Effects of piriprost (U-60, 257B) and leukotrienes on alkaline phosphatase activity of rat endometrial stromal cells in vitro.

The present study investigated the effects of piriprost (U-60,257B; an inhibitor of LT synthesis) and various LTs on alkaline phosphatase (ALP) activity of rat endometrial stromal cells in vitro. Mature ovariectomized rats were pretreated with hormones to sensitize their uteri for the decidual cell reaction. Endometrial stromal cells were isolated and cultured for up to 72 hr with various treatments. The ALP activity in all experiments was significantly (p less than 0.01) higher at 72 hr than at 24 hr, irrespective of treatment. We examined the effects of 100 microM piriprost, with or without 1 microM LTB4, 0.01 microM LTC4, 0.1 microM LTD4 or 0.001 microM LTE4 on ALP activity. At 72 hr, as indicated by analyses of variance, there were significant interactions (p less than 0.01) between the effects of piriprost and the LTs. Piriprost by itself increased (p less than 0.01) ALP activity in all experiments, and a further increase (p less than 0.01) in ALP activity was observed when either LTB4, LTC4, LTD4 or LTE4 was added with piriprost. LTB4, LTD4, or LTE4 alone had inhibitory effects (p less than 0.01) while LTC4 alone had no effect. These studies suggest LTs may be involved in decidualization which, in vitro, is accompanied by an increase in endometrial ALP activity. However the exact role of LTs is still unclear.     

Metabolic activity of cholesteryl esters in aortic smooth muscle cells is altered by prostaglandins I2 and E2

Among the biochemical processes associated with the atherogenic process are increased aortic cholesteryl ester (CE) accumulation and altered prostaglandin (PG) production. The precise physiological role of PG, particularly prostacyclin (PGI2), in the control of CE metabolism in intact aortic smooth muscle cells remains to be fully elucidated. We report here that cytosolic neutral cholesteryl ester hydrolytic activity (NCEH) in intact cultured aortic smooth muscle cells is significantly increased by 75-250 nM PGI2 at the end of a 2-hr incubation period. The effect was mediated by increased intracellular cAMP levels since the effect of PGI2 on NCEH activity was abolished in the presence of an inhibitor of adenylate cyclase activity, viz., dideoxyadenosine (DDA0. Although the addition of 20-100 microM dibutyryl cAMP (Bt2cAMP) and 50-100 microM sodium arachidonate also increased NCEH activity twofold, 6-keto PGF1 alpha, PGE1, and PGE2 did not increase the activity of this enzyme. In contrast to these findings, 75-250 nM PGE2 significantly inhibited CE synthetic activity (ACAT) approximately 60%. Arachidonate or Bt2cAMP did not affect ACAT activity. This decrease in ACAT activity induced by PGE2 does not appear to be mediated by cAMP. Taken together, these findings suggest that PGI2, a well known potent vasodilator and inhibitor of platelet aggregation, and PGE2 may have an important regulatory role in aortic CE metabolism.     

Cell-type specific responses in prostaglandin secretion by glandular and stromal cells from pig endometrium treated with catecholestrogens, methoxyestrogens and progesterone.

The pig conceptus and endometrium possess the ability to convert estrogens into catecholestrogens and catecholestrogens into methoxyestrogens. Experiments were carried out to evaluate the effect of catecholestrogens, methoxyestrogens and progesterone on the secretion of prostaglandin (PG) E and F2 alpha by porcine endometrial glandular and stromal cells in vitro. Both 2-hydroxyestradiol (2-OH-E2, 0-20 microM) and 4-hydroxyestradiol (4-OH-E2, 0-20 microM) increased (P less than .05) PGE and PGF2 alpha secretion by stromal cells in a dose response manner. Two-hydroxyestradiol tended (P less than .1) to decrease PGF2 alpha production by glandular cells. Two-methoxyestradiol (20 microM) suppressed (P less than .05) PGF2 alpha secretion by glandular and stromal cells. Four-methoxyestradiol (20 microM) stimulated (P less than .05) PGE production and PGE:PGF2 alpha ratio. Progesterone (.1 microM) suppressed (P less than .05) PG secretion in both cell types. We conclude that catecholestrogens, methoxyestrogens, and progesterone may participate in the establishment of pregnancy by modulating PG production in the endometrium.     

