The Future of Neurocritical Care Research: Proceedings and Recommendations from the Fifth Neurocritical Care Research Network Conference.

The Fifth Neurocritical Care Research Network (NCRN) Conference held in Boca Raton, Florida, in September of 2018 was devoted to challenging the current status quo and examining the role of the Neurocritical Care Society (NCS) in driving the science and research of neurocritical care. The aim of this in-person meeting was to set the agenda for the NCS's Neurocritical Care Research Central, which is the overall research arm of the society. Prior to the meeting, all 103 participants received educational content (book and seminar) on the 'Blue Ocean Strategy®,' a concept from the business world which aims to identify undiscovered and uncontested market space, and to brainstorm innovative ideas and methods with which to address current challenges in neurocritical care research. Three five-member working groups met at least four times by teleconference prior to the in-person meeting to prepare answers to a set of questions using the Blue Ocean Strategy concept as a platform. At the Fifth NCRN Conference, these groups presented to a five-member jury and all attendees for open discussion. The jury then developed a set of recommendations for NCS to consider in order to move neurocritical care research forward. We have summarized the topics discussed at the conference and put forward recommendations for the future direction of the NCRN and neurocritical care research in general.

An acquired distaste: sugar discrimination by the larval parasitoid Microplitis croceipes (Hymenoptera: Braconidae) is affected by prior sugar exposure.

We examined gustatory responses of the larval parasitoid Microplitis croceipes to determine whether the adults discriminate among common sugars, including fructose, glucose, maltose and sucrose, found in plants. When given single sugar solutions of sucrose, glucose, fructose and maltose at concentrations of 0.008-2.0 mol l(-1), the estimated concentrations at which 50% of wasps initiated feeding ranged between 0.054 and 0.085 mol l(-1) for sucrose, glucose and fructose, which was significantly lower than for maltose. Wasps showed a strong decrease in feeding time for maltose or fructose following a brief exposure to other sugars, suggesting that wasps can distinguish maltose and fructose from the other sugars tested. The higher acceptance threshold and short feeding time in the case of maltose appears adaptive in light of the relatively poor nutritional quality of the sugar in the longevity trial. The pronounced feeding inhibition seen for fructose following exposure to other sugars is not linked with lower nutritional performance. This feeding inhibition was even seen in wasps that had fed on glucose at the lowest acceptance threshold (0.031 mol l(-1)) and persisted for 24 h. This study is the first to show feeding inhibition of otherwise phagostimulant sugars such as maltose and fructose after gustatory stimulation on other sugars.

The application of epiphenotyping approaches to DNA methylation array studies of the human placenta.

BACKGROUND: Genome-wide DNA methylation (DNAme) profiling of the placenta with Illumina Infinium Methylation bead arrays is often used to explore the connections between in utero exposures, placental pathology, and fetal development. However, many technical and biological factors can lead to signals of DNAme variation between samples and between cohorts, and understanding and accounting for these factors is essential to ensure meaningful and replicable data analysis. Recently, "epiphenotyping" approaches have been developed whereby DNAme data can be used to impute information about phenotypic variables such as gestational age, sex, cell composition, and ancestry. These epiphenotypes offer avenues to compare phenotypic data across cohorts, and to understand how phenotypic variables relate to DNAme variability. However, the relationships between placental epiphenotyping variables and other technical and biological variables, and their application to downstream epigenome analyses, have not been well studied. RESULTS: Using DNAme data from 204 placentas across three cohorts, we applied the PlaNET R package to estimate epiphenotypes gestational age, ancestry, and cell composition in these samples. PlaNET ancestry estimates were highly correlated with independent polymorphic ancestry-informative markers, and epigenetic gestational age, on average, was estimated within 4 days of reported gestational age, underscoring the accuracy of these tools. Cell composition estimates varied both within and between cohorts, as well as over very long placental processing times. Interestingly, the ratio of cytotrophoblast to syncytiotrophoblast proportion decreased with increasing gestational age, and differed slightly by both maternal ethnicity (lower in white vs. non-white) and genetic ancestry (lower in higher probability European ancestry). The cohort of origin and cytotrophoblast proportion were the largest drivers of DNAme variation in this dataset, based on their associations with the first principal component. CONCLUSIONS: This work confirms that cohort, array (technical) batch, cell type proportion, self-reported ethnicity, genetic ancestry, and biological sex are important variables to consider in any analyses of Illumina DNAme data. We further demonstrate the specific utility of epiphenotyping tools developed for use with placental DNAme data, and show that these variables (i) provide an independent check of clinically obtained data and (ii) provide a robust approach to compare variables across different datasets. Finally, we present a general framework for the processing and analysis of placental DNAme data, integrating the epiphenotype variables discussed here.

