PGE2 Alpha Research Articles

Effects of oestrogen on progesterone synthesis and arachidonic acid metabolism in human luteal cells.

Locally produced oestrogens and prostaglandins (PGs) are implicated in the regulation of luteal lifespan in the human ovary. This study (1) assesses direct effects of these factors on progesterone synthesis in isolated luteal cells, and (2) explores interactions between luteal age and treatment with gonadotrophin or oestrogen on the metabolism of arachidonic acid (prostaglandin precursor) by steroidogenic luteal cells in vitro. Primary monolayer cultures of human luteal cells obtained at different stages of the luteal phase were used to investigate the effect of oestradiol, catechol oestrogens (2- and 4-hydroxyoestradiol), diethylstilboestrol, PGE2 and PGF2 alpha on basal and human chorionic gonadotrophin (hCG) stimulated progesterone production in vitro. The role of PGs as modulators of luteal cell function was further investigated by studying the metabolic fate of radioactively labelled arachidonic acid in hormone treated (oestradiol and hCG) and control cultures, assessed by high performance liquid chromatography. Corpora lutea were enucleated from nine women with regular ovulatory cycles undergoing microsurgical reversal of tubal sterilization. Granulosa cell aspirates were obtained from three patients undergoing in-vitro fertilization treatment. PGE2 and PGF2 alpha at various concentrations did not have a consistent effect, whereas oestradiol, diethylstilboestrol (and 2-hydroxyoestradiol in early luteal cell cultures) significantly inhibited basal and hCG stimulated progesterone biosynthesis. Evidence for direct inhibition of 3 beta-hydroxysteroid dehydrogenase enzymic activity by oestradiol was obtained. Both major metabolic pathways of arachidonic acid (lipoxygenase and cyclo-oxygenase) were operative in steroidogenic luteal cells recovered throughout the luteal phase. The ratio of PGE2 to PGF2 alpha synthesis in vitro by human luteal cells from endogenously incorporated arachidonic acid did not change significantly with corpus luteum age, with PGE2 tending to predominate. Oestradiol treatment shifted arachidonic acid metabolism from the lipoxygenase towards the cyclooxygenase pathway in cells isolated from ageing corpora lutea. Oestradiol, at relatively high concentrations, is a potent inhibitor of basal and hCG induced luteal cell steroidogenesis in vitro. No support is provided for the concept that luteolysis is mediated by local production of PGF2 alpha. The putative luteolytic effect of oestradiol may entail reduced metabolism of arachidonic acid to lipoxygenase derived products by luteal cells rather than direct stimulation of prostaglandin production by itself.

Effect of cytokines and prostaglandins on the growth of chick retinal pigment epithelial cells.

The effect of cytokines and prostaglandins on the growth of chick retinal pigment epithelial (RPE) cells was studied in vitro. The growth of RPE cells was evaluated by [3H]-thymidine uptake. Human recombinant interleukin-1 beta, interferon-beta and tumor necrosis factor-alpha stimulated the growth of RPE cells at 50-200 units/ml. Prostaglandins such as PGE1, PGE2 and PGF2 alpha suppressed RPE cell growth at 10(-5) approximately (-6)M. Indomethacin, an inhibitor of cyclooxygenase, did not affect the spontaneous RPE cell growth at 10(-7) approximately (-9) M. However, 10(-7) M indomethacin enhanced RPE cell growth stimulated by cytokines synergistically. The addition of 10(-6) M PGF2 alpha suppressed the enhanced growth of RPE cells which was induced with cytokines and indomethacin. Furthermore, RPE cells stimulated with cytokines produced PGE2. These results suggest that RPE cells produce prostaglandins which have a negative regulatory role in RPE cell growth.

Prostaglandin E2 enhances human endometrial stromal cell differentiation.

Endometrial stromal differentiation (decidualization) is essential for implantation of the developing blastocyst. Because prostaglandins (PGs) are synthesized in the endometrium, and PG-binding sites have been demonstrated in the proliferative endometrial stromal cell (the precursor of the decidual cell), experiments were performed to determine whether PGs are involved in the process of decidualization. Human endometrial stromal cells were cultured for 18 days in Dulbeco's Modified Eagle's Medium-2% fetal bovine serum with 1 microM medroxyprogesterone acetate (MPA) and 10 nM estradiol, with and without PGE2 or PGF2 alpha. Expression of PRL was used as a marker of decidualization. In the presence of estradiol and MPA alone (control), PRL was detected beginning on day 9 and gradually increased through day 18. In contrast, PRL was detected on day 3 in the PGE2-treated cells, and the magnitude of stimulation in these cells on days 9-12 was 1300-1400% of that in control cells. Furthermore, the PRL mRNA content of the PGE2-treated cells on day 12 was 4.6-fold greater than that in the control cells. The effect of PGE2 on PRL production was dose dependent, with a minimal effective dose of 10(-10) M. PGE2 in the absence of steroids had a minimal effect on PRL production. In contrast to PGE2, PGF2 alpha treatment had no effect on PRL expression in steroid-treated cells. These results indicate that there are synergistic effects among PGE2, estradiol, and MPA, resulting in acceleration of endometrial stromal cell differentiation and enhanced PRL expression.

