Secretion of progesterone and prostaglandins by cells of bovine corpora lutea from three stages of the luteal phase

The secretion of prostaglandins (PGs) by bovine corpora lutea was investigated. Corpora lutea from the early, early-mid and late-mid stages of the luteal phase were dissociated by collagenase treatment and cultured in monolayer in Dulbecco's modified Eagle's medium containing 10% (v/v) fetal calf serum. Treatment with either LH (100 ng/ml) or dibutyryl cyclic AMP (dbcAMP; 1 mmol/l) had no effect on progesterone secretion by early luteal phase cells but stimulated progesterone secretion two- to fourfold by cells from the latter stages. The secretion rates, per microgram cell protein, of 6-keto-PGF1 alpha, PGE2 and PGF2 alpha were substantially greater in cells from the early luteal phase than in those from the latter stages, however, all changes in PG secretion in response to treatments were qualitatively similar between cells from the three stages of the luteal phase. The secretion rate of 6-keto-PGF1 alpha was greater than that of PGE2 or PGF2 alpha and was inhibited by treatment with indomethacin (28 mumol/l) but unaltered by treatment with LH, dbcAMP or butyrate (1 mmol/l). Secretion of PGE2 was inhibited by indomethacin but stimulated two- to threefold by treatment with either dbcAMP or butyrate. Secretion of PGF2 alpha was minimal and not inhibited further by treatment with indomethacin, but was stimulated 10- to 40-fold with dbcAMP. Indomethacin treatment inhibited the stimulatory effect of dbcAMP; butyrate had no effect on PGF2 alpha secretion. Treatment with LH had no effect on any of the PGs measured. In these experiments the secretion of progesterone appeared unrelated to any changes in the secretion of PGs.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of prostaglandin F2 alpha on bone formation and resorption in cultured neonatal mouse calvariae: role of prostaglandin E2 production

Although most studies show that prostaglandin E2 (PGE2) is the most potent and effective of the prostanoids in bone, recent data in cell culture suggest that PGF2 alpha may have unique effects, particularly on cell replication. The present study was undertaken to compare the effects of PGF2 alpha and PGE2 in cultured neonatal mouse parietal bones by simultaneous measurement of bone resorption as release of previously incorporated 45Ca, bone formation as incorporation of [3H]proline into collagenase-digestible (CDP) and noncollagen protein, and DNA synthesis as incorporation of [3H]thymidine. PGF2 alpha was less effective than PGE2 as a stimulator of bone resorption, and its effects were partially inhibited by indomethacin and markedly inhibited by glucocorticoids. In contrast, the resorptive response to PGE2 was unaffected by indomethacin and only partially inhibited by cortisol. PGF2 alpha had little effect on bone formation, in contrast to the biphasic effect of PGE2, which inhibited labeling of CDP in the absence of cortisol and stimulated CDP labeling in the presence of cortisol. PGF2 alpha increased thymidine incorporation into DNA, but the effect was smaller than that of PGE2 and was inhibited by indomethacin. These observations suggested that PGF2 alpha might act in part by stimulating PGE2 production. By RIA, PGE2 concentrations were increased in the medium of bones treated with PGF2 alpha, and this increase was blocked by indomethacin. By HPLC, bones prelabeled with [3H]arachidonic acid showed an increase in labeled PGE2 release, and RIA showed an increase in PGE2 after PGF2 alpha treatment. These results indicate that PGF2 alpha is a relatively weak agonist in bone compared to PGE2 and that some of the effects of PGF2 alpha on bone resorption, formation, and cell replication may be mediated by an increase in endogenous PGE2 production.     

