Pregnancy outcomes in women with primary biliary cholangitis and primary sclerosing cholangitis: a retrospective cohort study.

OBJECTIVE: To determine maternal, obstetric and neonatal outcomes in a cohort of women with primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC). DESIGN: Retrospective cohort study. SETTING: Ten specialist centres managing pregnant women with liver disease. POPULATION: Women with a diagnosis of PBC and PSC and a pregnancy of ≥20 completed weeks of gestation. METHODS: Retrospective case notes review. MAIN OUTCOME MEASURES: Adverse outcomes were defined as: maternal - development of ascites, variceal bleeding, encephalopathy and jaundice; obstetric events - gestational hypertension, pre-eclampsia and postpartum haemorrhage; and neonatal - stillbirth, preterm delivery and admission to neonatal unit. The relationship of alanine transferase (ALT) and bile acid levels with gestation at delivery was studied. RESULTS: The first recorded pregnancies of 34 women with PSC and 27 women with PBC were analysed. There were 60 live births and one intrapartum stillbirth that did not occur in the context of maternal cholestasis. The overall median gestation of delivery was 38 weeks but the rate of preterm birth was 28% (17/61 deliveries), 76% (13/17) of which were spontaneous. Gestation at birth negatively correlated with maternal serum ALT concentration at booking (P = 0.017) and serum bile acid concentration during pregnancy (P = 0.016). There were no other significant correlations and maternal and neonatal outcomes were good. CONCLUSIONS: Pregnancy in PBC and PSC is well tolerated, but women should be counselled regarding the increased risk of preterm birth. Measurement of maternal ALT and bile acids may help identify women at risk of preterm delivery. TWEETABLE ABSTRACT: Pregnancy in women with PBC and PSC is well tolerated; however, rates of preterm birth are high and are related to maternal bile acid levels.

Alternative role for prolactin-releasing peptide in the regulation of food intake.

Prolactin-releasing peptide (PrRP) is a peptide ligand for the human orphan G-protein-coupled receptor hGR3/GPR10 and causes the secretion of prolactin from anterior pituitary cells. However, the lack of immunoreactive staining for PrRP in the external layer of the median eminence seems to rule out this peptide as a classical hypophysiotropic hormone and, furthermore, PrRP is less effective than another inducer of prolactin secretion, thyrotropin-releasing hormone, both in vitro and in vivo. Here we show a reduction in the expression of PrRP mRNA during lactation and fasting and an acute effect of PrRP on food intake and body weight, supporting the hypothesis of an alternative role for the peptide.

Reduced expression of the KATP channel subunit, Kir6.2, is associated with decreased expression of neuropeptide Y and agouti-related protein in the hypothalami of Zucker diabetic fatty rats.

The link between obesity and diabetes is not fully understood but there is evidence to suggest that hypothalamic signalling pathways may be involved. The hypothalamic neuropeptides, pro-opiomelanocortin (POMC), neuropeptide Y (NPY) and agouti-related protein (AGRP) are central to the regulation of food intake and have been implicated in glucose homeostasis. Therefore, the expression of these genes was quantified in hypothalami from diabetic Zucker fatty (ZDF) rats and nondiabetic Zucker fatty (ZF) rats at 6, 8, 10 and 14 weeks of age. Although both strains are obese, only ZDF rats develop pancreatic degeneration and diabetes over this time period. In both ZF and ZDF rats, POMC gene expression was decreased in obese versus lean rats at all ages. By contrast, although there was the expected increase in both NPY and AGRP expression in obese 14-week-old ZF rats, the expression of NPY and AGRP was decreased in 6-week-old obese ZDF rats with hyperinsulinaemia and in 14-week-old rats with the additional hyperglycaemia. Therefore, candidate genes involved in glucose, and insulin signalling pathways were examined in obese ZDF rats over this age range. We found that expression of the ATP-sensitive potassium (K(ATP)) channel component, Kir6.2, was decreased in obese ZDF rats and was lower compared to ZF rats in each age group tested. Furthermore, immunofluorescence analysis showed that Kir6.2 protein expression was reduced in the dorsomedial and ventromedial hypothalamic nuclei of 6-week-old prediabetic ZDF rats compared to ZF rats. The Kir6.2 immunofluorescence colocalised with NPY throughout the hypothalamus. The differences in Kir6.2 expression in ZF and ZDF rats mimic those of NPY and AGRP, which could infer that the changes occur in the same neurones. Overall, these data suggest that chronic changes in hypothalamic Kir6.2 expression may be associated with the development of hyperinsulinaemia and hyperglycaemia in ZDF rats.