Effect of the choline acetyltransferase inhibitor (2-benzoylethyl)- trimethylammonium iodide (BETA) on human placental prostaglandin release and phospholipase A 2 activity

We investigated the effect of an inhibitor of acetylcholine (ACh) synthesis, (2-benzoylethyl)trimethylammonium iodide (BETA), on prostaglandin (PG)E2 and PGF2 alpha release from incubated placental explants in the presence or absence of ACh or arachidonic acid (AA). BETA alone (100 microM) significantly reduced both PGE2 and PGF2 alpha release. However, this inhibitory effect of BETA was not reversed in the presence of ACh (10 microM to 1 mM). The addition of AA (10 to 100 microM) increased both PGE2 and PGF2 alpha release, and simultaneously overcame the inhibition of PGE2 but not PGF2 alpha release by BETA. These results are compatible with the hypothesis that BETA may be inhibiting the phospholipase A2 (PLA2) step rather than prostaglandin H synthase (PGHS) step in the enzymatic pathway of PG generation. This hypothesis was supported by evidence showing a lack of effect of BETA (10 and 100 microM) on ovine placental microsome PGHS activity. Moreover, human placental homogenate PLA2 activity was reversibly inhibited by BETA (100 microM). In the presence of BETA (100 microM), addition of exogenous ACh (100 microM) had no significant effect on placental PLA2 activity. These results indicate that the inhibitory effect of BETA on placental PG release was unlikely to be via an action on ACh synthesis, but rather via a reversible effect on PLA2 activity.

Locally produced angiotensin II induces ovulation by stimulating prostaglandin production in in vitro perfused rabbit ovaries.

The present study was undertaken to investigate the role of exogenous and endogenous angiotensin II (Ang II) in ovarian steroidogenesis and production of prostaglandin (PG) in in vitro perfused rabbit ovaries. The addition of 100 or 10 micrograms Ang II at 2-h intervals to the perfusate did not stimulate progesterone production, but significantly stimulated estradiol (E2) production by perfused rabbit ovaries. When the specific antagonist of Ang II, saralasin at 2 x 10(-6) M, was added to the perfusate 30 min before the onset of Ang II administration, Ang II-stimulated production of E2 was significantly blocked. Ang II also significantly stimulated both PGE2 and PGF2 alpha production, while the addition of saralasin to the perfusate significantly inhibited the Ang II-stimulated production of PG. The levels of PGs in ovaries perfused with saralasin plus 100 micrograms Ang II did not differ significantly from those in control ovaries perfused with medium alone. Exposure to human CG (hCG) significantly stimulated production of progesterone and E2 by perfused rabbit ovaries, while the concomitant administration of 2 x 10(-6) M saralasin significantly reduced only E2 production. Addition of saralasin to the perfusate inhibited hCG-stimulated PG production in a dose-dependent manner. The ovulatory efficiency in ovaries treated with hCG alone or hCG plus saralasin was significantly correlated with PG production by perfused rabbit ovaries at 12 h after exposure to hCG. The production of PG stimulated by Ang II was completely reduced by indomethacin treatment during the entire perfusion period. Indomethacin completely blocked Ang II-induced ovulation, but not Ang II-stimulated oocyte maturation. Concurrent administration of staurosporine, a protein kinase C inhibitor, at 10(-6) M significantly inhibited Ang II-stimulated meiotic maturation of ovulated ova and follicular oocytes. In conclusion, these results indicate that Ang II has a direct role in ovarian production of E2 and PG. An intrinsic renin-angiotensin system in the rabbit ovary may act as an intermediary of gonadotropin-stimulated PG production. Locally produced Ang II may induce ovulation in the rabbit ovary, at least in part, by stimulating PG production.