Effect of carprofen and indomethacin on gastric function and the content of prostaglandins E2 and F2 alpha in human gastric juice

The effect of indomethacin (2 X 50 mg daily) and carprofen (2 X 150 mg daily) on gastric secretion and the generation of prostaglandins PGE2 and PGF2 alpha in gastric juice, was investigated in a single blind cross-over study in eight healthy volunteers lasting one week. We observed no statistically significant change in basal and pentagastrin-stimulated gastric secretory parameters (outputs of gastric acid, N-acetyl-neuraminic acid and pepsin) before and after treatment with indomethacin and carprofen. However, an inhibitory effect was found on the output of PGE2 and PGF2 alpha after pentagastrin stimulation. While both drugs diminished the output of PGF2 alpha to a similar extent, carprofen exerted a markedly weaker inhibitory effect on the output of PGE2 than did indomethacin. It is suggested that the gastric tolerability of non-steroidal anti-inflammatory drugs (NSAIDs) is related to their inhibitory potency on PGE2 formation, in the sense that weak inhibitors of PGE2 cause less damage to the gastric mucosa than do strong inhibitors.     

Mechanism of the rapid antigonadotropic action of prostaglandins in cultured luteal cells

A reproducible method for dissociation and culture of rat luteal cells is described. The concentration of LH required to produce half-maximal stimulation of progesterone secretion was 50 ng/ml. The effects of prostaglandin E(2) (PGE(2)) and prostaglandin F(2alpha) (PGF(2alpha)) on basal and luteinizing hormone (LH)-stimulated progesterone production were examined. Both prostaglandins stimulated basal progesterone production but PGE(2) was about twice as active, showing a 2-fold maximal stimulation at 0.75 muM. When either prostaglandin was incubated simultaneously with LH, a dose-dependent inhibition of progesterone secretion occurred; PGF(2alpha) was 4 times more active than PGE(2), showing 50% inhibition at a concentration of 40 x nM. Thus, both prostaglandins are more active as antagonists than as agonists of LH with respect to progesterone secretion. PGF(2alpha) also inhibited LH-stimulated adenylate cyclase activity and cyclic AMP accumulation. The block in progesterone secretion was reversed by addition of dibutyryl cyclic AMP (1 mM) but not by theophylline (5 mM) alone. These data and the finding that PGF(2alpha) did not affect the specific binding activity of the LH receptor in intact luteal cells indicate that the rapid action of prostaglandins in luteal cells is due to a block of LH-dependent production of cyclic AMP which results in a decrease in progesterone secretion.     

Hormonal responsiveness by immature rabbit uterine epithelial cells polarized in vitro.

This paper reports the development of an in vitro cell culture system of polarized uterine epithelial (UE) cells from the immature rabbit. UE cells from immature rabbit were cultured on EHS (Engelbreth-Holm-Swarm) matrix-coated semipermeable filter inserts in a serum-free, phenol red-free defined medium. The cells in primary culture attached, proliferated, formed monolayers, and exhibited structural and functional polarity. Structural differentiation was validated by ultrastructural features, such as apical microvilli, intercellular junctions, desmosomes, and polar organization of apical vs. basal membrane domains. Trans-monolayer epithelial resistance, a reflection of functional tight junctions, preferential basal uptake of [35S]methionine, polarized distribution of labeled secretory proteins, and increased secretory activity marked by preferential apical secretion (greater than 90%) of proteins were indices of functional polarity. With the development and maintenance of polarized state by the UE cells, some of the newly synthesized proteins were sorted and vectorially secreted into their distinct compartments. De novo synthesis and exclusive apical secretion of uteroglobin (a progesterone-induced protein in vivo) by the polarized UE cells occurred in response to progesterone treatment in vitro. The hormone responsiveness was maintained for a prolonged period under these conditions. A culture system in which the UE cells maintain functional and morphological polarity and sustain their hormonal responsiveness facilitates the study of regulation of specialized functions by the UE cells that are regulated by steroid hormones and their interactions with the other uterine cell types.     

