Two isoforms of the leptin receptor are enhanced in pregnancy-specific tissues and soluble leptin receptor is enhanced in maternal serum with advancing gestation in the baboon

Leptin is a polypeptide hormone produced by adipose and other endocrine tissues. Although it has been linked to receptor-mediated pathways that directly influence human conceptus development, mechanisms that regulate the leptin receptor in pregnancy-specific tissues remain unclear. Therefore, we assessed leptin-receptor ontogeny and regulation in the baboon (Papio sp.), a primate model for human pregnancy. Placentae, decidua, and amniochorion were collected from baboons in early (Days 54-63, n = 4), mid (Days 98-103, n = 4), and late (Days 159-165, n = 4) gestation. Regulation by estrogen was assessed by elimination of androgen precursors via removal of the fetus (fetectomy) at midgestation and collection of tissues in late gestation (n = 4; term, approximately 184 days). Maternal serum was sampled with advancing gestation, and the abundance of soluble leptin receptor (solLepR), a potential mediator of gestational hyperleptinemia, was determined. Two placental leptin-receptor isoforms (130 and 150 kDa) increased (P < 0.04 and P < 0.02, respectively) in abundance with advancing gestation. Similarly, the 130-kDa isoform increased approximately fourfold (P < 0.0025) in decidua and approximately 10-fold (P < 0.015) in amniochorion between early and late gestation. Following fetectomy, maternal serum estradiol levels declined approximately 85% (P < 0.03), and the 150-kDa placental leptin-receptor isoform was reduced by more than half (P < 0.002). Maternal serum solLepR concentrations were correlated with gestational age (r = 0.52, P < 0.01) and were unaffected by fetectomy. The presence of leptin-receptor isoforms in pregnancy-specific tissues further denoted leptin's potential to directly influence conceptus development, whereas the 130-kDa solLepR identified in maternal serum suggested a means to facilitate the hyperleptinemia typical of primate pregnancy. Although estrogen did not appear to be the principal regulator of solLepR, it and other factors linked to advancing gestation may be implicated in the regulation of leptin-receptor synthesis.     

细胞色素P450scc在人早期胎盘中的表达（简报）

人妊娠期间,胎盘合成大量的类固醇激素,与妊娠的启动、维持、分娩以及胎儿的发育均存在密切的关系。阐明胎盘类固醇激素特别是孕酮合成与分泌的调节机制对于寻找理想的生育调控技术和生殖保健方法具有重要的意义。因此,胎盘类固醇激素合成与分泌的调节向来是生殖生物学与妇产科学领域所关注的焦点问题之一,     

Characterization of an insulin-like growth factor binding protein, analogous to human pregnancy-associated secreted endometrial alpha 1-globulin, in decidua of the baboon (Papio anubis) placenta

The major secreted protein of the human decidua (pregnancy-associated endometrial alpha 1-globulin [alpha 1-PEG]), is an insulin-like growth factor-binding protein (IGF-BP) that is immunologically and biochemically similar to placental protein 12 (PP12) extracted from term human placenta. Since previous studies have demonstrated that the baboon and human endometrium synthesize and release a number of biochemically and immunologically related polypeptides in culture, this study was undertaken to further characterize a related IGF-BP in baboon placental tissues. Decidua, chorio-amniotic membranes with adhering decidua (CAM-D), and placental villi were obtained from pregnant baboons between Days 134 and 160 of gestation by Cesarean sections. Portions of tissue were either cultured in the presence of 35S-methionine, fixed for immunocytochemistry, or frozen in liquid nitrogen for cytosol extraction. Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) of tissue culture media (TCM) revealed that the major secretory product of the decidua and CAM-D was an acidic polypeptide (Mr 33,000). Western blot analysis and immunoprecipitation of TCM with murine monoclonal antibody (B2H10) against human alpha 1-PEG demonstrated that this molecule, secreted by the baboon decidua and CAM-D, but not the placental villi, was immunologically identical to the human IGF-BP. Immunocytochemical localization of IGF-BP was intense in the cytoplasm of stromal cells in decidua and CAM-D and absent in the placenta. Gel filtration of TCM and cytosol followed by screening of eluates for 125I-IGF-I binding resolved two peaks (Mr greater than 100,000 and 35,000) of specific IGF-BP in decidua and CAM-D. The 35,000 peak had 100-200 times the binding capacity of the Mr greater than 100,000 peak and a Kd of 1.14-1.83 nM. The eluates contained in the Mr 35,000 peak were also immunoreactive to alpha 1-PEG, as accessed by a polyclonal radioimmunoassay. Affinity cross-linking with 125I-IGF-I followed by sodium dodecyl sulfate-PAGE revealed an immunoreactive complex of Mr 36,000, confirming that the baboon protein represents a high affinity IGF-BP. These studies indicate that the hypertrophied stromal cells of the baboon decidua and CAM-D synthesize and release an IGF-BP as their major secretory product, analogous to the situation in humans. The results of this study suggest that this protein may play a role in the regulation of IGF action during pregnancy.     

