Untargeted analysis of plasma samples from pre-eclamptic women reveals polar and apolar changes in the metabolome.

INTRODUCTION: Pre-eclampsia is a hypertensive gestational disorder that affects approximately 5% of all pregnancies. OBJECTIVES: As the pathophysiological processes of pre-eclampsia are still uncertain, the present case-control study explored underlying metabolic processes characterising this disease. METHODS: Maternal peripheral plasma samples were collected from pre-eclamptic (n = 32) and healthy pregnant women (n = 35) in the third trimester. After extraction, high-resolution mass spectrometry-based untargeted metabolomics was used to profile polar and apolar metabolites and the resulting data were analysed via uni- and multivariate statistical approaches. RESULTS: The study demonstrated that the metabolome undergoes substantial changes in pre-eclamptic women. Amongst the most discriminative metabolites were hydroxyhexacosanoic acid, diacylglycerols, glycerophosphoinositols, nicotinamide adenine dinucleotide metabolites, bile acids and products of amino acid metabolism. CONCLUSIONS: The putatively identified compounds provide sources for novel hypotheses to help understanding of the underlying biochemical pathology of pre-eclampsia.

Anti-acetylcholinesterase activity and antioxidant properties of extracts and fractions of Carpolobia lutea.

CONTEXT: There is an unmet need to discover new treatments for Alzheimer's disease. This study determined the anti-acetylcholinesterase (AChE) activity, DPPH free radical scavenging and antioxidant properties of Carpolobia lutea G. Don (Polygalaceae). OBJECTIVE: The objective of this study is to quantify C. lutea anti-AChE, DPPH free radical scavenging, and antioxidant activities and cell cytotoxicity. MATERIALS AND METHODS: Plant stem, leaves and roots were subjected to sequential solvent extractions, and screened for anti-AChE activity across a concentration range of 0.02-200 µg/mL. Plant DPPH radical scavenging activity, reducing power, and total phenolic and flavonoid contents were determined, and cytotoxicity evaluated using human hepatocytes. RESULTS: Carpolobia lutea exhibited concentration-dependent anti-AChE activity. The most potent inhibitory activity for the stem was the crude ethanol extract and hexane stem fraction oil (IC50 = 140 µg/mL); for the leaves, the chloroform leaf fraction (IC50 = 60 µg/mL); and for roots, the methanol, ethyl acetate and aqueous root fractions (IC50 = 0.3-3 µg/mL). Dose-dependent free radical scavenging activity and reducing power were observed with increasing stem, leaf or root concentration. Total phenolic contents were the highest in the stem: ∼632 mg gallic acid equivalents/g for a hexane stem fraction oil. Total flavonoid content was the highest in the leaves: ∼297 mg quercetin equivalents/g for a chloroform leaf fraction. At 1 µg/mL, only the crude ethanol extract oil was significantly cytotoxic to hepatocytes. DISCUSSION AND CONCLUSIONS: Carpolobia lutea possesses anti-AChE activity and beneficial antioxidant capacity indicative of its potential development as a treatment of Alzheimer's and other diseases characterized by a cholinergic deficit.

Regulation of GTP-binding protein (Gαs) expression in human myometrial cells: a role for tumor necrosis factor in modulating Gαs promoter acetylation by transcriptional complexes.

The onset of parturition is associated with a number of proinflammatory mediators that are themselves regulated by the nuclear factor κB (NF-κB) family of transcription factors. In this context, we previously reported that the RelA NF-κB subunit represses transcription and mRNA expression of the proquiescent Gαs gene in human myometrial cells following stimulation with the proinflammatory cytokine TNF. In the present study, we initially defined the functional consequence of this on myometrial contractility. Here we show that, contrary to our initial expectations, TNF did not induce myometrial contractility but did inhibit the relaxation produced by the histone deacetylase inhibitor trichostatin A, an effect that in turn was abolished by the NF-κB inhibitor N(4)-[2-(4-phenoxyphenyl)ethyl]-4,6-quinazolinediamine. This result suggested a role for TNF in regulating Gαs expression via activating NF-κB and modifying histone acetylation associated with the promoter region of the gene. In this context, we show that the -837 to -618 region of the endogenous Gαs promoter is occupied by cAMP-response element-binding protein (CREB), Egr-1, and Sp1 transcription factors and that CREB-binding protein (CBP) transcriptional complexes form within this region where they induce histone acetylation, resulting in increased Gαs expression. TNF, acting via NF-κB, did not change the levels of CREB, Sp1, or Egr-1 binding to the Gαs promoter, but it induced a significant reduction in the level of CBP. This was associated with increased levels of histone deacetylase-1 and surprisingly an increase in H4K8 acetylation. The latter is discussed herein.

