A randomized prospective study of misoprostol and dinoproston for induction of labor.

BACKGROUND: Misoprostol, a prostaglandin E1 analog registered for the prevention of gastric ulcers in NSAID-drug users, has been reported to be more effective for labor induction than the standard prostaglandin, dinoproston after vaginal application. There have been some concerns about possible hyperstimulation of the uterine activity and about the safety for the fetus with this new drug. METHODS: Two hundred and ten patients, 36 weeks pregnant or more, with an unfavorable cervix, single pregnancy, and intact membranes were randomized to receive misoprostol, 50 micrograms intravaginally every 6 hours, or dinoproston 0.5 mg intracervically every 12 hours for a maximum of 24 hours, for labor induction. RESULTS: Time from induction to delivery was shorter in both primigravidae and multigravidae after the application of misoprostol than dinoproston and failed induction was more common after dinoproston than after misoprostol. There were no differences in the condition of the newborns in the two groups, according to Apgar score, umbilical artery pH or referral to the neonatal unit, even if there were more operative deliveries for suspected fetal asphyxia after misoprostol than after dinoproston. We did not find any significant difference in the frequency of uterine hyperstimulation between the two groups. CONCLUSIONS: In the dosage used, misoprostol seems to be an effective agent for induction of labor in patients with an unfavorable cervix. Low cost and ease of administration make this drug a promising alternative for this purpose. Surprisingly, there is little interest from the manufacturer to promote the use of this drug for labor induction.

[Large fetus--a retrospective study]. / Stort foster--en retrospektiv studie.

BACKGROUND: Large foetus is associated with increased risk of complications during labour and delivery. MATERIAL AND METHODS: Data on deliveries and outcome of 504 macrosomic newborns (> or = 4,500 g) born 1995-97 at Rogaland Central Hospital, Norway. RESULTS: 74% of the mothers were multiparae. Mean weight increase during pregnancy was 17.6 kg. In 76% of the pregnancies delivery occurred after term. There were large margins of error both in terms of measurement of fundal height and after estimation with ultrasound. 12% of the newborns had shoulder dystocia. Six children were diagnosed with a brachial plexus injury; three injuries were still present three months after birth. Induction of labour did not increase the Apgar score or diminish the frequency of shoulder dystocia, but it did increase the frequency of operative deliveries and transfer to the neonatal unit. INTERPRETATION: The study confirms that large foetus is associated with risk of complications.

Phosphorylation and activation of hormone-sensitive adipocyte phosphodiesterase type 3B.

Phosphodiesterases (PDEs) include a large group of structurally related enzymes that belong to at least seven related gene families (PDEs 1-7) that differ in their primary structure, affinity for cAMP and cGMP, response to specific effectors, sensitivity to specific inhibitors, and regulatory mechanism. One characteristic of PDE3s involves their phosphorylation and activation in response to insulin as well as to agents that increase cAMP in adipocytes, hepatocytes, and platelets and in response to insulin-like growth factor 1 in pancreatic beta cells. In adipocytes, activation of the membrane-associated PDE3B is the major mechanism whereby insulin antagonizes catecholamine-induced lipolysis. PDE3B activation results in increased degradation of cAMP and, thereby, a lowering of the activity of cAMP-dependent protein kinase (PKA). The reduced activity of PKA leads to a net dephosphorylation and decreased activity of hormone-sensitive lipase and reduced hydrolysis of triglycerides. Activation of the rat adipocyte PDE3B by insulin is associated with phosphorylation of serine-302. The mechanism whereby insulin stimulation leads to phosphorylation/activation of PDE3B is only partly understood. In rat adipocytes, lipolytic hormones and other agents that increase cAMP, including isoproterenol, also induce rapid phosphorylation, presumably catalyzed by PKA, of serine-302 of PDE3B. The phosphorylation is associated with activation of the enzyme, most likely representing "feedback" regulation of cAMP, presumably allowing close coupling of the regulation of steady-state concentrations of both cAMP and PKA and, thereby, control of lipolysis. In the review we describe methods and strategies used in the authors' laboratories to study phosphorylation and activation of PDE3B in adipocytes and in vitro.

Insulin-induced phosphorylation and activation of phosphodiesterase 3B in rat adipocytes: possible role for protein kinase B but not mitogen-activated protein kinase or p70 S6 kinase.