Locally produced angiotensin II induces ovulation by stimulating prostaglandin production in in vitro perfused rabbit ovaries

The present study was undertaken to investigate the role of exogenous and endogenous angiotensin II (Ang II) in ovarian steroidogenesis and production of prostaglandin (PG) in in vitro perfused rabbit ovaries. The addition of 100 or 10 micrograms Ang II at 2-h intervals to the perfusate did not stimulate progesterone production, but significantly stimulated estradiol (E2) production by perfused rabbit ovaries. When the specific antagonist of Ang II, saralasin at 2 x 10(-6) M, was added to the perfusate 30 min before the onset of Ang II administration, Ang II-stimulated production of E2 was significantly blocked. Ang II also significantly stimulated both PGE2 and PGF2 alpha production, while the addition of saralasin to the perfusate significantly inhibited the Ang II-stimulated production of PG. The levels of PGs in ovaries perfused with saralasin plus 100 micrograms Ang II did not differ significantly from those in control ovaries perfused with medium alone. Exposure to human CG (hCG) significantly stimulated production of progesterone and E2 by perfused rabbit ovaries, while the concomitant administration of 2 x 10(-6) M saralasin significantly reduced only E2 production. Addition of saralasin to the perfusate inhibited hCG-stimulated PG production in a dose-dependent manner. The ovulatory efficiency in ovaries treated with hCG alone or hCG plus saralasin was significantly correlated with PG production by perfused rabbit ovaries at 12 h after exposure to hCG. The production of PG stimulated by Ang II was completely reduced by indomethacin treatment during the entire perfusion period. Indomethacin completely blocked Ang II-induced ovulation, but not Ang II-stimulated oocyte maturation. Concurrent administration of staurosporine, a protein kinase C inhibitor, at 10(-6) M significantly inhibited Ang II-stimulated meiotic maturation of ovulated ova and follicular oocytes. In conclusion, these results indicate that Ang II has a direct role in ovarian production of E2 and PG. An intrinsic renin-angiotensin system in the rabbit ovary may act as an intermediary of gonadotropin-stimulated PG production. Locally produced Ang II may induce ovulation in the rabbit ovary, at least in part, by stimulating PG production.

Prostaglandin secretion by guinea pig tracheal epithelial cells caused by eosinophil major basic protein

We examined the effect of eosinophil major basic protein (MBP) on prostaglandin (PG) secretion from guinea pig tracheal epithelial (GPTE) cells. Primary cultures of GPTE cells were incubated with 10(-6) M MBP for up to 6 h and then stimulated with 10(-6) M bradykinin (BK). PGE2, 6-ketoprostaglandin F1 alpha (PGF1 alpha), PGF2 alpha, and thromboxane B2 (TxB2) concentrations in media were determined by enzyme-linked immunoabsorbent assay (EIA). Incubation with MBP for 6 h caused secretion of both PGE2 (17,614 +/- 4,416 vs. 1,426 +/- 555 pg/10(6) cells at baseline, P < 0.001, n = 7) and PGF2 alpha (20,303 +/- 5,724 vs. 3,790 +/- 1.075 pg/10(6) cells at baseline, P < 0.002, n = 7). Secretion of PGE2 and PGF2 alpha stimulated by MBP required at least 2 h. Incubation with MBP for 6 h also augmented the subsequent response to BK: PGE2 secretion was 29,215 +/- 6,853 vs. 3,445 +/- 1,041 pg/10(6) cells for BK alone (P < 0.0001), and PGF2 alpha secretion was 25,407 +/- 6,237 vs. 5,213 +/- 1,535 pg/10(6) cells for BK alone (P < 0.0001). MBP did not change 6-keto-PGF1 alpha and TxB2 secretion. Incubation of GPTE cells from seven animals with polylysine, a protein with mass and ion charge similar to MBP, for 2 h, both caused secretion of PGE2 (8,579 +/- 3,244 vs. 788 +/- 419 pg/10(6) cells at baseline, P < 0.01) and augmented the response to BK (12,732 +/- 4,788 vs. 1,653 +/- 680 pg/10(6) cells after BK alone, P < 0.005).(ABSTRACT TRUNCATED AT 250 WORDS)

Does endogenous peripheral arginine vasopressin have a role in the febrile responses of conscious rabbits?

1. The actions of peripheral arginine vasopressin (AVP) on the febrile responses of conscious rabbits induced by peripherally administered polyinosinic:polycytidylic acid (poly(I).poly(C)) have been studied using an AVP V1 receptor antagonist ([deamino-Pen1, O-Me-Tyr2, Arg8]-vasopressin). 2. Temperature responses were monitored continuously using rectal thermistor probes. Test substances were administered intravenously (i.v.). Blood samples were taken at timed intervals from a marginal ear vein and plasma PGE2 and PGF2 alpha levels determined by radioimmunoassay. 3. Poly(I).poly(C) (2.5 micrograms/kg) stimulated a reproducible biphasic rise in body temperature with a lag phase of 45-60 min and peaks at 90 and 225 min. The febrile response was accompanied by a 5-fold rise in circulating immunoreactive (ir) PGE2, which peaked after 90 min and remained elevated up to 300 min. Poly(I).poly(C) also stimulated a 2.5-fold rise in circulating irPGF2 alpha, which peaked after 150 min and was followed by a return to basal levels after 300 min. 4. The overall magnitude of the febrile response to poly(I).poly(C) (2.5 micrograms/kg, i.v.) was significantly antagonized by the AVP V1 receptor antagonist (250 micrograms/kg, i.v.) administered 5 min prior to the pyrogen. 5. The irPGE2 response to poly(I).poly(C) (2.5 micrograms/kg, i.v.) was significantly antagonized by the AVP V1 receptor antagonist (250 micrograms/kg, i.v.) administered 5 min prior to the pyrogen. The irPGF2 alpha response was only reduced at the peak 150 min time point measurement. 6. In conclusion, these results show a modulatory role for a peripherally administered AVP V1 antagonist in the febrile responses to poly(I).poly(C), suggesting a possible propyretic role for endogenous peripheral AVP. This modulatory role appears to be mediated via actions on prostaglandin E2.

