(W675) A COMPARIOSN OF IFN- MESSENGER RNA EXPRESSION BETWEEN SOMATIC NUCLEAR TRANSFER AND IN VITRO PRODUCED FEMALE BOVINE BLASTOCYSTS.
Kimura, Koji1, Akagi, Satoshi2, Takahashi, Seiya2, Iwata, Hisataka3, Kadokawa, Hiroya1, Aoki, Mari4, Hirako, Makoto5, 1 National Institute of Livestock and Grassland Science, Reproductive Physiology Lab, Nasushiobara, Tochigi, Japan2 National Institute of Livestock and Grassland Science, Reproductive Cell Biology Lab, Tsukuba, Ibaraki, Japan3 Tokyo University of Agriculture, Atsugi, Kanagawa, Japan4 National Agriculture Research Center for Hokkaido Region, Sapporo, Hokkaido, Japan5 National Agriculture and Bio-oriented Research Organaization, Tsukuba, Ibaraki, Japan
ABSTRACT- Somatic nuclear transfer (SNT) has been developed in several mammalian species. The fetal and neonatal survival rate of SNT embryos however remains low. IFN- is critical in the establishment of pregnancy in cattle and its production is sexually dimorphic, with female embryos producing more IFN- than male embryos. In this study, expression of IFN- mRNA was investigated in SNP embryos and compared to levels in in vitro fertilized (IVF) embryos. Abattoir derived bovine oocytes were matured and either fertilized (insemination on d-0) to produce IVF embryos, or used to produce SNT embryos via the fusion of enucleated oocytes and cumulus (donor) cells prior to activation. All zygotes were cultured until day 8, when expanded IVF and SNT blastocysts were collected and cultured individually for 48 hrs. IVF but not SNT blastocysts, since these were derived from female donor cells, were biopsied for retrospective sex determination. Total RNA was extracted from all blastocysts individually and then reverse transcribed. Quantification of IFN- mRNA abundance was performed by real-time RT-PCR and the expression level of the gene was normalized to the abundance of GAPDH. A total of 96 (male:55, female:41) IVF and 30 SNT embryos were analyzed. As expected, sexual dimorphic IFN- mRNA expression was determined between male (141.5±63.6) and female (336.8±69.6) IVF blastocysts (P<0.05). No significant difference (P>0.05) was evident between the average IFN- mRNA levels of IVF (224.9±47.8) and SNT (173.6±29.5) blastocysts. Notably however, female IVF blastocysts had significantly (p<0.05) greater IFN- mRNA expression than SNT blastocysts derived from female donor cells. These results suggested that SNT blastocysts have reduced IFN- mRNA expression that may, along with other factors, explain the aberrant placental development and poor pregnancy rates attained with SNT blastocysts. Supported by JLTA grant.
KEY WORDS: somatic nuclear transfer embryo, IFN-, sexual dimorphism, bovine