Regional expression of a MCD-peptide and dendrotoxin I-sensitive voltage-dependent potassium channel in rat brain.

In situ hybridization histochemistry has been used to analyze the regional expression of a class of voltage-dependent K+ channel that is sensitive to two polypeptide toxins (MCD peptide and dendrotoxin I) that produce spectacular effects on brain function. A heterogeneous expression of this K+ channel was observed throughout the brain. High mRNA contents were observed in the granule cells of the gyrus dentatus as well as in pyramidal cells of the Ammon horn (CA3 greater than CA1) and in the cerebellum. Conversely, low levels of expression were found in basal ganglia (caudate putamen, globus pallidus, and ventral pallidum).

Activated protein C sensitivity, protein C, protein S and coagulation in normal pregnancy.

A prospective study of activated protein C sensitivity, protein C, protein S, and other coagulation factors in 239 women during normal pregnancy was carried out. Protein C activity appeared unaffected by gestation, although an elevation of protein C activity was observed in the early puerperium. A fall in total and free protein S with increasing gestation was observed. Activated protein C sensitivity ratio (APC:SR) showed a progressive fall through pregnancy. This fall correlated with changes in factor VIIIc, factor Vc and protein S. 38% of subjects, with no evidence of Factor V Leiden or anticardiolipin antibodies, showed a low APC:SR (APC:SR <2.6) in the third trimester of pregnancy. Aside from a significant reduction in birth weight, no difference in pregnancy outcome was observed between these subjects and those with a normal APC:SR. Activated protein C sensitivity ratio, modified by pre-dilution of patient samples with factor V depleted plasma, showed no consistent trend with gestation.

Agouti-related protein (83-132) is a competitive antagonist at the human melanocortin-4 receptor: no evidence for differential interactions with pro-opiomelanocortin-derived ligands.

Interactions between pro-opiomelanocortin (POMC)-derived peptides, agouti-related protein (AGRP) and the melanocortin-4 receptor (MC4-R) are central to energy homeostasis. In this study we have undertaken comprehensive pharmacological analysis of these interactions using a CHOK1 cell line stably transfected with human MC4-R. Our main objectives were (1) to compare the relative affinities and potencies of POMC-derived peptides endogenously secreted within the hypothalamus, (2) to investigate the potency of AGRP(83-132) antagonism with respect to each POMC-derived peptide and (3) to determine whether AGRP(83-132) and POMC-derived peptides act allosterically or orthosterically. We have found that beta melanocyte-stimulating hormone (betaMSH), desacetyl alpha MSH (da-alphaMSH) and adrenocorticotrophic hormone all have very similar affinities and potencies at the MC4-R compared with the presumed natural ligand, alphaMSH. Moreover, even MSH precursors, such as beta lipotrophic hormone, showed significant binding and functional activity. Therefore, many POMC-derived peptides could have important roles in appetite regulation and it seems unlikely that alphaMSH is the sole physiological ligand. We have shown that AGRP(83-132) acts as a competitive antagonist. There was no significant difference in the potency of inhibition by AGRP(83-132) or agouti(87-132) at the MC4-R, regardless of which POMC peptide was used as an agonist. Furthermore, we have found that AGRP(83-132) has no effect on the dissociation kinetics of radiolabelled Nle4,D-Phe7 MSH from the MC4-R, indicating an absence of allosteric effects. This provides strong pharmacological evidence that AGRP(83-132) acts orthosterically at the MC4-R to inhibit Gs-coupled accumulation of intracellular cAMP.

Changes in prostaglandin synthesis and metabolism associated with labour, and the influence of dexamethasone, RU 486 and progesterone.

