Stimulation of hCG and inhibition of hPL in isolated human trophoblast cells in vitro by glycodelin A.

The immunosuppressive protein glycodelin A (formerly named PP14) is produced by human decidua and secreted in the maternal circulation. Glycodelin A concentrations in serum have been used as indicators of endometrial function. The purpose of this study was to investigate the effect of glycodelin A on human chorionic gonadotropin (hCG) and human placental lactogen (hPL) release by freshly isolated cytotrophoblasts (in vitro). Cytotrophoblasts have been prepared from human term placenta by the three-step trypsin-DNase dispersion method of villous tissue followed by a percoll gradient centrifugation step. When placed in culture, the isolated mononuclear trophoblasts differentiated into syncytial counterparts within 12-72 h after plating. Trophoblasts were incubated with varying concentrations (60-300 microg/ml) of glycodelin A. Glycodelin A was isolated and purified by chromatographic methods from amnion fluid. Supernatants of the trophoblast cell cultures were assayed for hCG and hPL by immunological methods. The release of hCG is increased in glycodelin A-treated trophoblast cell cultures compared to untreated trophoblast cells. Glycodelin A inhibits the production of hPL in vitro. Differences in Glycodelin A stimulated cells and untreated controls are statistical significant. hCG and hPL are markers for the differentiation process of trophoblast cells to syncytial trophoblasts. The results imply that glycodelin A secreted by decidualised endometrium modulates endocrine function, as well as the differentiation of trophoblasts in culture.

Lipopolysaccharides selectively inhibit mouse placental lactogen-II secretion through stimulation of interleukin-1 alpha (IL-1 alpha) and IL-6 production.

To determine whether lipopolysaccharides (LPS) regulate mouse placental lactogen-I (mPL-I), mPL-II, and mouse GHRF (mGHRF) secretion, mouse placental tissue from days 7, 9, and 12 of pregnancy was dispersed with collagenase and the purified trophoblast cells were cultured in a serum-free medium with or without LPS for 5 days. LPS significantly inhibited mPL-II secretion by cells from days 9 and 12 of pregnancy. However, LPS did not affect mPL-II secretion by cells from day 7 of pregnancy, mPL-I secretion by cells from days 7 and 9 of pregnancy, or mGHRF secretion by cells from day 12 of pregnancy. The inhibitory effect of LPS on mPL-II secretion by cells from day 12 of pregnancy was dose-dependent. Steady-state levels of mPL-II mRNA were significantly reduced by incubation of placental cells from day 12 of pregnancy with LPS. The inhibitory effect of LPS on mPL-II secretion was abolished by the addition of antibodies to IL-1 alpha and IL-6. These findings suggest that LPS selectively inhibit mPL-II secretion, at least partly through increases in IL-1 and IL-6 production, after midpregnancy.

Identification of a tyrosine residue in ovine placental lactogen as essential for its binding to receptors.

Nitration of ovine placental lactogen (oPL) with a 10-fold molar excess of tetranitromethane over protein content resulted in the modification of 0.8 tyrosine residue. No conformational changes were observed by either fourth-derivative spectral analysis or circular dichroism. Nitration significantly decreased the binding capacity of the hormone to lactogenic and somatogenic rat liver receptors. This binding capacity was not restored by reduction of the nitro groups, thus indicating that the decrease was not due to the difference in pK between tyrosine and nitrotyrosine. The nitrotyrosine-containing peptide was isolated from a tryptic digest by HPLC and its modification extent was of 67%, which is consistent with the decrease in binding capacities (65% and 70%). Its amino acid sequence was determined and the modified tyrosine residue was identified as Tyr-46. These results provide the first evidence of the involvement of a tyrosine residue in the binding of oPL to both lactogenic and somatogenic receptors. This tyrosine appears to be a shared binding epitope between oPL and the prolactins.

Posttranscriptional inhibition of mouse placental lactogen-II secretion by transforming growth factor beta 1: synergistic effects with epidermal growth factor and interleukin-6.