A role for two-pore potassium (K2P) channels in endometrial epithelial function.

The human endometrial epithelium is pivotal to menstrual cycle progression, implantation and early pregnancy. Endometrial function is directly regulated by local factors that include pH, oxygen tension and ion concentrations to generate an environment conducive to fertilization. A superfamily of potassium channels characterized by two-pore domains (K2P) and encoded by KCNK genes is implicated in the control of the cell resting membrane potential through the generation of leak currents and modulation by various physicochemical stimuli. The aims of the study were to determine the expression and function of K2P channel subtypes in proliferative and secretory phase endometrium obtained from normo-ovulatory women and in an endometrial cancer cell line. Using immunochemical methods, real-time qRT-PCR proliferation assays and electrophysiology. Our results demonstrate mRNA for several K2P channel subtypes in human endometrium with molecular expression of TREK-1 shown to be higher in proliferative than secretory phase endometrium (P < 0.001). The K2P channel blockers methanandamide, lidocaine, zinc and curcumin had antiproliferative effects (P < 0.01) in an endometrial epithelial cancer cell line indicating a role for TASK and TREK-1 channels in proliferation. Tetraethylammonium- and 4-aminopyridine-insensitive outwards currents were inhibited at all voltages by reducing extracellular pH from 7.4 to 6.6. Higher expression of TREK-1 expression in proliferative phase endometrium may, in part, underlie linked to increased cell division. The effects of pH and a lack of effect of non-specific channel blockers of voltage-gated potassium channels imply a role for K2P channels in the regulation of human endometrial function.

Inhibition of tissue transglutaminase 2 attenuates contractility of pregnant human myometrium.

Premature delivery remains a serious risk factor in pregnancy, with currently licensed tocolytics unable to offer significant improvement in neonatal outcome. Further understanding of the regulators of uterine contractility is required to enable the development of novel and more effective tocolytic therapies. The transglutaminase family is a class of calcium-dependent, transamidating enzymes, of which tissue transglutaminase 2 is a multifunctional enzyme with roles in cell survival, migration, adhesion, and contractility. The aim of the present study was to investigate the role of this enzyme in regulating the contractility of pregnant human myometrium. Tissue strips from biopsy samples obtained at elective cesarean section were either allowed to contract spontaneously or induced to contract with oxytocin, phenylephrine, or bradykinin. Activity integrals, used to measure contractile activity, were taken following cumulative additions of the reversible, polyamine transglutaminase inhibitors cystamine and mono-dansylcadaverine and the irreversible, site-specific transglutaminase inhibitors N-benzyloxycarbonyl-l-phenylalanyl-6-dimethylsulfonium-5-oxo-L-norleucine and 1,3-dimethyl-2[(oxopropyl)thio]imidazolium. The ability of cystamine and mono-dansylcadaverine to affect oxytocin-mediated calcium mobilization within primary cultured myometrial cells was also measured utilizing a calcium indicator. All inhibitors attenuated myometrial contractions in a concentration-dependent manner independent of the method of contraction stimulus. Similarly cultured myometrial cells preincubated with cystamine and mono-dansylcadaverine displayed an altered calcium response to oxytocin stimulation. Our findings demonstrate a potential role for tissue transglutaminase 2 in regulating uterine contractility in pregnant human myometrium that may be associated with the calcium signaling cascade required for contraction.

The role of Eag and HERG channels in cell proliferation and apoptotic cell death in SK-OV-3 ovarian cancer cell line.