Interleukin-1 beta, -1 alpha, and -6 and prostaglandins in vaginal/cervical fluids of pregnant women before and during labor.

Interleukin-1 beta (IL-1 beta) is not detected in the amniotic fluid of normal human pregnancies before the initiation of parturition, but during labor, both at term and preterm, this cytokine is present in the amniotic fluid of 25-40% of pregnancies. A critical question, however, is whether this finding is indicative of a role for IL-1 beta (directly or indirectly) in the initiation of parturition or is the result of IL-1 beta formation and entry into amniotic fluid as a natural sequela of normal labor. The forebag of the amniotic sac is formed during labor in response to cervical dilatation, and on the decidual surface, the tissues of this structure become exposed and bathed by vaginal fluids as the cervix opens. Microorganisms and bacterial toxins are present in vaginal fluid before labor begins; these agents should act upon the exposed tissues of the forebag to cause inflammation and evoke an inflammatory response. This study was conducted to examine the likelihood that the inflammatory mediators found in amniotic fluid in increased amounts at parturition are produced in forebag tissues after the onset of labor because of obliged inflammation in these tissues. Vaginal/cervical fluids were collected by lavage from nonpregnant women and from pregnant women at term before and during labor. The amount of immunoreactive IL-1 beta in vaginal/cervical fluids of pregnant women during labor (mean +/- SEM, 91.5 +/- 16.9 ng; n = 17) was significantly greater (P < 0.001) than that in fluids collected before labor (7.8 +/- 3 ng; n = 14). The in vivo rate of IL-1 beta secretion directly from the decidua lining the forebag during labor was brisk (1.71 +/- 0.88 ng/cm2.min; n = 4), consistent with previous observations of higher levels of pro-IL-1 beta mRNA in decidual tissues adherent to the forebag compared with those in decidua adherent to chorion laeve of the upper compartment of the amnionic sac. The vaginal fluid content of prostaglandins (PGs) during labor [PGE2, 82.1 +/- 16.4 ng; PGF2 alpha, 141.5 +/- 30.5 ng; PGFM, 35.2 +/- 5.8 ng (mean +/- SEM; n = 13)] was significantly greater for PGE2 and PGF2 alpha (P < 0.05 and 0.004, respectively) than that before labor (PGE2, 42.6 +/- 9.4 ng; PGF2 alpha, 35.3 +/- 8.5 ng; PGFM, 21.7 +/- 4.6 ng; n = 12). In addition, there was a significant increase in the ratio of PGF2 alpha to PGE2 (P < 0.03) in vaginal fluids during labor.(ABSTRACT TRUNCATED AT 250 WORDS)

Prostaglandins stimulate the secretion of inhibin from human placental cells.

In this study, the effects of prostaglandins (PGs) on inhibin secretion were investigated in primary culture of human placental cells. Isolated trophoblast cells were cultured for 2-5 days with PGE1, PGE2, 6-keto-PGF1 alpha, PGF2 alpha, and thromboxane B2. Inhibin levels in the culture medium were measured by immunoenzymatic assay. PGs significantly stimulated inhibin secretion in the cell culture. The addition of PGs (1 nM to 10 microM) to the culture resulted in dose-dependent increases in inhibin levels in the medium. At a dose of 10 microM, inhibin levels in the medium were increased by 35.2-172.5% compared to the control value. Compared with the efficacies of these PGs, PGE2 and PGF2 alpha enhanced inhibin secretion more potently than other tested PGs. A close correlation between the effects of PGs and the number of trophoblasts seeded in the culture was observed, with an optimal response of the cells to PGs at 1.0-2.0 x 10(6) cells/well. Time-course studies showed that a significant increment in inhibin levels in the PG-treated culture occurred after 48-120 h, but not during the first 24 h, of the culture, indicating a possible involvement of PGs in inhibin biosynthesis in the cells. Epidermal growth factor (EGF) could enhance PG-stimulated inhibin releases in the cell culture. Treatment with EGF (100 ng/mL) increased inhibin in the medium by 83.3% (ED50) in the presence of PGE2 or by 70.3% (ED50) in the presence of PGF2 alpha compared to the control values. The addition of 8-bromo-cAMP (100 microM) and cholera toxin (1000 ng/mL) to the culture also enhanced basal and PG-induced inhibin secretion, but the addition of the 4-bromo-calcium ionophore A23187 (1 microM) did not alter inhibin releases, suggesting that the stimulatory effects of PGs on inhibin secretion in placental cells may be mediated through the cAMP pathway. In the cell cultures treated with PGE and PGF2 alpha, there was no change in cell growth or intracellular DNA content, as measured by 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide assay and DNA determination with fluorescence spectrophotometry. Morphological studies showed that PGE2 alpha (1 microM), alone or combined with EGF (100 ng/mL), significantly accelerated the process of differentiation from cytotrophoblasts to syncytiotrophoblasts, indicating that the actions of PGs may be related to their effects on syncytial transformation, rather than cellular proliferation.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of elevated glucose on cyclic GMP and eicosanoids produced by porcine aortic endothelium.

