Albumin concentration in human amniotic fluid from normal pregnancies

The present study is an attempt to estimate the albumin concentration in the human amniotic fluid of normal pregnancy. The study has been conducted from 12th to 20th week and from 30th to 43rd weeks of gestation. The results revealed a characteristic gestational profile with an increase in the concentration with the advancement of pregnancy from the 12th till the 20th week and an inverse relationship between the albumin level and the gestational age after the 30th week till term. The relationship between the albumin levels and the order of pregnancy indicated that the albumin concentration was not influenced by the birth order.     

Changes in surfactant phospholipids in fetal rat lungs from normal and diabetic pregnancies

The purposes of this study were to adapt and evaluate further a pulmonary surfactant isolation method applicable to unperfused fetal rat lung, to quantitate key phospholipids phosphatidylcholine (disaturated phosphatidylcholine, and phosphatidylglycerol) of the isolated material during the last 3 days of gestation, and to determine if abnormalities in surfactant phospholipids were present in fetuses of diabetic pregnancies. A simplified scheme of sucrose gradient centrifugation proved useful for small scale preparations of material enriched in the phospholipids most characteristic of pulmonary surfactant. It was shown that fetal blood phospholipids did not contaminate the surfactant fraction and therefore would not produce artifacts in assessment of lung maturational changes. Analyses of subcellular fractions isolated at 19.5, 20.5, and 21.5 days revealed that the percentages of disaturated phosphatidylcholine relative to total phospholipids were 23-44% in the surfactant preparations and 14-21% in the residual (nonsurfactant) fractions, while the disaturated phosphatidylcholine/phosphatidylcholine ratios were 0.62 +/- 0.06 and 0.41 +/- 0.03, respectively. Summation of the amounts of individual phospholipids in the two fractions yielded data that were nearly identical to the concentrations of these compounds in whole fetal lung samples analyzed independently, implying that losses during the surfactant isolation technique were negligible. The concentrations of phosphatidylcholine, disaturated phosphatidylcholine, phosphatidylglycerol, and total phospholipids increase markedly (more than 10-fold) and progressively in surfactant fractions prepared from normal fetal rat lung at 19.5, 20.5, and 21.5 days of gestation. In contrast, the residual fractions showed no changes from 19.5 to 20.5 days and then relatively modest increases from 20.5 to 21.5 days, except for phosphatidylglycerol, which increased markedly.(ABSTRACT TRUNCATED AT 250 WORDS)     

Nutrient levels in amniotic fluid from women with normal and neural tube defect pregnancies.

We analyzed nutrient levels in amniotic fluid obtained during the second trimester of normal, uncomplicated pregnancies from 221 women who delivered apparently healthy infants and from 8 with neural tube defect (NTD) pregnancies. Folate was measured by microbiological assay, vitamin B12 by a radiobinding method, and zinc, copper and iron by atomic absorption spectrophotometry. We found that the mean amniotic fluid nutrient levels of normal pregnancies were 24.7 nmol/l for folate, 600 pmol/l for vitamin B12, and 1.7, 1.9, and 9.0 mumol/l for zinc, copper and iron, respectively. Amniotic fluid folate, zinc, copper and iron levels of NTD pregnancies were similar to those found during normal pregnancy, however, vitamin B12 levels were markedly lower than those of normal pregnancies.     

Magnesium concentration in the amniotic fluid

Using a direct colorimetric method the concentration of magnesium in amniotic liquid was determined in 150 pregnant women at gestational ages ranging from 9 to 41 weeks. Results revealed a men value of 1.7 mg/dl (s = 0.39).     

Renin activity and concentrations of angiotensin I and II in amniotic fluid of normal and Rh-sensitized pregnancies.

Renin activity and the concentrations of angiotensin I and angiotensin II in amniotic fluid of second- and third-trimester pregnancies were determined by radioimmunoassay. Between the 28th and 38th wk of gestation, the mean renin activity in the amniotic fluid was higher than during early pregnancy (before the 18th wk of gestation). Both renin activity and the concentrations of angiotensin I and II were increased on some cases of Rh-incompatibility. One to two weeks after the administration of betamethasone to the mother with threatened premature delivery, the intra-amniotic renin--angiotensin system was slightly suppressed. In urine samples of newborns, angiotensin concentrations were in the same range as those found in the amniotic fluid; renin activity was very low or undetectable in the urine of male neonates (1--7 days of age). Thus, angiotensin II in the amniotic fluid may be derived both from fetal urine and/or as the product of enzymatic reactions in the amniotic sac; the latter is dependent not only on the presence of renin and converting enzyme but also on the local renin substrate (angiotensinogen) concentration.     