BACKGROUND: The voltage gated potassium (K+) channels Eag and HERG have been implicated in the pathogenesis of various cancers, through association with cell cycle changes and programmed cell death. The role of these channels in the onset and progression of ovarian cancer is unknown. An understanding of mechanism by which Eag and HERG channels affect cell proliferation in ovarian cancer cells is required and therefore we investigated their role in cell proliferation and their effect on the cell cycle and apoptosis of ovarian cancer cells. METHODS: The presence of Eag and HERG was determined in SK-OV-3 cells using immunofluorescence and western blotting. The effect of the Eag blockers (imipramine and clofilium) and HERG blockers (E-4031 and ergtoxin) on cell proliferation was assessed using the MTS assay with further investigation of their role in the cell cycle and apoptosis determined by flow cytometry. RESULTS: Eag and HERG channels were present in the cytoplasm and nuclei of SK-OV-3 cells. There was significant inhibition of proliferation of SK-OV-3 cells by imipramine (P < 0.001) and ergtoxin (P < 0.05) at 72 hours of culture. Incubation of cells with ergtoxin led to the accumulation of cells in the S and G2/M phase, while cells accumulated in S phase after incubation with E-4031, with no effect on apoptosis. Imipramine did not affect the cell cycle but increased the proportion of SK-OV-3 cells undergoing early apoptosis. CONCLUSION: Both Eag and HERG channels are expressed in SK-OV-3 ovarian cancer cells and have a role in cell proliferation. HERG channels affect the cell cycle while Eag channels are implicated in the inhibition of apoptosis of ovarian cancer cells. The family of Eag channels may represent a new therapeutic target for the treatment of ovarian cancer.

Detection of EETs and HETE-generating cytochrome P-450 enzymes and the effects of their metabolites on myometrial and vascular function.

Cytochrome P-450 (CYP450) enzymes of the CYP2 and -4 family in humans metabolize arachidonic acid to generate bioactive epoxyeicosatrienenoic acids (EETs) and hydroxyeicosatetrenoic acids (HETEs). We report significantly higher levels of CYP 2J2 protein expression following the onset of labor (n = 6, P < 0.05), implying increased EET-generating capacity within the uterus. Myometrial relaxation to 8,9-EET and 5,6-EET was observed, with the latter being inhibited by preincubation with 1 muM paxilline and is supported by whole cell recordings showing a modest effect of 5,6-EET on myometrial outward-current density (n = 4, P < 0.05). Only 5,6-EET of the EETs tested affected vascular reactivity (n = 6). Both 12- and 20-HETE (n = 5-6) caused vasoconstriction of partially depolarized blood vessels, with glibenclamide (n = 5) enhancing the effect of 12-HETE alone. Our findings signify a role for CYP2C9/19, -2J2, and -4A11/22 in late pregnancy, possibly related to the synthesis of lipid metabolites and downstream effects on vascular remodeling in the term pregnant uterus. The presence of CYP4A11/22 and their resultant procontractile metabolites could argue either a role in the control and initiation of labor and/or modification of the vascular delivery system to influence blood flow to the laboring uterus. The differential effects of the EETs and HETEs in the pregnant human uterus identify the CYP pathway as a novel modulator of myometrial and vascular physiology during late pregnancy.

Pharmacological profile of vascular activity of human stem villous arteries.

INTRODUCTION: The function of the placental vasculature differs considerably from other systemic vascular beds of the human body. A detailed understanding of the normal placental vascular physiology is the foundation to understand perturbed conditions potentially leading to placental dysfunction. METHODS: Behaviour of human stem villous arteries isolated from placentae at term pregnancy was assessed using wire myography. Effects of a selection of known vasoconstrictors and vasodilators of the systemic vasculature were assessed. The morphology of stem villous arteries was examined using IHC and TEM. RESULTS: Contractile effects in stem villous arteries were caused by U46619, 5-HT, angiotensin II and endothelin-1 (p ≤ 0.05), whereas noradrenaline and AVP failed to result in a contraction. Dilating effects were seen for histamine, riluzole, nifedipine, papaverine, SNP and SQ29548 (p ≤ 0.05) but not for acetylcholine, bradykinin and substance P. DISCUSSION: Stem villous arteries behave differently to vessels of the systemic vasculature and results indicate that the placenta is cut off from the systemic maternal vascular regulation. Particularly, endothelium-dependent processes were attenuated in the placental vasculature, creating a need to determine the role of the endothelium in the placenta in future studies.