Absence of prostacyclin involvement in angiotensin-induced aldosterone secretion in rat adrenal cells

The role of prostaglandins (PGs) in aldosterone secretion was studied in isolated rat adrenal glomerulosa cells. [14C]Arachidonic acid was metabolized to [14C]6-keto-PGF1 alpha, [14C]PGF2 alpha, [14C]PGE2, and [14C]PGD2. Pretreatment with indomethacin (5 X 10(-5) M) or U-51605 (5 micrograms/ml) inhibited the synthesis of these metabolites. Angiotensin II (AII) stimulated a concentration-related release of aldosterone and 6-keto-PGF1 alpha, but not PGE2. Significant increases in aldosterone and 6-keto-PGF1 alpha occurred at AII concentrations of 0.2 and 2 nM. The increases in 6-keto PGF1 alpha concentrations after AII treatment were small, however (278 +/- 33 pg/10(6) cells X h for control vs. 581 +/- 90 after 2 nM AII). At higher concentrations, AII further stimulated aldosterone, but 6-keto PGF1 alpha levels declined. AII stimulated the synthesis of aldosterone and 6-keto PGF1 alpha in parallel with time of incubation. Indomethacin (3 microM) decrease basal and AII-stimulated aldosterone release by 40% and 23%, respectively, and inhibited the synthesis of PGs. U-51605 (5 micrograms/ml) failed to alter aldosterone release. Arachidonic acid increased the synthesis of PGE2 and 6-keto-PGF1 alpha in a concentration-related manner without altering the synthesis of aldosterone. In contrast, PGH2 stimulated the release of PGE2, 6-keto-PGF1 alpha, and aldosterone. PGI2 and PGE2 stimulated aldosterone secretion, which was concentration related. Threshold stimulation by PGI2 and PGE2 occurred at 0.5 and 5 nM, respectively. Maximal stimulation occurred at 5 nM for PGI2 and at 5000 nM for PGE2, with PGE2 producing the greater maximal response. Treatment of the cells with trypsin eliminated the steroidogenic response to PGE2. These findings indicate that PGI2 and PGE2 are produced by the adrenal glomerulosa cells, and the synthesis of PGI2 may be stimulated by AII. However, the concentrations of PGI2 synthesized are not adequate to stimulate aldosterone secretion. Thus, PGI2 does not appear to mediate angiotensin-induced aldosterone secretion.     

Prostaglandins and steroidogenesis in isolated bovine adrenal cells. Effects of ACTH, prostaglandin-synthesis inhibitors, prostaglandins and prostaglandin analogs

In bovine adrenal cortex cells, dispersed without preferential loss of cells, we investigated (1) whether endogenous prostaglandins (PGs) are involved in ACTH-induced adrenal steroidogenesis, and (2) the steroidogenic effects of PGs and PG analogs. Free cells produced considerable amounts of PGE2, whereas only minute quantities of PGF2 alpha and PGA1 were synthesized. PGE2 synthesis, however, was not significantly increased when ACTH elicited a steroidogenic response in free cells. High concentrations of PG-synthesis inhibitors such as indomethacin affected both PG synthesis and steroidogenesis, whereas intermediate concentrations (10(-6) M) inhibited production of both PGE2 and aldosterone even after cAMP and cortisol response to ACTH had returned to normal values. It is concluded that endogenous PGE2 is not a link in the acute mechanism of action of trophic hormones in which cAMP is involved. Of the prostanoid structures, PGs of the E series were the most potent stimulating agents of cortisol production, although less active than ACTH. On the other hand, PGA1 induced an ACTH-like aldosterone synthesis. PGE2 was less active, and other prostanoid structures were without effect on aldosterone production. It is suggested that in pathological circumstances, PGA1 regulates aldosterone production and PGE2 increases both aldosterone and cortisol production.     

Prostaglandin synthesis by isolated rat renal glomeruli.

The PGE2, PGF2 alpha and 6-keto-PGF1 alpha contents of the incubation medium of glomeruli isolated from rat kidney were measured at different times with or without addition of arachidonic acid. These prostaglandins accumulated progressively with time and reached equilibrium after 60--120 min incubation. Synthesis of the 3 prostaglandins was inhibited when indomethacin was added whereas it was markedly enhanced, mainly for PGE2, at increasing doses of arachidonic acid. Plateaus were reached above 5 micrograms/ml and concentrations corresponding to 50% of the maximum values were 2 micrograms/ml for PGE2 and PGF2 alpha, and 0.8 microgram/ml for 6-keto-PGF1 alpha. There were strictly linear relationships between PGE2 or PGF2 alpha productions and the concentration of glomerular protein. PGE2 and PGF2 alpha synthesis with or without arachidonic acid were maximum at 30--37 degrees C. PGE2 glomerular content was almost undetectable initially and increased with time. These data demonstrate that PGE2, PGF2 alpha and PGI2, in order of decreasing abundance, are synthesized by the glomerular cells and suggest that PGE2 and PGI2-sensitive glomerular adenylate cyclase activities and PGE2-sensitive renin synthesis may be stimulated by prostaglandins formed in the glomeruli themselves.     