Neurocritical care nursing research priorities.

The science of nursing has long been discussed as a blending of the art and science of caring, and nursing research builds the evidence of support for nursing practice. Nurses and nursing care are key to successful neurocritical care research endeavors. Ideally nursing care should be evidence based and supported by solid research. The goal of nursing research is to expand the knowledge of caring for patients. Within the scope of nursing research, the priorities for research in neurocritical care should support this goal. In this manuscript, we discuss what we believe are the priorities of neurocritical care nursing research, the obstacles, and some possible solutions.

Ecology. Giant pandas in a changing landscape.

The giant panda has been restricted to several disjunct montane forest populations, and habitat loss and fragmentation are the primary threats to its survival. For pandas to survive, conservation efforts must focus on larger landscapes rather than individual nature reserves. China recently initiated several policies, including the Natural Forest Conservation Program and Grain-to-Green Policy, which provide a historic opportunity to integrate panda conservation into national policies. Simultaneously, China is promoting the Western China Development Program, which calls for substantial infrastructure and hydropower development and economic investments. Integrating panda conservation into these development policies will be a critical challenge.

Growth hormone-dependent changes in the rat lung proteome during alveorization.

Growth hormone (GH) mRNA and protein have recently been demonstrated in the rat lung throughout the period of alveolarization (day 4-14 postnatally). The functional significance of this finding was therefore assessed, by determining the effects of GH mRNA knockout using aerosolized antisense oligodeoxynucleotides (ODN) directed against the GH gene. In a preliminary experiment, the effectiveness of the antisense GH ODN was demonstrated in a lung Type II epithelial cell line (L2 cells), in which constitutive GH mRNA expression was completely abolished by GH ODN transfection. Administration of the aerosolized GH ODN to 4-day-old rats for 10 days was accompanied by a widespread presence of its delivery liposomes within lung cells. Aerosolized GH ODN treatment decreased lung concentrations of IGF (insulin-like growth factor)-1 and increased concentrations of albumin, calcyclin binding protein, superoxide dismutase, RNA binding protein motif 3, and the alpha- and beta-subunits of ATP synthase and electron transfer flavoprotein. At least 32 other proteins (identified by 2D gel electrophoresis) were also significantly affected by the antisense GH ODN treatment. By changing the lung proteome, these results indicate hitherto unsuspected autocrine/paracrine actions of GH in developmental lung function.

Recent Canadian efforts to develop population-level pregnancy intervention studies to mitigate effects of natural disasters and other tragedies.

The preconception, pregnancy and immediate postpartum and newborn periods are times for mothers and their offspring when they are especially vulnerable to major stressors - those that are sudden and unexpected and those that are chronic. Their adverse effects can transcend generations. Stressors can include natural disasters or political stressors such as conflict and/or migration. Considerable evidence has accumulated demonstrating the adverse effects of natural disasters on pregnancy outcomes and developmental trajectories. However, beyond tracking outcomes, the time has arrived for gathering more information related to identifying mechanisms, predicting risk and developing stress-reducing and resilience-building interventions to improve outcomes. Further, we need to learn how to encapsulate both the quantitative and qualitative information available and share it with communities and authorities to mitigate the adverse developmental effects of future disasters, conflicts and migrations. This article briefly reviews prenatal maternal stress and identifies three contemporary situations (wildfire in Fort McMurray, Alberta, Canada; hurricane Harvey in Houston, USA and transgenerational and migrant stress in Pforzheim, Germany) where current studies are being established by Canadian investigators to test an intervention. The experiences from these efforts are related along with attempts to involve communities in the studies and share the new knowledge to plan for future disasters or tragedies.