Regulation of functional differentiation of the placental villous syncytiotrophoblast by estrogen during primate pregnancy.

By using the baboon as an in vivo model for the study of the endocrinology of human pregnancy, studies in the authors' laboratories have shown that the primate placenta is an estrogen target tissue and that estrogen, via interaction with the estrogen receptor, regulates functional differentiation of the syncytiotrophoblast, which is manifest as an upregulation of key components of the progesterone biosynthetic pathway and the metabolism of corticosteroids critical to placental-fetal development. Thus, estrogen exerts specific stimulatory effects on the receptor-mediated uptake of low density lipoprotein by, and expression of, the P-450 cholesterol side-chain cleavage enzyme within the syncytiotrophoblast, thereby promoting the production of progesterone. Concomitantly, there is an estrogen-dependent developmental regulation of the 11beta-hydroxysteroid dehydrogenase enzyme system in the syncytiotrophoblast, which enhances transplacental oxidation of maternal cortisol to cortisone and leads to maturation of the fetal hypothalamic pituitary adrenocortical axis late in gestation. Consequently, estrogen has a central, integrative role in modulating the dialogue and signaling system operating between the placenta and fetus that results in the maintenance of pregnancy and the development of adrenocortical self-sufficiency that are essential for maturation of the fetus and neonatal survival after birth.     

Source and regulation of 17 alpha-hydroxyprogesterone during baboon pregnancy

The present study determined the source and regulation of 17 alpha-hydroxyprogesterone (17-OHP4) during mid-late baboon pregnancy. Serum 17-OHP4 (ng/ml) in 5 untreated baboons increased from low values at mid-late gestation to a mean (+/- SEM) of 0.49 +/- 0.02 during the final 20 days of gestation. Fetectomy of 5 baboons resulted in serum 17-OHP4 concentrations which declined to and remained at baseline. Serum 17-OHP4 concentrations were 5- to 10-fold greater (P less than 0.001) in the uterine, utero-ovarian, and umbilical veins than peripherally. Apparently the fetal adrenal provides precursors for placental 17-OHP4 formation because the fetal adrenal gland develops delta 5-3 beta-hydroxysteroid dehydrogenase only late in gestation, and because the fetal adrenal and not the placenta has the capacity for 17-hydroxylation. Thus, at mid-late gestation the placenta appears to supply a major, and at term the corpus luteum a minor portion of the total 17-OHP4. Administration of the estrogen antagonist ethamoxytriphetol (MER-25, 15 mg/kg BW) to 4 baboons did not affect 17-OHP4 during mid-late gestation, when the placenta was the only source of 17-OHP4. However, MER-25 resulted in serum 17-OHP4 concentrations (ng/ml) at term which were greater (1.08 +/- 0.10, P less than 0.001) than in untreated baboons (0.49 +/- 0.02). Prior removal of the corpus luteum of pregnancy in 4 animals subsequently given MER-25 prevented this rise in 17-OHP4. This suggests that the marked elevation in 17-OHP4 observed near term after MER-25 administration was of luteal origin and that antiestrogen enhanced 17-OHP4 secretion by the corpus luteum.     