Insulin stimulation of adipocytes results in serine phosphorylation/activation of phosphodiesterase 3B (PDE 3B) and activation of a kinase that phosphorylates PDE 3B in vitro, key events in the antilipolytic action of this hormone. We have investigated the role for p70 S6 kinase, mitogen-activated protein kinases (MAP kinases), and protein kinase B (PKB) in the insulin signaling pathway leading to phosphorylation/activation of PDE 3B in adipocytes. Insulin stimulation of adipocytes resulted in increased activity of p70 S6 kinase, which was completely blocked by pretreatment with rapamycin. However, rapamycin had no effect on the insulin-induced phosphorylation/activation of PDE 3B or the activation of the kinase that phosphorylates PDE 3B. Stimulation of adipocytes with insulin or phorbol myristate acetate induced activation of MAP kinases. Pretreatment of adipocytes with the MAP kinase kinase inhibitor PD 98059 was without effect on the insulin-induced activation of PDE 3B. Furthermore, phorbol myristate acetate stimulation did not result in phosphorylation/activation of PDE 3B or activation of the kinase that phosphorylates PDE 3B. Using Mono Q and Superdex chromatography, the kinase that phosphorylates PDE 3B was found to co-elute with PKB, but not with p70 S6 kinase or MAP kinases. Furthermore, both PKB and the kinase that phosphorylates PDE 3B were found to translocate to membranes in response to peroxovanadate stimulation of adipocytes in a wortmannin-sensitive way. Whereas these results suggest that p70 S6 kinase and MAP kinases are not involved in the insulin-induced phosphorylation/activation of PDE 3B in rat adipocytes, they are consistent with PKB being the kinase that phosphorylates PDE 3B.

Regulation of protein kinase B in rat adipocytes by insulin, vanadate, and peroxovanadate. Membrane translocation in response to peroxovanadate.

Protein kinase B (PKB) (also referred to as RAC/Akt kinase) has been shown to be controlled by various growth factors, including insulin, using cell lines and transfected cells. However, information is so far scarce regarding its regulation in primary insulin-responsive cells. We have therefore used isolated rat adipocytes to examine the mechanisms, including membrane translocation, whereby insulin and the insulin-mimicking agents vanadate and peroxovanadate control PKB. Stimulation of adipocytes with insulin, vanadate, or peroxovanadate caused decreased PKB mobility on sodium dodecyl sulfate-polyacrylamide gels, indicative of increased phosphorylation, which correlated with an increase in kinase activity detected with the peptide KKRNRTLTK. This peptide was found to detect activated PKB selectively in crude cytosol and partially purified cytosol fractions from insulin-stimulated adipocytes. The decrease in electrophoretic mobility and activation of PKB induced by insulin was reversed both in vitro by treatment of the enzyme with alkaline phosphatase and in the intact adipocyte upon removal of insulin or addition of the phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor wortmannin. Significant translocation of PKB to membranes could not be demonstrated after insulin stimulation, but peroxovanadate, which appeared to activate PI 3-kinase to a higher extent than insulin, induced substantial translocation. The translocation was prevented by wortmannin, suggesting that PI 3-kinase and/or the 3-phosphorylated phosphoinositides generated by PI 3-kinase are indeed involved in the membrane targeting of PKB.

Protein kinase A-dependent activation of PDE4 (cAMP-specific cyclic nucleotide phosphodiesterase) in cultured bovine vascular smooth muscle cells.

Incubation of cultured bovine vascular smooth muscle cells (VSMC) with forskolin increased cAMP as measured by an increase in cAMP-dependent protein kinase (PKA) activation (PKA ratio). Forskolin also produced a concentration- and time-dependent increase in activity (3-5-fold within 15 min) of a PDE4 (cAMP-specific cyclic nucleotide phosphodiesterase). The increase in PDE4 activity was not affected by cycloheximide and thus not likely due to increased synthesis of the enzyme. Activation, which was preserved during partial purification of the enzyme by chromatography on Sephacryl S-200 and MonoQ, was most likely due to a covalent modification. Incubation of cell homogenates with the catalytic subunit of PKA (PKA(c)) induced a approximately 5-fold activation of PDE4 with a time course similar to that in intact cells after forskolin addition. The forskolin-mediated activation was reversed during incubation of homogenates at room temperature for two hours. Addition of PKA(c) resulted in rapid reactivation of PDE4. These data are consistent with the hypothesis that rapid, reversible activation of PDE4 in cultured VSMC is mediated by PKA.

Identification of the site in the cGMP-inhibited phosphodiesterase phosphorylated in adipocytes in response to insulin and isoproterenol.

