Human serum Zn-alpha2-glycoprotein in amniotic fluid

Human serum Zn-alpha2-glycoprotein (Zn-alpha2-GP) was found to be present in the amniotic fluid in the mean concentration of 0.98 +/- 0.40 mg/100 ml, which represents about one-tenth of its concentration in the maternal serum (9.65 +/- 1.18 mg/100 ml). Its concentration in the amniotic fluid was proportional to the amniotic fluid total protein and very approximately to the maternal serum Zn-alpha2-GP. The relationship between the maternal serum Zn-alpha2GP and the maternal serum total protein as well as between the amniotic fluid total protein and the maternal serum total protein was found to be not significant. The amniotic fluid Zn-alpha2-GP as well as the amniotic fluid total protein showed some increase during gestation to reach the highest values at the end of the second trimester. At present both the origin and significance of the amniotic fluid Zn-alpha2-GP are not known.     

Pancreatic secretory trypsin inhibitor from human amniotic fluid and fetal and neonatal urine: concentrations and physicochemical characterization

The levels and physicochemical properties of the pancreatic secretory trypsin inhibitor, also known as Kazal type trypsin inhibitor, were studied in human amniotic fluid. In the second trimester, the median concentration was 160 ng/ml, which exceeds the maternal serum levels 20-fold. Towards term, the amniotic fluid levels declined about 5-fold, whereas the maternal serum values remained constant. In fetal urine, the concentration of the trypsin inhibitor was similar to that in amniotic fluid in early gestation, whereas in newborn urine, the median level was 4-to 5-fold higher than in term amniotic fluid. The physiochemical characteristics of the trypsin inhibitor in amniotic fluid, neonatal urine and cancer urine from an ovarian cancer patient were similar, as studied by gel filtration, high performance reverse phase liquid chromatography, and complete immunological identity in immunodiffusion. The physicochemical similarity and levels in various compartments suggest fetal contribution to amniotic fluid levels of the trypsin inhibitor.     

The amnion regulates movement of fetally derived alpha-fetoprotein into maternal blood.

The present investigation documents that, under normal conditions, most fetally produced AFP reaches the maternal circulation via diffusion across the amnion from amniotic fluid. This has been determined by comparing maternal serum AFP levels with amniotic fluid albumin concentrations in paired samples. The proportionally demonstrated between them indicates a proportional, transamniotic exchange of the two proteins, each originating on opposite sides of the amnion. Albumin is known to reach amniotic fluid by transamniotic diffusion from maternal blood. All amnions restrict AFP movement into maternal serum, but some are distinctly more restrictive than others; in such cases, a relatively greater increase in amniotic fluid AFP concentration would likely have to occur from a fetal lesion before being reflected in maternal serum. Inconsistencies found in several paired samples identify that other variables may also influence passage of AFP to the mother.     

Prostate-specific antigen in amniotic fluid of normal and abnormal pregnancies

Objective: To examine if prostate-specific antigen (PSA) is present in amniotic fluid or maternal serum during pregnancy and if its presence is associated with fetal abnormalities.

Methods: Samples tested included amniotic fluids from 853 pregnant women for whom amniocentesis was performed; 312 nonpregnant women who donated blood; 259 pregnant women who donated blood at various gestational ages. Amniotic fluid or serum PSA was measured with an ultrasensitive time-resolved immunofluorometric procedure. 372 pregnancies were studied for the presence of genotypic or phenotypic fetal abnormalities.

Results: PSA was present in most amniotic fluids; the median PSA concentration increased from gestational week 11 to 22 and stabilized thereafter until delivery. The most prominent PSA concentration change occurred during gestational weeks 13–14. Pregnant women had significantly higher serum PSA concentrations than nonpregnant women; the pattern of serum fSA concentration change during pregnancy was similar to that of amniotic fluid; however, serum PSA concentrations were lower by a factor of 20–40. No association existed between amniotic fluid F'SA and maternal age, gender of fetus, or length of abstinence of mother from sexual intercourse. After gestational week 15, fetuses with trisomy 21 or 18, anencephaly, or renal disorders were associated with low amniotic fluid PSA levels.

