Estradiol-17 beta and 2-hydroxyestradiol-17 beta-induced differential production of prostaglandins by cells dispersed from human intrauterine tissues at parturition

Prostaglandin (PGE, 6-keto PGF1 alpha) output by cells dispersed from human amnion and decidua in the presence of increasing levels (0-5000 ng/ml) of estradiol-17 beta (E2) or 2-hydroxyestradiol-17 beta (2-OH E2) was studied in relation to parturition. Tissues were obtained from women at term either before (CS) or after (SL) spontaneous labor and vaginal delivery. In the absence of estrogens, the output of both PGs from amnion increased significantly with labor. No significant increase in decidua PG output occurred with labor. Neither estrogen influenced CS amnion PG output. However, both E2 and 2-OH E2 stimulated SL amnion PGE output (2-OH E2 greater than E2) while having no affect on 6-keto PGF1 alpha output. Only the highest dose of 2-OH E2 stimulated PGE output in CS decidua, but both estrogens significantly inhibited 6-keto PGF1 alpha output in this tissue. In SL decidua only 2-OH E2 significantly stimulated PGE, and neither estrogen affected 6-keto PGF1 alpha output. These results might suggest that estrogens modulate PG biosynthesis at the level of endoperoxide to primary PG conversion.     

In-vitro production of prostaglandins by intrauterine tissues from pregnant goats

In the fetal cotyledons and membranes, PGE production (determined in a continuous superfusion system) was significantly greater than that of PGF (P less than 0.01 and P less than 0.02 respectively). The fetal cotyledon also produced more PGE than the maternal cotyledon (P less than 0.01) and more PGF after parturition than before (P less than 0.01).     

Nuclear factor-kappa B localization and function within intrauterine tissues from term and preterm labor and cultured fetal membranes

Background  

The objective of this study was to quantify the nuclear localization and DNA binding activity of p65, the major transactivating nuclear factor-kappa B (NF-kappaB) subunit, in full-thickness fetal membranes (FM) and myometrium in the absence or presence of term or preterm labor.     

Estradiol-17β and 2-hydroxyestradiol-17β-induced differential production of prostaglandins by cells dispersed from human intrauterine tissues at parturition

Prostaglandin (PGE, 6-keto PGF_1α) output by cells dispersed from human amnion and decidua in the presence of increasing levels (0–5000 ng/ml) of estradiol-17β (E₂) or 2-hydroxyestradiol-17β (2-OH E₂) was studied in relation to parturition. Tissues were obtained from women at term either before (CS) or after (SL) spontaneous labor and vaginal delivery. In the absence of estrogens, the output of both PGs from amnion increased significantly with labor. No significant increase in decidua PG output occurred with labor. Neither estrogen influenced CS amnion PG output. However, both E₂ and 2-OH E₂ stimulated SL amnion PGE output (2-OH E₂>E₂) while having no affect on 6-keto PGF_1α output. Only the highest dose of 2-OH E₂ stimulated PGE output in CS decidua, but both estrogens significantly inhibited 6-keto PGF_1α output in this tissue. In SL decidua only 2-OH E₂ significantly stimulated PGE, and neither estrogen affected 6-keto PGF_1α output. These results might suggest that estrogens modulate PG biosynthesis at the level of endoperoxide to primary PG conversion.     

Progesterone and testosterone production by dispersed rat placental cells

Isopycnic separation and unit gravity sedimentation were employed to identify the rat placental cell types capable of producing progesterone and testosterone. Subdivision of Day 12-dispersed placental cells in Percoll gradients revealed that fractions (less than 1.048 g/ml) containing giant cytotrophoblast cells produced greater quantities of progesterone (p less than 0.01) than did fractions (greater than 1.048 g/ml) with equal numbers of placental cells but void of giant cytotrophoblasts. Unit gravity sedimentation of Day 16-dispersed placental cells revealed that when incubated, isolated giant cytotrophoblast cells were capable of producing both progesterone and testosterone. Both of the separation studies strongly suggested that other cell types also produce steroids. However, the biosynthetic capacity of the giant cytotrophoblast cell appeared to be 1000-fold greater than that of the other cell types. Incubation of Day 12-dispersed placental cells with human chorionic gonadotropin or 3',5'-cyclic adenosine monophosphate did not further increase progesterone production as compared to untreated control incubates, suggesting rat placental steroidogenesis is not under trophic hormone control. Electron microscopic observations of giant cytotrophoblast cells revealed a complex ultrastructure suggesting a variety of physiological functions.     