Stimulation of rat adipocytes with insulin and isoproterenol results in serine phosphorylation and activation of the adipocyte cGMP-inhibited phosphodiesterase (cGI PDE), events believed to be important in the antilipolytic action of insulin (Degerman, E., Smith, C.J., Tornqvist, H., Vasta, V., Manganiello, V.C., and Belfrage, P. (1990) Proc. Natl. Acad. Sci. U.S.A. 87,533-537). Here we demonstrate, by two-dimensional phosphopeptide mapping, that the major phosphopeptide generated by trypsin, or trypsin followed by Asp-N protease digestion of [32P]cGI PDE phosphorylated in adipocytes in response to isoproterenol and/or insulin, in each case co-migrates with the phosphopeptide released by the same treatment of M297FRRPS(P)LPCISREQ310. This peptide was synthesized based on the deduced sequence of the cloned rat adipocyte cGI PDE and phosphorylated by cAMP-dependent protein kinase (protein kinase A). Radiosequencing of authentic and synthetic tryptic 32P-peptides showed that a single site in cGI PDE (Ser302) was phosphorylated in adipocytes incubated with isoproterenol and/or insulin. The more than additive phosphorylation and activation of cGI PDE in response to the two hormones found in this report and previously (Smith, C.J., Vasta, V., Degerman, E., Belfrage, P., and Manganiello, V.C. (1991) J. Biol. Chem. 266, 13385-13390) is proposed to reflect cross-talk between their respective signal transduction pathways at the level of the cGI PDE serine protein kinase or upstream regulatory component(s).

Type III cGMP-inhibited cyclic nucleotide phosphodiesterases (PDE3 gene family).

Seven different but related cyclic nucleotide phosphodiesterase (PDE) gene families have been identified. Type III cGMP-inhibited (cGI) PDEs, the PDE3 gene family, are found in many tissues. cGI PDEs exhibit a high affinity for both cAMP and cGMP, and are selectively and relatively specifically inhibited by certain agents which augment myocardial contractility, promote smooth muscle relaxation and inhibit platelet aggregation. Adipocyte, platelet, and hepatocyte cGI PDE activities are regulated by cAMP-dependent phosphorylation. Insulin-induced phosphorylation/activation of adipocyte and hepatocyte cGI PDEs is thought to be important in acute regulation of triglyceride and glycogen metabolism by insulin. Two distinct cGI PDE subfamilies, products of distinct but related genes, have been identified. They exhibit the domain structure common to PDEs with a carboxyterminal region, conserved catalytic domain and divergent regulatory domain. In their catalytic domains cGI PDEs contain a 44 amino acid insertion not found in other PDE families. The expression of cGIP1 and cGIP2 mRNAs differs in different rat tissues, suggesting distinct functions for the two cGI PDE subfamilies, i.e., cGIP1 in adipose tissue, liver, testis and cGIP2 in myocardium, platelets and smooth muscle.

Evidence for the key role of the adipocyte cGMP-inhibited cAMP phosphodiesterase in the antilipolytic action of insulin.

Enhancement of cAMP degradation by increased cGMP-inhibited cAMP phosphodiesterase (cGI-PDE) activity is thought to be an important component of the mechanism whereby insulin counteracts catecholamine-induced lipolysis in adipocytes. In this study the selective cGI-PDE inhibitor OPC3911 was used to evaluate this role of cGI-PDE activation in intact rat adipocytes with special reference to changes in cAMP levels measured as cAMP-dependent protein kinase (cAMP-PK) activity ratios. OPC3911 completely blocked (IC50 = 0.3 microM) the maximal inhibitory effect of insulin on noradrenaline-induced lipolysis and the net dephosphorylation of hormone-sensitive lipase and other intracellular target proteins for insulin action, whereas insulin-induced lipogenesis was not changed. The effect of OPC3911 on cAMP-PK activity ratios at different levels of lipolysis achieved by noradrenaline stimulation revealed that the reduction of cAMP-PK caused by 1 nM insulin was completely blocked by 3 microM OPC3911. The effect of OPC3911 was not due to an excessive increase in cellular cAMP resulting in 'supramaximal' lipolysis unresponsive to insulin. These data demonstrate that reduction in cAMP levels by the activation of cGI-PDE may be sufficient to account for the antilipolytic action of insulin.

Essential role of phosphatidylinositol 3-kinase in insulin-induced activation and phosphorylation of the cGMP-inhibited cAMP phosphodiesterase in rat adipocytes. Studies using the selective inhibitor wortmannin.

Incubation of rat adipocytes with wortmannin, a potent and selective phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor, completely blocked the antilipolytic action of insulin (IC50 = 100 nM), the insulin-induced activation and phosphorylation of cGMP-inhibited cAMP phosphodiesterase (cGI-PDE) as well as the activation of the insulin-stimulated cGI-PDE kinase (IC50 = 10-30 nM). No direct effects of the inhibitor on the insulin-stimulated cGI-PDE kinase, the cGI-PDE and the hormone-sensitive lipase were observed. These data suggest that activation of PI 3-kinase upstream of the insulin-stimulated cGI-PDE kinase in the antilipolytic insulin signalchain has an essential role for insulin-induced cGI-PDE activation/phosphorylation and anti-lipolysis.