Purification of natural killer-like Kurloff cells and arachidonic acid metabolism.

Pulmonary and splenic Kurloff cells have been purified from estrogen-treated guinea pig. Enzymatic digestion of lung tissue and mechanical dispersion of cells yielded about 650 x 10(6) viable cells. After centrifugal elutriation and centrifugation on continuous Percoll gradient, a population of high-density (1,100 g/ml) pulmonary Kurloff cells were obtained with high viability (approximately 99%) and purity (approximately 99%). Splenic Kurloff cells have been isolated by disruption of spleen tissue and centrifugation on continuous Percoll gradient. High-density splenic Kurloff cells (150 x 10(6) cells per spleen) were also obtained with high purity (approximately 99%) and viability (approximately 99%). Pulmonary and splenic Kurloff cells were incubated with various concentrations of arachidonic acid (10, 30 and 100 microM) in the absence or presence of 2 microM ionophore A23187. With 10 microM arachidonic acid the relative production of cyclooxygenase products was the following: TxB2 greater than PGE2 approximately PGI2. For an arachidonic acid concentration superior to 10 microM, the profile of release was PGE2 much greater than TxB2 greater than PGI2. Arachidonic acid metabolism through the 5-lipoxygenase pathway was also studied by incubating pulmonary or splenic Kurloff cells with 10 microM arachidonic acid in the absence or presence of 2 microM ionophore A23187, or in some experiments, with 2.5 microM leukotriene A4. Reverse phase HPLC profiles clearly indicated that high-density Kurloff cells did not express 5-lipoxygenase activity. However, these cells showed the ability to convert exogenous leukotriene A4 into leukotriene B4 suggesting the presence of LTA4 hydrolase activity. These data have been confirmed by a sensitive RIA method. This study constitutes the first report on the purification of pulmonary Kurloff cells and on arachidonic acid metabolism by these cells. The possible implications of Kurloff cells in various biological events are discussed.     

Role of cyclic adenosine 3',5'-monophosphate in mediating the effect of prostaglandin E2 on decidualization in vitro.

When rat endometrial stromal cells from uteri sensitized for decidualization are cultured in vitro, there is an increase in alkaline phosphatase (ALP) activity paralleling that seen in vivo during decidualization. The addition of indomethacin to the culture medium decreases the endogenous production of prostaglandin E2 (PGE2) to below detectable levels and substantially reduces the increase in ALP activity. The addition of either PGE2 or its analog 16,16-dimethyl-PGE2, but not PGF2 alpha or its analog 15(S),15-methyl-PGF2 alpha, overrides this inhibitory effect, suggesting that PGE2 has a specific stimulatory effect upon ALP activity. This in vitro system was used to investigate the role of the cAMP pathway in mediating the stimulatory effect of PGE2 on ALP activity. The data indicate that PGE2 causes an increase in cAMP accumulation by the cells and that the addition of an analog of cAMP or substances which increase the level of cAMP in the cells (1-methyl-3-isobutyl xanthine, cholera toxin, forskolin) causes an increase in ALP activity. Collectively, the results suggest that the stimulatory effect of PGE2 is at least partially mediated by the cAMP pathway.     

