High concentrations of plasma immunoreactive inhibin during normal pregnancy in women.

The plasma inhibin concentrations in 190 normal pregnant women at 5-40 weeks gestation and in 4 puerperal women were measured by a specific RIA for human inhibin. The average plasma inhibin concentrations in pregnant women throughout pregnancy (minimum, 2.25 +/- 0.48 IU/mL at 17 weeks gestation; maximum, 24.15 +/- 6.99 IU/mL at 39 weeks gestation) were much higher than those in nonpregnant women with a normal menstrual cycle (0.46 +/- 0.04 IU/mL in the midfollicular phase and 2.02 +/- 0.47 IU/mL in the midluteal phase). The inhibin concentrations were already high at 5 weeks gestation (7.54 +/- 1.10 IU/mL) and rose to peak at 8-10 weeks gestation. The concentrations then decreased and remained relatively low during 14-30 weeks gestation, but rose again during the third trimester. The inhibin concentrations decreased to undetectable levels after delivery. Immunoreactive inhibin was demonstrated in the corpus luteum and term placental extracts, and the dose-response curves were parallel to an inhibin preparation from human follicular fluid. Immunoreactive inhibin concentrations were also high in both the umbilical vein and artery (7.77 +/- 0.80 and 7.84 +/- 0.78 IU/mL, respectively). These observations suggest that both the corpus luteum and placenta are likely sources of inhibin.

Human chorionic gonadotropin localization and morphometric characterization of human granulosa-luteal cells obtained during in vitro fertilization cycles.

The area and cytoplasmic-to-nuclear ratio (C/N) of cells aspirated from follicles with mature oocytes was determined using a computerized image analysis system. The presence of human chorionic gonadotropin (hCG) on the surface membrane and/or within the cytoplasm of each cell also was determined using a horseradish peroxidase immunocytochemical procedure. Based on morphometric characteristics, follicular cells were classified as granulosa or luteal. Granulosa cells were less than 75 micron 2 in area with a C/N of approximately 0.5. Luteal cells were classified as small (less than 75 micron 2, C/N approximately 1.5), midluteal (76 to 100 micron 2, C/N greater than 1.5) and large luteal (greater than 100 micron 2, C/N greater than 1.5). Compared with aspirates from follicles containing fertilizable oocytes, aspirates from follicles with nonfertilizable oocytes had fewer granulosa cells and more large luteal cells. HCG was localized on the membranes of granulosa and small luteal cells and within the cytoplasm of midluteal cells. Human chorionic gonadotropin was generally not observed on either the membranes or cytoplasm of luteal cells over 120 micron 2. These data support the concept that granulosa cells bind hCG to membrane receptors, internalize hCG, and begin to luteinize in response to hCG stimulation. Since the aspirates from follicles containing nonfertilizable oocytes possessed a higher percentage of large luteal cells, it is postulated that the cells from these aspirates began the luteinization process earlier than those from follicles containing fertilizable oocytes.

Effects of prostaglandin F2 alpha on degranulation of bovine luteal cells on days 4 and 12 of the estrous cycle.

Corpora lutea were collected from 18 beef heifers on day 4 or 12 of the estrous cycle, 1 hour after prostaglandin (PG) F2 alpha or saline (control) treatment. Five heifers also were treated with PGF2 alpha on day 4, but their corpora lutea were not collected until day 12. The relative percentage of cytoplasm occupied by granules decreased only in large luteal cells (LLC) in heifers given PGF2 alpha on day 12, compared with the percentage in controls. Small luteal cells (SLC) were not as affected. The luteal concentration of progesterone was similarly decreased only in heifers given PGF2 alpha on day 12. Treatment of heifers with PGF2 alpha on day 4 had no marked effect on progesterone values or on the relative percentage of cytoplasm occupied by granules in LLC or SLC. Seemingly, LLC were more responsive to PGF2 alpha than were SLC, and PGF2 alpha treatment of beef heifers at day 4 did not markedly impair luteal function.

Neurophysin in the large luteal cell of the nonpregnant water buffalo (Bubalus bubalis): immunohistochemical localization.

Light microscopy immunohistochemistry was used to localize neurophysin in the corpus luteum of the mid-luteal phase of the estrous cycle of the water buffalo (Bubalus bubalis). Corpora lutea weighing 0.39-0.65 g from a recent ovulation showed no staining. Corpora lutea identified with the late luteal phase showed only weak evidence of staining. The neurophysin staining was confined to a specific region of large oval-shaped cells (20-30 microns diameter), which had a very eosinophilic cytoplasm. The intense localization of staining to a distinct area of the cytoplasm was previously only observed in the corpus luteum of the cow. Corpora lutea obtained from all quadrants of pregnancy did not stain. Controls in which the neurophysin antiserum was substituted with serum from an unimmunized rabbit (normal rabbit serum) or neurophysin antiserum preabsorbed with bovine oxytocin-associated neurophysin I also did not stain. These data indicate the neurophysin is present in the mature corpus luteum of the nonpregnant water buffalo as it is in other nonpregnant ruminants, the ewe and cow.

