Amniotic fluid and plasma concentrations of pregnancy-associated plasma protein-A (PAPP-A) throughout pregnancy: comparison with other fetoplacental products

The development of a sensitive radioimmunoassay has enabled measurements of pregnancy-associated plasma protein-A (PAPP-A) to be performed from early pregnancy. The present paper compares the plasma concentrations of PAPP-A with the levels of two trophoblastic proteins, human placental lactogen (hPL) and the beta-subunit of human chorionic gonadotrophin (beta-hCG), with a steroid of fetoplacental origin, total oestriol (total E3), and with a fetal protein, alpha-fetoprotein (AFP). PAPP-A was also measured in amniotic fluid and in maternal urine. In contrast with the secretion of the other substances studied, which either reach a plateau or even decrease during the last 4 weeks of pregnancy, PAPP-A steadily increased in the maternal circulation from 7 to 40 weeks gestation. It is proposed that PAPP-A production is either not related to placental mass or that PAPP-A is not of trophoblastic origin. The increase of PAPP-A in amniotic fluids parallels the increase in maternal blood; virtually no PAPP-A is excreted in urine.     

First trimester maternal serum free beta human chorionic gonadotrophin and pregnancy associated plasma protein A as predictors of pregnancy complications

Objective To examine the value of first trimester maternal serum free β human chorionic gonadotrophin (β hCG) and pregnancy associated plasma protein A (PAPP-A) as predictors of pregnancy complications.
Design Screening study.
Setting Antenatal clinics.
Population Singleton pregnancies at 10–14 weeks of gestation.
Methods Maternal serum free β hCG and PAPP-A were measured at 10–14 weeks of gestation in 5584 singleton pregnancies. In the 5297 (94.9%) pregnancies with complete follow up free β hCG and PAPP-A were compared between those with normal outcome and those resulting in miscarriage, spontaneous preterm delivery, pregnancy induced hypertension or fetal growth restriction and in those with pre-existing or gestational diabetes.
Results Maternal serum PAPP-A increased and β hCG decreased with gestation. The multiple of median maternal serum PAPP-A was significantly lower in those pregnancies resulting in miscarriage, pregnancy induced hypertension, growth restriction and in those with pre-existing or gestational diabetes mellitus, but not in those complicated by spontaneous preterm delivery. The level was < 10th centile of the reference range in about 20% of the pregnancies that subsequently resulted in miscarriage or developed pregnancy induced hypertension or growth restriction, and in 27% of those that developed gestational diabetes. Maternal serum free β hCG was < 10th centile of the reference range in about 15% of the pregnancies that subsequently resulted in miscarriage or developed pregnancy induced hypertension or growth restriction, and in 20% of those that developed gestational diabetes.
Conclusion Low maternal serum PAPP-A or β hCG at 10–14 weeks of gestation are associated with subsequent development of pregnancy complications.     

Does ultrasound examination render biochemical tests obsolete in the prediction of early pregnancy failure?

Summary. Serum levels of fetal, placental and maternal hormones and proteins [β-fetoprotein (AFP), human chorionic gonadotrophin, human placental lactogen. schwangerschaftsprotein 1, pregnancy associated plasma protein-A (PAPP-A), oestradiol-17β, progesterone, pregnancy zone protein] were measured in 108 women with bleeding during the first half of pregnancy. Ultrasound examination at the time of each blood sampling revealed a fetal heart action on at least one occasion in 77 women. Spontaneous abortion occurred in 42 pregnancies, 31 of these showed no ultrasound sign of fetal life, whilst the fetal heart action was observed repeatedly until abortion in the remaining 11 women. Abnormally low levels of PAPP-A were most likely to indicate pregnancy failure, in particular if the fetal heart action was seen at the time of blood sampling. The predictive value, sensitivity and relative risk of a single depressed PAPP-A level were respectively 49, 89 and 41%, the predictive value of a normal result being 99%. With the exception of AFP, all other biochemical indices examined were consistently in the normal range in this group of women. If ultrasound findings were not considered, the biochemical indices were of comparable value in the prediction of spontaneous abortion. PAPP-A levels were uniformly depressed in all patients who spontaneously aborted, frequently weeks before this event, in the presence of a live fetus.     