Binding to prostaglandin (PG) receptors and activation of adenyl cyclase by 20-isopropylidene-PGS in rabbit platelet membranes

20-Isopropylidene-PGE1 (Isop-PGE1) was about 10 times more potent than PGE1 in inhibition of thrombin-induced aggregation of rabbit washed platelets. Likewise, 20-isopropylidene-17(R)-methyl-carbacyclin (CS-570), a stable PGI2 analogue, was more potent than carbacyclin in the anti-aggregatory activity. In order to define the platelet-prostaglandin interactions, a binding assay was done using platelet membranes with [3H]-PGE1 as a radioligand. Isop-PGE1 (IC50 = 0.18 microM) bound to the PG receptors more potently than PGE1 (IC50 = 2.1 microM). CS-570 (IC50 = 0.39 microM) was more potent than carbacyclin (IC50 = 1.9 microM). These indicate that introduction of an isopropylidene group to the carbon 20 of PGs increases the binding ability to the receptors. These PGE1 and PGI2 analogues activated platelet membrane adenyl cyclase and increased intracellular cAMP levels with the same potency series obtained in the binding experiments. All these results suggest that the binding to the receptors by these PGs is coupled to the activation of adenyl cyclase, followed by the increase in cAMP levels in platelets and the inhibition of platelet aggregation. Thus, the increased anti-aggregatory activity of 20-isop-PGs may be explained by their increased affinity for the PG receptors and stimulation of adenyl cyclase. 15-Epimeric-20-isopropylidene-PGE1 (15-Epi-isop-PGE1), which has an unnatural configuration of the 15-hydroxyl group, was much less potent than isop-PGE1 in the binding experiment and the other three investigations. This indicates that the configuration of the 15-hydroxyl group is important for the binding to the PG receptors and the consequent activities in platelets.     

Prostaglandin E and E2 alpha secretion by glandular and stromal cells of the pig endometrium in vitro: effects of estradiol-17 beta, progesterone, and day of pregnancy.

Prostaglandins (PGs) are believed to play important roles in the establishment of pregnancy. Glandular and stromal cells were isolated from pig endometrium on days 11 through 19 of pregnancy and cultured in the presence of estradiol-17 beta (E2) and progesterone (P4) to determine the effect of day of pregnancy and steroids on the secretion of PGE and PGF2 alpha. Estradiol at concentrations between .01 and 1 microM did not affect PGE and PGF2 alpha secretion into the medium by glandular and stromal cells. Progesterone (.1 microM) suppressed (P less than .001) PGE and PGF2 alpha production from both cell types. Glandular cells secreted more (P less than .01) PGF2 alpha than PGE, whereas stromal cells collected on days 11, 12, 13, and 19 secreted more (P less than .05) PGE than PGF2 alpha. Stromal cells isolated from tissues collected on day 13 of pregnancy produced PGs with higher (P less than .01) PGE:PGF2 alpha ratio than those from tissues harvested on other days of pregnancy. Glandular cells isolated from tissues collected on days 13 and 19 and stromal cells isolated from tissue collected on day 13 of pregnancy secreted more (P less than .05) PGE and PGF2 alpha than cells isolated on other days of pregnancy. We conclude that: 1) P4 has a suppressing effect on PG secretion; 2) endometrial glandular and stromal cells each produce a unique profile of PGs; and 3) endometrial cells harvested on different days of pregnancy secrete different amounts of PGE and PGF2 alpha.     

Rat mammary preadipocytes in culture produce a trophic agent for mammary epithelia-prostaglandin E2