Decidual activation: abundance and localization of prostaglandin F2alpha receptor (FP) mRNA and protein and uterine activation proteins in human decidua at preterm birth and term birth.

OBJECTIVE: To study how the decidua contributes to parturition, we examined prostaglandin F(2alpha) concentrations as well as prostaglandin 15-hydroxy dehydrogenase, prostaglandin F(2alpha) receptor, matrix metalloproteinases 2 and 9, oxytocin receptor, prostaglandin-H synthase-2, and the prostaglandin E(2) receptor expression in human decidua. MATERIALS AND METHODS: Decidual samples were isolated from placentas collected from patients at preterm not in labor (PTNIL), preterm labor (PTL), term not in labor (TNIL), and term labor (TL). For immunohistochemistry, fresh membranes which included chorion, amnion and decidua from term patients were collected. RESULTS: Prostaglandin F(2alpha) receptor mRNA was low in all preterm patients and then significantly increased towards term (p=0.049). Prostaglandin F(2alpha) receptor protein was identified in the amnion epithelium and mesoderm, chorion trophoblasts and decidua by immunohistochemistry, and levels were highest at TNIL (p=0.007) as measured by western blot. Prostaglandin F(2alpha) levels were higher at PTNIL than TNIL. Matrix metalloproteinases 2 and 9 protein and pro-enzyme activities were higher at TL than TNIL. There were no significant changes among the groups for any of the other factors measured. CONCLUSIONS: These results suggest that the induction of Prostaglandin F(2alpha) receptor at term may facilitate the decidua contribution to parturition, and its regulation and role should be examined further.

Changes in matrix metalloproteinase (MMP)-2 and MMP-9 in the fetal amnion and chorion during gestation and at term and preterm labor.

Increased matrix metalloproteinase (MMP)-9 proteolytic activity is associated with term birth, preterm birth and premature rupture of membranes. However, most studies show no changes with MMP-2, which binds tightly to cell and matrix proteins. We hypothesized better protein extraction would reveal new MMP patterns. Human amnion and chorion were collected from 25 patients at preterm or term, extracted with 2% SDS (a high concentration), and the MMP protein levels and pro-enzyme activities were determined by Western immunoblotting and zymography. MMP-2 protein and MMP-2 and -9 pro-enzyme activities in the amnion increased significantly (p<0.05) with labor at term, and were higher than at preterm labor (p<0.05), when extracted with high SDS concentration. There were no changes in chorion MMPs under any condition. These associations suggest MMP-2 may be another regulator of membrane rupture and other labor-associated mechanisms at term parturition, and its role(s) should be examined further.

Production of prostaglandins by fetal rat lung type II pneumonocytes and fibroblasts.

The output of prostaglandins I2, E2, F2 alpha and 13,14-dihydro-15-keto-PGF2 alpha (PGFM) from third passage day 20 rat fetal fibroblasts and type II alveolar pneumonocytes was studied. In 2 h incubations, the output levels for each cell type were: PGI2 greater than PGE2 much greater than PGF2 alpha = PGFM when cells were incubated with Ca2+ ionophore A23187 (10 microM) or arachidonic acid (1 microgram/ml).

Glucocorticoid stimulation of amnion cell prostaglandin synthesis: suppression by protein kinase C inhibitors and independence of phorbol ester-sensitive protein kinase C.