Regulation of placental villous angiopoietin-1 and -2 expression by estrogen during baboon pregnancy

We recently showed an increase in vascular endothelial growth factor (VEGF), decrease in angiopoietin-1 (Ang-1) and unaltered Ang-2 expression by the villous placenta with advancing baboon pregnancy. Moreover, placental VEGF expression was increased by estrogen in early pregnancy. In the present study, we determined whether placental Ang-1 and Ang-2 are regulated by estrogen. Ang-1 and Ang-2 mRNA and protein were determined by RT-PCR and immunocytochemistry in the placenta of baboons on Day 60 of gestation (term is 184 days) after administration of estrogen precursor androstenedione on Days 25-59 or on Day 54 after acute estradiol administration. Chronic androstenedione treatment increased serum estradiol levels three-fold (P < 0.001) and decreased (P < 0.05) villous cytotrophoblast Ang-1 mRNA to a level (0.36 +/- 0.08 relative to 18S rRNA) that was one-third of that in untreated animals (0.98 +/- 0.26). Within 2 hr of estradiol administration, cytotrophoblast Ang-1 mRNA was decreased to a level (0.24 +/- 0.05) one-fifth (P < 0.05) of that in untreated animals (1.14 +/- 0.23). However, Ang-2 mRNA levels were unaltered. Ang-1, Ang-2 and estrogen receptors alpha and beta protein were localized within villous cytotrophoblasts providing a mechanism for estrogen action at this site. In summary, estrogen increased VEGF, decreased Ang-1, and had no effect on Ang-2 expression within placental cytotrophoblasts during early baboon pregnancy. We propose that the estrogen-dependent differential regulation of these angioregulatory factors underpins the unique pattern of neovascularization established within the villous placenta during primate pregnancy.     

Leptin in pregnancy

Leptin is a polypeptide hormone that aids in the regulation of body weight and energy homeostasis and is linked to a variety of reproductive processes in both animals and humans. Thus, leptin may help regulate ovarian development and steroidogenesis and serve as either a primary signal initiating puberty or as a permissive regulator of sexual maturation. Perhaps significantly, peripheral leptin concentrations, adjusted for adiposity, are dramatically higher in females than in males throughout life. During primate pregnancy, maternal levels that arise from adipose stores and perhaps the placenta increase with advancing gestational age. Proposed physiological roles for leptin in pregnancy include the regulation of conceptus growth and development, fetal/placental angiogenesis, embryonic hematopoiesis, and hormone biosynthesis within the maternal-fetoplacental unit. The specific localization of both leptin and its receptor in the syncytiotrophoblast implies autocrine and/or paracrine relationships in this endocrinologically active tissue. Interactions of leptin with mechanisms regulating pre-eclampsia and maternal diabetes have also been suggested. Collectively, therefore, reports suggest that a better understanding of the regulation of leptin and its role(s) throughout gestation may eventually impact those causes of human perinatal morbidity and mortality that are exacerbated by intrauterine growth retardation, macrosomia, placental insufficiency, or prematurity.     

Synthesis of 'decidualization-associated protein' in tissues of the rat uterus and placenta during pregnancy.