We studied the effect of transforming growth factor beta 1 (TGF beta 1) on mouse placental lactogen (mPL)-I and mPL-II secretion by primary cultures of placental cells from Days 7, 9, and 12 of pregnancy. We also studied the effects of co-incubation of epidermal growth factor (EGF) or interleukin-6 (IL-6) with TGF beta 1 on mPL-I and mPL-II secretion. TGF beta 1 at 10 ng/ml did not affect mPL-I secretion by cells from Days 7 or 9 of pregnancy or mPL-II secretion by cells from Day 7 of pregnancy but significantly inhibited mPL-II secretion by cells from Days 9 or 12 of pregnancy. The lowest concentration of TGF beta 1 that significantly inhibited mPL-II secretion by cells from Days 9 or 12 of pregnancy was 1 ng/ml. Immunocytochemistry for mPL-II indicated that treatment of placental cells from Day 12 of pregnancy with 10 ng/ml TGF beta 1 significantly reduced the number of mPL-II-containing cells. Inhibition of mPL-II secretion by TGF beta 1 was eliminated completely by addition of an anti-TGF beta 1 antibody. Northern analysis showed that steady state levels of mPL-II mRNA were not reduced by incubation of placental cells from Day 12 of pregnancy with 10 ng/ml TGF beta 1 for 5 days. EGF at 10 ng/ml significantly inhibited mPL-II secretion by cells from Day 7 of pregnancy, and addition of 10 ng/ml TGF beta 1, which did not itself inhibit mPL-II secretion by those cells, enhanced the inhibition by EGF of mPL-II secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Cytokines that use gp130 as a signal transducer stimulate mouse placental lactogen-I (mPL-I) but inhibit mPL-II production in vitro.

Interleukin-11 (IL-11), leukemia inhibitory factor (LIF), and oncostatin M (OM), all of which use gp130 as a signal transducer, significantly inhibited mouse placental lactogen-II (mPL-II) secretion by cultured placental cells from Days 7, 9, and 12 of pregnancy. These cytokines significantly stimulated mPL-I secretion by cells from Day 9, but not Day 7, of pregnancy. An antibody to LIF completely blocked the stimulatory and inhibitory effects of LIF on mPL-I and mPL-II secretion, respectively. LIF and OM decreased the abundance of mPL-II mRNA in placental cells. Double immunocytochemistry for mPL-I and mPL-II indicated that LIF, OM, and IL-11 significantly increased the number of giant cells containing only mPL-I or both mPL-I and mPL-II but decreased the number of giant cells containing only mPL-II. IL-6, which also uses gp130 as a signal transducer, inhibits mPL-II secretion only after midpregnancy; however, addition of soluble IL-6 receptor (sIL-6R) together with IL-6 resulted in a significant inhibition of mPL-II secretion before midpregnancy. Treatment of cells from Day 12 of pregnancy with IL-6 during the first 2 days of culture resulted in significant inhibition of mPL-II secretion by the third day of culture.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin 6 inhibits mouse placental lactogen II but not mouse placental lactogen I secretion in vitro.

The mouse placenta produces several polypeptides belonging to the prolactin-growth hormone gene family, including mouse placental lactogen (mPL) I and mPL-II. The present study was undertaken to determine whether the secretion of mPL-I and mPL-II is regulated by interleukin 6 (IL-6), which is present in the placenta and has previously been reported to stimulate the secretion of pituitary members of this gene family. Effects of human and mouse IL-6 on mPL-I and mPL-II secretion were examined in primary cultures of placental cells from days 7, 9, and 12 of pregnancy. IL-6 caused a dose-dependent reduction in the mPL-II concentration in the medium of cells from days 9 and 12 of pregnancy but did not affect the mPL-II concentration in the medium of cells from day 7 of pregnancy or the mPL-I concentration in the medium of cells from days 7 or 9 of pregnancy. The lowest concentration of human IL-6 that significantly inhibited mPL-II secretion was 250 pM. The effect of IL-6 on the mPL-II concentration in the medium was due primarily to inhibition of mPL-II synthesis, which resulted at least partly from a decrease in the steady-state level of mPL-II mRNA. These data raise the possibility that IL-6 may regulate mPL-II production after midpregnancy in vivo.

Increased intestinal uptake of cobalamin in pregnancy does not require synthesis of new receptors.

Increased intrinsic factor cobalamin binding to receptors present in ileal mucosa from mice in the late stages of pregnancy is regulated by placental lactogen. In mice at day 18-20 of pregnancy given an intraperitoneal injection of cycloheximide, 0.5 mg/kg, receptor binding was reduced from 0.42 ng/mg protein to 0.18 ng/mg protein 4 h later. Intestinal mucosal protein synthesis was less than 20% of control values after this dose of cycloheximide. Although this result could be interpreted to mean that the increase in receptors in pregnancy was due to new protein synthesis, cycloheximide-treated mice also had reduced concentrations of placental lactogen in serum. Supplementation with the hormone in cycloheximide-treated mice maintained receptor binding at pregnant levels. Analysis of binding data showed receptor number to be 3.1 X 10(11)/mg protein and the binding constant (Ka) to be 0.5 X 10(12) M-1, which were similar to values found in untreated pregnant mice. It is concluded that, because the increase in receptors cannot be explained on the grounds of new protein synthesis, placental lactogen may recruit cryptic intrinsic factor cobalamin receptors.