The short-term effects of elevated glucose on cyclic GMP (cGMP) and eicosanoid production in pig aortic endothelial cell monolayers was determined by incubating cells in 5.5 mM or 44 mM glucose for 6 hours. Bradykinin- or A23187-stimulated cGMP production was significantly reduced in cells incubated in 44 mM glucose compared with 5.5 mM glucose. Stimulation of cGMP levels with exogenously added nitric oxide (NO) was also decreased to a similar extent in cells exposed to 44 mM glucose. These data suggest that NO production stimulated by bradykinin or A23187 was unchanged by elevated glucose. Assayed eicosanoids, including 6-ketoprostaglandin (PG) F1 alpha, PGE2 alpha, and 15(S)-hydroxy-(5Z, 8Z, 11Z, 13E)-eicosatetraenoic acid, stimulated by bradykinin or A23187, were increased in cells exposed to 44 mM glucose. These eicosanoid products formed from exogenously added arachidonic acid did not differ between cells incubated in 5.5 mM or 44 mM glucose. Hyperosmolar concentrations of mannose or sucrose had no effect on cGMP levels but did mimic the effect of elevated glucose on eicosanoid production. These data suggest that hyperglycemia in diabetes may interfere with NO-induced guanylate cyclase activation but not NO production in the endothelium and that increased phospholipase activity, secondary to hyperosmolarity, may account for elevated eicosanoid levels.

Renal vasodilator activity of 5,6-epoxyeicosatrienoic acid depends upon conversion by cyclooxygenase and release of prostaglandins

The 5,6-epoxyeicosatrienoic acid (5,6-EET), a renal vasodilator metabolite of arachidonic acid via cytochrome P450 (P450) requires cyclooxygenase for expression of its vasoactivity as the responses are inhibited by indomethacin and other aspirin-like drugs. We now report on the metabolism of 5,6-EET by rabbit kidneys in order to characterize those metabolites that may account for its vasoactivity. The 5,6-EET was injected close-arterially into the rabbit isolated Krebs-Henseleit perfused kidney, preconstricted with phenylephrine, and the effluent collected throughout the response period. Basal collections, following injection of 100 microliters of vehicle, were made at 20-min intervals before each 5,6-EET injection. Prior to acidic extraction, deuterated 6-keto-prostaglandin (PG) F1 alpha and PGE2 were added as internal standards. The extracts were separated by TLC and prostaglandins were derivatized for gas chromatography-mass spectrometry analysis using a negative ion chemical ionization mode. Injection of 0.5, 1, 5, 10, and 20 micrograms of 5,6-EET (n = 4) resulted in dose-related decreases in perfusion pressure of 6 +/- 2, 12 +/- 4, 21 +/- 4, 26 +/- 4, and 27 +/- 7 mm Hg, respectively. Basal perfusates contained 6-keto-PGF1 alpha and PGE2, levels of which were increased by 2-fold or more by 5,6-EET. Perfusates, collected during 5,6-EET administration, also contained 5-hydroxy-PGI1 and 5,6-epoxy-PGE1, cyclooxygenase metabolites of 5,6-EET. This is the first report of the recovery and identification of these 5,6-EET metabolites from an intact organ. Since the responses to 5,6-EET are endothelial-dependent, we also studied the profile of eicosanoids formed following incubation of 5,6-EET with cultured bovine pulmonary endothelial cells. Endothelial cells metabolized 5,6-EET to products with a similar radioactive profile on reverse-phase high pressure liquid chromatography compared to kidney perfusates. We compared the vasodilator activity of 5,6-epoxy-PGE1 and 5-hydroxy-PGI1, chemically synthesized by us from PGE2 and PGF2 alpha, respectively, with PGE2 and PGI2 in the rabbit kidney. The 5,6-epoxy-PGE1 was equipotent to PGE2 as a vasodilator. The ED50 values for 5,6-EET, 5,6-epoxy-PGE1, and PGE2 were 4.69, 0.43, and 0.42 nmol, respectively. Although PGI2 was a potent vasodilator (ED50, 0.24 nmol), 5-hydroxy-PGI1 was devoid of activity. Thus, the cyclooxygenase-dependent vasoactivity of 5,6-EET in the rabbit kidney has two components: release of vasodilator prostaglandins, PGE2 and PGI2, and metabolism of 5,6-EET to a prostaglandin analog, 5,6-epoxy-PGE1.