The objective was to compare the changes in prostaglandin synthesis and metabolism occurring within the fetal membranes that are associated with the onset of parturition and to study the effect of steroid hormones on prostaglandin metabolism. A tissue explant study was made of discs of amnion and chorion obtained from 24 pregnant women at 37-42 weeks' gestation following spontaneous labour and delivery (12 women) and elective caesarean section (12 women). Significantly more prostaglandin E2 (PGE2) and PGF2 alpha were synthesized by amnion obtained following spontaneous labour than elective caesarean section. Arachidonic acid stimulated both PGE2 and PGF2 alpha synthesis by amnion in both groups. Phorbol myristoyl acetate stimulated PGE2 synthesis in both groups. There was no difference between the groups in the capacity of the chorion to metabolize prostaglandins. Mifepristone (RU 486) reduced the metabolism of added PGE2 following spontaneous labour, while dexamethasone and progesterone had no effect on prostaglandin metabolism. In conclusion, the increase in concentration of PGE2 and PGF2 alpha associated with the onset of spontaneous labour is the result of an increase in synthesis rather than a reduction in metabolism. There was no decrease in metabolism to account for the increase in prostaglandin concentrations and, with the exception of mifepristone, metabolism was not altered by the addition of steroid hormones.

Direct regulation of GnRH transcription by CRF-like peptides in an immortalized neuronal cell line.

The existence of a CRF-dependent inhibition of GnRH transcription was investigated using a neuronal GnRH-expressing cell line (Gn11) stably transfected with mouse (-611 bp) or chicken (-3000 bp) GnRH promoter/luciferase reporter constructs. The presence of the CRF-R1 receptor was established using a specific CRF-R1 antiserum. After 7 h of incubation, urotensin-I and sauvagine increased the mouse GnRH-reporter bioluminescence by 1.3- and 1.2-fold, respectively, compared with control cells. Subsequently, CRF, urotensin-I and sauvagine decreased luciferase reporter activity to about 60% of the control values after 14 h. Similar trends occurred with the chicken GnRH promoter with UI increasing reporter gene activity 2.4-fold over the controls after 14 h incubation. These data provide additional evidence for the direct regulation of GnRH transcription by CRF-like peptides.

The effects of mifepristone on cervical ripening and labor induction in primigravidae.

OBJECTIVE: To compare the effects of 50 mg or 200 mg of oral mifepristone with placebo on cervical ripening and induction of labor in primigravid women at term with unfavorable cervices. METHODS: This was a double-blind study in which 80 primigravidae at term with a modified Bishop score of 4 or less were randomly assigned to one of three treatment groups. They were assessed at 24-hour intervals for 72 hours, after which labor was induced if it had not occurred spontaneously. RESULTS: Two hundred milligrams of mifepristone resulted in a favorable cervix (with a Bishop score greater than 6 or in spontaneous labor) in significantly more women than placebo (P = .01). An improvement in cervical ripening was seen in the group given 50 mg of mifepristone, but this was not statistically significant. There were more cesarean deliveries performed for fetal distress in the group treated with 200 mg of mifepristone than placebo, but this was not statistically significant and was not associated with any differences between groups in terms of neonatal outcome. CONCLUSION: Mifepristone, a progesterone antagonist, is known to cause softening and dilation of the human early pregnant cervix and an increase in uterine activity. It is theoretically attractive for use as an adjunct in cervical priming and labor induction. In this study, 200 mg of mifepristone was significantly more likely to result in a favorable cervix than placebo.

Cloning and characterization of corticotropin-releasing factor and urocortin in Syrian hamster (Mesocricetus auratus).

Corticotropin-releasing factor and urocortin belong to a superfamily of neuropeptides that includes the urotensins-I in fishes and the insect diuretic peptides. Sequence analysis suggests that urocortin is the mammalian ortholog of urotensin-I, although the physiological role for this peptide in mammals is not known. Within the Rodentia, hamsters belong to a phylogenetically older lineage than that of mice and rats and possess significant differences in hypothalamic organization. We have, therefore, cloned the coding region of the Syrian hamster (Mesocricetus auratus) corticotropin-releasing factor and urocortin mature peptide by polymerase chain reaction. Hamster urocortin was prepared by solid-phase synthesis, and its pharmacological actions on human corticotropin-releasing factor R1 and R2 receptors were investigated. The deduced hamster corticotropin-releasing factor amino acid sequence and cleavage site is identical to that in rat, whereas the urocortin sequence is unique among the urocortin/urotensin-I/sauvagine family in possessing asparagine and alanine in positions 38 and 39, respectively. The hamster urocortin carboxy terminus sequence bears greater structural similarity to the insect diuretic peptide family, suggesting either retrogressive mutational changes within the mature peptide or convergent sequence evolution. Despite these changes, human and hamster urocortin are generally equipotent at cAMP activation, neuronal acidification rate, and R1/R2 receptor affinities.

Mechanisms involved in the stimulatory effect of amniotic fluid on prostaglandin production by human fetal membranes.

