Intraplatelet cyclic guanosine-3',5'-monophosphate levels during pregnancy and preeclampsia.

OBJECTIVE: To investigate the intraplatelet cyclic guanosine-3',5'-monophosphate (cGMP) levels during normal pregnancy and preeclampsia. STUDY DESIGN: Pregnant women (n = 15), women with preeclampsia (n = 15), and nonpregnant, normotensive women (n = 15) were included. Intraplatelet cyclic guanosine-3',5'-monophosphate levels were measured by an enzyme-linked immunosorbent assay. RESULTS: Intraplatelet cGMP levels were significantly different among all groups (p < 0.02). The values were higher in normal pregnant women (mean 19.8 SD 2.6 fmol/10(5) platelets) in comparison to nonpregnant women (mean 7.6 SD 0.3 fmol/10(5)platelets; p = 0.001) and women with preeclampsia (mean 11.3 SD 1.8 fmol/10(5) platelets; p = 0.05). Plasma nitric oxide levels did not reveal differences between all groups. CONCLUSIONS: The results of this study in a high-risk Andean population demonstrated that intraplatelet cyclic guanosine-3',5'-monophosphate levels are decreased during preeclampsia compared to normal pregnancy, suggesting a lack in action of nitric oxide.     

Urinary cyclic guanosine 3',5'-monophosphate and cyclic adenosine 3',5'-monophosphate changes in spontaneous and induced onset active labor

BACKGROUND. The aim of this prospective, randomized study was to investigate the changes in urinary cyclic guanosine 3',5'-monophosphate (cGMP) and cyclic adenosine 3',5'-monophosphate (cAMP) between the latent and the active phases of spontaneous and prostaglandin E(1) (PGE(1))-induced labor. METHODS. Seventy singleton pregnant women at 36-41(+) weeks' gestation without signs of fetal distress were enrolled. The first group consisted of 35 pregnant women in whom labor was induced by PGE(1) applied intravaginally. The second group consisted of 35 women who had spontaneous active labor. Clinical data of the two groups were assessed as labor progressed. RESULTS. After the onset of active labor, urinary cGMP/creatinine (U cGMP/Cr) decreased in both groups with the percentage decline of 35.2 and 9.7, respectively, but this difference was only significant in the PGE(1)-induced group (P=0.033). After the onset of active labor, urinary cAMP/creatinine (U cAMP/Cr) decreased in both groups with the percentage decline of 36.5 and 15.6, respectively, but this difference was only significant in the PGE(1)-induced group (P=0.001). The duration of the latent phase was significantly shortened in the PGE(1)-induced group compared with the spontaneous labor group (P<0.05). CONCLUSIONS. Decreased U cGMP/Cr and U cAMP/Cr may be a transition from the latent to the active phase in PGE(1)-induced labor. Our results suggest that U cGMP/Cr and U cAMP/Cr can serve as easily obtained secondary messenger markers of myometrial contractility and cervical ripening at the onset of active labor. The NO-cGMP system and the G-protein alpha-cAMP system in the human uterus may concomitantly contribute to uterine quiescence during pregnancy and show downregulation in U cGMP/Cr and U cAMP/Cr at the initiation of active labor.     

Plasma cAMP in normal and abnormal human pregnancy

Plasma cAMP was determined using the method of Tovey et al. in normal pregnant women with a mean concentration of 18.9 +/- 0.8 pmol/ml (x- +/- SEM). Between weeks 9-12 and 33-36 of gestation, there were two peaks, with a mean cAMP of 22.5 +/- 2.4 which were significantly increased in comparison to the other weeks of pregnancy. Significantly decreased values were found in patients with threatened abortion (weeks 12-28) which terminated in abortion (11.6 +/- 2.4; p < 0.01). In premature labor no differences were found. During therapy with fenoterol there were highly significantly increased plasma cAMP levels (48.2 +/- 2.8; p < 0.0005). During thyroid hormone therapy in euthyroid goiter, cAMP was significantly decreased (14.0 +/- 1.4; p < 0.05). 1 week after cessation of therapy a highly significant increase of cAMP was observed (38.2 +/- 6.9; p < 0.0005). There was a negative linear regression between T3 and cAMP (2p < 0.01). In pregnancy with hypertension cAMP was significantly elevated (30.5 +/- 3.8 p < 0.0005), but nearly normal under antihypertensive therapy. In pregnancy with edema only, no difference was found. Induction of labor with PGE2 alpha was followed by a decrease of plasma cAMP.     