Tissue and plasma concentrations of pregnancy-associated plasma protein-A (PAPP-A): comparison with other fetoplacental products

Summary. Human placental lactogen (hPL), α-fetoprotein (AFP), prolactin (PRL) and pregnancy-associated plasma protein-A (PAPP-A) were measured by radioimmunoassay in plasma, in homogenates of trophoblast, decidua, chorion, amnion and in amniotic fluid from 10 patients after non-complicated term delivery. Plasma samples and homogenates of trophoblast and decidua were also collected from 10 patients undergoing surgical termination of pregnancy between 7 and 12 weeks gestation. In addition, plasma and endometrial samples from 10 patients undergoing hysterectomy for other indications than malignancy were analysed for comparison. The highest tissue concentrations of hPL, AFP and PRL corresponded in each case to the known site of synthesis. For PAPP-A the highest concentration was found in maternal plasma at term [238·8±75·6 (SEM)μg/ml]. The highest tissue concentration was found at term in the decidua (57·0±2·0 μg/g), more than three times higher than that in the trophoblast (16·9±5·4 μg/g). The concentrations of PAPP-A in endometrial samples from non-pregnant women (1·9±0·6 μg/g) was 40 times higher than that in the corresponding plasma samples (0·05±0·02 μg/ml). These observations point to the decidua as a possible source of PAPP-A.     

In vitro production of pregnancy-associated plasma protein-A (PAPP-A) by trophoblastic cells

Pregnancy-associated plasma protein-A (PAPP-A) a high molecular weight glycoprotein, is found in high concentration in the maternal circulation during pregnancy. Immunohistochemically, PAPP-A can be localized in the trophoblast and in the decidua. Short term cultures of trophoblastic and decidual explants produce PAPP-A in vitro. It was thus of interest to see if long term cultures of primary monolayers of human trophoblast cells were capable of producing PAPP-A. Under our in vitro conditions, trophoblastic monolayers were producing both PAPP-A and beta hCG. During the first 3 days of culture PAPP-A levels increased in the medium whereas beta hCG levels decreased. The production of both proteins could be inhibited by cycloheximide. These results strongly suggest that the trophoblast is a source of PAPP-A in vivo.     

Relation of obstetric parameters to the concentrations of four pregnancy-associated plasma proteins at term in normal gestation.

The relations between normal obstetric parameters and the maternal levels of four pregnancy-associated plasma proteins (PAPP) were studied, with the use of plasma samples taken from 187 normal pregnant women within seven days before delivery. PAPP-A levels were correlated with placental and newborn weights. The levels of this pregnancy protein was higher in primigravid women and in groups with higher diastolic blood pressure than in other groups. Women with extremely high PAPP-A concentrations were likely to have extremely large placental weight was not necessarily associated with a high PAPP-A level. PAPP-C was not correlated with placental or newborn weight. The relationship between PAPP-C and maternal age, as well as maternal weight, was significant by one but not in the other three statistical analyses employed. The pregnancy zone protein was found to be correlated with parity. In primigravid women, this protein additionally showed an inverse correlation with the placental weight. Human chorionic somatomammotropin was significantly related to placental weight and inversely related to maternal weight. Its relationship with newborn weight was best seen in primigravid subjects. Other parameters (systolic blood pressure, one- and five-minute Apgar scores, weeks of gestation, days before delivery, newborn sex, and newborn bilirubin level) were not related to any of these pregnancy proteins.     