Micelles of pulmonary surfactant in human amniotic fluid at term

Studies using in vitro analysis have shown that the interaction between pulmonary surfactant and vernix caseosa could explain the appearance of amniotic fluid turbidity. That phenomenon is interpreted based on the "roll-up" hypothesis. We tested the roll-up hypothesis by examining the presence of micelles of pulmonary surfactant in human amniotic fluid at term. Amniotic fluid samples were collected from each of six healthy pregnant women at term and at 16 wk of gestation. These samples were stained negatively and analyzed using an electron microscope. Ultrastructures present in amniotic fluid were compared with the structure of micelles derived from suspended surfactant TA isolated from bovine lung. Surfactant TA formed spheroidal and rod-shaped micelles 10-70 nm in diameter above the critical micelle concentration. Identical micelle particles were described in human amniotic fluid at term. In addition, surfactant protein B was identified in the micelle fraction of amniotic fluid. However, no micelles were found in human amniotic fluid taken at 16 wk of gestation. Our results support the view that pulmonary surfactant could induce the detachment of vernix caseosa and increase the turbidity of the amniotic fluid.     

Total body water measurement in normal and diabetic pregnancy: evidence for maternal and amniotic fluid equilibrium

The H2[18O] tracer dilution method was applied to quantify total body water in 6 normal nonpregnant women, 8 normal pregnant women (gestation 29.3 +/- 6.0 weeks, range 21-37 weeks), and 8 insulin-dependent diabetic pregnant subjects (gestation 29.4 +/- 6.1 weeks, range 20-37 weeks). Plateau or equilibrium enrichment of 18O in the breath CO2 was achieved at approximately 2 h in nonpregnant normal subjects. In contrast, in pregnant subjects equilibrium plateau was reached later, at approximately 3 h. In order to examine whether the amniotic fluid/fetal compartment had also reached equilibrium with the maternal water compartment, amniotic fluid samples were obtained from 5 women undergoing elective cesarean section, approximately 5 h after the administration of tracer H2[18O]. The 18O enrichment of amniotic fluid was similar to that of the mother. Total body water was similar in normal and diabetic pregnant subjects (40.4 +/- 4.4 vs. 40.1 +/- 5.8 kg) and represented 55.8 +/- 5.4 and 58.7 +/- 5.5% of body weight, respectively. These data demonstrate the usefulness of the H2[18O] tracer method to measure total body water in pregnancy and emphasize the importance of an adequate equilibrium period 4-5 h in order to measure the dilution of administered tracer. The measurements by H2[18O] tracer are similar to those previously reported with deuterium oxide. Insofar as total body water reflects lean body mass, the data suggest that body compositional changes in normal and diabetic pregnancy are qualitatively similar.     

Determination and characterization of immunoreactive trypsin in amniotic fluid from normal and cystic fibrosis fetuses

High concentrations of immunoreactive trypsin (IRT) in the blood, and low concentrations of trypsin activity in fecal specimens have been found in newborn infants with cystic fibrosis (CF). The amniotic fluid concentrations of IRT and of IRT in complex with alpha 1-antitrypsin (alpha 1AT) were studied in 39 samples taken in about the 17th gestational week, and in 7 samples taken because the mothers had previously given birth to children with CF. The midtrimester samples contained trypsin in complex with alpha 1AT in a concentration of 30-200 micrograms/liter, and small amounts of trypsinogen, 0-50 micrograms/liter. Three of four amniotic fluid samples from CF fetuses had very low concentrations of trypsin in complex with alpha 1AT (less than 10 micrograms/liter), and only small amounts of trypsinogen (less than 10 micrograms/liter). Further prospective studies are needed to ascertain whether the determination of IRT in amniotic fluid may be of use in prenatal diagnosis of CF.     

Ascorbic acid concentration in amniotic fluid in late pregnancy.

Amniotic fluid and venous blood specimens were obtained from 34 pregnant women and analyzed for the ascorbic acid concentration. The mean amniotic ascorbic acid concentration of smoking pregnant women was less than 50% of that of non-smoking women. Pregnant women who smoked had a significantly lower serum and amniotic fluid ascorbic acid concentration than those who did not smoke. No differences were observed between the groups with or without premature rupture of the fetal membrane. The results suggest that ascorbic acid in the amniotic fluid reflects the ascorbic acid status in the blood of pregnant women and smoking had a greater effect in decreasing the ascorbic acid concentration in amniotic fluid than in serum.     