The synthesis of a soluble protein (referred to as 'decidualization-associated protein', DAP), has been examined in uterine and placental tissues of rats during pregnancy by means of polyacrylamide gel electrophoretic analysis of [3H]leucine-labelled soluble proteins. No synthesis of the protein was detected in non-implantation regions of the uterus. In implantation site tissue, no synthesis was detected on Days 6 or 7 of pregnancy. Only slight synthesis was present in the endometrium on Day 8, but synthesis rose rapidly from Days 9 to 12 in both the endometrium and myometrium although differences in the rates of increase were observed. Synthesis fell from Day 12 to 14 in both tissues. Synthesis by the myometrium was entirely localized in the mesometrial region, which contains the metrial gland. After Day 12, when the endometrium is represented by the chorioallantoic placenta, synthesis was examined in the labyrinthine and the decidua basalis/basal zone placenta tissues. No synthesis of 'DAP' was detected in the labyrinthine placenta from Day 16 of pregnancy. Synthesis observed in the decidua basalis/basal zone placenta fell dramatically from Day 14 to 20. The pattern of synthesis of 'DAP' during pregnancy suggests a role in the establishment of the chorioallantoic placenta and metrial gland in the rat.     

Differential localization of heat shock proteins 90, 70, 60 and 27 in human decidua and placenta during pregnancy

Little is known about the localization of heat shock proteins (HSPs) in the decidua and placenta during the course of normal pregnancy. In this study, we have examined the localization of the HSPs in decidual and placental tissues obtained from women during the first, second and third trimesters of pregnancy (five in each trimester) by immunohistochemistry using highly specific antisera. HSPs 90, 70, 60 and 27 were detected in decidual stromal cells during each trimester. The intensity of staining did not change during gestation for HSPs 60 and 27, whereas it decreased with advancing gestation for HSPs 90 and 70. HSPs 90 and 60 were localized primarily in the nucleus, whereas HSP 70 was present equally in the nucleus and the cytoplasm; HSP 27 was primarily in the cytoplasm. In the placenta, HSPs 90, 70 and 60 were localized in cytotrophoblast, syncytiotrophoblast, intermediate trophoblast, Hofbauer and endothelial cells. HSPs 90 and 60 were localized primarily in the nucleus, while HSP 70 was in the nucleus and the cytoplasm. In the placenta, HSP 27 was detected only in the intermediate trophoblast and syncytiotrophoblast cells and only in the first two trimesters. These results indicate that there are striking differences in the subcellular localization of HSPs in the decidua and the placenta during normal pregnancy.     

Serum leptin in nonpregnant and pregnant women and in old and new world nonhuman primates

Leptin is a hormone that is produced during mammalian pregnancy in the placental trophoblast and other tissues, including! fetal and maternal adipocytes. Synthesis of the polypeptide and the presence of its specific receptors throughout the human maternal fetoplacental unit suggest direct effects on conceptus growth and development. However, both the physiologic roles of leptin and the mechanisms regulating leptin synthesis in human pregnancy differ from those in laboratory and domestic species, necessitating the development of non-human primate research models. Therefore, we compared serum leptin concentrations in nonpregnant and pregnant women with those in both old world nonhuman primates (i.e., baboon, rhesus monkey, cynomolgus monkey) and new world nonhuman primates (i.e., squirrel monkey, titi monkey). As expected, maternal leptin levels were elevated in human and baboon pregnancies (P < 0.05 and P < 0.001, respectively). Levels in both species of old world monkeys were also greatly enhanced (P < 0.001). Although maternal serum concentrations were slightly elevated compared to nonpregnant levels in both species of new world monkeys, overall concentrations were dramatically lower than for either old world primates or humans. Results provide comparisons of serum leptin concentrations in pregnant and nonpregnant humans and baboons with those in both old and new world monkeys and further characterize these nonhuman primates as models for the investigation of leptin dynamics in pregnancy.     

Developmental maturation of primate placental trophoblast: placental cytosolic and secretory phospholipase A2 expression after estrogen suppression of baboons