Dietary lipids influence insulin action.

Insulin binding and insulin responsiveness are altered by dietary fat-induced changes in the fatty acid composition of the adipocyte plasma membrane. Feeding a high P/S diet increased polyunsaturated fatty acid content of major membrane phospholipids of adipocyte plasma membrane in normal and diabetic animals, increased membrane linoleic acid content, and prevented a decrease in arachidonic acid level in diabetic animals. The high P/S diet increased insulin binding in control animals. Animals fed the high P/S diet had significantly higher rates of insulin-stimulated glucose transport and lipogenesis than did animals fed the low P/S diet. Feeding a high P/S diet significantly increased the amount of glucose transported when expressed as a function of the specific amount of insulin bound. To determine if dietary fat-induced alterations in the fatty acid composition of skeletal muscle lipid alter insulin-dependent and basal muscle metabolism, contralateral epitrochlearis and extensor digitorum longus muscles were isolated and incubated in vitro. High levels of dietary omega-3 fatty acids reduced PGE2 and PGF2 alpha synthesis in extensor digitorum longus and epitrochlearis muscle. Insulin increased glucose and amino acid transport; the increase in glucose transport by insulin was significantly greater after consumption of the high omega-3 fatty acid diet. Rats fed high levels of omega-3 fatty acids showed reduced net protein degradation in the presence and absence of insulin due to decreased rates of protein degradation and synthesis. These experiments indicate that high levels of dietary omega-3 fatty acids alter muscle membrane composition, glucose transport, and metabolism of muscle protein. To determine if dietary fatty acids alter the onset of diabetes and insulin binding to liver nuclei in spontaneously diabetic rats, weanling rats were fed chow or semipurified diets containing 20% (w/w) fat of either high or low P/S ratio. Feeding a high P/S diet increased insulin binding to liver nuclei of control and diabetic animals. Although diet did not alter the onset of diabetes, insulin binding to liver nuclei is higher in animals at the onset of diabetes than in highly diabetic animals. Eight-week-old female C57 B 6J lean and ob/ob mice were fed semipurified diets containing 20% (w/w) fat of either high or low P/S ratio to investigate the effect of diet on specific binding of insulin to liver nuclei. Insulin binding was highest in nuclei from lean mice fed a high P/S diet. Specific binding of insulin to nuclei from obese mice was also increased by the high P/S diet, but to a lesser extent.(ABSTRACT TRUNCATED AT 250 WORDS)

The influence of growth factors on the prostaglandin synthesis in cultured rabbit luteal cells.

Isolated rabbit luteal cells from day 12 of pseudopregnancy were cultured with a variety of growth factors, hCG and arachidonic acid (AA) in serum-free culture medium for 96 hours. The time course of prostaglandin (PG) E2, PGF2 alpha, and progesterone was determined. TNF-alpha, IL-2, IGF-I, IGF-II, and EGF alone were not able to change significantly the production of either progesterone or PGE2 and PGF2 alpha when compared with the controls. AA as the precursor of PG biosynthesis increased the production of PGE2 and PGF2 alpha significantly (1.2-fold and 2.6-fold, respectively; p < 0.05; p < 0.01). AA together with EGF increased the level of PG more pronounced when compared with AA alone (1.5-fold for PGE2, 5-fold for PGF2 alpha; p < 0.05 and p < 0.005). The progesterone production of the cells remained unchanged during growth factor exposure. We conclude that EGF together with high amounts of AA is able to stimulate the PG biosynthesis in cultured rabbit luteal cells. Since the level of PGF2 alpha increases more pronounced than PGE2, an activation of the PGE2 9-ketoreductase may be possible. Therefore, EGF could be relevant as an additional regression factor controlling the PGE2/PGF2 alpha-ratio in the corpus luteum of rabbits. This mechanism seems to be independent from the progesterone synthesis.

Respiratory and cardiovascular effects of inhaled and intravenous bradykinin, PGE2, and PGF2 alpha in dogs.