Polarized secretion of androgen-binding protein and transferrin by Sertoli cells grown in a bicameral culture system

Sertoli cells in vivo are believed to secrete androgen-binding protein (ABP) as well as other proteins in a polarized manner, primarily into the adluminal (apical) compartment of the seminiferous tubules. It is now possible to examine polarized secretion by Sertoli cells in vitro by growing them in dual environment (bicameral) culture chambers such that there is a separation of the apical and basal compartments of the cells. Sertoli cells obtained from 17-day-old rats were grown in these chambers on Millipore filters coated with a reconstituted basement membrane matrix. Within 3 days a confluent epithelial sheet of Sertoli cells (30-40 microns in height) is established, and these cells are joined along their baso-lateral plasma membranes by typical Sertoli-Sertoli tight junctions. We have previously shown that this epithelial sheet of Sertoli cells establishes an electrical resistance, as well as a permeability barrier to small molecules, between the basal and apical compartments of the culture chamber. In the absence of Sertoli cells, proteins equilibrate within 8 h across the matrix-coated filter support. Between 12 and 48 h of culture the Sertoli cell monolayers secrete approximately 4- and 2-fold more ABP and transferrin (Tf), respectively, into the apical compartment than into the basal compartment when grown in serum-free defined medium. ABP secretion is diminished 10-fold by the removal of testosterone from the serum-free defined medium. In addition, the apical/basal ratio of ABP secretion decreases from 4.1 to 1.7 in the absence of testosterone. In contrast, neither the amount nor the direction of Tf secretion is altered by the removal of testosterone. These results demonstrate the bidirectional secretion of ABP and Tf by Sertoli cells grown in bicameral chambers. In addition, the polarity of secretion of these two proteins by Sertoli cells appears to be under differential regulation by testosterone.     

Inhibition by prostaglandin E2 of the release of vasopressin and beta-endorphin from rat pituitary neurointermediate lobe or medial basal hypothalamus in vitro

The present study was performed to examine the effect of the cyclo-oxygenase inhibitor, indomethacin, and that of various prostaglandins on the release of vasopressin and beta-endorphin-like immunoreactivity (beta-EI) from the rat neurointermediate lobe of the hypophysis, which was superfused in vitro. Indomethacin (2.8 and 28 mumol/l) changed neither basal secretion of vasopressin nor that evoked by electrical stimulation, whereas the resting release of beta-EI was enhanced by indomethacin (28 mumol/l). Prostaglandin (PG) E2 did not influence resting release of vasopressin but markedly inhibited (by about 50%) electrically induced release of vasopressin (least effective concentration: 300 nmol/l) as well as spontaneous secretion of beta-EI (least effective concentration: 100 nmol/l) in the presence of indomethacin (28 mumol/l). Prostaglandin F2 alpha (5 mumol/l) also inhibited the evoked release of vasopressin, whereas PGD2 (5 mumol/l) did not. Prostaglandin F2 alpha (5 mumol/l), D2 and I2 (1.5 mumol/l each) produced no effects on beta-EI release. As observed in the neurohypophysis, PGE2 inhibited the electrically induced release of vasopressin from the medial basal hypothalamus in vitro. We conclude that prostaglandins (especially PGE2) can inhibit (1) the stimulated release of vasopressin when acting on vasopressin-containing nerve terminals of either neurosecretory system (neurohypophysis, median eminence region), and (2) the secretion of beta-EI and, as can be inferred, alpha-MSH, by a direct action on intermediate lobe cells.     

