Nuclear transfer from somatic cells still has limited efficiency in terms of live calves born due to high fetal loss after transfer. In this study, we addressed the type of donor cells used for cloning in in vivo development. We used a combination of repeated ultrasonography and maternal pregnancy serum protein (PSP60) assays to monitor the evolution of pregnancy after somatic cloning in order to detect the occurrence of late-gestation losses and their frequency, compared with embryo cloning or in vitro fertilization (IVF). Incidence of loss between Day 90 of gestation and calving was 43.7% for adult somatic clones and 33.3% for fetal somatic clones, compared with 4.3% after embryo cloning and 0% in the control IVF group. Using PSP60 levels in maternal blood as a criterion for placental function, we observed that after somatic cloning, recipients that lost their pregnancy before Day 100 showed significantly higher PSP60 levels by Day 50 than those that maintained pregnancy (7.77 +/- 3.3 ng/ml vs. 2.45 +/- 0.27 ng/ml for normal pregnancies, P < 0.05). At later stages of gestation, between 4 mo and calving, mean PSP60 concentrations were significantly increased in pathologic pregnancy after somatic cloning compared with other groups (P < 0.05 by Day 150, P < 0.001 by Day 180, and P < 0.01 by Day 210). In those situations, and confirmed by ultrasonographic measurements, recipients developed severe hydroallantois together with larger placentome size. Our findings suggest that assessing placental development with PSP60 and ultrasonography will lead to better care of recipient animals in bovine somatic cloning.

Implantation is crucial for placental development that will subsequently impact fetal growth and pregnancy success with consequences on postnatal health. We postulated that the pattern of genes expressed by the endometrium when the embryo becomes attached to the mother uterus could account for the final outcome of a pregnancy. As a model, we used the bovine species where the embryo becomes progressively and permanently attached to the endometrium from day 20 of gestation onwards. At that stage, we compared the endometrial genes profiles in the presence of an in vivo fertilized embryo (AI) with the endometrial patterns obtained in the presence of nuclear transfer (SCNT) or in vitro fertilized embryos (IVF), both displaying lower and different potentials for term development. Our data provide evidence that the endometrium can be considered as a biological sensor able to fine-tune its physiology in response to the presence of embryos whose development will become altered much later after the implantation process. Compared with AI, numerous biological functions and several canonical pathways with a major impact on metabolism and immune function were found to be significantly altered in the endometrium of SCNT pregnancies at implantation, whereas the differences were less pronounced with IVF embryos. Determining the limits of the endometrial plasticity at the onset of implantation should bring new insights on the contribution of the maternal environment to the development of an embryo and the success of pregnancy.bovine ͉ implantation ͉ microarray ͉ nuclear transfer

The developmental competence of oocytes from 3-month-old calves was studied through in vitro maturation, fertilization and culture up to the blastocyst stage and by embryo transfer into a foster mother. Oocytes were recovered from antral follicles of calves after or without ovarian stimulation with exogenous FSH and their developmental potential was compared with that of oocytes recovered from cow ovaries. Fertilization and cleavage rates from calf oocytes did not differ significantly from those of cow oocytes. However, after 7 days of culture, the blastocyst formation rate was significantly lower for calves (9% and 11% for nontreated and treated animals, respectively) than for cows (over 20%). Transfer of blastocysts obtained from calf oocytes resulted in a lower pregnancy rate (1 of 23 recipients; 4%) than that achieved with cow oocytes (10 of 26; 38%). The recipient cow that was pregnant from calf embryos delivered a full-term live calf. These data show that some key regulative event that determines the ability to form blastocysts in cattle has not been fully achieved in oocytes from 3-month-old calves.

Somatic nuclear transfer (NT) in cattle is often complicated by fetal oversize (i.e., large offspring syndrome), hydrallantois, and placentomegaly in late gestation. The aims of this work were to obtain data on the placentome structure in NT-recipient cows with hydrallantois (NTH) and to relate these with fetal and placental weights to better understand the abnormalities observed in NTH pregnancies during the third trimester. Pregnant cows were slaughtered between Gestation Days 180 and 280. The fetuses were weighed, and the placentomes were numbered and weighed. Placentomes were examined by histologic and stereological techniques. Macroscopic data showed that placental overgrowth preceded fetal overgrowth, and the ratio of the fetal to the total placentome weight in the NTH group was lower than that in controls after Gestation Day 220. This suggests that placental overgrowth is due to placental default rather than due to fetal overgrowth, as shown also by stereological analysis showing primary deregulation of the growth of cotyledonary tissues. Observed alterations, such as thinning of the maternal epithelium within placentomes and increased trophoblastic surface, could be secondary adaptations. Thus, placental growth deregulations would be due to modifications of the expression of placental factors. Various examples of placental deficiency were observed, suggesting that some fetal abnormalities observed in NTH calves, such as enlarged heart, enlarged umbilical cord, and abdominal ascites, are consequences of placental dysfunction. Therefore, the condition described by the term "large offspring syndrome" might better be described by "large placenta syndrome," because this syndrome affects an average of 50% of late-gestation NT pregnancies. No conclusion can be drawn from this work on apparently normal pregnancies.