Oxytocin is hydrolyzed by an enzyme in human placenta that is identical to the oxytocinase of pregnancy serum

doi:10.1016/0196-9781(95)02124-8

Peptides

Volume 17, Issue 2, 1996, Pages 257-261

https://doi.org/10.1016/0196-9781(95)02124-8 Get rights and content

Abstract

The hydrolysis of oxytocin (OT) by human placental subcellular fractions and pregnant sera was studied in the presence of bestatin, a potent inhibitor of aminopeptidases, and the antibody against pregnant serum oxyotocinase (P-LAP) (EC 3.4.11.3) by measuring liberated amino acids by high performance liquid chromatography (HPLC). Our immunotitration study and the effect of bestatin on the oxytocin-degrading protease showed that the initiating and responsible protease in oxyotocin degradation in human placenta and pregnant serum is P-LAP.

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Cited by (16)

The effect of augmentation of labour with syntocinon on the fetal CTG using objective computerised analysis: A nested case-control study
2014, European Journal of Obstetrics and Gynecology and Reproductive Biology
Citation Excerpt :
These observations are consistent with available evidence which suggests that oxytocin acts on uterine smooth muscle receptors and does not cross the placenta [12] to enter the fetal circulation. It is rapidly broken down by placental oxytocinases [13] and so any effect on the fetal heart rate is likely to be mediated by uterine activity rather than by a direct effect on the fetus. The effect of syntocinon on the PRSA of babies born with acidaemia is very interesting.
To investigate the effect of syntocinon augmentation on the fetal cardiotocogram (CTG) using computerised analysis. We hypothesised that syntocinon will have no direct effects on the fetal heart rate if used correctly.
A retrospective, nested case–control study.
Intrapartum CTG records from the digital archive at the John Radcliffe Hospital, Oxford, UK.
110 women with singleton pregnancies of >36 weeks gestation, no known congenital abnormality, spontaneous onset of labour and syntocinon augmentation for failure to progress, with start time of syntocinon recorded, from between August 1998 and December 1999, extensively matched to 110 controls who had normally progressing labours.
Eight different CTG features were measured during four time points with OxSys, a computerised numerical analysis system.
Differences in the CTG features over time in cases and controls using ANOVA and Friedman's ANOVA and at each time point between case–control pairs using Student's t-test and the Wilcoxon signed rank test.
After administration, syntocinon increased the frequency, decreased the duration and decreased the resting time between contractions (p < 0.001), resulting in no significant difference between normally progressing labours and those requiring augmentation. The case group had a significantly higher signal stability index (SSI) and fewer decelerations compared to the control group – differences which disappeared after augmentation was commenced (p = 0.025 and 0.033 respectively). Syntocinon did not affect the baseline heart rate, short term variability (STV) or phase rectified signal averaging (PRSA) (p = 0.518, 0.215 and 0.138) in comparison with controls. There was a significant increase in the PRSA in babies born with acidaemia (arterial pH≤7.05) 60–120 min after syntocinon was commenced that was not seen with in babies with a normal pH (p = 0.002).
Syntocinon “normalises” ineffective uterine activity without any direct effect on the fetal heart rate. Therefore its administration does not confound objective computerised analysis. There may be a specific response in PRSA shortly after commencing syntocinon augmentation in the fetus which is subsequently born acidaemic which requires further investigation.
A new approach regarding the treatment of preeclampsia and preterm labor
2011, Life Sciences
Citation Excerpt :
P-LAP degrades the ring structure of OT at the peptide bond between Cys1 and Tyr2 and hydrolyzes the remaining structure in a stepwise fashion from the amino terminal. Prolylendopeptidase, and neutral endopeptidase are not involved in opening the ring structure of OT (Mizutani et al., 1985a; Naruki et al., 1996) indicating that only P-LAP should be regarded as an oxytocinase. Our laboratory was the first to purify P-LAP from retro-placental blood (Sakura et al., 1981).
Both preeclampsia and preterm delivery are important complications in pregnancy and are leading causes for maternal and perinatal morbidity and mortality. The underlying molecular mechanisms of both diseases remain unknown, thus treatments (beta2-stimulants and magnesium sulfate) are essentially symptomatic. Both molecules have molecular weights less than 5–8 kDa and cross the placental barrier thus exerting their effects on the fetus. In addition, the fetus produces peptide hormones that are highly vasoactive and uterotonic and increase in response to maternal stress and with continued development. Fetal peptides are also small molecules that inevitably leak across into the maternal circulation. Aminopeptidases such as placental leucine aminopeptidase (P-LAP) and aminopeptidase A (APA) are large molecules that do not cross the placental barrier. We have shown that APA acts as an antihypertensive agent in the pregnant spontaneously hypertensive rat by degrading vasoactive peptides and as a result returns the animal to a normotensive state. We have also noted that P-LAP acts as an anti-uterotonic agent by degrading uterotonic peptides, and as a result prolongs gestation in the pregnant mouse. Thus, P-LAP and APA represent promising agents for the treatment of preeclampsia and preterm labor by degrading bioactive hormones derived from the feto-placental circulation.