The effect of LH/FSH, dibutyryl cyclic AMP, and prostaglandins on the production of estrogens by rabbit granulosa cells in vitro.

The purpose of this study was to establish the effects of trophic hormones on the production of estrogens by rabbit granulosa cells. A pure population of these cells was isolated from preovulatory follicles (1-15 mm in diameter) of estrous rabbits, and cultured for 6 days with either one or a combination of the following hormones: androstenedione, Pergonal (LH/FSH), dibutyryl cyclic AMP (Bu2cAMP), prostaglandin F2alpha (PGF2alpha), or prostaglandin E2 (PGE2). The medium was collected every 2 days and progesterone (P), estrone (E1) and estradiol-17 beta (E2beta) were measured by radioimmunoassay. Granulosa cells cultured as controls (i.e., without exogenous trophic hormones) secreted P spontaneously and its secretion was stimulated 100 to 1,000 fold with LH/FSH and Bu2cAMP, but not with PGF2alpha or PGE2. Androstenedione, either alone or with trophic hormones had no apparent effect on the cytology of the granulosa cells or their ability to secrete P. In the absence of exogenous androstenedione, the cultures produced very small amounts of E1 or E2beta (smaller than 100 pg/ml), either spontaneously or in response to LH/FSH, B12cAMP, PGF2alpha, or PGE2. Incubating granulosa cells with exogenous androstenedione (1 mug/ml) resulted in a 30- to 150-fold increase in E2beta production, which was stimulated an additional 3- to 5-fold with LH/FSH and Bu2cAMP, but not with PGF2alpha or PGE2. In most cultures, E2beta production was restricted to the first 2 days in vitro. Bu2cAMP, however, maintained E2beta production at relatively high levels throughout the duration of the experiment, but there was a progressive decrease in its production. The production of E1 was only 5 percent of E2beta, but the pattern of secretion was similar for both estrogens. These results suggest that cyclic AMP could have a role in regulating the synthesis of estrogens by rabbit granulosa cells.     

In vitro effects of serotonin and prostaglandins on release of eggs from the ovary of the scallop, Patinopecten yessoensis

When ovarian pieces of the scallop, Patinopecten yessoensis, were incubated in media containing serotonin (5-HT), the number of released eggs increased significantly. These eggs developed normally after fertilization. The threshold of the 5-HT effect reached a maximum at a concentration of 10(-6) M, and decreased sharply with increasing concentrations. Methysergide, a 5-HT antagonist, completely inhibited the 5-HT-induced egg release. Addition of aspirin or indomethacin to the incubation media also inhibited the 5-HT effect. However, neither aspirin nor indomethacin inhibited the 5-HT-induced egg release when 10(-6) M prostaglandin E2 (PGE2) was also present. PGE2 did not induce egg release, but enhanced the 5-HT effect. In contrast to PGE2, PGF2 alpha significantly inhibited the 5-HT effect. These results show that 5-HT induces spawning of female P. yessoensis via specific 5-HT receptors in the ovary, and that the effect of 5-HT may be regulated by PGE2 (progressive) and PGF2 alpha (suppressive).     

Prostaglandin E1 and F2 alpha stimulate differentiation and proliferation, respectively, of clonal osteoblastic MC3T3-E1 cells by different second messengers in vitro