Immunoreactive beta-endorphin in human ovaries.

Beta-endorphin (beta-EP) immunostainable cells were demonstrated in human ovarian tissue using a non-cross-reacting anti-beta-EP serum and the avidin-biotin-peroxidase detection technique. In ovaries from ovulating and premenopausal women, beta-EP immunoreactivity was localized in the luteinized cells of theca interna of maturing follicles with almost negligible staining in granulosa cells; cells of primary follicles did not stain. In corpora lutea, luteinized cells in both theca interna and granulosa, layers were equally positive. In postmenopausal ovaries, staining was detectable only in scattered luteinized stromal cells. This is the first report on the presence of immunoreactive beta-EP in human ovaries, in which beta-EP seems to be produced by the same sex cord cells engaged in active steroidogenesis and may be under gonadotropin central regulation. The significance of this finding is discussed.

[Effect of low density lipoprotein on intracellular cholesterol and progesterone production by monolayer cultured human luteal cells].

The purpose of the present study was to define the effect of low density lipoprotein (LDL) on progesterone (P) synthesis and on intracellular cholesterol content using monolayer cultured human luteal cells. When cultured in TC199 containing 10% lipoprotein poor serum (LPPS), P production decreased as culture time passed. In the presence of hCG, it also decreased, but slowly. In the presence of LDL, P production was kept at a higher level. Intracellular cholesterol content in luteal cells was measured by gas chromatography. The cholesterol content of luteal cells in a 4 day LPPS culture was higher than that in a 10 day LPPS culture. The cholesterol content in a 10 day LPPS culture with LDL was higher than that in a 10 day LPPS culture. The cholesterol content in a 10 day LPPS culture with hCG had a tendency to be lower than that in a 10 day LPPS culture, but there was no statistical difference (p less than 0.1). It was concluded that LDL was utilized in supplying intracellular cholesterol to luteal cells as a substrate for P synthesis. And in a state of LDL depletion in the medium, the luteal cells produced a small amount of P utilizing intracellular cholesterol. It was estimated from the results of the present study that hCG enhanced utilization of intracellular cholesterol.

Ultrastructural localisation of oxytocin and neurophysin in the ovine corpus luteum.

Polyclonal rabbit antisera raised against oxytocin, bovine neurophysin I and vasopressin were used, together with an immunogold complex, to localise the peptides in ultrathin sections of ovine corpus luteum. The only organelle which consistently showed gold labelling was the secretory granule of the large luteal cell. In non-consecutive sections of the same large luteal cell all the granules showed a similar level of labelling after oxytocin or neurophysin I antisera: however no immunolabelling was detected for vasopressin. Oxytocin and neurophysin seem to be rapidly lost after secretion since exocytosed granule cores showed no labelling above background levels.

Ultrastructural localization of relaxin in the corpus luteum of the nonpregnant, pseudopregnant, and pregnant pig.

Relaxin was localized in luteal cells of ovaries from nonpregnant, pseudopregnant, and pregnant pigs using porcine relaxin antiserum and peroxidase-antiperoxidase light microscopy immunohistochemistry. The number of immunoreactive cells seemed to increase from Days 17 to 106 of gestation. Luteal cells from pseudopregnant (Day 110) and nonpregnant (Day 14 of the estrous cycle) pigs were also positive for relaxin. However, less than 3% of the luteal cells in the nonpregnant animals were immunoreactive. Electron microscopy immunocytochemistry using porcine relaxin antiserum and goat antirabbit immunoglobulin G-colloidal gold demonstrated that relaxin was packaged in the small membrane-bound granules in luteal cells of pregnant as well as pseudopregnant and nonpregnant pigs. The intensity of labeling (number of gold particles) of the granules increased with pregnancy. There was a 10-fold increase in labeling of granules with the 10-nm versus 25-nm diameter gold. The goat antirabbit labeled with the smaller 10-nm gold particles was necessary to demonstrate the apparent low levels of relaxin in the luteal cells of the nonpregnant pigs. These data further indicate that pregnancy is not required for relaxin synthesis. However, physiologic significance of relaxin in corpora lutea of nonpregnant pigs has not been determined.