Caspase-1-mediated cytokine release from gestational tissues, placental, and cord blood.

Distinguishing between fetal and maternal inflammatory responses is necessary for understanding the immune interplay either side of the placenta. Fetal immunity reaches maturity during extrauterine life and while basic inflammatory responses afford a certain degree of protection, fetuses are vulnerable to infection. With the discovery of inflammasomes-intracellular scaffolds that facilitate the elaboration of reactions resulting in the release of mature interleukin-1ß (IL-1ß)-it is necessary to consider how inflammatory stimuli are processed. The purinergic P2X7 receptor located on haematopoietic cells is a key intermediary in signal transduction initiated at Toll-like receptors (TLR) terminating in release of the mature IL-1ß product. We demonstrate herein that IL-1ß release from fetal membranes and mononuclear cells isolated from cord, placental, and maternal blood, obtained at term, is P2X7- and caspase-1 dependent. The P2X7-dependent release of the cytokine, which was highest from choriodecidua, was attenuated by progesterone (P4), prolactin and an NFkB inhibitor. The NLRP3 inflammasome appears necessary for the processing of IL-1ß in gestational tissues and leukocytes.

Functional characteristics of chorionic plate placental arteries from normal pregnant women and women with pre-eclampsia.

OBJECTIVES: We aimed to compare placental small artery function from women with pre-eclampsia and normal pregnancy. In particular, we wished to test the hypothesis that these arteries respond differently to an endothelium-dependent vasodilator, to the presence of nitric oxide, and to the presence of cyclic monophosphate nucleotides. METHODS: A small vessel wire myograph was used to study placental arteries (200 to 550 microm). Contractile function was assessed with vasopressin. Relaxation was assessed with the endothelium-dependent vasodilator, bradykinin, and the endothelium-independent vasodilators sodium nitroprusside (a nitric oxide donor) and papaverine (a phosphodiesterase inhibitor). RESULTS: The constrictor response to vasopressin did not differ between patient groups (p=0.79; repeated measures ANOVA). For both normal pregnancy and pre-eclampsia, the response of pre-constricted arteries to the endothelium-dependent vasodilator, bradykinin, was minimal. Vasorelaxation to sodium nitroprusside and papaverine was attenuated in pre-eclampsia compared to normal pregnancy (p=0.03 and p<0.001, respectively; repeated measures ANOVA). CONCLUSIONS: In pre-eclampsia, placental arteries exhibit an attenuated vasodilatory response to nitric oxide.

Expression and prognostic significance of the oncogenic K2P potassium channel KCNK9 (TASK-3) in ovarian carcinoma.

BACKGROUND/AIMS: The TWIK-related acid sensitive K(+) channel-3 (TASK-3) is an oncogenic potassium channel. We investigated the expression of TASK-3 in human ovaries, examined its prognostic significance, and determined effects of TASK-3 blockers on cell proliferation and apoptosis. MATERIALS AND METHODS: Immunofluorescence and western blotting were used to investigate TASK-3 expression in two ovarian cancer cell lines, normal ovarian surface epithelium and cancer. Immunohistochemistry quantified expression in an ovarian cancer tissue microarray. The effect of TASK-3 blocking agents on cell proliferation was investigated with the CellTiter 96® Aqueous Non-Radioactive Cell Proliferation assay and on apoptosis with flow cytometry. RESULTS: TASK-3 expression was confirmed by immunofluorescence in the SKOV-3 and OVCAR-3 cell lines, normal ovaries (n=4) and ovarian tumours (n=4) and by western blotting in normal ovaries (n=6) and ovarian tumours (n=22). Immunohistochemistry demonstrated immunostaining in 99% of tumours (n=230). Increased immunostaining conferred a survival advantage (p=0.002; median survival of >24 months). TASK-3 blockers caused a significant reduction in cell proliferation and an increase in apoptosis in the SKOV-3 and OVCAR-3 cell lines. CONCLUSION: TASK-3 is expressed in epithelial ovarian cancer, conferring a significant survival advantage on patients with increased expression. TASK-3-modulating agents have a significant effect on cell proliferation and apoptosis. Based on these results, we propose that TASK-3 could prove to be both a novel tumour marker and a new therapeutic target in ovarian cancer, but further investigation is required.