The effect of several prostaglandins (PGs) on osteoblastic cells was investigated using clone MC3T3-E1 under serum-free conditions. PGA1, A2, B1, and B2 had little effect on intracellular cAMP, alkaline phosphatase (ALP) activity, and DNA synthesis in the cells. At 4-2000 ng/ml, PGE1 among PG analogs tested had a dose-dependent stimulatory effect on ALP activity in the cells, and this effect was amplified by isobutyl methylxanthine. Also, PGE1 strongly augmented the amount of intracellular cAMP over the same concentration range. However, PGE1 had little effect on ornithine decarboxylase activity and DNA synthesis, and at high doses it rather depressed DNA synthesis. Furthermore, PGE1 did not affect the intracellular cGMP level. The effect of PGE1 on the cells closely mimics that of forskolin, suggesting that the PG stimulates the differentiation of the osteoblastic cells predominantly via the stimulation of adenylate cyclase. In contrast with PGE1, PGF2 alpha strongly increased ornithine decarboxylase activity and DNA synthesis in the cells in a dose-related fashion at low concentrations (4-100 ng/ml), at which concentrations it had little effect on the intracellular cAMP or cGMP level and depressed ALP activity. Moreover, PGF2 alpha depressed the stimulatory effect of PGE1 on ALP activity but did not affect the elevation of cAMP level by PGE1. The accumulation of inositol phosphates was greatly increased by PGF2 alpha in the concentration range effective in stimulating DNA synthesis, but was increased little by PGE1, suggesting that PGF2 alpha is a potent stimulator of phosphatidyl inositol turnover in the cells. In addition, A23187, a Ca ionophore, alone did not influence the DNA synthesis, but the effects of tetradecanoyl phorbol acetate, a direct activator of protein kinase C, were very similar to those of PGF2 alpha. Moreover, the stimulation of DNA synthesis or the inhibition of ALP activity by PGF2 alpha was partially counteracted by H-7, a strong inhibitor of protein kinase C. These results suggest that PGF2 alpha stimulates the proliferation of osteoblastic cells predominantly through the phosphatidyl inositol turnover system following in part the activation of protein kinase C. Our data presented here indicate that PGE1 and PGF2 alpha are closely involved in the differentiation and proliferation, respectively, of osteoblasts in vitro and that their action may be mediated by second messengers which differ from each other.     

Stimulation of renin release from rabbit renal cortex by arachidonic acid and prostaglandin endoperoxides.

The mechanism by which renal prostaglandins stimulate renin secretion in vivo is unknown. In this in vitro study we measured the effects of activation of the prostaglandin (PG) system on renin release from slices of rabbit renal cortex. The PG precursor arachidonic acid (C20:4), a natural PG endoperoxide (PGG2), two stable synthetic PG endoperoxide analogues (EPA I and II), PGE2, PGF2alpha, and two different PG synthesis inhibitors [indomethacin and 5,8,11,14-eicosatetraynoic acid (ETA)] were used to evaluate the possibility of a direct action of the cortical PG system on renin secretion. Renin release increased significantly with time after addition of C20:4, PGG2, EPA I, and EPA II to the incubation medium. Stimulation of renin release was se-related for C20:4 in concentrations of 0.6 to 4.5 X 10(-6) M, for EPA I in concentrations of 0.7 to 2.8 X 10(-6) M, and for EPA II in concentrations of 1.4 to 14.0 X 10(-6) M. Indomethacin (10(-4) M) and ETA (10(-4) M) significantly decreased basal renin release as well as the renin release stimulated by C20:4 and EPA I. PGE2(10(-12) to 10(-6) M) had no effect on renin release, whereas PGF2alpha (10(-12) to 10(-6) M) decreased renin release in a dose-dependent manner. These data raise the possibility of a direct action of the renal cortical PG system on renin secretion. The results further indicate that stimulation of renin release by C20:4 may depend more specifically on the action of PG endoperoxides than on the primary prostaglandins.     

The effect of the antiprogestins RU 486 and ZK 98734 on the synthesis and metabolism of prostaglandins F2 alpha and E2 in separated cells from early human decidua

