Expression and possible role of 20alpha-hydroxysteroid dehydrogenase in the placenta of the goat.

20Alpha-hydroxysteroid dehydrogenase (20alpha-HSD) catalyzes the conversion of progesterone to its inactive form 20alpha-dihydroprogesterone (20alpha-OHP). 20Alpha-HSD is expressed in the murine placenta, suggesting a role, yet unidentified, played by this enzyme during the course of pregnancy. To elucidate the possible roles of 20alpha-HSD during pregnancy, 20alpha-HSD gene expression in the placenta was examined by Northern blot analysis, and progestin (progesterone and 20alpha-OHP) concentrations in the maternal and fetal sera and the amniotic fluid were measured by radioimmunoassay in pregnant Shiba goats. The expression of 20alpha-HSD mRNA was observed in the placenta and the intercaruncular part of the uterus during mid to late pregnancy. Analysis by in situ hybridization revealed that 20alpha-HSD mRNA was mainly localized in the endometrial epithelium of the caruncle side of the placenta. Considerable enzyme activity of 20alpha-HSD was also detected in the cytosolic fraction of the placenta and intercaruncular part of the uterus. Although concentrations of progesterone and 20alpha-OHP in the maternal serum showed similar profiles, progesterone levels in the fetal serum stayed extremely low throughout the pregnancy. The 20alpha-OHP concentration in the fetal serum was always higher than that in the maternal serum. In the amniotic fluid, the concentrations of both progesterone and 20alpha-OHP remained at very low levels throughout the pregnancy. These results support the notion that 20alpha-HSD protects the fetus from the cytotoxic effects of progesterone, and thereby maintains the normal development of the fetus.

Molecular cloning of goat 20alpha-hydroxysteroid dehydrogenase cDNA.

20Alpha-hydroxysteroid dehydrogenase (20alpha-HSD), which catalyzes the conversion of progesterone to its inactive form 20alpha-dihydroprogesterone, is expressed in murine placenta and has been suggested to play roles in maintaining pregnancy. To understand the role of 20alpha-HSD during pregnancy in the goat, as a first step, cloning and sequencing of 20alpha-HSD cDNA were performed. The full nucleotide sequence of 20alpha-HSD cDNA was determined on samples obtained from the corpus luteum at the luteal phase of the estrous cycle and the placenta in late pregnancy by RT-PCR and 3' and 5' RACE systems. Cloned 20alpha-HSD cDNA consisted of 1124 bp and belonged to the aldo-keto reductase superfamily. From the start codon to stop codon there were 323 amino acids, the same as in other species. To verify whether the protein derived from goat 20alpha-HSD cDNA had 20alpha-HSD activity, the cDNA was expressed by bacteria. Bacterially expressed goat 20alpha-HSD protein showed 20alpha-HSD enzyme activity. A tissue distribution study demonstrated that 20alpha-HSD was expressed in the placenta, but not in the adrenal gland, liver and spleen during pregnancy. The present study suggests that goat 20alpha-HSD is another member of the aldo-keto reductase superfamily and that it plays a role in the placenta during pregnancy.