The Eag potassium channel as a new prognostic marker in ovarian cancer.

BACKGROUND: Ovarian cancer is the second most common cancer of the female genital tract in the United Kingdom (UK), accounting for 6% of female deaths due to cancer. This cancer is associated with poor survival and there is a need for new treatments in addition to existing chemotherapy to improve survival. Potassium (K+) channels have been shown to be overexpressed in various cancers where they appear to play a role in cell proliferation and progression. OBJECTIVES: To determine the expression of the potassium channels Eag and HERG in ovarian cancer tissue and to assess their role in cell proliferation. METHODS: The expression of Eag and HERG potassium channels was examined in an ovarian cancer tissue microarray. Their role in cell proliferation was investigated by blocking voltage-gated potassium channels in an ovarian cancer cell line (SK-OV-3). RESULTS: We show for the first time that high expression of Eag channels in ovarian cancer patients is significantly associated with poor survival (P = 0.016) unlike HERG channel expression where there was no correlation with survival. There was also a significant association of Eag staining with high tumour grade (P = 0.014) and presence of residual disease (P = 0.011). Proliferation of SK-OV-3 cells was significantly (P < 0.001) inhibited after treatment with voltage gated K+ channel blockers. CONCLUSION: This novel finding demonstrates a role for Eag as a prognostic marker for survival in patients with ovarian cancer.

Measurement of vasoactive metabolites (hydroxyeicosatetraenoic and epoxyeicosatrienoic acids) in uterine tissues of normal and compromised human pregnancy.

BACKGROUND: Preeclampsia and intrauterine growth restriction define two disorders of a multifactorial etiology that compromise maternal and fetal well being as well as cardiovascular health in later life. Many of the overt symptoms of preeclampsia are attributable to the systemic endothelial dysfunction observed in the uteroplacental and systemic circulation, leading to a generalized vasoconstriction, hypertension and inadequate placental perfusion. Mounting evidence implicates nonprostanoid eicosanoids, epoxyeicosatrienoic acids (EETs) and hydroxyeicosatetraenoic acids (HETEs) in the control of vascular function and dysfunction. OBJECTIVE: To determine whether levels of EETs and HETEs are altered in preeclampsia and intrauterine growth restriction compared with normal term pregnancy. METHODS: An analytical liquid chromatography-tandem mass spectrometry profiling method was utilized in order to analyze differential levels of EETs and HETEs in intrauterine tissues of term nonlaboring, laboring and preeclamptic women as well as women with a growth-restricted pregnancy. RESULTS: Placentae of preeclamptic women contained significantly (P < 0.05) larger amounts of 5-HETE, 12-HETE and 15-HETE known to possess either vasoconstrictive or proinflammatory actions. Laboring tissues were characterized by significantly higher (P < 0.05) EET levels in the amnion compared with the other clinical groups. EET and HETE levels in preeclampsia and intrauterine growth restriction were positively correlated (P < 0.05), whereas in normal and laboring pregnancies, EETs and HETEs were negatively correlated. CONCLUSION: Increased production of 5-HETE, 12-HETE and 15-HETE metabolites in preeclamptic placentae indicates an important role for this family of eicosanoids in the cause of this disease.

Interleukin-1 beta secretion from cord blood mononuclear cells in vitro involves P2X 7 receptor activation.

The purinergic receptor P2X(7) is activated by extracellular adenosine 5'-triphosphate (ATP) and promotes the efficient release of interleukin-1 beta. The authors examine protein and molecular expression of the P2X(7) receptor and its ability to stimulate interleukin-1 beta release in cord blood mononuclear cells (CBMCs) from placentae of term nonlaboring and laboring women. They show both mRNA and protein (78 kDa) expression for the P2X( 7) receptor in CBMCs of parturient and nonparturient women. Costimulation of CBMCs in vitro with bacterial endotoxin and ATP resulted in significantly (P < .05) enhanced interleukin-1 beta release in laboring (54.17 +/- 24.78 pg/mL(-1); n = 8) compared with nonlaboring (13.60 +/- 3.20 pg/mL(-1); n = 7) deliveries. Release of interleukin-1 beta in both groups was blocked by preincubation with oxidized ATP, a P2X(7) receptor antagonist. The authors provide evidence for a novel inflammatory pathway in the release of interleukin-1 beta, which may be linked to the onset of labor.