Enriched preparations of glandular and stromal cells were obtained from early human decidua and incubated for 24 h in the presence of two progesterone antagonists, RU 486 (17 beta-hydroxy-11 beta-[4-dimethylaminophenyl]17 alpha-[1-propynyl]-estra-4,9-dien-3-one) and ZK 98734 (17 beta-hydroxy-11 beta-4[4-dimethylaminophenyl]17 alpha-[3-hydroxy-1-propynyl]estra-4,9-dien-3-one) to determine the effect of the antiprogestins on the release of prostaglandin F2 alpha (PGF2 alpha) and PGE2 and their subsequent conversion to 15-keto-13,14-dihydro-PGF2 alpha and 15-keto-13,14-dihydro-PGE2. In the presence of exogenous arachidonic acid (AA, 30 microM), both steroids stimulated PGF2 alpha release by glandular, but not stromal, cells (P less than 0.001) and inhibited the metabolism of PGF2 alpha by the glandular fraction (P less than 0.005 and P less than 0.001 respectively). In the absence of exogenous AA, RU 486 and ZK 98734 stimulated the release of PGF2 alpha from glandular, but not stromal, cells (P less than 0.001 and P less than 0.005, respectively). Neither steroid altered the release or metabolism of PGE2 when the cells were incubated with AA, but both RU 486 and ZK 98734 increased the release of PGE2 by glandular, but not stromal, cells when incubated without AA (P less than 0.005 and P less than 0.001, respectively). Both steroids inhibited the metabolism of PGE2 under these conditions (P less than 0.05). These results suggest that 1) antiprogestins stimulate the synthesis of PGs by glandular cells in early human decidua, but do not alter the synthesis of PGs by stromal cells; 2) this stimulation of PG synthesis involves an effect on cyclooxygenase activity and is not a consequence of increased availability of endogenous AA; 3) the metabolism of PGs by glandular cells is altered by RU 486 and ZK 98734; 4) as RU 486 has greater antiglucocorticoid activity than ZK 98734, these results suggest that both steroids act on decidua by antagonizing endogenous progesterone rather than glucocorticoid activity.     

Prostaglandin secretion by the neocortex and fetal zones of the human fetal adrenal gland

Previously, we reported that the human fetal adrenal (HFA) gland secretes various prostaglandins (PGs) in vitro and that PG secretion is inhibited by endogenously synthesized glucocorticosteroids. In this investigation, the neocortex (NC) and fetal zone (FZ) of the HFA gland were separated by microdissection and maintained as tissue fragments in organ culture. The rate of PG secretion into the culture medium was determined by measuring various PGs using specific RIAs in media collected at 24-h intervals. During the first 24 h in culture, the secretion rates of PGF2 alpha and PGE2 were 6- and 7-fold greater by NC [14 +/- 5 and 9.9 +/- 3 ng mg protein-1 24 h-1 (mean +/- SE)], respectively, than by FZ tissue (2.5 and 1.4 ng mg protein-1 24 h-1). The secretion rates of PGFM and PGD2 were 2-fold greater in NC tissue than in FZ tissue, but the secretion rates of thromboxane B2 were similar in both zones of HFA tissue. In another study, the patterns of secretion of PGF2 alpha and PGE2 were determined as a function of days in culture. The secretion rates of PGF2 alpha and PGE2 fell rapidly in NC from 19.0 +/- 11 and 38.3 +/- 9.7 ng mg protein-1 24 h-1, respectively, to 1.3 +/- 7.2 and 4.8 +/- 3.3 by day 4. In contrast, the secretion rates of PGF2 alpha and PGE2 rose 8- and 3-fold in FZ tissue (from 0.7 +/- 0.2 and 0.9 +/- 0.6 ng mg protein-1 24 h-1, respectively, to 5.9 +/- 0.5 and 3.1 +/- 1.2 by day 4). The addition of ACTH or dexamethasone inhibited PG secretion in both zones, but to a greater degree in FZ tissue than in NC tissue. In summary, the NC secretes larger quantities of PG than the FZ, and the patterns of secretion are different in the two zones. The secretion of PGs is inhibited more in FZ than in NC tissue by ACTH and glucocorticosteroids.     

Role of prostaglandins in adrenal steroidogenesis

Using an in vitro rat glomerulosa and fasciculata cell preparation, we studied the role of porstaglandins (PGs) as potential mediators of angiotensin II-, ACTH-, and potassium-induced adrenal steroidogenesis. The glomerulosa cells primarily produced prostacyclin under basal and stimulated conditions, whereas the fasciculata cells primarily produce PGF2 alpha. Using 6-keto-PGF1 alpha (the stable hydrolysis product of prostacyclin) and PGF2 alpha as indices of PG production in glomerulosa and fasciculata cells, respectively, we noted no significant changes in PGs during angiotensin II-, ACTH-, or potassium-induced steroidogenesis. Pretreatment with indomethacin significantly (P less than 0.01) inhibited PG production, but did not alter adrenal steroidogenesis. Furthermore, the administration of exogenous PGs did not affect steroidogenesis. We conclude that PGs do not mediate steroidogenesis from rat glomerulosa or fasciculata cells.     