Testosterone production by collagenase-dispersed cells from baboon fetal testis

We determined whether dehydroepiandrosterone (DHA) and androstenedione (A) were converted to testosterone (T) by the midgestation primate fetal testis in the absence of gonadotropins. Testes from six baboon (Papio anubis) fetuses, obtained by cesarean section at Day 100-107 of gestation (term = Day 184) were dispersed with 0.2% collagenase. Cells (1.1 X 10(6)) were suspended in 4 ml Eagle's Minimum Essential Medium containing penicillin/streptomycin (MEM) and incubated for 20 h (37 degrees C) with or without DHA, A, pregnenolone (P5), 17 alpha-hydroxypregnenolone (17OH-P5), progesterone (P4) or 17 alpha-hydroxyprogesterone (17OH-P4). Concentrations of T, A, P4, and 17OH-P4 in the medium and cells were measured by radioimmunoassay. Mean secretions of T and A, in the absence of exogenous substrates, were 0.5 +/- 0.2 and 0.8 +/- 0.3 ng/mg testis, respectively, and were not elevated by human chorionic gonadotropin (hCG). Addition of DHA at 100, 500, or 1000 ng/4 ml increased (p less than 0.05) the production of T to 6 +/- 0.6, 33 +/- 10, and 64 +/- 26 ng/mg testis and the production of A to 13 +/- 5.5, 54 +/- 10, and 67 +/- 22 ng/mg testis, respectively. Similarly, addition of A at 100, 500, or 1000 ng/4 ml increased (p less than 0.05) production of T to 27 +/- 5.3, 155 +/- 29, and 254 +/- 79 ng/mg testis, respectively. In contrast, production of T and A remained near baseline concentrations when cells were incubated with 1000 ng/4 ml of P5, P4, 17OH-P5, or 17OH-P4.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prostaglandin production by dispersed granulosa and theca interna cells from porcine preovulatory follicles

Prepubertal gilts were treated with 750 IU pregnant mares' serum gonadotropin (PMSG) and 72 h later with 500 IU human chorionic gonadotropin (hCG) to induce follicular growth and ovulation. Dispersed granulosa (GC) and theca interna (TIC) cells were prepared by microdissection and enzymatic digestion from follicles obtained 36, 72 and 108 h after PMSG treatment and incubated for up to 6 h in a chemically defined medium in the presence or absence of arachidonic acid, follicle-stimulating hormone (FSH), luteinizing hormone (LH) and indomethacin. Production of prostaglandin E2 (PGE) and prostaglandin F2 alpha (PGF) was measured by radioimmunoassay. Both GC and TIC had the capacity to produce prostaglandins, with production by each cell type increasing markedly with follicular maturation. PGE was the major prostaglandin produced by both cellular compartments. Only PGE production by GC was consistently enhanced by addition of arachidonic acid to the incubation medium. Neither cell type was responsive to FSH and LH in vitro. Indomethacin inhibited the production of PGE and PGF by both cell types. These results provide convincing evidence for an intrafollicular source of prostaglandins and indicate that both cellular compartments contribute significantly to the increased production of prostaglandins associated with follicular rupture.     

Steroid production from 17alpha-hydroxypregnenolone and dehydroepiandrosterone by human granulosa cells in vitro.

Granulosa cells were aspirated 3--4 h before the expected time of ovulation from 10 follicles of 4 patients treated with gonadotrophins: 4 of the follicles were immediately preovulatory. The granulosa cells were cultured for 10 h with 17alpha-hydroxypregnenolone or dehydroepiandrosterone and samples of medium removed at 3 and 10 h were assayed for 6 steroids. Granulosa cells were unable to synthesize androgens from endogenous substrate or undertake conversions via the delta5 pathway, but cells from all follicles were capable of aromatizing exogenous androgens to oestrogens although this capability was reduced in cells from follicles beginning to luteinize. Granulosa cells from preovulatory follicles synthesized more progesterone from endogenous substrate than cells from follicles which had not begun to luteinize. The results provide further support for the two-cell theory of oestrogen biosynthesis whereby granulosa cells aromatize androgens which are synthesized by the thecal cells in vivo.     

