, cDNA macroarray analysis revealed a twofold increased expression of IGF2, 75 overexpression of which in transgenic mice is embryonic lethal, 98 and less than 50% reduction of hemoglobin alpha 1, hemoglobin alpha 2, and the bovine homologues of spectrin, beta, erythrocytic and spectrin, beta, nonerythrocytic 1. 75 Genetic studies of the underlying mechanisms of placental defects in SCNT sheep have not been as extensive as those in cattle or mice because of the limited knowledge of sheep genetics.…”

A comparative study on expression profile of developmentally important genes during pre-implantation stages in buffalo hand-made cloned embryos derived from adult fibroblasts and amniotic fluid derived stem cells

“…Although, the morphologic appearance of SCNT embryos does not differ from that of in vitro fertilized embryos, the potential to develop to term dramatically differs between SCNT and IVF embryos (Li et al 2005(Li et al , 2006a(Li et al , b, 2008Kato et al 2007). Research using SCNT technology suggested that abnormal expression of genes during the earliest stages of development could be the reason of embryonic losses (Arnold et al 2006). Abnormal gene expression might be accountable for the observed phenotypes in embryos produced by SCNT, not only during early embryonic development, but also during gestation and parturition leading in many cases to the so called abnormal offspring syndrome (AOS; Garry et al 1996;Ranilla et al 1998;Zhang et al 2004).…”