This study aims to investigate potential mechanisms involved in the stimulatory effect of amniotic fluid on prostaglandin production by fetal membranes. A cell culture study of amnion and chorion was obtained following elective caesarean section, incubated with amniotic fluid collected at term (37-42 weeks' gestation) following either spontaneous labour (n = 6) or elective caesarean section (n = 6). The effect of addition of cycloheximide and actinomycin D (inhibitors of translation and transcription respectively), and staurosporine and genistein (inhibitors of protein kinase C and tyrosine kinase respectively) to these cultures was investigated. ANOVA was employed for statistical analysis. Cycloheximide and staurosporine significantly inhibited the stimulatory effect of spontaneous labour and elective section amniotic fluid on PGE2 production by amnion, and PGEM production by chorion. Genistein significantly inhibited the stimulatory effect of spontaneous labour amniotic fluid on PGE2 and PGEM production by amnion and chorion respectively. The stimulatory effect of amniotic fluid on prostaglandin production is dependent on new protein synthesis, presumably cyclooxygenase (COX), and stimulation of cell signal transduction pathways involving protein kinase C and tyrosine kinase.

The application of the human beta-globin gene locus control region and murine erythroleukemia cell system to the expression and pharmacological characterization of human endothelin receptor subtypes.

The cDNAs encoding both A and B subtypes of the human endothelin receptor have been inserted into mammalian cell expression vectors that utilize the human globin gene, locus control region. These constructs have been introduced into murine erythroleukemia cells and inducible high level expression of the receptors has been achieved (approximately 1.5-pM/mg membrane protein and approximately 13,500 binding sites/cell for both receptor subtypes). Cell lines expressing these receptors were obtained on a rapid time scale (3-4 weeks), facilitated by the need for the analysis of only small numbers of cell clones/receptor (approximately 6). Competitive binding assays with endothelin-1 gave IC50s of 130 +/- 30 pM for endothelin-A receptor and 160 +/- 30 pM for endothelin-B receptor. Similar studies with the different isoforms of endothelin, sarafatoxin-S6b and -S6c, BQ123 and BQ3020, all gave the expected selectivity profiles. The IC50s for all compounds were in close agreement with those reported for native receptors. Thus, this expression system, which has several advantages over other described expression systems, is capable of rapidly providing large quantities of receptor for detailed pharmacological analyses or drug screening. In addition, the expressed receptors display the expected pharmacological profiles in the absence of any complicating, competing interactions from other subtypes or binding sites.

Effects of metabolic inhibitors on cel lethality and mutation induction in Chinese hamster cells. II. The effect of posttreatment with non-toxic concentrations of thymidine.

The ability of posttreatment exposure to non-toxic concentrations of thymidine (TdR) to enhance the lethal effects of a number of alkylating agents, X-rays and UV and the lethal and mutagenic effects of N'-ethyl-N-nitrosourea (ENU) and N-methyl-N-nitrosourea (MNU) has been examined in V79 cell lines. TdR posttreatment enhanced the cytotoxic effects of ethyl methanesulphonate (EMS), MNU and ENU but not of UV or X-rays and increased both the spontaneous and MNU- and ENU-induced frequencies of azaguanine resistant (AZR) mutants. No significant effect of TdR on the spontaneous frequency of thioguanine resistant (TGR) mutants was demonstrated but the frequency of MNU-induced mutants to TGR premutagenic was enhanced. The effects on expression of both potentially lethal and premutagenic damage were reversed by addition of an equimolar concentration of deoxycytidine (dCdR). The enhancement in spontaneous and induced mutant frequency (IMF) at the HGPRT locus appears to be due to an alteration in the selective efficiency of urine analogous due to alteration in growth kinetics of cells exposed to TdR or treated with alkylated agents or posttreated with thymidine after alkylation damage and not to an alteration in the miscoding potential of alkylated bases.

Relative mutagenicity of antineoplastic drugs and other alkylating agents in V79 Chinese hamster cells, independence of cytotoxic and mutagenic responses.