Conclusion: Our data suggest that PSA may play a role in fetal development, especially at gestational ages between 13–20 weeks. The diagnostic usefulness of PSA in identifying fetal abnormalities remains to be determined.     

N-Acetyl-beta-hexosaminidase isoenzymes of amniotic fluid and maternal serum. Their relevance to prenatal diagnosis of the GM2 gangliosidoses.

The isoenzymes of N-acetyl-beta-hexosaminidase were quantitated in 30 amniotic fluid and 13 maternal serum samples collected between 11 and 40 weeks of gestation using DEAE-Sephadex A-25 chromatography. Isoenzymes A and B consitituted the major components of most amniotic fluids but seven samples characterized by high N-acetyl-beta-hexosaminidase activities contained high proportion of an isoenzyme apparently identical to isoenzyme P of maternal serum. This passage of maternal serum N-acetyl-beta-hexosaminidase into the amniotic cavity could lead to false negative diagnosis of type B and type O GM2 gangliosidoses if the diagnosis rely solely upon isoenzyme analysis in amniotic fluid. However, the release of maternal enzyme into amniotic fluid seems to be restricted to the third trimester of gestation and should not interfere with prenatal diagnosis of the GM2 gangliosidoses ususally performed at an earlier stage of gestation.     

Binding of corticotropin-releasing hormone (CRH) in maternal and fetal plasma and in amniotic fluid

Placenta secretes corticotropin-releasing hormone (CRH) into the maternal and fetal circulation, but a CRH binding protein in plasma may decrease its biological activity. Using a charcoal adsorption method we found that 92% of added 125I-Tyr-CRH was bound to a binding protein in the nonpregnant plasma, 72% in the plasma at term pregnancy, 90% in umbilical cord plasma, 82% in the amniotic fluid in the second and 25% in the third trimester. CRH added to plasma inhibited the binding of 125I-Tyr-CRH over the concentration range of 0.1-8.8 nmol/l in plasma and of 0.1-2.2 nmol/l in amniotic fluid. There was a significant negative correlation (R = -0.80) between the binding capacity of the CRH-binding protein and CRH concentration in maternal plasma. Plasma or amniotic fluid was incubated with 125I-Tyr-CRH and subjected to gel filtration on Sephadex G-50. The bound radioactivity was eluted at the region of Mr 25-40 kDa and the unbound radioactivity at the location of synthetic CRH. Bound and unbound CRH concentrations were determined using charcoal adsorption method and gel filtration on Sephadex G-50 in ten maternal plasma samples at the third trimester of pregnancy. Following mean percentages were found to be bound: charcoal method 61.9 +/- 6.80% (SE) and gel filtration 62.8 +/- 6.33%. We conclude that the bulk of CRH is bound to a binding protein in maternal and fetoplacental circulation, whereas at term pregnancy the role of the binding is small in amniotic fluid.     

Immunoassay for the determination of surfactant apoprotein (SP-A) in human amniotic fluid: comparison with other indices of assessing foetal lung maturity.

The concentration of the major surfactant-associated protein SP-A (28-36 kDa) was determined in 73 amniotic fluid samples obtained from normal (n = 40) and complicated (n = 33) pregnancies. Lecithin/sphingomyelin (L/S) ratio and phosphatidylglycerol (PG) levels were also determined in all the samples by one-dimensional step-wise thin-layer chromatography. An enzyme-linked immunosorbent assay was used to determine human lung surfactant apoprotein SP-A. The amount of SP-A in human amniotic fluid increased as a function of gestational age from 8 mg l-1 at 36 weeks to 11.75 mg l-1 at 40-41 weeks of gestation. There was a significant difference (p less than 0.01) in amniotic fluid SP-A concentration from female (9.93 +/- 0.60 micrograms ml-1) compared to male (9.10 +/- 0.52 micrograms ml-1) foetuses. In amniotic fluid samples obtained from a group of complicated pregnancies, SP-A levels were significantly lower than in the normal group when adjusted for gestational age and sex of the foetus (p less than 0.05).     