Quantitative profiling of epoxyeicosatrienoic, hydroxyeicosatetraenoic, and dihydroxyeicosatetraenoic acids in human intrauterine tissues using liquid chromatography/electrospray ionization tandem mass spectrometry.

A reversed-phase liquid chromatography negative ion electrospray tandem mass spectrometry (LC/ESI-MS/MS) method was developed and validated to quantify a range of physiologically relevant eicosanoids, including 5,6-, 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acids (EETs); 5-, 8-, 9-, 12-, and 15-hydroxyeicosatetraenoic acids (HETEs), and 5,6-, 8,15-, and 12,20-dihydroxyeicosatetraenoic acids (DiHETEs) in human intrauterine tissues. A solid-phase extraction method was employed to extract the eicosanoids, and gradient LC separation was performed on a Kromasil C(18) column. Mass spectrometric detection was performed by multiple reaction monitoring over a 31-min run time. The calibration curves were linear over the range of 4-400pmol/g tissue, and the intra- and interday precision and accuracy were within a coefficient of variation of 2.0 to 27.4% and 4.6 to 17.9%, respectively. The lower limit of quantitation was 1.0pmol/g tissue. The method was applied successfully to the characterization and quantitation of eicosanoids in the different compartments of human intrauterine tissues. Our results demonstrate significantly greater amounts of HETEs than of either the EETs or DiHETEs (P<0.001), irrespective of tissue type. Specifically, the metabolite 12-HETE was significantly more abundant (P<0.001) than all other HETEs. Of the EET metabolites, 5,6-EET predominated (P<0.001). A significant negative correlation between EETs and HETEs for all tissues (rho=-0.390, P<0.001) was identified, implying a biological feedback mechanism between these two arachidonate metabolite classes.

Progesterone enhances the tocolytic effect of ritodrine in isolated pregnant human myometrium.

OBJECTIVE: To evaluate the effect of natural progesterone on the relaxant effect of ritodrine on pregnant human oxytocin-induced myometrial contractility. STUDY DESIGN: Isometric tension recordings were performed under physiologic conditions on isolated myometrial strips taken from low-risk term pregnant women undergoing elective cesarean section. Cumulative effects of natural progesterone (10 (-11) to 10 (-5) mol/L) on oxytocin-induced myometrial contractility were evaluated. Contractile activity following ritodrine exposure was also investigated in myometrium pretreated with natural progesterone. RESULTS: Natural progesterone alone exerted a concentration-dependent relaxant effect on myometrial contractions. The concentration-response curve for ritodrine from natural progesterone pretreated myometrium was shifted to the left with a significant reduction ( P < .01) of 50% of the maximal response, contraction amplitude ( P < .05), and frequency ( P < .05). However, there was no significant difference in the mean maximal inhibition achieved ( P = .95). CONCLUSION: Natural progesterone increased the relaxant effect of ritodrine by reducing 50% of the maximal response, amplitude, and frequency of myometrial contraction, most likely through nongenomic actions. These results suggest that natural progesterone may be beneficial for preventing preterm birth in a low-risk population.

Hydrogen peroxide and superoxide anion modulate pregnant human myometrial contractility.

