Excess Estrogen Sulfoconjugation Causes a Weak Sign of Delivery in Somatic Cell Clone Recipient Cows.

Abstract

This study was conducted to elucidate a factor causing a weak sign of delivery and a prolonged gestation, which are frequently observed in cows carrying somatic clone fetus, in spite of the maturity of fetal size or the occurrence of large offspring syndrome. Till date, several investigators have suggested morphological and functional abnormalities of the placenta in clone fetus in the various gestational stages. However, the roles for the placenta in the final process of clone pregnancy, namely placental estrogen synthesis and regulation of estrogen activity with sulfoconjugation, have not been fully understood. In IETS (2007 in Kyoto, Japan), we reported that prepartum rise of estrone and estradiol-17b concentrations in clone pregnancy was subtle, but estrone-sulfate concentration in clone pregnancy was higher than that in cows carrying fetus derived from in vivo embryo. In this study, we analyzed mRNA expression levels of estrogen-related genes, estrogen sulfotransferase (SULT1E1), estrogen sulfatase (STS) and aromatase (CYP19), in placental tissues, cotyledon (COT) and caruncle (CAR). Placentomes were obtained immediately after purtrition from cows, which delivered clone and in vivo calves after induction of parturition (gestation length was 284 to 289 days). Correlation analyses between mRNA abundances in placental tissues and maternal estrogen concentrations at parturition were performed using the data obtained from clones. Localization of gene expression in placentome was examined with in situ hybridization (SULT1E1 and STS) and immunohistochemistry (CYP19). Number of binucleate cells (BNC), existing in placentome, was counted from sections stained with hematoxylin-eosin. mRNA abundance of SULT1E1 was tended to be high in COT compared with CAR, clone pregnancy showed significantly higher (p<0.05) expression level than in vivo pregnancy. In situ hybridization signal of SULT1E1 was detected in binucleate cells. SULT1E1 mRNA abundance was negatively and positively correlated with maternal estrone (p<0.05) and estrone-sulfate (p<0.05) at parturition, respectively. mRNA abundances of STS and CYP19 were significantly higher in CAR and COT, respectively. In situ hybridization and immunohistochemistry showed that BNC and carunclar epithelial cells expressed both genes. Expression levels of STS and CYP19 mRNA were similar between clone and in vivo pregnancy, and were positively correlated (p<0.05) with maternal estradiol-17b at parturition of clones. Population of BNC in placentome did not differ between clone (vaginally delivered clone; 18.8}1.4/0.1mm2, cesarean section delivered clone; 18.6}1.7/0.1mm2) and in vivo (18.9}1.8/0.1mm2) pregnancy. Therefore, it can be surmised that increased SULT1E1 mRNA abundance in clone placental tissue was due to an increased transcription of the gene, rather than to an increased proportion of SULT1E1 producing BNC. The results obtained here suggested that excess estrogen sulfoconjugation in placenta was reasonable for the confusion of the ratio of unconjugated (active) and sulfoconjugated (inactive) estrogens. The hormonal abnormality would cause the weak sign of delivery and the prolonged gestation in cows carrying clone.