Cell strains and cell lines rat mammary (Rama) 350-353 have been isolated from the slowly adherent stromal fraction of enzymatically digested rat mammary glands. Primary cultures of this fraction yield fat cels on extended culture. Their proportion can be increased with horse serum or growth hormone in the medium, and this increase is associated with a 100-fold or more increase in the release of radioimmunoassayable prostaglandins of the E type (PGE). The stromal cell strains and lines that are capable of yielding fat cells also secrete elevated levels (greater than 100 ng/mg/24 hr) of PGE; the fast-sticking epithelial fraction in primary cultures and the epithelial cell lines derived from it secrete 10-100 times less. Chromatography and radioisotopic labeling of the culture media from Rama 352 cells identify the PG as PGE2. PGE2 with insulin and hydrocortisone maximally stimulates [3H]DNA synthesis of epithelial cell lines and primary cultures from normal and tumorous glands by 2-4-fold at concentrations (10-20 ng/ml) well below those released by the preadipocytic stromal cells (20-100 ng/ml). Medium exposed to most cultured cells stimulates [3H]DNA synthesis of one epithelial cell line, Rama 25, by 2-4-fold. Prevention of the synthesis of PGE2 in Rama 352 cultures with indomethacin or flurbiprofen abolishes the mitogenic activity present in the culture medium, and the PG receptor antagonist polyphloretin phosphate inhibits completely the mitogenic activity for Rama 25 cells. Myoepithelial-like cell lines normally secrete moderate levels of PGE (10-100 ng/mg/24 hr) but the mitogenic activity for Rama 25 cells released from one such line, Rama 29, is not abolished by preventing the synthesis of PG's nor by PG-receptor antagonists.     

Factors regulating the production of prostaglandin E2 and prostacyclin (prostaglandin I2) in rat and human adipocytes.

The regulation of PGE2 (prostaglandin E2) and PGI2 (prostaglandin I2; prostacyclin) formation was investigated in isolated adipocytes. The formation of both PGs was stimulated by various lipolytic agents such as isoproterenol, adrenaline and dibutyryl cyclic AMP. During maximal stimulation the production of PGE2 and PGI2 (measured as 6-oxo-PGF1 alpha) was 0.51 +/- 0.04 and 1.21 +/- 0.09 ng/2 h per 10(6) cells respectively. Thus PGI2 was produced in excess of PGE2 in rat adipocytes. The production of the PGs was inhibited by indomethacin and acetylsalicylic acid in a concentration-dependent manner. The half-maximal effective concentration of indomethacin was 328 +/- 38 nM and that of acetylsalicylic acid was 38.5 +/- 5.3 microM. The PGs were maximally inhibited by 70-75% after incubation for 2 h. In contrast with their effect on PG production, the two agents had a small potentiating effect on the stimulated lipolysis (P less than 0.05). The phospholipase inhibitors mepacrine and chloroquine inhibited both PG production and triacylglycerol lipolysis and were therefore unable to indicate whether the PG precursor, arachidonic acid, originates from phospholipids or triacylglycerols in adipocytes. Angiotensin II significantly (P less than 0.05) stimulated both PGE2 and PGI2 production in rat adipocytes without affecting triacylglycerol lipolysis. Finally, it was shown that PGE2 and PGI2 were also produced in human adipocytes, although in smaller quantities than in rat adipocytes. It is concluded that the production of PGs in isolated adipocytes is regulated by various hormones. Moreover, at least two separate mechanisms for PG production may exist in adipocytes: (1) a mechanism that is activated concomitantly with triacylglycerol lipolysis (and cyclic AMP) and (2) an angiotensin II-sensitive, but lipolysis (and cyclic AMP)-independent mechanism.     

Stimulatory and inhibitory effects of prostaglandins on the gonadotropin-sensitive adenylate cyclase in the monkey corpus luteum