Steroid production in vitro by granulosa, theca, and luteal cells from goat ovaries

Basal progesterone (P4) production by isolated goat ovarian cells in vitro was in the order corpus luteum (CL) greater than granulosa (G) greater than theca (TH), while estradiol (E2) production was in the order TH greater than G greater than CL. In G cells, various concentrations (0.01 to 100 micrograms/ml) of luteinizing hormone (LH), human chorionic gonadotropin (hCG) and follicle-stimulating hormone (FSH) increased P4 and E2 secretion. Testosterone (T, 10(-9) to 10(-5) M) produced dose-dependent increases in P4 and E2 secretion. Testosterone and LH together had an additive effect on E2 secretion. The combined effect of the lower (less than 10(-6) M) concentrations of T and LH on P4 production was marginally higher than either agent alone, but the increase was statistically insignificant; at higher concentrations of T (10(-6) and 10(-5) M) in combination with LH, P4 secretion was similar to that with LH alone, but was significantly (p less than 0.01 and less than 0.001, respectively) less compared to that with T alone. Follicle-stimulating hormone and T together produced a synergistic effect on E2 and an additive effect on P4 production. In TH cells, a dose-dependent increase in P4 and E2 production was observed with LH and hCG, but the effect of FSH was not significant. Testosterone produced a dose-dependent increase in P4 and E2 secretion. Testosterone and LH together induced higher steroid production than either agent alone. However, the increase was not statistically significant compared to T alone.(ABSTRACT TRUNCATED AT 250 WORDS)     

The endogenous production of hydrogen sulphide in intrauterine tissues

Background  

Hydrogen sulphide is a gas signalling molecule which is produced endogenously from L-cysteine via the enzymes cystathionine beta-synthase (CBS) and cystathionine gamma-lyase (CSE). The possible role of hydrogen sulphide in reproduction has not yet been fully investigated. It has been previously demonstrated that hydrogen sulphide relaxes uterine smooth muscle in vitro. The aim of the present study was to investigate the endogenous production of hydrogen sulphide in rat and human intrauterine tissues in vitro.     

Catecholamine-induced stimulation of testosterone production by Leydig cells from fetal mouse testis

In contrast to the strong stimulation of testosterone production by hCG, L-isoproterenol had little effect on freshly isolated Leydig cells from 18-day-old mouse fetuses. However, the ability of fetal Leydig cells to respond to L-isoproterenol exposure increased during culture (0-24 h). The response of the cultured cells to L-isoproterenol was dose-dependent with an ED50 at 2 X 10(-7) M. Adrenaline and noradrenaline at a concentration of 10(-5) M also increased testosterone production by cultured fetal Leydig cells. DL-Propranolol, a beta-antagonist, inhibited L-isoproterenol-stimulated testosterone production in a dose-dependent manner, while phentolamine, an alpha-adrenergic antagonist, had no effect. These results suggest that catecholamines may play an essential role in the control of testicular steroidogenesis during fetal development.     

Differential production of steroids by dispersed granulosa and theca interna cells from developing preovulatory follicles of pigs

Dispersed granulosa and theca interna cells were recovered from follicles of prepubertal gilts at 36, 72 and 108 h after treatment with 750 i.u. PMSG, followed 72 h later with 500 i.u. hCG to stimulate follicular growth and ovulation. In the absence of aromatizable substrate, theca interna cells produced substantially more oestrogen than did granulosa cells. Oestrogen production was increased markedly in the presence of androstenedione and testosterone in granulosa cells but only to a limited extent in theca interna cells. The ability of both cellular compartments to produce oestrogen increased up to 72 h with androstenedione being the preferred substrate. Oestrogen production by the two cell types incubated together was greater than the sum produced when incubated alone. Theca interna cells were the principal source of androgen, predominantly androstenedione. Thecal androgen production increased with follicular development and was enhanced by addition of pregnenolone or by LH 36 and 72 h after PMSG treatment. The ability of granulosa and thecal cells to produce progesterone increased with follicular development and addition of pregnenolone. After exposure of developing follicles to hCG in vivo, both cell types lost their ability to produce oestrogen. Thecal cells continued to produce androgen and progesterone but no longer responded to LH in vitro. These studies indicate that several functional changes in the steroidogenic abilities of the granulosa and theca interna compartments occur during follicular maturation.