Atrial natriuretic peptide and cyclic guanosine-3',5'-monophosphate in normal and pathological pregnancies

The concentrations of atrial natriuretic peptide (ANP) and its presumed second messenger, cyclic guanosine-3',5'-monophosphate (cGMP) were determined in maternal plasma and amniotic fluid. Samples were obtained from normal and pathological pregnancies revealing hydramnios or severe Rh incompatibility between week 16 of gestation and delivery. For analysis of ANP and cGMP, radioimmunoassays were used. ANP and cGMP concentrations in maternal plasma did not differ in normal and pathological pregnancies. In amniotic fluid, we found ANP levels of about 120 pg/ml at 16 weeks of gestation which then decreased below the detection limit of 15.6 pg/ml. In contrast, cGMP levels were low at the beginning of pregnancy (4 pmol/ml) and rose significantly (14 pmol/ml), at the end of pregnancy. Pregnancies revealing severe Rh incompatibility exhibited the same levels as normal pregnancies when regular prenatal transfusions resulted in sufficient blood volume substitution. ANP, a volume-homeostasis-regulating hormone, is already produced in early pregnancy. The meaning of the presumed second messenger, cGMP, requires further investigation.     

Quantitative analysis of the effects of caffeine on sperm motility and cyclic adenosine 3',5'-monophosphate (AMP) phosphodiesterase

Caffeine in high concentrations (2 to 6 mM) has been shown to increase sperm motility and metabolism when added to semen. High concentrations of caffeine have also been demonstrated to inhibit cyclic adenosine 3',5'-monophosphate (AMP) phosphodiesterase isolated from sperm. Since cyclic AMP is known to increase sperm motility, some investigators have attributed the stimulatory effect of caffeine on sperm motility to the inhibition of phosphodiesterase. The authors used semen from healthy donors to quantitate the effects of caffeine on sperm motility by turbidimetric analysis and to analyze kinetically the effect of caffeine on phosphodiesterase isolated from sperm preparations. Caffeine (0.012 to 2.0 mM) produced a dose-dependent increase in sperm motility. The ED50 for motility stimulation was 300 microM of caffeine; a significant increase in motility was observed with 60 microM; and maximal stimulation (a threefold increase) occurred with 1 mM. The phosphodiesterase activity of sperm homogenates displayed linear kinetics with a Vmax of 0.5 nmoles/mg of protein per minute and a Km of 125 microM. Caffeine (0.1 to 40 mM) produced a dose-dependent inhibition of sperm phosphodiesterase. Kinetically, caffeine was a mixed inhibitor of sperm phosphodiesterase, displaying a Ki of 1.2 mM. These results suggest that caffeine may stimulate sperm motility by a mechanism other than phosphodiesterase inhibition.     

Synergistic effect of relaxin and progesterone on cyclic adenosine 3',5'-monophosphate levels in the rat uterus

Cyclic adenosine 3',5'-monophosphate is known to modulate smooth muscle contractility. Because both relaxin and progesterone have been demonstrated to affect myometrial cyclic adenosine monophosphate activity, we questioned whether the previously observed synergism of these two hormones in inhibiting uterine contractility is mediated via cyclic adenosine monophosphate. Immature rats were treated with estradiol benzoate (n = 7) or a combination of estradiol benzoate and progesterone (n = 7). Uterine horns were isolated, each horn was divided into two segments, and these horn segments were incubated in Ringer-Locke solution, either alone (control) or with 3-isobutyl-1-methylxanthine (MIX) 0.5 mM, MIX 0.5 mM + relaxin 10 ng/ml, or MIX 0.5 mM + relaxin 50 ng/ml. When compared with uterine segments incubated in MIX alone, treatment with MIX + relaxin 50 ng/ml significantly increased cyclic adenosine monophosphate levels in animals treated with estradiol benzoate alone or in combination with progesterone. Relaxin 10 ng/ml was sufficient to significantly elevate mean (+/- SEM) uterine cyclic adenosine monophosphate levels above that of control MIX-treated uteri in animals receiving both estradiol benzoate and progesterone (2.49 +/- 0.39 pm/micrograms deoxyribonucleic acid [DNA] versus 1.08 +/- 0.16 pm/microgram DNA, p less than 0.05) but not in animals receiving estradiol benzoate alone (2.08 +/- 0.32 pm/micrograms DNA versus 1.28 +/- 0.16 pm/micrograms DNA, NS). Compared with treatment with MIX only, MIX + relaxin 10 ng/ml and MIX + relaxin 50 ng/ml produced greater increases in uterine cyclic adenosine monophosphate in the steroid combination group than in the estradiol benzoate controls (144.8% and 233.7% versus 71.7% and 156.6%, respectively). These results suggest that the synergism of relaxin and progesterone in inhibiting uterine contractility may be mediated by intracellular cyclic adenosine monophosphate.     