Detailed analysis of the action of prostaglandins (PGs) on the corpus luteum in primate species is very limited. In this study we examined the response of the adenylate cyclase system to PGs in homogenates prepared from the corpus luteum of rhesus monkeys at midluteal phase of the menstrual cycle. The conversion of [alpha 32p] ATP to [32p] cyclic AMP (cAMP) was assessed in the absence (control activity; 50 microM GTP) and presence of various concentrations of seven PGs and arachidonic acid, either alone or in combination with 250 nM hCG. Cyclic AMP production increased up to three-fold in the presence of PGD2, PGE2, PGI2 or PGF2 alpha; however PGA2, PGB2, 13, 14-dihydro-15-keto PGE2 and arachidonic acid alone did not alter cAMP levels. In dose-response studies, adenylate cyclase was 10 and 100-fold more sensitive to PGD2 (Vmax at 1 X 10(-5) M) than to PGE2 or to PGI2 and PGF2 alpha, respectively. Activity in the presence of hCG plus either PGD2, PGE2, PGI2 or PGF2 alpha did not differ from that for hCG (or the PG) alone. In contrast, addition of PGA2 or arachidonate inhibited (p less than 0.05) hCG-stimulated cAMP production by 50 and 100 percent. We conclude that the gonadotropin-sensitive adenylate cyclase of the macaque corpus luteum is also modulated by several PGs. These factors may either mimic (e.g., PGD2, PGE2, PGI2) or suppress (PGA2) gonadotropin-stimulated cAMP production and possibly cAMP-mediated events in luteal cells.     

Prostaglandin production in cultured gastric mucosal cells: role of cAMP on its modulation

The effect of cAMP on prostaglandin production may depend on cell types. To clarify the relationship between PG and cAMP, we examined arachidonate's effects on PG synthesis and intracellular cAMP accumulation in monolayers of rat gastric mucosal cells. These cells produced PGE2, PGI2 and thromboxaneA2 (TXA2) in amounts of 316 +/- 18, 100 +/- 7 and 30 +/- 5 pg per 10(5) cells in 10 min, respectively, in response to 10 microM arachidonic acid (AA). The production of these PG, however, leveled off subsequently. Cells initially exposed to AA responded poorly to a subsequent stimulation by AA. AA simultaneously stimulated intracellular cAMP accumulation; this stimulatory effect on cAMP production was abolished by the pretreatment with indomethacin. Nevertheless, the pretreatments with dibutyryl cAMP (0.1-5 mM) did not alter the amount of subsequent AA-induced PGE2 production. Furthermore, the preincubation with 1mM isobutyl methyl xanthine also failed to affect PGE2 synthesis, while it increased intracellular cAMP accumulation. Our studies suggest AA stimulates intracellular cAMP formation in cultured gastric mucosal cells, linked with conversion of AA to cyclooxygenase metabolites, AA-induced PG production is limited in these cells, and it seems, however, unlikely that intracellular cAMP modulates AA metabolism to PG.     

Prostacyclin production by rat aorta "in vitro" is increased by the combined action of dipyridamole plus pentoxifylline

The present study evaluates the effect of dipyridamole and pentoxifylline, individually and in combination, on PGI2-like production and arachidonic acid metabolism of rat aorta "in vitro". Pentoxifylline 100 microM and dipyridamole 92 and 184 microM increased PGI2-like activity, as measured by the platelet aggregation inhibitory capacity of the aortic ring incubates, by 71%, 46% and 60% respectively; a greater increase in PGI2-like activity was observed with the combination of the drugs than when they were used separately. This effect was observed even at the lowest doses assayed. In fact, dipyridamole 9.2 microM plus pentoxifylline 1 microM increased the PGI2-like activity by 30% while the individual increase was 4.5% and 10.6% respectively. To obtain more information on the effect of the dipyridamole-pentoxifylline combination on arachidonic acid metabolism, arteries were incubated with (1-14C) arachidonic acid, and the 6-keto-PGF1 alpha and PGE2 quantified. Dipyridamole 92 microM plus pentoxifylline 1 and 10 microM increased 6-keto-PGF1 alpha and PGE2 production by about 30% and 48% respectively while the combination with pentoxifylline 100 microM increased the 6-keto-PGF1 alpha 76.5% and the PGE2 50%. The possible biological effect and therapeutic implications of increased PGI2 production by the arteries due to the dipyridamole-pentoxifylline combination remains to be ascertained.     