Tissue and plasma concentrations of pregnancy-associated plasma protein-A (PAPP-A): comparison with other fetoplacental products

Human placental lactogen (hPL), alpha fetoprotein (AFP), prolactin (PRL) and pregnancy-associated plasma protein-A (PAPP-A) were measured by radioimmunoassay in plasma, in homogenates of trophoblast, decidua, chorion, amnion and in amniotic fluid from 10 patients after non-complicated term delivery. Plasma samples and homogenates of trophoblast and decidua were also collected from 10 patients undergoing surgical termination of pregnancy between 7 and 12 weeks gestation. In addition, plasma and endometrial samples from 10 patients undergoing hysterectomy for other indications than malignancy were analysed for comparison. The highest tissue concentrations of hPL, AFP and PRL corresponded in each case to the known site of synthesis. For PAPP-A the highest concentration was found in maternal plasma at term [238.8 +/- 75.6 (SEM) micrograms/ml]. The highest tissue concentration was found at term in the decidua (57.0 +/- 2.0 micrograms/g), more than three times higher than that in the trophoblast (16.9 +/- 5.4 micrograms/g). The concentrations of PAPP-A in endometrial samples from non-pregnant women (1.9 +/- 0.6 micrograms/g) was 40 times higher than that in the corresponding plasma samples (0.05 +/- 0.02 microgram/ml). These observations point to the decidua as a possible source of PAPP-A.     

Studies on pregnancy-associated plasma protein A in the third trimester of pregnancy.

The plasma concentration of pregnancy-associated plasma protein A (PAPP-A) was measured in 34 women during the last 10 weeks of pregnancy. From 30 to 36 weeks the concentration of this protein increased steadily. Thereafter the concentration of PAPP-A rose more steeply, the highest amounts being found in early labour. The concentration of PAPP-A in peripheral venous blood and in the uterine vein was much the same. It was less in the retroplacental blood and a great deal less in the peritoneal fluid. The day-to-day variation was small; the coefficient of variation at 38 weeks was only 7.3 per cent. After delivery, the concentration of PAPP-A fell more slowly than other placental proteins and steroids, the average half-life being 51 hours. Although there is no doubt that PAPP-A is a product of the syncytiotrophoblast, our findings suggest that it is not simply secreted by the chorionic villi directly into the intervillous space but makes its way into the maternal circulation by a more circuitous route.     

The effect of cigarette or sheesha smoking on first-trimester markers of Down syndrome

Objective  To investigate the influence of cigarette or sheesha smoking on first-trimester markers of Down syndrome.
Design  A prospective observational study.
Setting  Primary care centres and antenatal clinics of Maternity and Children Hospital, King Abdulaziz University Hospital and New Jeddah Clinic Hospital, Jeddah, Saudi Arabia.
Population  Women with a singleton pregnancy who were either nonsmokers ( n = 1736) or cigarette smokers ( n = 420) or sheesha smokers ( n = 181).
Methods  Fetal nuchal translucency thickness (fetal NT), maternal serum free beta-human chorionic gonadotrophin (free β-hCG) and pregnancy-associated plasma protein-A (PAPP-A) were measured at 11 weeks 0 days to 13 weeks 6 days of gestation in all women. Women were grouped according to smoking status, confirmed by maternal serum cotinine measurements, and analyte levels between groups were compared.
Main outcome measures  Fetal NT, maternal serum free β-hCG, PAPP-A and cotinine measurements.
Results  Compared with nonsmoking women, fetal NT was significantly increased and free β-hCG and PAPP-A levels were significantly decreased in both cigarette and sheesha smokers. There were significant relationships between all three markers and the number of sheeshas consumed per day.
Conclusions  Cigarette and sheesha smoking significantly affect first-trimester markers of Down syndrome (fetal NT, free β-hCG and PAPP-A). Correction for this effect in women who smoke might improve the effectiveness of first-trimester screening for Down syndrome in these women. The underlying mechanism(s) relating smoking to the changes in first-trimester markers require further studies.     