Congenital cytomegalovirus infection in twin pregnancies: viral load in the amniotic fluid and pregnancy outcome

Human cytomegalovirus (CMV) is the most common cause of viral intrauterine infection and fetal damage largely attributable to maternal primary infection. Most cases of congenital CMV infection in twins reported in the literature involved only 1 twin. We assessed the validity of polymerase chain reaction (PCR) and quantitative PCR on amniotic fluid (AF), at 21 to 22 weeks' gestation and at least 6 to 8 weeks after seroconversion, to predict the outcome of newborns in twin pregnancies. Two pregnant women with twin pregnancies and 1 woman with a triple pregnancy with primary CMV infection defined by the presence of immunoglobulin (Ig) M and low IgG avidity and/or by the presence of clinical symptoms and abnormal liver enzyme values were evaluated. CMV infection was found in 6 fetuses/newborns, 3 of whom were symptomatic. In the first twin pregnancy with diamniotic-dichorionic separate placentas, CMV symptomatic infection of the female twin was demonstrated by positive virus isolation and high viral load in AF. The male fetus was not infected as demonstrated by negative CMV culture and DNA detection in AF. In the triple pregnancy, the woman had a placenta with 2 monozygotic twins (females) and a separate placenta with a heterozygotic twin (male). The quantitative PCR results were 10(3) genome equivalents (GE)/mL of females AF and 1.9 x 10(5) GE/mL of male AF. Both female twins were asymptomatic at birth, whereas the male presented petechiae, thrombocytopenia, and cerebral ventriculomegaly. In the last twin pregnancy with fused dichorionic placentas, congenital CMV infection of both twins was diagnosed at birth in contrast with prenatal diagnosis. At time of amniocentesis, the left side twin was not infected as shown by negative results of CMV culture and DNA detection in the AF. CMV infection of the right side twin was demonstrated by positive CMV DNA detection with a CMV DNA load of 4.9 x 10(4) GE/mL and positive virus isolation in the AF. The morphologic and histologic examinations of the placentas strongly supported a prenatal horizontal acquisition of CMV infection. These twin pregnancies showed a marked difference in the quantity of virus load documented by the prenatal diagnosis suggesting that twin fetuses may react differently to primary maternal infection despite being exposed to the same maternal influences. A high viral load is correlated with congenital CMV infections symptomatic at birth. In such cases, with fetal infection of only 1 twin (at amniocentesis) and fusion of placentas, fetal outcome of both twins needs to be evaluated for the possibility of viral transfer from one fetus to the other.     

Insulin-like growth factor-1 receptor expression in the placentae of diabetic and normal pregnancies

BACKGROUND: Septal hypertrophic cardiomyopathy (sHCM) is a characteristic anomaly of the infant of diabetic mother (IDM). Insulin-like growth factor-1 (IGF-1) has been identified as a mediator of tissue overgrowth and we have previously shown that maternal IGF-1 levels were significantly elevated among neonates with asymmetrical sHCM. IGF-1 does not cross the placenta; it exerts physiologic action through binding to the IGF-1 receptor (IGF-1R). Localisation and expression of IGF-1R in term diabetic pregnancies are largely unclear. We have studied IGF-1R in the placentae of diabetic and normal pregnancies and this receptor expression in association with neonates with sHCM. METHODS: IGF-1R localization and expression in the placentae of six diabetic pregnancies associated with neonatal sHCM were compared with six each of randomly selected diabetic and normal pregnancies without neonatal sHCM by immunohistochemistry. The staining for IGF-1R in the deciduas, cytotrophoblasts, syncytiotrophoblasts and villous endothelium for these 18 samples were assessed and scored by two pathologists who were blinded to the respective diagnoses. RESULTS: Placental IGF-1R staining was negative in the villous endothelium for all three groups. IGF-1R staining was present in deciduas, cytotrophoblasts and syncytiotrophoblasts but the staining was weaker in the entire group of infants with sHCM compared to those without sHCM. CONCLUSIONS: IGF-1R is localized in all cell types of the placenta except in villous endothelium. Weaker placental IGF-1R staining in the placentae of diabetic pregnancies associated with sHCM suggests reduced expression of IGF-1R. This may be a down-regulatory response to elevated maternal IGF with neonatal sHCM outcome.     