Prostaglandins (PGs) and bradykinin act as potent respiratory irritants in both normal and asthmatic subjects, but their sites of action are unknown. We compared the cardiorespiratory effects of bradykinin, PGE2, and PGF2 alpha nebulized into the isolated "in situ" larynx, inhaled into the tracheobronchial tree, and injected intravenously in anesthetized spontaneously breathing dogs. Laryngeal administration only resulted in a brief burst of rapid shallow breaths produced by bradykinin (1,000 micrograms/ml) in one of five dogs. Tracheobronchial administration of bradykinin (1,000 micrograms/ml) increased breathing rate and tidal volume (VT) in four of seven dogs without changing cardiovascular parameters, whereas PGE2 (500 micrograms/ml) caused similar effects in two of six dogs. Lower concentrations of both agents were essentially without effect. PGF2 alpha (50-500 micrograms/ml) inhaled into the lower airway increased breathing rate, reduced VT, and caused a concentration-dependent bronchoconstriction that was significantly reduced by atropine. Inhaled PGF2 alpha only slightly increased arterial blood pressure (5.8 +/- 2.8%) and heart rate (12.0 +/- 6.4%). Intravenous PGF2 alpha (5 micrograms/kg) increased upper and lower airway resistances, which were accompanied by a decrease in breathing rate and VT, hypertension, and bradycardia. Bradykinin (1 micrograms/kg) and PGE2 (1 and 3 micrograms/kg) produced apnea followed by rapid shallow breathing, bradycardia, and hypotension. These results indicate that the tracheobronchial tree is considerably more responsive to aerosolized bradykinin, PGE2, and PGF2 alpha than the laryngeal region. Moreover, the stronger effects produced by intravascular administration suggest a greater accessibility of rapidly adapting stretch receptors and C-fiber endings from the vascular bed than from the airway lumen.

The accumulation of prostaglandins (PG) in amniotic fluid is an aftereffect of labor and not indicative of a role for PGE2 or PGF2 alpha in the initiation of human parturition.

The accumulation of prostaglandins (PGs) in amniotic fluid (AF) during labor is cited frequently as one line of evidence in support of a role for these eicosanoids in the initiation of human parturition. In this study, we evaluated an alternate possibility, viz. that PGs entering AF at parturition are produced as a sequela of labor-associated processes. During labor, the AF normally becomes separated into two compartments, viz. the forebag and the upper compartment, by the obstruction produced as the descending fetal presenting part is engaged in the maternal pelvis. We theorized that the PGs that enter AF are produced in traumatized tissues lining the forebag, which is formed as the result of labor-driven cervical dilatation. In addition, these traumatized tissues are exposed to and bathed by the vaginal fluid, which contains many potent stimuli of PG formation, viz. large numbers of microorganisms and bacterial toxins. AF was collected at term before labor (n = 50) and from the upper compartment during labor (n = 47) by transuterine amniocentesis, and AF was collected by direct needle aspiration of the forebag during labor (n = 143). PGF2 alpha, 13,14-dihydro-15-keto-PGF2 alpha (PGFM), and PGE2 were quantified by RIA. The concentrations (nanomoles per L mean +/- SEM) of PGs in AF of the forebag (PGF2 alpha, 85.6 +/- 10.6; PGFM, 20.8 +/- 2.58; PGE2, 26.9 +/- 2.73) were much greater than those in the AF before labor (PGF2 alpha, 0.56 +/- 0.05; PGFM, 0.9 +/- 0.08; PGE2, 5.89 +/- 1.13) or in AF of the upper compartment during labor (PGF2 alpha, 7.14 +/- 1.64; PGFM, 5.11 +/- 0.82; PGE2, 8.74 +/- 1.71). The concentrations of PGs in AF of the upper compartment during early labor (< or = 2.5-cm cervical dilatation) were no greater than those in AF before labor began. The concentration and total content of PGs in AF of the forebag increased as a function of cervical dilatation until delivery. At 3- to 5-cm cervical dilatation, the levels of PGs in AF of the upper compartment were greater than those before labor, but significantly less than those in AF of the forebag at the same stage of labor progress. After 3-5 cm, the levels of PGs in the upper compartment did not increase further. These findings indicate that PGF2 alpha, PGFM, and PGE2, which enter AF in increased amounts during parturition, are produced during, not before, labor in tissues (principally decidua) lining for forebag.(ABSTRACT TRUNCATED AT 250 WORDS)

Possible roles for prostaglandins E2 and F2 in seasonal changes in ovarian steroidogenesis in the frog (Rana esculenta)