Glucocorticoids stimulate the prostaglandin E2 production of confluent amnion cell cultures, but have no stimulatory effect on the PGE2 output of freshly isolated human amnion cells. Since protein phosphorylation may modify the responsiveness of target cells to steroids, and activators of protein kinase C (PKC), as well as corticosteroids, promote amnion cell PGE2 output by stimulating the synthesis of prostaglandin endoperoxide H synthase (PGHS), we investigated the possibility that PKC is involved in the glucocorticoid-induction of PGE2 synthesis in cultured amnion cells. The dexamethasone-induced PGE2 output of arachidonate-stimulated cells was blocked by the protein kinase inhibitors staurosporine, K-252a, H7, HA1004, and sphinganine, in a manner consistent with their effect on PKC. However, dexamethasone increased the PGE2 production of cultures treated with maximally effective concentrations of the PKC-activator compound TPA. Moreover, dexamethasone stimulated PGE2 synthesis in cultures which were desensitized to TPA-stimulation by prolonged phorbol ester treatment. Concentration-dependence studies showed that staurosporine completely (greater than 95%) blocked glucocorticoid-provoked PGE2 synthesis at concentrations which did not inhibit TPA-stimulated prostaglandin output, and that K-252a inhibited the effect of TPA by more than 95% at concentrations which decreased the effect of dexamethasone only moderately (approximately 40%). Dibutyryl cyclic AMP had no influence on the basal- or dexamethasone-stimulated PGE2 production, and on the staurosporine inhibition of the steroid effect. These results show that glucocorticoids and phorbol esters control amnion PGE2 production by separate regulatory mechanisms. It is suggested that the response of human amnion cells to glucocorticoids is modulated by protein kinase(s) other than phorbol ester-sensitive PKC and cyclic AMP-dependent protein kinase.

Response of fetal rat lung fibroblasts to elevated oxygen concentrations after liposome-mediated augmentation of antioxidant enzymes.

Cultured fetal rat lung fibroblasts are growth-inhibited and have increased lactate dehydrogenase release and prostaglandin synthesis in response to 50% and 95% oxygen exposure. Extended exposure to 50% oxygen, but not to 95% oxygen, was associated with tolerance to the cytotoxic effects of oxygen. Pretreatment of fibroblasts with liposome-encapsulated superoxide dismutase and catalase also conferred protection against the cytotoxic effects of 50% and 95% oxygen. Exogenous enhancement of intracellular superoxide dismutase and catalase specific activities did not attenuate the growth inhibition or increased prostaglandin synthesis associated with exposure to 50% or 95% oxygen. The growth inhibition could not be attributed to an autocrine prostaglandin effect, since inhibition of prostaglandin synthesis with 50 microM ibuprofen did not prevent O2-mediated inhibition of DNA synthesis.

Regulation of prostaglandin H2 synthase-2 expression in primary human amnion cells by tyrosine kinase dependent mechanisms.

Prostaglandin H2 synthase (PGHS)-1 and PGHS-2 expression was examined in primary cultures of human amnion cells, an in vitro model of amnion tissue. Epidermal growth factor (EGF), the protein kinase C (PKC) activating phorbol ester TPA, and the protein phosphatase inhibitor, okadaic acid (OA), stimulated PGHS activity and the level of PGHS-2 mRNA, but did not affect the level of PGHS-1 mRNA. In situ hybridization suggested that the same population of cells responded to EGF, TPA and OA. Okadaic acid promoted PGHS activity independently of PKC. EGF stimulated the activity of extracellular signal-regulated protein kinase (Erk) and N-terminal c-Jun kinase (Jnk). OA increased Jnk activity but had no effect on Erk activity, while TPA had no influence on either Erk or Jnk activity. PD098059, a selective inhibitor of the Erk-activating kinase MEK, blocked the stimulation of PGHS expression by EGF, but did not decrease stimulation in response to OA. Herbimycin A, a tyrosine kinase inhibitor, suppressed the stimulation of PGHS activity and PGHS-2 mRNA abundance by all three stimulants, and blocked signalling via the Erk and Jnk mitogen-activated protein kinase pathways. Thus, growth factor stimulation, PKC activation and protein phosphatase inhibition induced the expression of PGHS-2 in primary amnion cells by distinct regulatory mechanisms involving tyrosine kinase(s). Tyrosine kinase inhibitors may constitute a new category of PGHS-2 inhibitors that act by blocking the expression of the enzyme.

Glucocorticoids stimulate the expression of prostaglandin endoperoxide H synthase-2 in amnion cells.