Difference between inactive renins in amniotic fluid and in control or pregnancy plasma

Phenylethylamine-agarose column chromatography has been used to compare inactive renins in amniotic fluid, in control plasma during estrogen treatment and in maternal plasma during normal pregnancy. Inactive renin in all three was retained by the column and separated from renin substrate. Inactive renin in control and maternal plasma desorbed similarly and was different than that in amniotic fluid. The results suggest that during normal pregnancy the elevated concentrations of inactive renin in maternal plasma are more likely of renal than placental origin.     

Progesterone dependence and extratrophoblastic origin of pregnancy-associated plasma protein-A (PAPP-A) in early pregnancy

Pregnancy-associated plasma protein-A (PAPP-A) is a macromolecular glycoprotein produced in increasing concentration as pregnancy advances. PAPP-A is not specific to pregnancy since measurable levels have been found in non-pregnant females and in males. In non-pregnant females, PAPP-A is probably produced by the endometrium. The origin of PAPP-A in pregnant women is still controversial. In-vitro trophoblast and decidual explants both produce PAPP-A. So far, it is not known if the same applies to the in-vivo situation and to what extent these two tissues contribute to the circulating levels of PAPP-A. This study compares the circulating concentrations of PAPP-A and beta-hCG and progesterone in different pathological situations. In hydatidiform moles, beta-hCG levels are very high demonstrating an intense trophoblastic activity, whereas PAPP-A levels remain in the normal range. With spontaneous abortions, beta-hCG levels decline to very low values whereas PAPP-A continues to increase. These observations furnish indirect evidence for a major contribution to circulating PAPP-A levels by extratrophoblastic sites. Furthermore, PAPP-A levels decrease after administration of an anti-progesterone (RU486) either in-vivo or in-vitro. This is considered as a proof that PAPP-A levels in early pregnancy are progesterone dependent.     

Immunological heterogeneity of pregnancy-associated plasma protein-A (PAPP-A). Effects on the radioimmunoassay of PAPP-A

Pregnancy-associated plasma protein-A (PAPP-A) is a macromolecular glycoprotein produced by the trophoblast during pregnancy. Because the presence of PAPP-A in non-pregnant females is controversial, we re-evaluated our own radioimmunoassay technique for PAPP-A in the light of new observations about its immunological heterogeneity. Irrespective of the antibody (Geneva anti-PAPP-A or Dako anti-PAPP-A) the use of EDTA pregnancy plasma instead of pregnancy serum as a standard yielded different slopes of the standard curves and estimated significantly different amounts of PAPP-A in test samples. Moreover, highly purified tracers, isolated from maternal EDTA pregnancy plasma or pregnancy serum also produced significantly different results. So that the use of a tracer purified from EDTA plasma and EDTA pregnancy plasma as a standard will yield measurable levels of PAPP-A in non-pregnant female serum whereas the use of a tracer purified from maternal pregnancy serum and maternal pregnancy serum as a standard will not detect PAPP-A in the same samples. We conclude that, irrespective of the antiserum used, but depending on the biological origin of the tracer and the standard, different results will be obtained. PAPP-A is clearly immunologically heterogeneous, and the immunorecognition of PAPP-A will depend on whether or not blood coagulation has taken place.     

Immunological heterogeneity of pregnancy-associated plasma protein-A (PAPP-A). Effects on the radioimmunoassay of PAPP-A

Summary. Pregnancy-associated plasma protein-A (PAPP-A) is a macromolecular glycoprotein produced by the trophoblast during pregnancy. Because the presence of PAPP-A in non-pregnant females is controversial, we re-evaluated our own radioimmunoassay technique for PAPP-A in the light of new observations about its immunological heterogeneity. Irrespective of the antibody (Geneva anti-PAPP-A or Dako anti-PAPP-A) the use of EDTA pregnancy plasma instead of pregnancy scrum as a standard yielded different slopes of the standard curves and estimated significantly different amounts of PAPP-A in test samples. Moreover, highly purified tracers, isolated from maternal EDTA pregnancy plasma or pregnancy serum also produced significantly different results. So that the use of a tracer purified from EDTA plasma and EDTA pregnancy plasma as a standard will yield measurable levels of PAPP-A in non-pregnant female serum whereas the use of a tracer purified from maternal pregnancy serum and maternal pregnancy serum as a standard will not detect PAPP-A in the same samples. We conclude that, irrespective of the antiserum used, but depending on the biological origin of the tracer and the standard, different results will be obtained. PAPP-A is clearly immunologically heterogeneous, and the immunorecognition of PAPP-A will depend on whether or not blood coagulation has taken place.     