Amniotic fluid adenosine 3',5' - monophosphate in prostaglandin-induced midtrimester abortions.

Cyclic adenosine 3'5-monophosphate (cAMP) was serially measured in the amniotic fluid (AF) of 9 patients undergoing midtrimester abortions induced by intraamniotic prostaglandins. The results demonstrate an increase in AF cAMP ranging from 2- to 7-fold within the 6 hours of observation. The fetal heart tones were closely monitored by a Doppler instrument and the time from injection of abortifacient to fetal demise (IDT) and to fetal expulsion (IAT) was accurately recorded. No correlation between the rate of AF cAMP increase and IDT or IAT could be demonstrated. The concentration of cAMP in amniotic fluid obtained from patients with fetal demise in utero was lower than that found in AF of fetuses of corresponding gestational age where the fetus was alive. The significance of AF cAMP as a potential indicator of fetal distress is discussed.     

Effect of two phosphodiesterase inhibitors, cyclic adenosine 3':5'-monophosphate, and a beta-blocking agent on human sperm motility.

Mann-fructose fluid (MF), and MF plus caffeine, MF plus pentoxifylline, MF (dibutyryl cAMP), MF plus propranolol, and MF plus propranolol plus dibutyryl cAMP were individually added to aliquots of semen samples obtained from 18 normal men. These drugs were added to a final concentration of 0.6 mM. One aliquot with no addition served as control. Samples were incubated at 37 degrees C and observed by light microscopy at 30 minutes and at 1,2, and 4 hours after obtained the material. At each observation time, semen quality was evaluated by determinating the percentages of forwardly progresssive spermatozoa, slowly progressive spermatozoa, "in situ" motile spermatozoa, live and nonmotile spermatozoa, and dead spermatozoa. Mann-fructose fluid resulted in a decrease in motility and the duration of activity of spermatozoa, Caffeine seemed to neutralize the deleterus effect on the buffer, whereas pentoxifylline and cAMP seemed to increase the duration of activity of spermatozoa. Propranolol resulted in a dramatic decrease in motility, an effect that could not be neutralized by the simultaneous addition of cAMP.     

Stimulation of cyclic adenosine 3':5'-monophosphate accumulation in human testes in vitro by luteinizing hormone, follicle-stimulating hormone, and prostaglandins.

The gonadotropins luteinizing hormone (LH) and follicle-stimulating hormone (FSH) stimulate cyclic adenosine 3':5'-monophosphate (cyclic AMP) accumulation in human testicular incubates. The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine acts synergistically with LH and FSH to enhance cyclic AMP accumulation. These studies indicate that cyclic AMP accumulation may be involved in the mechanism of action of LH and FSH in the human testes, as has been proposed for rat testes. The prostaglandins (PG PGE1, PGE2, PGA1, and PGF2alpha stimulate cyclic AMP levels at 10(-4) M in human testes. The E type prostaglandins are the most potent. They induce half-maximal stimulation of cyclic AMP at 7 x 10(-7) M.     

In vitro recovery of human chorionic gonadotropin-stimulated cyclic adenosine 3',5'-monophosphate production in desensitized human granulosa-luteal cells

Human granulosa-luteal cell production of cyclic adenosine 3',5'-monophosphate (cAMP) and progesterone (P) were studied in response to purified human chorionic gonadotropin (hCG) in cultured cells from hyperstimulated follicles of in vitro fertilized patients. The hCG injection given to the patients 36 hours before laparoscopy caused partial desensitization of adenylate cyclase of these cells to gonadotropins. Preincubation of the cells in hormone free medium for 2 to 3 days significantly increased their cAMP responsiveness to hCG. P production was stimulated initially by hCG and showed no desensitization. In the cells preincubated for 72 hours without hCG, a subsequent stimulus of 50 ng/ml of hCG elicited maximal cAMP response, whereas 1 ng/ml of hCG was sufficient to bring about maximal P secretion. Time-course studies indicated that maximal cAMP response to hCG was obtained in 1 to 3 hours. Both basal and hCG-stimulated P accumulation continued to rise for up to 24 hours. Preincubation of granulosa-luteal cells from hyperstimulated follicles improves the cells' cAMP responsiveness to hCG, whereas P response remains unaltered.     