Corticosteroids increase the production of prostaglandin E2 (PGE2) and the activity of prostaglandin endoperoxide H synthase (PGHS) in cultured amnion cells, although they inhibit prostanoid biosynthesis in numerous other cell types. This suggests that glucocorticoids control the level of PGHS in amnion cells by a hitherto unexplored, positive regulatory mechanism. We have tested the possibility that corticosteroids act by stimulating the expression of messenger RNAs (mRNAs) encoding one or both isoforms of PGHS. Ribonuclease protection assays were used to determine the levels of PGHS-1 and -2 mRNAs and, for reference, gamma-actin mRNA levels in confluent primary cultures of human amnion cells. In untreated cultures, PGHS-1 and -2 mRNA levels were low, often not reaching the level of detection. Dexamethasone (DEX) treatment for 4 h resulted in a measurable level of PGHS-2 mRNA, which increased further 10-fold and 20-fold after incubation with the glucocorticoid for 8 h and 16 h, respectively. The stimulation was dependent on DEX concentration, and was concomitant with an increase in the capacity of the cells to metabolize arachidonic acid to PGE2. PGHS-1 mRNA levels remained low in DEX-treated cells, while the gamma-actin message level showed no change. Estradiol and progesterone had no influence on PGHS-2 mRNA expression, but cortisol increased the PGHS-2 mRNA abundance. The glucocorticoid antagonist RU486 blocked the effect of DEX. Conditioned media of DEX-treated cells did not contain steroid-induced factor(s) stimulating PGE2 production. Inhibition of protein synthesis by cycloheximide potentiated the effect of DEX, and raised the abundance of PGHS-1, PGHS-2, and gamma-actin mRNAs in untreated cells. DEX did not affect the stability of the PGHS-2 mRNA. These results show that glucocorticoids promote PGE2 synthesis by amnion cells by stimulating the expression of PGHS-2 mRNA in a receptor-dependent, selective, and immediate fashion.

Prostanoid concentrations in maternal/fetal plasma and amniotic fluid and intrauterine tissue prostanoid output in relation to myometrial contractility during the onset of adrenocorticotropin-induced preterm labor in sheep.

Prostanoid [prostaglandin (PG)] concentrations were measured in ovine maternal and fetal plasma and amniotic fluid during the onset of preterm labor induced by the administration of a pulsatile infusion of ACTH-(1-24) (P-ACTH; 66.7 ng/min for 15 min every 2 h) to the fetus and in saline-infused controls. P-ACTH administration stimulated a change in intra-uterine pressure from type A-activity, characterized by sustained increases of low amplitude, to type B labor-like activity of short duration, high amplitude (greater than or equal to 10 mm Hg) increases which occurred between 12 and 8 h before the onset of labor. PGF2 alpha and/or PGFM (13,14-dihydro-15-keto PGF2 alpha) concentrations increased consistently in all fluids 16 h or earlier before labor. All PGs increased in fetal carotid arterial plasma (PGE2 greater than PGF2 alpha) and amniotic fluid, and the relative increases in each PG were similar. However, PGF2 alpha and PGFM selectively increased in maternal vena caval and aortic plasma, whereas smaller or negligible increases in the prostacyclin hydrolysis metabolite 6-keto PGF1 alpha (6KF) and PGE2 were noted. The output of PGs E2 and F2 alpha (picograms per 10(5) cells/8 h) increased 1.6- and 1.7-fold, respectively, by cells dispersed from the chorioallantois of P-ACTH-treated animals compared to that in control animals infused with saline for 100 h. From fetal cotyledons, these increases were 2.4-fold (P less than 0.05) and 3.6-fold, respectively. No significant changes occurred in 6-keto PGF1 alpha output from any tissue or PGE2 or PGF2 alpha output from amnion or maternal cotyledons. We conclude 1) that PGs increase in all fluids before the increase in uterine mechanical activity during induced preterm labor, implying that PGs mediate this event and are not a result thereof; 2) that syntheses of PGs E2 and F2 alpha increase similarly in intrauterine tissues with the onset of labor; and 3) that a selective increase in PGF2 alpha, a myometrial stimulatory PG, occurs exclusively in maternal plasma, suggesting that endoperoxide conversion to PGF2 alpha is specifically enhanced during parturition or suggesting the existence of an intrauterine tissue source of 9-keto PG reductase.

Expression of myometrial activation and stimulation genes in a mouse model of preterm labor: myometrial activation, stimulation, and preterm labor.