Zinc concentration in amniotic fluid of zinc-deficient rats and its relation to fetal weight

Zinc concentration in amniotic fluid and its relation to fetal weight were investigated in three groups of pregnant rats: group A received a zinc-adequate diet, and the rats of groups B and C were fed a zinc-deficient diet. Group C also received zinc supplementation in water. The daily food consumption, weight and plasma zinc levels on days 1, 10 and 20, zinc concentration in amniotic fluid, the number of implantation sites, the number of resorptions, the number of live fetuses and fetal weight were determined. Plasma zinc concentrations were significantly different at the end of gestation between group B (Zn = 167.6 +/- 26.6 micrograms/dl) and the other two groups (group A = 199 +/- 18.6 micrograms/dl; group C = 204 +/- 13.7 micrograms/dl). The number of resorptions was significantly higher in group B and the number of live fetuses was significantly lower in this same group (p less than 0.025). Fetal weight was significantly lower in group B (p less than 0.001). The zinc concentrations of the amniotic fluid were significantly lower in group B (14 +/- 4.7 micrograms/dl) as compared to group A (83 +/- 11.4 micrograms/dl) and C (82 +/- 21 micrograms/dl; p less than 0.001). There was a positive linear correlation between zinc concentrations in amniotic fluid and fetal weight, being r: 0.7379 (p less than 0.001).     

Metabolic fingerprinting of amniotic fluid from the streptozotocin-induced diabetic pregnant rat using Fourier transform-ion cyclotron resonance mass spectrometry

Metabolic fingerprinting of amniotic fluid from streptozotocin-induced diabetic pregnant rats was performed using Fourier transform-ion cyclotron resonance mass spectrometry. Some of the fetuses from the diabetic pregnant rats exhibited ventricular septal defect. The positive ion profiles of amniotic fluids from diabetes were different to those of the control rats. The alteration of biochemical composition in the diabetic amniotic fluid suggests the presence of potential biomarkers to indicate progression of malformation under the diabetic pregnancy.     

Interaction between pulmonary surfactant and vernix: a potential mechanism for induction of amniotic fluid turbidity

The development of amniotic fluid turbidity during the third trimester is a known marker of fetal lung maturity. We hypothesized that this turbidity results from detachment of vernix caseosa from the fetal skin secondary to interaction with pulmonary-derived phospholipids in the amniotic fluid. To test this hypothesis, we exposed vernix to bovine-derived pulmonary surfactant over a physiologically relevant concentration range. Ten milligrams of vernix was evenly applied to the interior walls of 1.5-mL polypropylene microfuge tubes. Surfactant phospholipids were added to the tubes followed by slow rotation at 37 degrees C overnight. The liquid was decanted and spectrophotometrically analyzed at 650 nm to detect solution turbidity due to vernix detachment and/or emulsification. Increasing concentrations of surfactant phospholipids produced a dose-dependent increase in solution turbidity. A phospholipid mixture closely approximating natural pulmonary surfactant but devoid of surfactant-associated proteins yielded no increase. In other studies, the flow properties of vernix were studied in a Haake flow rheometer at 23 degrees C and 37 degrees C. There was a marked temperature-dependent effect with lower stress required to elicit flow at 37 degrees C compared with 23 degrees C. This temperature dependence was also demonstrated in the turbidity assay with a 124% increase in turbidity at body temperature compared with room temperature. We conclude that under in vitro conditions, pulmonary surfactant interacts with vernix resulting in detachment from a solid phase support. We speculate that in utero, this phenomenon contributes to the increase in amniotic fluid turbidity that is observed near term.     

Two-site "simultaneous" immunoassay with monoclonal antibodies for the determination of surfactant apoproteins in human amniotic fluid

Monoclonal antibodies against human surfactant apoproteins were prepared, which recognized 37, 34, and 62 kilodalton proteins in human lung lavage fluid and amniotic fluid. Two-site "simultaneous" immunoassay for the surfactant apoproteins was developed using the monoclonal antibodies. The assay was capable of measuring 10-640 ng of the apoproteins per ml of human amniotic fluid. The immunoassay was used to quantitate the apoproteins in 59 amniotic fluid samples from 23 to 41 wk gestation. The concentration of the surfactant apoproteins at less than 30 wk gestation was very low (mean 0.84 micrograms/ml). It then increased 6.5-fold from 34 to 36 wk gestation and 15.5-fold at more than 37 wk gestation. The simultaneous immunoassay with the monoclonal antibodies presented herein seems to be ideal for clinical use because of its high specificity, sensitivity, rapidity, simplicity, and a continuous unlimited supply of the antibodies. The results in this study show that the clinical use of the two-site simultaneous immunoassay with monoclonal antibodies to pulmonary surfactant apoproteins can predict fetal lung maturity more precisely.