Role of catecholestrogens on ovarian prostaglandin secretion in vitro in the catfish Heteropneustes fossilis and possible mechanism of regulation

Seasonal, periovulatory and 2-hydroxyestradiol-17β (2-OHE(2))-induced changes on ovarian prostaglandin (PG) E(2) and F(2α) were investigated under in vivo or in vitro in the female catfish Heteropneustes fossilis. Both PGE(2) and PGF(2α) increased significantly during ovarian recrudescence with the peak levels in spawning phase. The PGs showed periovulatory changes with the peak levels at 16 h after the hCG treatment. Incubation of postvitellogenic ovary fragments with estradiol-17β (E(2)), 2-OHE(2) or 2-methoxyE(2) produced concentration-dependent increases in PG levels; 2-OHE(2) was more effective. In order to identify the receptor mechanism involved in the 2-OHE(2)-induced PG stimulation, the ovarian pieces were incubated with phentolamine (an α-adrenergic antagonist), propranolol (a β-adrenergic antagonist) or tamoxifen (an estrogen receptor blocker) alone or in combination with 2-OHE(2). The incubation of the tissues with the receptor blockers alone did not produce any significant effect on basal PG levels. However, co- and pre-incubation of the tissues with the blockers resulted in inhibition of the stimulatory effect of 2-OHE(2) on the PGs. Phentolamine was more effective than propranolol. The signal transduction pathway(s) involved in the 2-OHE(2)-induced PG secretion was investigated. The incubation of the ovarian pieces with 3-isobutyl-1-methylxanthine (IBMX, a phosphodiesterase inhibitor), chelerythrine (a protein kinase C inhibitor) and PD098059 (a mitogen-activated protein kinase inhibitor) significantly lowered the basal secretion of PGF(2α) and PGE(2). In contrast, H89 (a protein kinase A inhibitor) increased the basal secretion of PGs at 1 and 5 μM concentration and decreased it at 10 μM concentration. The co- or pre-incubation with IBMX, H89, chelerythrine and PD098059 significantly inhibited the stimulatory effect of 2-OHE(2) on PGF(2α) and PGE(2) levels. The inhibition was higher in the pre-incubation groups. Chelerythrine was the most effective followed by PD098059, IBMX and H89. The results suggest that 2-OHE(2) may employ both adrenergic and estrogen receptors, or a novel receptor mechanism having properties of both adrenergic and estrogen receptors.     

Renal prostaglandins and hypertension

A total of 154 patients with essential hypertension (EH), 24 patients with renovascular hypertension (RVH), and 130 Wistar rats were investigated. PGE2 and PGF2 alpha levels were assayed radioimmunologically in renal venous blood and urine of the patients, and the synthesis of PGE2 and PGF2 alpha in renal tissue, renal PGE-9-ketoreductase activity and urinary PG excretion were measured in rats. It was demonstrated that the PGE2 synthesis was depressed in the vascular channel and the renal uropoietic system, with elevated F/E rations, in patients with arterial hypertension. Clinical and experimental studies showed prolonged and excessive salt consumption to be a cause of these changes, rooted in suppressed renal biosynthesis of both PGs and increased conversion of PGE2 to PGF2 alpha. In addition, renal PGE2 inactivation was increased in EH patients, as compared to those with RVH. PGE2 produced in the kidneys of EH patients is always a depressor natriuretic substance, whereas the role of PGF2 alpha is dependent on the water-salt balance. Furosemide and, to a smaller extent, other diuretics, as well as some hypotensive agents, increase urinary PG excretion and depress the F/E ratio in the urine. Repeated PGE2 infusions are shown to enhance the sensitivity of EH patients to hypotensive drugs, so they can be used for the treatment of refractory EH cases.