Myometrial contractions of labor result from an increase in myometrial activation and stimulation. Activation develops through the expression of contraction associated proteins (CAPs), including oxytocin receptors (OTR), connexin-43 (Cx-43), and prostaglandin F2 alpha, receptors (FP). Stimulation involves increases in contractile agonists including prostaglandin E2 (PGE2) and prostaglandin F2 alpha. (PGF2 alpha) that may result from increases in prostaglandin endoperoxide H synthase (PGHS)-2. A mouse model of preterm birth was used to study gene expression involved in myometrial activation and stimulation. To induce preterm birth, pregnant C57BL/6J mice were intubated with 6 g/kg ethanol on gestational day 16 and were killed every 6 h from treatment until birth. RIA was used to measure uterine PGE2 and PGF2 alpha, while PGHS-2, OTR, Cx-43, and FP messenger RNA levels were measured by ribonuclease protection assay. Increases in CAP mRNA were associated with term and preterm birth. There were differences in stimulation effectors associated with preterm and term birth. Uterine PGF2 alpha values were increased only at the time of term birth, but PGE2 was elevated during both preterm and term labor. These data suggest that existing levels of PGF2 alpha are sufficient for preterm birth when CAP expression is increased, but term labor requires increases in PGE2, PGF2alpha, and CAPs. The PGHS-2 messenger RNA expression pattern suggests that it is a CAP.

Stimulation of human amnion prostaglandin E2 production by activators of protein kinase-C.

We tested the possibility that the activation of protein kinase-C by the tumor-promoter phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) or diacylglycerol stimulates the production of prostaglandin E2 (PGE2) by the amnion. Confluent primary cultures of human amnion epithelial cells were adapted to serum-free medium and treated with the agonists for up to 8 h. Cumulative PGE2 output in the medium was measured by RIA. TPA, a potent activator of protein kinase-C, stimulated basal PGE2 output from less than 50 pg/well.5 h to 3 ng/well.5 h (P less than 0.01) in a time- and dose-dependent manner. 4-Methoxy-TPA, a weak tumor promoter derivative of TPA, was ineffective when tested in the same concentration range as TPA (1 nmol/L to 1 mumol/L). Neither calcium ionophore A23187 (20 nmol/L) nor arachidonate (1 mumol/L) stimulated PGE2 output alone, but each agonist potentiated the effect of TPA as much as 5-fold (P less than 0.01). 1,2-Dioctanoyl-sn-glycerol stimulated PGE2 output 4- to 7-fold (P less than 0.05), and this effect was potentiated by Ca ionophore and arachidonate. Studies involving actinomycin-D and cycloheximide indicated that the stimulatory effect of TPA was dependent on RNA synthesis during the first 60 min and on protein synthesis during the entire length of the phorbol ester treatment period (300 min). TPA was also able to stimulate PGE2 production after irreversible inactivation of PG endoperoxide synthase activity with acetylsalicylic acid. These results suggest that activation of protein kinase-C in amnion cells increases the de novo synthesis of the PG endoperoxide synthase enzyme in a RNA synthesis-dependent manner. Elevated intracellular calcium levels contribute to the stimulation apparently by increasing the availability of endogenous arachidonate for subsequent conversion to PGE2.

Arachidonic acid release from cultured human amnion cells: the effect of dexamethasone.

Glucocorticoids inhibit prostaglandin (PG) synthesis in several cell types, presumably by inhibiting arachidonic acid (AA) deacylation from phospholipids. We studied the effects of glucocorticoids on cultured term human amnion cell AA release. Confluent monolayer cultures of amnion cells were adapted to serum-free medium, and phospholipids were labeled for 18 h with [14C]AA. The calcium ionophore A23187 (0.2-5.0 mumol/L) stimulated [14C]AA release (up to 2.2-fold) in a dose- and time-dependent manner. The apparent sources of the liberated [14C]AA were phosphatidylcholine and phosphatidylethanolamine. Pretreatment for 24 h with the synthetic glucocorticoid dexamethasone (0.1-1000 nmol/L) significantly inhibited (P less than 0.01) basal (unstimulated) [14C]AA release by 69% in subsequent 1-h experiments. The sole apparent source of free [14C]AA during this inhibitory state was phosphatidylethanolamine. Dexamethasone pretreatment slightly inhibited (13%; P less than 0.05) calcium ionophore-stimulated [14C]AA release; however, it was still 3.8-fold greater than basal release, suggesting that the glucocorticoid effect on stimulated AA release was not biologically relevant. Further characterization of the glucocorticoid effect revealed that preincubation of the cultures with dexamethasone for as little as 20 min inhibited basal [14C]AA release. Furthermore, studies involving actinomycin-D and cycloheximide demonstrated that inhibition of RNA and protein synthesis failed to block the glucocorticoid inhibition of basal AA liberation. The glucocorticoid receptor antagonist RU 38486, alone or in the presence of dexamethasone, also inhibited unstimulated [14C]AA release. Cortisol, dehydroisoandrosterone sulfate, 17 beta-estradiol, and progesterone all inhibited basal [14C]AA liberation. We conclude that glucocorticoids inhibit unstimulated AA release from cultured amnion cells, but do not prevent calcium ionophore from stimulating a large increase in AA release.(ABSTRACT TRUNCATED AT 250 WORDS)

