Successful somatic cloned animal production has been reported in various domesticated species, including cattle; however, it is associated with a high rate of pregnancy failure. The low cloning yield could possibly arise from either an abnormal and/or poorly developed placenta. In comparison to control cows, fewer placentomes were found in somatic cell nuclear recipient (NT) cows at day 60 of gestation, suggesting a retardation of fetal/placental growth in these animals. NT cows not only had fewer numbers of chorionic villi but also had poorly developed caruncles. Macroscopic examination revealed atypical development of the placentome in terms of shape and size. Histological disruption of chorionic villi and caruncular septum was found in NT cows. Of particular interest was that the expression of genes, as well as proteins in the placentome, was disparate between NT and artificially inseminated cows, especially placental lactogen (PL) and pregnancy-associated glycoprotein (PAG). In contrast, prolactin-related protein-1 (PRP-1) signals were comparable across cows, including NT cows carrying immotile fetuses. The expression of extracellular matrix degrading molecule, heparanase (HPA), in NT cows was divergent from that of control cows. Microarray data suggest that gene expression was disorientated in early stages of implantation in NT cows, but this was eliminated with progression of gestation. These findings strongly support a delay in trophoblast development during early stages of placentation in NT cows, and suggest that placental specific proteins, including PLs, PAGs, and HPA, are key indicators for the aberration of gestation and placental function in cows.

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Changes of histone H1 kinase activity before and after electrical stimulation were connected with the ability of cytoplasm of pig oocytes to be activated parthenogenetically when matured and aged in vitro. Cumulus-oocyte complexes were collected from prepubertal gilts and cultured in a modified Waymouth's MB752/1 medium. The first mature oocytes were observed after 30 h of culture. After 36 h of culture, about 65% of oocytes had matured (reached metaphase II stage with the first polar body). When oocytes matured after 36 h of culture were stimulated with an electric pulse and subsequently cultured for 10 h, only 7% became parthenogenetically activated (formation of a female pronucleus). When oocytes matured for 60 h and 72 h underwent the same treatment, significantly more became activated parthenogenetically (46% and 57%, respectively). Oocytes matured for 72 h but not stimulated electrically also exhibited high spontaneous parthenogenetic activation (24%). Activation of oocytes resulted either in the formation of a female pronucleus(ei) or in fragmentation of oocytes. Fragmentation in stimulated and nonstimulated oocytes increased significantly after 72 h of culture (37% and 18%, respectively). Histone H1 kinase activity in immature oocytes at the germinal vesicle stage was low (17.2 fmol h-1 per oocyte). However, when oocytes were cultured for 36 and 48 h, histone H1 kinase activity increased significantly (168.2 and 138.5 fmol h-1 per oocyte, respectively). Prolonged culture (60 h and 72 h) resulted in a significant decrease in histone H1 kinase activity (94.3 and 49.3 fmol h-1 per oocyte, respectively). When oocytes cultured for up to 72 h were electrically stimulated, histone H1 kinase activity in activated oocytes (oocytes that formed a female pronucleus and fragmented oocytes) was significantly lower (24.7 mol h-1 per oocyte) than that in nonactivated oocytes (99.9 mol h-1 per oocyte). The present data clearly indicate that the gradual decrease of histone H1 kinase activity is correlated with ageing of oocytes matured in vitro and with their ability to be parthenogenetically activated.

Gene expression analysis comparing nonpregnant with pregnant bovine uteri, including placenta, was performed with a custom cDNA microarray containing 1,933 independent genes. These genes were classified into six categories according to biological function, as follows: cell and tissue structural dynamics (108 genes), intercellular communication (221), intracellular metabolism (265), cell cycle and apoptosis (26), regulation of gene expression (113), expressed sequence tag (EST) and function unknown (617), and uncomplemented genes (583 clones). This array possessed bovine placental/endometrial specificity, as it included many pregnancy-specific molecules, such as pregnancy-associated glycoprotein-1 (PAGs), placental lactogen (PLs), and prolactin-related protein-1 (PRPs). A total of 77 genes were induced and 12 repressed in the placenta/endometrium. Our results point to a fundamental role for bovine placental-specific genes such as PAGs, PLs, and PRPs, in implantation and placentogenesis, and document that cDNA microarray analysis from bovine placenta/endometrium is possible and is a specific tool for monitoring genome-wide gene expression during the establishment and maintenance of pregnancy.

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