Reactive oxygen species (ROS) have the propensity to cause macromolecular damage with consequent modification of cellular function. We investigated the effects of two particular oxidants, superoxide (O2(-)) anions and hydrogen peroxide (H2O2), on oxytocin-induced myometrial contractility using biopsies from women undergoing Caesarean section at term gestation. Isometric tension recordings were performed and concentration-response curves derived after addition of test agents. A maximal reduction in myometrial contractility to 27.2 +/- 4.5% of control was observed followed application of H2O2. The enzyme scavenger catalase (CAT) reduced the inhibitory effect of H2O2 but had little effect at 10-fold lower concentrations. Addition of dialysed xanthine oxidase +/- hypoxanthine significantly inhibited contractility to 23.8.0 +/- 4.2% compared with control. Pre-incubation with superoxide dismutase and CAT diminished this effect. The non-specific potassium channel blocker, tetraethylammonium chloride (1 mM), had no effect on myometrial contractility. We conclude that human myometrium is susceptible to the effects of ROS, which may be produced by reperfusion-ischaemic episodes during labour. Our findings could, in part, explain the weak or prolonged depression of contractions characteristic of myometrial dysfunction culminating in difficult labours.

Relaxant action of sildenafil citrate (Viagra) on human myometrium of pregnancy.

OBJECTIVE: The purpose of this study was to investigate the effect of the cyclic guanosine monophosphate phosphodiesterase 5-specific inhibitor, sildenafil citrate, on the contractions of isolated pregnant human myometrium. STUDY DESIGN: Myometrial samples were obtained from women who underwent elective cesarean delivery. Myometrial contractions that were recorded in response to sildenafil in the absence and presence of the potassium channel blocker, tetraethylammonium or the guanylate cyclase inhibitor, methylene blue (10 micromol/L) were studied. One-way analysis of variance with post-hoc analysis was used to test differences among groups. RESULTS: Sildenafil caused relaxation of myometrium in a concentration-dependent manner. The log(10) EC(50) value for this relaxation in the presence of 20 mmol/L tetraethylammonium was significantly different (P<.01) than values that were obtained with sildenafil alone or sildenafil in the presence of either methylene blue or 5 and 10 mmol/L tetraethylammonium. CONCLUSION: Myometrial relaxation that is evoked by the direct application of sildenafil occurs independently of cyclic guanosine monophosphate. Potassium channels appear to be the likely candidates in mediating this response.

Vasoactive effects of neurokinin B on human blood vessels.

OBJECTIVES: Preeclampsia (PE) is a multisystem disease unique to human pregnancy. Abnormal placentation results in placental hypoperfusion leading to the secretion of a factor(s) by the placenta. Our aim was to investigate whether neurokinin B (NKB) is the circulating factor associated with PE. STUDY DESIGN: Vascular effects of NKB were assessed in blood vessels dissected from myometrial and omental biopsy specimens obtained at caesarean section from normal pregnant women (n = 26) or in mesenteric blood vessels obtained from nonpregnant female Wistar rats (n = 4). RESULTS: Incubation with NKB did not alter endothelial-dependent relaxation of omental or myometrial arteries. NKB produced a dose-dependent relaxation in preconstricted omental arteries and veins. NKB did not affect vasoactive responsiveness of rat mesenteric arteries. CONCLUSION: We conclude from these observations that NKB is not the circulating factor associated with increased vascular resistance in PE.

Effects of a phosphodiesterase-5 (PDE5) inhibitor on endothelium-dependent relaxation of myometrial small arteries.

OBJECTIVE: In preeclampsia, endothelium-dependent function is markedly aberrant. Myometrial resistance arteries from women with preeclampsia show a minimal, wholly nitric oxide-mediated, bradykinin-induced relaxation. Our aim was to test that phosphodiesterase 5 (PDE5) inhibition could improve endothelium-dependent function in preeclampsia. Study design Small arteries dissected from myometrial biopsies obtained at cesarean section from normal pregnant women (N=22) or women with preeclampsia (N=24) were mounted on wire or pressure myographs. Vessels were constricted (arginine vasopressin or U46619) and relaxed (bradykinin) before and after incubation with a PDE5 inhibitor, UK-343664. RESULTS: Endothelium-dependent vasodilatation was decreased in vessels from women with preeclampsia. 100 nmol/L UK-343664 did not affect normal pregnant but significantly improved relaxation of the vessels from women with preeclampsia. CONCLUSION: A PDE5 inhibitor enhances endothelial function of myometrial vessels from women with preeclampsia, such that the behavior of these arteries approximates to those from normal women. These agents offer a potential therapeutic strategy for the management of preeclampsia.