Glucocorticoids stimulate prostaglandin synthesis in human amnion cells by a receptor-mediated mechanism.

Prostaglandin E2 (PGE2) synthesis by human amnion increases with the onset of labor and is thought to participate in the initiation and maintenance of parturition. Since cortisol levels increase in amniotic fluid in late pregnancy, we studied the effects of glucocorticoids on cultured term human amnion cell PGE2 output. In 24-h studies, the synthetic glucocorticoid dexamethasone stimulated basal PGE2 output 2-fold over control levels at 16-500 nmol/L. PGE2 output was dramatically stimulated (greater than 10-fold) when, after dexamethasone pretreatment, the cells were incubated with calcium ionophore A23187 or arachidonic acid (AA) for 2 h. Maximum effects were achieved at 31 nmol/L dexamethasone. Basal PGE2 output was stimulated at 12 h of dexamethasone treatment, whereas A23187- or AA-stimulated PGE2 output was enhanced after 3-6 h of dexamethasone pretreatment. Cortisol (50 and 500 nmol/L) also enhanced basal and stimulated PGE2 output, while dehydroepiandrosterone sulfate, 17 beta-estradiol, and progesterone were ineffective. The glucocorticoid receptor antagonist RU 38486 attenuated dexamethasone-enhanced basal and stimulated PGE2 output. Dexamethasone pretreatment had no effect on basal or stimulated PGE2 output from cultured term chorion cells, suggesting tissue specificity. We conclude that glucocorticoids specifically enhance PGE2 output from cultured amnion cells via a receptor-mediated mechanism. We speculate that the action of glucocorticoids is to increase the capacity of the cells to convert AA to PGE2.

Prostaglandin output in relation to parturition by cells dispersed from human intrauterine tissues.

Collagenase-dispersed cells from human amnion, chorion, decidua, and placenta have been maintained in short term cultures to study prostaglandin (PG) biosynthesis in relation to parturition. Cells retained metabolic function during the 6-h incubation period, as determined by the apparently linear utilization of radioactive glucose and the formation of tritiated water. All cells synthesized PGs (E, F, and 6-keto F1 alpha) from endogenous precursors. The output of all three PGs significantly increased in amnion and chorion, but not in decidua or placenta, obtained from women who had entered labor spontaneously at term and delivered vaginally (SL) when compared to women at term, but not in labor, delivered by elective cesarean section (CS). For example, PGE output (picograms per 10(5) cells/6 h) increased from 207 +/- 77 (n = 5) to 908 +/- 334 with labor (mean +/- SEM). The addition of indomethacin (10(-7)-10(-5) M) to SL amnion cells significantly decreased (P less than 0.001, by analysis of variance) PGE and PGF, but not 6-keto PGF1 alpha output. Comparison of PGF output with its metabolite, 13,14-dihydro-15-keto PGF2 alpha (PGFM); indicated that PGF values were similar to or higher than PGFM for all tissues other than CS chorion, where PGFM output was significantly greater (142 +/- 63 vs. 486 +/- 95; n = 5; P less than 0.05). PGFM levels in amnion increased with labor [104 +/- 51 (n = 5) to 341 +/- 96 (n = 5); P less than 0.05], suggesting that PG output increased with labor in the fetal membranes as a result of increased synthesis and not decreased metabolism.