Characterization of a secretase activity for placental leucine aminopeptidase
2001, Archives of Biochemistry and Biophysics
Placental leucine aminopeptidase (P-LAP) is believed to play an important role in the inactivation of small regulatory peptides. P-LAP exists in both membrane-bound and soluble forms and cDNA cloning has demonstrated that P-LAP is a type II membrane protein, which means that its soluble form is released by a specific proteolytic cleavage. In this report, we studied this process in COS7 cells. Inhibitors of serine or aspartic proteases did not affect the secretion of P-LAP, while EDTA and 1,10-phenanthroline inhibited it. In addition, we transfected P-LAP expression vectors that have point mutations of the cleavage site or deletion of the juxtamembrane stalk. Point mutations of the cleavage site resulted in significantly lower secretion of P-LAP. On the contrary, the distance to cleavage site showed no relation to P-LAP secretion. These results suggest that P-LAP secretase has a metalloprotease activity which depends on the amino acid sequence of the cleavage site.
Maternal serum placental leucine aminopeptidase (P-LAP)/oxytocinase and preterm delivery
2001, International Journal of Gynecology and Obstetrics
Objective: To test the applicability of maternal serum placental leucine aminopeptidase (P-LAP) as a parameter for predicting preterm delivery. Patients and Methods: Maternal serum P-LAP activity of 61 consecutive women with preterm labor at admission were assayed and examined to assess the relationship of levels to the incidence of preterm and term deliveries. Results: When P-LAP activity of study patient serum was at or below the 10th percentile, we found a greater ratio of preterm delivery (P=0.0085) and an approximately 2.3-fold increase risk. The sensitivity, specificity, positive predictive value and negative predictive value were 29, 97, 89 and 62%, respectively. Conclusion: Maternal serum P-LAP activity decreases in cases with spontaneous preterm delivery and may be a clinically potential marker for some cases of preterm labor.
Placental leucine aminopeptidase/oxytocinase in maternal serum and placenta during normal pregnancy
2000, Life Sciences
Placental leucine aminopeptidase (P-LAP), which is identical with cystine aminopeptidase as oxytocinase, was found to be homologous with rat insulin-regulated membrane aminopeptidase (IRAP) by sequence comparison. In the current study, we determined the P-LAP levels in maternal serum and placenta during healthy pregnancy. P-LAP activities in maternal serum increased with gestation and rose to the peak of 80 IU/ml at 38 weeks of gestation. Northern blot analysis revealed the increase of P-LAP mRNA levels in placenta in the third trimester compared to the first trimester. P-LAP protein and related activities could be detected in the conditioned medium of placental tissue, while they could not be detecetd in that of human umbilical vein endothelial cells. Immunohistochemically P-LAP was positively stained in the apical membrane of syncytiotrophoblast cells throughout the gestation. These results established the normal range of serum and tissue P-LAP levels during pregnancy and the possible source of serum P-LAP, which will be helpful to elucidate the physiological and clinical roles of P-LAP/oxytocinase/IRAP.
Immunoaffinity purification and characterization of native placental leucine aminopeptidase/oxytocinase from human placenta
2000, Placenta
cDNA cloning of placental leucine aminopeptidase (P-LAP)/cystinyl aminopeptidase (CAP)/oxytocinase demonstrated that this enzyme is a type II integral membrane protein, which means that native P-LAP, found in placenta, is membrane-bound and that the soluble form of this enzyme, found in maternal sera, is most likely derived from the native form. The presence of the different forms of the protein makes it difficult to purify homogeneously. In the current study we prepared antibody specific to native P-LAP and used it to purify native P-LAP from microsomal fractions of human placenta to homogeneity, 5039-fold within 4 h, by immunoaffinity chromatography. Zn²⁺and Cu²⁺strongly inhibited the enzyme but Ca²⁺did not. Amastatin was a more potent inhibitor than bestatin and leupeptin. Using antibodies against native P-LAP, protein having 83 per cent of $l$ -methionine insensitive Leu-p-nitroanilide cleaving activity, was immunoprecipitated from the microsomal fraction of human placenta.
The availability of a specific antibody against native P-LAP permits the rapid purification and the preliminary immunoassay of the enzyme. Establishment of simple purification and assay methods for the native, membrane bound form of P-LAP pave the way to elucidating the roles and processing systems of this enzyme.

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ArticleOxytocin is hydrolyzed by an enzyme in human placenta that is identical to the oxytocinase of pregnancy serum

Abstract

Am. J. Obstet. Gynecol.

J. Biol. Chem.

J. Chromatogr.

J. Biol. Chem.

Am. J. Obstet. Gynecol.

Biochim. Biophys. Acta

Regul. Pept.

Clin. Biochem.

Early Hum. Dev.

Am. J. Obstet. Gynecol.

Peptides

Arch. Biochem. Biophys.

Arch. Biochem. Biophys.

Article
Oxytocin is hydrolyzed by an enzyme in human placenta that is identical to the oxytocinase of pregnancy serum