Expression of pregnancy-associated plasma protein-A (PAPP-A) during human villous trophoblast differentiation in vitro

Pregnancy-associated placental protein-A (PAPP-A), first isolated from maternal serum, has been identified as a metalloprotease cleaving insulin-like growth factor binding protein-4 (IGFBP-4). The source of PAPP-A during pregnancy is unclear. We therefore investigated PAPP-A expression during in vitro human villous cytotrophoblast cell (CT) differentiation into syncytiotrophoblast (ST). CT were isolated from normal first trimester, second trimester and term placentae (n=10) and cultured to form ST. PAPP-A mRNA was quantified by real-time PCR, and PAPP-A protein expression was studied by immunocytochemistry and TRACE technology with specific monoclonal antibodies. PAPP-A mRNA expression in total placental extracts increased during the course of pregnancy. PAPP-A protein was detected in the cytoplasm of both CT and ST. ST formation in vitro was associated with a 19-fold increase in PAPP-A mRNA expression and an 8-fold increase in PAPP-A secretion into the culture medium. No significant difference in PAPP-A production was observed between cultured cells isolated from early and term placentae. In conclusion, PAPP-A production in vitro, is associated to the differentiation of villous cytotrophoblast cells into syncytiotrophoblast, independently of the age of gestation.     

Measurement of plasma placental proteins and estriol in the detection of intrauterine growth retardation

This study was designed as an assessment of the clinical value of total estriol, placental lactogen (hPL), pregnancy specific glycoprotein (SP1), pregnancy-associated plasma protein-A (PAPP-A) and placental protein 5 (PP5), measured in a single sample of maternal serum, in the detection of intrauterine growth retardation (IUGR) in a large series of high risk pregnancies. Our results show the highest detection rate with SP1 (51%) and second highest with hPL (35%); estriol was less useful while PAPP-A and PP5 were of no value. However, there was 31% of false positives with SP1 compared with 14% for hPL. The best combination of the five parameters for the diagnosis of IUGR was achieved by measurement of both SP1 and hPL, with a low concentration of either protein indicating IUGR (sensitivity 63%, specificity 63%).     

High levels of human chromogranin A in umbilical cord plasma and amniotic fluid at parturition

OBJECTIVE: The human placenta is a neuroendocrine organ that produces several hypothalamic and pituitary hormones that are secreted during pregnancy and parturition into maternal and fetal circulation and amniotic fluid. Human chromogranin A (CgA) is a glycoprotein mainly localized to the adrenal medulla and released in response to several stressful events. During pregnancy, intrauterine tissues express and synthesize CgA mRNA and peptide and secret it into the biologic fluids of pregnancy, so we investigated whether maternal, umbilical, and amniotic human CgA levels are affected by the stress of parturition. METHODS: We measured pregnancy CgA levels in maternal and umbilical cord plasma and in amniotic fluid at term (39-40 weeks), by enzyme-linked immunosorbent assay at elective cesarean (n = 16), after spontaneous vaginal delivery (n = 12), and longitudinally throughout labor and 2 hours postpartum. RESULTS: CgA levels were highest in umbilical cord blood (P <.001). Umbilical cord plasma and amniotic fluid CgA levels were significantly higher at spontaneous vaginal delivery than at cesarean (P <.001), and the levels were not changed in maternal plasma according to cervical dilatation and postpartum. CONCLUSIONS: The present findings showed that the stress of parturition increased CgA levels in umbilical cord plasma and amniotic fluid and was probably of fetal origin. Whatever the mode of delivery, CgA levels in infants were much more elevated than in mothers, providing evidence for an unusual and sustained high level of sympathoadrenal stimulation in full-term neonates.     