Changes in plasma adenosine concentrations during normal pregnancy

The aim of this study was to measure changes in plasma adenosine concentration [ADO] during a normal pregnancy and to evaluate the possible role of platelets and red blood cells (RBC) as causes of changes in plasma [ADO]. We measured the plasma [ADO] in normal pregnant women (n = 11) during the first, second and third trimesters. The mean plasma [ADO] in the third trimester was 0.41 +/- 0.08 microM (means +/- SEM), significantly higher than in the first and second trimesters (p < 0.05). In pregnant women, platelet and RBC counts, hematocrit and hemoglobin concentration decreased slightly throughout the pregnancy. The elevation in the plasma [ADO] correlated inversely with the platelet count (r = -0.43, p < 0.05). These results suggest that an increase in the plasma [ADO] in the third trimester may be attributed to the enhanced adenosine release from activated platelets.     

Alterations of polyamines in body fluids during pregnancy in rats

Polyamines in plasma, urine, and amniotic fluid during pregnancy in rats were measured using high performance liquid chromatography (HPLC). Putrescine and spermidine in the plasma, and putrescine and spermine in the urine were elevated during pregnancy. The sharp increase in putrescine in the late stage of pregnancy in body fluids was particularly characteristic. Slight increases in plasma and urinary spermine levels were observed in the early stage of pregnancy. It was found that the administration of the steroid hormones, progesterone, estradiol-17 beta, and estriol, to virgin rats did not influence the concentrations of urinary polyamines, and that urinary putrescine levels did not rise in experimental IUGR rats, but they rose in the urine of the sham operation group. These data suggest that changes in polyamines in plasma, urine, and amniotic fluid were possibly subject to fetal growth and/or functional differentiation more than hormonal regulation during rat pregnancy.     

Glucagon stimulated plasma cyclic adenosine-3',5'-monophosphate in the differential diagnosis of jaundice.

The amount of plasma cyclic adenosine-3',5'-monophosphate was estimated before and 15 minutes after the intravenous injection of 1 milligram of glucagon in 34 patients with obstructive and in 23 patients with nonobstructive jaundice. After stimulation with glucagon, the median adenosine-3',5'-monophosphate concentration in obstructive jaundice rose fortyfold or more, while in nonobstructive jaundice, the increase was twentyfold or less. The difference was highly significant. The results indicate that this test can be useful in the differential diagnosis of obstructive and nonobstructive jaundice.     

Haemostasis in normal and abnormal pregnancy

Haemostasis is a complex and dynamic equilibrium involving pro-coagulants, the natural anticoagulation system and fibrinolysis. Normal human pregnancy is associated with profound alterations to the process of haemostasis such that the pro-coagulant effect becomes dominant. There are very few studies which have attempted to elucidate the adaptations that take place in the uteroplacental circulation where the haemostatic system faces the conflicting tasks of maintaining blood fluidity during pregnancy while preparing for the haemostatic challenge of delivery. It is hypothesised that excessive thrombosis within the uteroplacental circulation provides the mechanistic basis for the reported associations between the inherited thrombophilias and major pregnancy complications. The evidence underpinning this widely quoted hypothesis is weak.     

Changes in the protein conformation of human spermatozoal membranes after treatment with cyclic adenosine 3':5'-monophosphate and human follicular fluid.

Infrared spectra in the amide I and amide II regions of acrosomal membranes isolated from ejaculated human spermatozoa indicate the presence of a high proportion of the constitutive proteins in the most stable protein configuration, the antiparallel beta-conformation. Since the infrared spectra obtained with the membranes suspended in D2O or after extraction of the lipid components do not show any significant change, it can be postulated that the antiparallel pleated sheet conformation of human spermatozoal membrane proteins is independent of the hydrated state and of the lipid constitution of the membrane.