The role of prostaglandins, cyclic nucleotides and tricarboxylic acids in the regulation of the human placental 20 alpha-hydroxysteroid dehydrogenase in vitro

The in vitro effect of non-steroidal regulators (prostaglandins, cyclic nucleotides and tricarboxylic acids) on the cytoplasmic 20 alpha-hydroxysteroid dehydrogenase (20 alpha-HSDH, EC 1.1.1.149) isolated from human early gestational and term placentas was investigated. When prostaglandins (PG) were tested at 100 microM concentrations, an inhibition of the human placental 20 alpha-HSDH by PGE1 (80% inhibition), PGE2 (70%), PGF1 alpha (40%) PGF2 alpha (30%) and 13,14-dihydro-15-keto-PGF2 (PGFM) (20%) was observed. This effect was shown to be dose-dependent. The I50 (concentration at 50% inhibition) was determined for PGE1 and PGE2 to be 11 microM and 38 microM, respectively. No effect on the activity of the 20 alpha-HSDH could be demonstrated for PGI2 and its stable metabolite 6-keto-PGF1 alpha, for the cyclic nucleotides (dbcAMP, dbcGMP) and for the tricarboxylic acids (citrate, ketoglutarate, lactate, malonate, pyruvate and succinate) when added to the incubation at 100 microM concentration. The 20 alpha-HSDH isolated from early gestational and term placentas did not respond differently to the substances tested. These results suggest that prostaglandins can have a direct, dose-dependent effect on the isolated human placental 20 alpha-HSDH without cyclic nucleotides as intermediates and thereby play a role in the regulation of human progesterone synthesis and metabolism during pregnancy and near term.     

Prostaglandin synthesis by the cochlea of the guinea pig. Influence of aspirin, gentamicin, and acoustic stimulation

This study describes the synthesis of prostaglandins (PGs) by the vascular structures of the inner ear (lateral wall = stria vascularis and spiral ligament) in vitro. The main PGs produced were PGI2, PGF2 alpha and PGE2. PGI2 and PGF2 alpha were also found in the perilymph. A 350 mg/kg ip injection of aspirin decreased PG synthesis by the lateral wall and PG levels in perilymph. This effect was reversed after 3 days. Gentamicin (10(-9) to 10(-5) M) decreased significantly and reversibly PG synthesis in vitro, as did 100 mg/kg ip injection. Acoustic stimulation increased ex vivo PGI2 and PGE2 synthesis without modifying PG levels in perilymph. Results suggest that PGs could be one humoral mediator of the cochlear microcirculation homeostasis, and, possibly, of the circulatory disturbances reported after acoustic stimulation. The decreased PG synthesis after gentamicin treatment could account for the angiotoxic component observed in aminoglycoside ototoxicity.     

Human follicular dendritic cells interact with T cells via expression and regulation of cyclooxygenases and prostaglandin E and I synthases

PGE(2) inhibits mature T cell proliferation and protects T cells from activation-induced cell death (AICD). We have previously demonstrated that human follicular dendritic cells (FDC) strongly express PGI synthase. In this study, the hypothesis that FDC have regulatory roles on germinal center T cells by controlling production of PGE(2) and PGI(2) was tested. Confocal microscopic analyses of human tonsil tissues revealed that FDC indeed expressed PGE synthase in addition to PGIS. To confirm these results, we studied the regulation mechanism of PG production in FDC, using an established human FDC-like cell line, HK. Specifically in response to TNF-alpha, TGF-beta, and LPS, protein expression of cyclooxygenase (COX)-2 and downstream PGE synthase was up-regulated with coordinate kinetics, whereas COX-1 and PGIS were constitutively expressed. The increase of these enzymes was reflected in actual production of PGE(2) and PGI(2). Interestingly, IL-4 almost completely abrogated the stimulatory activity of TNF-alpha, TGF-beta, and LPS in PG production. Furthermore, the up-regulation of PGE(2) and PGI(2) production was markedly down-regulated by indomethacin and a selective COX-2 inhibitor. PGI(2) analog and PGE(2) inhibited proliferation and AICD of T cells in dose- and time-dependent manners. Finally, coculture experiments revealed that HK cells indeed inhibit proliferation and AICD of T cells. Put together, these results show an unrecognized pathway of FDC and T cell interactions and differential mechanisms for PGE(2) and PGI(2) production, suggesting an important implication for development and use of anti-inflammatory drugs.     