Serum PAPP-A in normal pregnancy: relationship to fetal and maternal characteristics

Pregnancy associated plasma protein A (PAPP-A) was measured in maternal serum by electroimmunoassay in 414 women (with subsequent normal pregnancy outcome) during the 38th week of pregnancy. PAPP-A levels were examined in relation to maternal and fetal characteristics, revealing a statistically significant relationship with maternal weight, placental weight, fetal sex and gravidity. The correlation of PAPP-A levels to fetal sex and gravidity could not be explained by differences in placental or maternal weight. Mean levels of PAPP-A were also not significantly different in relation to fetal sex.     

Radioimmunoassay of placental protein 12: levels in amniotic fluid, cord blood, and serum of healthy adults, pregnant women, and patients with trophoblastic disease

A radioimmunoassay for placental protein 12 (PP12) is described and the levels in amniotic fluid, cord blood, and serum of nonpregnant individuals, pregnant women, and patients with trophoblastic disease are presented. During pregnancy, the highest PP12 levels were found at 22 to 23 weeks (mean +/- SD, 169 +/- 123 ng/ml), and there was a transient decline at 32 to 33 weeks (63 +/- 23 ng/ml). In amniotic fluid, the levels were 100 to 1,000 times higher than in maternal serum. In cord blood at birth, the values were of the same magnitude as in maternal serum. Also healthy nonpregnant women and men had PP12-like immunoreactivity in serum. Nonpregnant women (9 to 47 ng/ml) had higher levels than men (undetectable to 21 ng/ml). Elevated levels up to 84 ng/ml were occasionally observed in trophoblastic disease, both hydatidiform mole and choriocarcinoma, but they bore no correlation with the human chorionic gonadotropin levels. On the basis of these results PP12 is not a suitable marker for trophoblastic disease. PP12 values in normal pregnancy provide the basis for the evaluation of PP12 levels in abnormal pregnancy.     

Relationship of obstetric parameters to the concentration of pregnancy-associated plasma protein A

Pregnancy-associated plasma protein A (PAPP-A) was measured by rocket immunoelectrophoresis in 272 patients at 34 weeks of pregnancy and within 48 hours before delivery. The values obtained have been compared to maternal parameters (age, weight, weight gain, blood group, rhesus factor, and parity) and to fetal parameters (sex, weight, Apgar score, rhesus factor, blood group, and placental weight). Maternal age, increased parity, and increased body weight are related to decreased PAPP-A levels. On the other hand, mothers carrying male fetuses, rhesus-negative children, and babies with Apgar scores higher than 7 at 1 minute have increased PAPP-A concentrations. These findings are discussed in relation to PAPP-A's involvement in the maternal immunologic system.     

Origin of the alkaline phosphatases in amniotic fluid.

The nature and origins of amniotic fluid ALP activity were investigated early (14 to 22 weeks) and late (25 to 44 weeks) in pregnancy. The total enzyme activities for both stages were significantly different and considerable changes in the activities of individual enzyme components occurred. Early activity consists of intestinal (81%), bone/liver/kidney (15%), and placental (4%) ALP. In late fluids, the values are 5%, 69% and 27%, respectively. The intestinal enzyme is shown to be of fetal origin, presumably arising from the direct entry of desquamated intestinal mucosal cells into the amniotic fluid. The bone/liver/kidney enzyme is mostly of maternal serum origin early with an increased fetal contribution toward term. Early in pregnancy, the placental ALP activity is probably derived both directly from the placenta and from the placental activity in maternal serum. The latter source contributes more toward term.