We have demonstrated that the baboon placenta expressed the mRNAs and proteins for secretory and cytosolic phospholipase A2 (PLA2) enzymes and that cPLA2 expression increased with advancing gestation in association with the increase in placental estrogen production. To determine whether estrogen regulates placental PLA2 expression, as it does other aspects of syncytiotrophoblast functional differentiation, we compared sPLA2 and cPLA2 mRNA levels in placentas obtained on day 165 of gestation (term = day 184) from baboons that were untreated or treated during the second half of gestation with the aromatase inhibitor CGS 20267 or CGS 20267 and estradiol. Maternal saphenous and uterine vein estradiol levels were reduced (P < 0.05) by approximately 95% by treatment with CGS 20267 and restored by concomitant administration of CGS 20267 and estrogen. However, sPLA2 and cPLA2 mRNA levels expressed as a ratio of beta-actin were similar in whole villous placenta from baboons that were untreated or treated with CGS 20267 or CGS 20267 plus estrogen. PLA2 expression in an enriched fraction of nontrophoblast cells of the baboon placenta was also not altered by CGS 20267 treatment. Collectively these findings indicate that placental cPLA2 and sPLA2 expression is not estrogen-dependent. Because estrogen has been shown to regulate other aspects of placental steroidogenesis, we suggest that the regulatory role of estrogen on syncytiotrophoblast functional maturation is specific.     

Passive immunization against the MHC class I molecule Mamu-AG disrupts rhesus placental development and endometrial responses

The unique MHC phenotype of the human and nonhuman primate placenta has suggested a potential role in maternal-fetal immune tolerance, pregnancy success, and maternal as well as fetal well-being. In the rhesus monkey (Macaca mulatta) a nonclassical MHC class I molecule, Mamu-AG, is a putative homologue of HLA-G and is hypothesized to play a role in maternal-fetal immune interactions during pregnancy. Rhesus monkeys were passively immunized during the second week after implantation with a mAb against Mamu-AG. Passive immunization altered the growth and vascularization of the fetal placenta, the placental modification of maternal endometrial vessels, the maternal leukocyte response to implantation, and the differentiation of epithelial and stromal cells in the endometrium. These data are the first to demonstrate in vivo the importance of MHC class I molecules expressed on primate trophoblasts in establishing an important environment for pregnancy success through coordinated interactions between endometrial and fetal tissues.     

The size of the corpus luteum during pseudopregnancy and pregnancy in the rat.

The corpus luteum in mature Sprague Dawley rats was weighted at the various stages of pseudopregnancy and pregancy. The average size of these corpora lutea was 1.0 +/- 0.10 mg, 1.61 +/- 0.69 mg, 1.90 +/- 0.25 mg, 3.69 +/- 0.36 mg, and 4.37 +/- 0.50 mg on day 2 of diestrus, on days 10-15 of psuedopregnancy, on days 9-10, 14, and 20 of pregnancy, respectively. The fact that the average size of the corpus luteum on days 10-15 of pseudopregnancy was larger than that on day 2 of diestrus is thought to drive from prolonged exposure of the corpus luteum to prolactin. The average size of the corpus luteum on days 9-10 of pregnancy had a tendency to be larger than that on days 10-15 of pseudopregnancy and this seems to demonstrate that the placenta secreted placental lactogen by this stage of pregnancy. The average size of the corpus luteum on day 14 of pregnancy was larger than that on days 9-10 of pregnancy. This phenomenon might be attributed to the presence of large amounts of placental lactogen secreted from the placenta between days 10 and 14 of pregnancy. Furthermore, it was noted that the size of the corpus luteum on day 20 of pregnancy was larger than that of day 14, which suggests that further secretion of placental lactogen continued after day 14 of pregnancy. As there was a remarkable decrease in the number of fetuses on day 20 of pregnancy when overiectomy was performed on day 14 of pregnancy, the ovary was considered indispensable in maintaining pregnancy in the rat.     

细胞色素P450scc在人早期胎盘中的表达(简报)

人妊娠期间,胎盘合成大量的类固醇激素,与妊娠的启动、维持、分娩以及胎儿的发育均存在密切的关系。阐明胎盘类固醇激素特别是孕酮合成与分泌的调节机制对于寻找理想的生育调控技术和生殖保健方法具有重要的意义。因此,胎盘类固醇激素合成与分泌的调节向来是生殖生物学与妇产科学领域所关注的焦点问题之一,并为此开展了大量的研究,但迄今仍不清楚,其主要原因之一是