Endogenous synthesis of prostaglandins E1 and I2 in 3T3 fibroblasts transformed by polyoma virus. Effects on adenosine 3':5'-monophosphate production

Prostaglandins (PG)E1, E2 and I2 were produced by polyoma virus transformed (py) 3T3 fibroblasts. The levels of PGE1, PGE2 and 6-keto-PGF1 alpha (degradation product of PGI2) were 22.7, 225 and 33.2 ng/ml medium, respectively, 72 h after medium change. The stimulatory potencies of exogenous PGE1, PGE2 and PGI2 on adenosine 3':5'-monophosphate (cyclic AMP) formation were similar. Therefore, the prostaglandin mediated increase in cyclic AMP levels observed during growth of these cells (Claesson, H.-E., Lindgren, J.A. and Hammarstr?m, S. (1977) Eur. J. Biochem. 74, 13) is largely (greater than 80%) mediated by PGE2 and to lesser extents by PGE1 and PGI2.     

Gamma interferon inhibits basal and interleukin 1-induced prostaglandin production and bone resorption in neonatal mouse calvaria

Production of the osteolytic arachidonic acid metabolites, prostaglandin (PG) E2, PGI2 and PGF2 alpha, by neonatal mouse calvariae was quantitated by gas chromatography/mass spectrometry. Mouse recombinant interleukin 1 (rIL-1) raised medium levels of PGE2 and PGI2 (measured as 6-keto-PGF1 alpha) in the dose range tested (1.0-10.0 U/ml culture medium), while an effect on PGF2 was only observed at 10 U/ml. Bone resorption in response to rIL-1 reached a plateau at 3.0 U/ml. Mouse recombinant gamma-interferon (rIFN-gamma) between 100-500 U/ml suppressed basal PG synthesis and spontaneous resorption of cultured bone. In addition, IFN-gamma at 100 U/ml prevented stimulation of PG synthesis by 3.0 U/ml rIL-1 and thereby reduced the bone resorbing activity of the cytokine by at least 60%. 5 X 10(-7) M indomethacin was equally effective in suppression of PG synthesis and bone resorption. The present study provides evidence that IFN-gamma inhibits PG synthesis and consequently resorption of cultured bone.     

Conceptus-derived prostaglandins regulate endometrial function in sheep

In sheep, the trophectoderm of the elongating conceptus secretes interferon tau (IFNT) and prostaglandins (PGE2, PGF2alpha, PGI2). The PGs are derived from PG synthase 2 (PTGS2), and inhibition of PTGS2 in utero prevents conceptus elongation. IFNT increases expression of many genes in the endometrial epithelia that regulate conceptus elongation. This study tested the hypothesis that PGs secreted by the conceptus regulate endometrial functions that govern conceptus elongation. Cyclic ewes received intrauterine infusions of control vehicle or early pregnancy levels of IFNT, PGE2, PGF2alpha, or PGI2 from Days 10-14 postestrus. Expression levels of endometrial GRP, IGFBP1, and LGALS15, whose products stimulate trophectoderm cell migration and attachment, were increased by PGE2, PGI2, and IFNT. All PGs and IFNT increased expression of the HEXB protease gene, but only IFNT increased the CST6 protease inhibitor gene. Differential effects of PGs were observed for expression of the CTSL protease gene and its inhibitor, CST3. IFNT, PGF2alpha, and PGI2 increased ANGPTL3 expression, but only IFNT and PGE2 increased HIF1A expression, both of which regulate angiogenesis. For glucose transporters, IFNT and all PGs increased SLC2A1 expression, but only PGs increased SLC2A5 expression, whereas endometrial SLC2A12 and SLC5A1 expression levels were increased by IFNT, PGE2, and PGF2alpha. Infusions of all PGs and IFNT increased the amino acid transporter SLC1A5, but only IFNT increased SLC7A2 expression. In the uterine lumen, only IFNT increased glucose levels, and only PGE2 and PGF2alpha increased total amino acids. These results indicate that PGs and IFNT from the conceptus coordinately regulate endometrial functions important for growth and development of the conceptus during the peri-implantation period of pregnancy.     