The mutagenic and cytotoxic effects of 4 antineoplastic drugs, vinblastine, vincristine, adriamycin and nitrogen mustard and of several monofunctional alkylating agents have been assayed in V79 Chinese hamster cells. Vincristine, vinblastine and nitrogen mustard did not significantly increase the frequency of TGRHGPRT- mutants but were were all highly cytotoxic. Adriamycin and the monofunctional alkylating agents were all significantly mutagenic even at the lowest doses tested (approx. 70% survival level). Induced mutant frequency increased linearly with increasing dose whereas dose-response curves for cytotoxicity for these effective mutagens invariably showed a shoulder followed by an exponential decline. At equitoxic doses the relative mutagenic effectiveness was MNU > ENU > EMS > MNNG > MMS greater than or equal to DMS. MNU was approx. 20 times more effective than MMS and DMS. Measurement of the total amount of alkylation and the relative amounts of reaction with individual DNA bases at approx. equitoxic doses of MNU and DMS indicated a significantly higher O6/N7 ratio after MNU (0.15) than after DMS (0.005). However, approx. equal numbers of mutants/10(5) cells/microM O6-Meguanine were induced by these 2 agents. These results support previous conclusions, that mutagenic and cytotoxic responses are independent in V79 cells.

Prawn sandwiches, red herrings and abdominal pregnancy.

An unusual presentation of abdominal pregnancy is reported. The difficulty in diagnosis of this form of ectopic pregnancy, and the potential risks of delayed intervention are highlighted. The association with uterine anomaly, in this case uterus didelphys, is discussed.

Induction of labour: new horizons.

Induction of labour may be indicated in the maternal or the fetal interest, and is more likely to be successful if physiological mechanisms are replicated. Induction of labour in the presence of an unfavourable cervix presents the greatest challenge and pharmacological techniques must encourage cervical ripening if we are to advance our management of this common obstetric intervention.

Human parvovirus B19 in pregnancy.

Parvovirus B19 infection can result in an adverse outcome when acquired during pregnancy. However, in the majority of cases a successful outcome can be anticipated. Public awareness of this condition is essential and obstetricians should be familiar with the options available to them if they are presented with this clinical problem.

Short-term outcomes following intrauterine transfusion in Scotland.

AIM: To describe neonatal outcomes following intrauterine transfusion (IUT) for severe Rhesus isoimmunisation from 1993 to 2004. RESULTS: 116 neonates who had undergone 457 IUTs (median 4, range 1-9) were identified. Three neonates died, all before 1995 (two because of hypoxic ischaemic multiorgan failure and one because of overwhelming Escherichia coli sepsis). 13 neonates (11%) were delivered by emergency Caesarean section following either IUT complication or spontaneous onset of preterm labour. They were more likely to require intubation (p<0.0001), on-going respiratory support (p=0.0007) and an exchange transfusion (p=0.007). 23 (20%) required an exchange transfusion and 63 (54%) at least one top-up transfusion. CONCLUSIONS: Management of severe Rhesus disease is associated with encouraging neonatal outcomes and most infants can be managed with phototherapy and a few top-up transfusions. IUT complications are rare but significantly increase neonatal mortality and morbidity. Antenatal counselling should address the likely postnatal course for these infants.

Reduced appetite and body mass index with delayed puberty in a mother and son: association with a rare novel sequence variant in the leptin gene.

INTRODUCTION: Leptin deficiency caused by mutations within the leptin gene (LEP) results in severe early onset obesity, hypogonadism, pubertal delay and immune system abnormalities. Constitutional delay in growth and puberty (CDGP) is a common condition seen in paediatric clinics, in which children present with delayed growth and puberty but usually also have a slim body habitus. We hypothesized that LEP variants may play a role in the phenotype seen in CDGP. AIM: To screen a group of children with CDGP for pathogenic sequence variants in LEP. PATIENTS AND METHODS: Denaturing HPLC was used to screen for LEP sequence variants in DNA samples from 78 children with CDGP (predominantly white males) and 112 control subjects. DNA fragments with a WAVE pattern deviant from wild type were directly sequenced. A STAT3 luciferase reporter assay in human embryonic kidney (HEK293) cells transiently transfected with the leptin receptor was used to test activity of mutant leptin. RESULTS: One child with CDGP was identified to be heterozygous for a novel missense variant (c.68C>G), which results in a proline to arginine substitution (p.P23R). This sequence variant was not identified in any of the other control subjects, but was identified in his mother who shared a similar phenotype of slim body habitus, reduced appetite and pubertal delay (menarche aged 15 years). The leptin variant showed similar stability in serum compared with wild type and did not demonstrate increased activity in an in vitro reporter gene assay. CONCLUSIONS: This is the first report of a sequence variant within the LEP gene associated with reduced body mass index rather than obesity. We hypothesize that this variant has increased bioactivity in vivo.