Effect of Nutrition of Oocyte Donor on the Outcomes of Somatic Cell Nuclear Transfer in the Sheep1

Abstract

The purpose of this study was to determine if the nutrition of the oocyte donor ewe influenced the success of somatic cell cloning. Merino ewes were fed at either a high- or a low-nutrition level for 3-5 mo before superovulation treatments. Freshly ovulated oocytes were enucleated and fused with serum-starved adult granulosa cells, and resulting reconstructed embryos were cultured for 6 days in modified synthetic oviduct fluid. Embryo cleavage and development to blastocysts were recorded, and good-quality embryos were transferred to synchronized recipient ewes either fresh or, on a few occasions, after vitrification. Pregnancies were monitored by ultrasonography from Day 40 of pregnancy, and offspring were delivered by either cesarean section or vaginal delivery. No differences occurred in the numbers of follicles aspirated, of oocytes recovered, or of oocytes utilizable for cloning between the high and low groups. Neither were there treatment differences in development to the blastocyst stage. However, transfer of embryos from the high group led to significantly more pregnancies and implanted fetuses. Also, more of the established pregnancies from the high group were carried to term, although this difference was not statistically significant. Lamb mortality was high, with half the live-born perishing soon after birth and more succumbing to various infections within days or weeks of birth, but no clear association between the offspring fate and the treatment group could be established. These results suggest that more research into the effect of nutrition on oocyte quality and its subsequent effect on cloning is warranted.

Use of cross-species in-situ hybridization (ZOO-FISH) to assess chromosome abnormalities in day-6 in-vivo- or in-vitro-produced sheep embryos

“…Adult ewes (n = 9) were superovulated as described elsewhere (Peura et al 2003). Briefly, on day 1 each ewe received an intravaginal sponge containing 45mg medroxyprogesterone acetate (MPA, Veramix; Upjohn).…”

Expression of Innate Immune Response Genes in Liver and Three Types of Adipose Tissue in Cloned Pigs

“…The increased mortality has been attributed to a range of causes, including metabolic and cardiopulmonary abnormalities (Hill et al, 1999;Wells et al, 1999), lymphoid hypoplasia (Renard et al, 1999), and neonatal respiratory distress (Hill et al, 1999). Interestingly, increased susceptibility of such cloned animals to bacterial infections has also been described repeatedly (Carter et al, 2002;Keefer et al, 2001;Peura et al, 2003). Surviving cloned pigs are generally found to be healthy and production and reproduction properties in adult cloned pigs are generally normal; however, there are several reports of cloned pigs having deviant phenotypes.…”

The differential requirement of albumin and sodium citrate on the development of in vitro produced bovine embryos

“…Abe et al [5] reported a significant accumulation of intracellular granules and lipid vesicles in blastocysts when serum was included in the culture medium. These data are indicative of an adverse effect of serum on the morphology of embryos and may be related to subsequent abnormal fetal development, such as the large offerspring syndrome [2,3,50,51], and fetal loss following conception [21,52]. Replacing fetal calf serum with BSA as a protein source in a variety of culture media, along with using somatic-cell free culture, has provided an effective alternative for the successful development of IVF and nuclear transfer embryos [10,52,53].…”

“…These data are indicative of an adverse effect of serum on the morphology of embryos and may be related to subsequent abnormal fetal development, such as the large offerspring syndrome [2,3,50,51], and fetal loss following conception [21,52]. Replacing fetal calf serum with BSA as a protein source in a variety of culture media, along with using somatic-cell free culture, has provided an effective alternative for the successful development of IVF and nuclear transfer embryos [10,52,53]. The nuclear and cell counting revealed that embryos derived from serumfree CR1aa culture either with supplement of continuous Sigma-BSA or ICPbio-BSA/ Sigma-BSA regime possessed a similar cell number to those embryos produced from serum additive culture or somatic co-culture.…”