Interactions of prostaglandins with subpopulations of ovine luteal cells. I. Stimulatory effects of prostaglandins E1, E2 and I2

Preparations of small and large steroidogenic cells from enzymatically dispersed ovine corpora lutea were utilized to study the in vitro effects of luteinizing hormone (LH) and prostaglandins (PG) E1, E2 and I2. Cells were allowed to attach to culture dishes overnight and were incubated with either LH (100 ng/ml), PGE1, PGE2, or PGI2 (250 ng/ml each). The secretion of progesterone by large cells was stimulated by all prostaglandins tested (P less than 0.05) while the moderate stimulation observed after LH treatment was attributable to contamination of the large cell population with small cells. Prostaglandins E1 and E2 had no effect on progesterone secretion by small cells, while LH was stimulatory at all times (0.5 to 4 hr) and PGI2 was stimulatory by 4 hr. Additional studies were conducted to determine if the effects of PGE2 upon steroidogenesis in large cells were correlated with stimulated activity of adenylate cyclase. In both plated and suspended cells PGE2 caused an increase (P less than 0.05) in the rate of progesterone secretion but had no effect upon the activity of adenylate cyclase or cAMP concentrations within cells or in the incubation media. Exposure of luteal cells to forskolin, a nonhormonal stimulator of adenylate cyclase, resulted in marked increases in all parameters of cyclase activity but had no effect on progesterone secretion. These data suggest that the actions of prostaglandins E1, E2 and I2 are directed primarily toward the large cells of the ovine corpus luteum and cast doubt upon the role of adenylate cyclase as the sole intermediary in regulation of progesterone secretion in this cell type.     

Arachidonic acid metabolism in thrombocytes and vascular tissues of turkeys

Turkeys are hypertensive compared to mammals of similar size. In vitro synthesis of thrombocyte thromboxane B2 (TxB2), 12L-hydroxy-5,8,10 heptadecatrienoic acid (HHT), 12L-hydroxy-5,8,10,14-eicosatetraenoic acid (HETE) and aortic prostaglandin (PG) production was studied in one to ten month old domestic white turkeys. Compared to normal human platelets, TxB2 production was increased (55.4 vs. 31.4%) and HETE production was markedly reduced (6.5 vs. 34.6%) in control thrombocytes. Similar to human platelets in which cyclooxygenase inhibition with aspirin results in an increase in HETE production, block of the thrombocyte enzyme with aspirin doubled the production of HETE. In vitro conversion of radiolabeled arachidonic acid (AA) showed that the primary PG produced by turkey aorta was PGE2. A 6-keto immunoreactive PG was present which comigrated with authentic 6-keto PGF1 alpha, but failure of the aortic supernatant to inhibit adenosine diphosphate or AA induced platelet aggregation suggested that PGI2 was not produced. The vasodepressor potency of PGE1, PGE2 and PGI2 was altered in awake turkeys with PGE1 and PGE2 having five times the hypotensive effect as PGI2. In addition, conversion of AA to PGE2 by aorta in one month turkeys was greater (17.3 vs. 9.2%) than in ten month old turkeys. Systemic arterial pressure was increased in the ten month old turkeys (188 mmHg) compared to one month old turkeys (143 mmHg). Thus, both vascular AA metabolism and the vasodepressor potencies of PGE2 and PGI2 are altered and the activity of the lipoxygenase pathway in thrombocytes is limited in the turkey.