Concentrations of PGE2, PGF2 alpha, androgens and oestradiol in plasma, and ovary weights were measured in the female frog, Rana esculenta, during the annual breeding cycle. Experiments were carried out in vivo to study the effects of PGE2 and PGF2 alpha on plasma sex steroids during the following stages: pre-reproduction (April), reproduction (May), post-reproduction (June) and recovery (October). Experiments were performed in vitro during these stages to evaluate the effects of these two prostaglandins on the secretion of ovarian steroids. Concentrations of PGE2 were low in plasma during winter hibernation, the reproduction and post-reproduction stages, whereas they were high during the pre-reproduction and recovery stages. PGE2 treatment in vivo increased androgen secretion in April, whereas PGF2 alpha treatment increased oestradiol secretion in June and October. In experiments in vitro, PGE2 increased androgen secretion and decreased oestradiol secretion from ovaries collected in April, whereas PGF2 alpha increased oestradiol secretion from ovaries collected in October. These results suggest that a seasonal increase in plasma PGE2 may inhibit breeding activity, probably by stimulating ovarian androgen secretion, whereas, as previously reported, a seasonal increase in plasma PGF2 alpha may inhibit breeding, by stimulating ovarian oestradiol secretion.

The accumulation of prostaglandins (PG) in amniotic fluid is an aftereffect of labor and not indicative of a role for PGE2 or PGF2 alpha in the initiation of human parturition

The accumulation of prostaglandins (PG) in amniotic fluid is an aftereffect of labor and not indicative of a role for PGE2 or PGF2 alpha in the initiation of human parturition

Products of cyclooxygenase mediate the responses of the guinea pig trachea to hydrogen peroxide.

The role of products of cyclooxygenase was investigated in the responses of isolated airways to H2O2. Strips of guinea pig trachea, in some of which the epithelium had been removed mechanically, were suspended in organ chambers, and isometric tension was recorded. Under basal conditions, H2O2 induced indomethacin-sensitive contractions, which were larger in preparations without than in those with epithelium; the difference was abolished by inhibitors of thromboxane synthase or thromboxane A2 receptors. During contractions to acetylcholine, low concentrations of H2O2 induced relaxation in preparations with but had no significant effect in those without epithelium. At higher concentrations of H2O2, the epithelium-dependent relaxation was attenuated but an epithelium-independent relaxation appeared. The epithelium-dependent but not the epithelium-independent responses to H2O2 were blocked by indomethacin. Under basal conditions, prostaglandin E2 (PGE2; < or = 10(-7) M), U-46619, prostaglandin PGF2 alpha (PGF2 alpha), prostaglandin PGD2 (PGD2), and prostacyclin (PGI2) caused contractions. During contractions to acetylcholine, PGE2 induced larger relaxations in preparations with than in those without epithelium. Radioimmunoassay revealed that lower concentrations of H2O2 predominantly increased the release of PGE2 and 6-ketoprostaglandin F1 alpha (6-keto-PGF1 alpha); in preparations without epithelium, the release of thromboxane B2 was augmented also. At higher concentrations of H2O2, the release of PGE2, PGF2 alpha, PGD2, 6-keto-PGF1 alpha, and thromboxane B2 increased in preparations with and without epithelium. These findings demonstrate that the responses of the guinea pig trachea to H2O2 are mediated mainly by products of cyclooxygenase and that the effects of H2O2 are modulated by the epithelium.

Prevention of postasphyxial increase in lipid peroxides and retinal function deterioration in the newborn pig by inhibition of cyclooxygenase activity and free radical generation.

Free radicals have been implicated in the development of injury to the immature retina. Asphyxia increases free radicals as well as prostaglandins (PG) in neural tissues. We assessed whether in the retina the cyclooxygenase pathway contributes to free radical formation after oxidative insults such as asphyxia, which in turn disrupts retinal function. Newborn pigs were treated with either saline, ibuprofen (194 mumol/kg i.v.), or allopurinol (1 mmol/kg i.v.), and retinal malondialdehyde (MDA), hydroperoxides, PGE2 and PGF2 alpha levels, and the amplitudes and implicit times of the a- and b-waves of the full-field electroretinogram were measured before and 1 h after a 5-min period of asphyxia. In saline-treated animals, asphyxia caused a marked increase (p < 0.01) in MDA, hydroperoxides, PGE2, and PGF2 alpha concentrations in the retina. This was associated with a significant decrease (p < 0.01) in the b-wave amplitude measured under scotopic and photopic conditions and an increase in the b-wave implicit times. Ibuprofen and another cyclooxygenase inhibitor, indomethacin (28 mumol/kg i.v.), decreased PGE2 and PGF2 alpha levels and prevented the increase in MDA and hydroperoxides after asphyxia. Allopurinol maintained low concentrations of MDA and hydroperoxides after asphyxia. Both ibuprofen and allopurinol prevented the postasphyxial changes in the b-wave amplitude and diminished the delay in implicit time observed after asphyxia in saline-treated pigs. Our findings suggest that in the retina after asphyxia free radicals appear to originate primarily from the cyclooxygenase pathway and contribute to the deterioration in retinal electrophysiologic function of the newborn animal. Cyclooxygenase inhibitors, like free radical scavengers, may protect retinal function from deteriorating after oxidative stresses.