Foetal and maternal magnesium metabolism: effect of magnesium deficiency and isoproterenol

Pregnant rats were fed diets with an Mg2+ content of 40, 12, 6 and 3 mmol/kg from day 10-19 of pregnancy. There was a linear correlation of non-protein bound Mg2+ between foetal and maternal serum, between amniotic fluid and maternal serum, and between foetal serum and amniotic fluid, the ratios being 2.7, 2.0 and 1.3 respectively, indicating active transport of Mg2+ up to a constant concentration gradient by the placenta. In hearts, increases of Na+ and Ca2+, and decreases of Mg2+ and K+ were observed only in the group receiving the lowest Mg2+ supply. After i.v. injection of MgCl2 to pregnant rats, Mg2+ was slowly transported from maternal to foetal serum and more slowly into the amniotic fluid. The effect of isoproterenol on cardiac electrolyte content in pregnant rats was less than in non-pregnant rats, and the effect of isoproterenol in foetal rats was smaller than in maternal rats. These results are explained by inactivation of isoproterenol in the placenta, by the small diaplacental transport of isoproterenol and by a smaller isoproterenol-stimulation of foetal cardiac adenylate cyclase.     

High molecular weight angiotensinogen: a pregnancy associated protein

Although it was previously recognized that human amniotic fluid contained appreciable quantities of angiotensinogen, it remained unknown what fraction of the total is high molecular weight angiotensinogen. Fractionation of human amniotic fluid showed that high molecular weight angiotensinogen represents the major component of total angiotensinogen; 58% for 11 normotensive pregnant women and 67% for 12 hypertensive pregnant women. In contrast to plasma where high molecular weight angiotensinogen is elevated in hypertensive pregnant women as compared to normotensive pregnant women, no such difference exists for amniotic high molecular weight angiotensinogen. Serum and amniotic fluid high molecular weight angiotensinogen were compared in six subjects, and no significant correlation was found. In fetal cord blood, high molecular weight angiotensinogen represented only 5.8% of the total angiotensinogen. The site of synthesis of high molecular weight angiotensinogen remains unknown but these data suggest that it is produced in the placenta or in the maternal uterine tissue. Therefore, we propose that human high molecular weight angiotensinogen should be classified as a pregnancy-associated protein.     

Study of soluble proteins from human amniotic fluid by fraction chromatography (author's transl]

The nature and origin of soluble proteins from human amniotic fluid have been investigated by gel filtration chromatography on Sepharose 6B and ion-exchange chromatography on Ecteola-cellulose. Each fraction has been studied by immunoelectrophoresis. Specific antisera against serum proteins, amniotic fluid proteins, fetal proteins and salivary proteins have been used. These various antisera showed that the amniotic fluid contains maternal proteins, but also more specific proteins from fetal origin such as beta2-microglobulin, urinary mucopolysaccharides, salivary proteins and carcinoembryonic antigen. These results not only confirm that amniotic proteins are essentially from maternal origin, but prove the important fetal contribution and emphasize the importance of the fetoplacental unit in the monitoring of high risk pregnancies.     

白细胞介素-8、肿瘤坏死因子-α水平与胎膜早破合并宫内感染的相关性研究

目的：探讨孕妇血清和羊水白细胞介素-8（IL-8）、肿瘤坏死因子-α（TNF-α）水平与胎膜早破合并宫内感染的相关性和临床意义.方法：选取我院 2005年1月至2006年12月住院胎膜早破孕妇52例（研究组）,以及产科门诊定期产检的正常孕妇50例为对照组;采用酶联免疫法（ELISA）测定血清与羊水IL-8、TNF-α水平;同步取胎盘胎膜组织进行病理组织学检查.结果：研究组血清和羊水IL-8水平均高于对照组（P＜0.05）,而研究组TNF-α仅羊水水平高于对照组（P＜0.05）.研究组绒毛膜羊膜炎的发生率高于对照组（37%vs4%,P＜0.01）.羊水IL-8水平与血清 IL-8水平呈较好的正性相关（r=0.677,P＜0.01）;羊水TNF-α与羊水IL-8水平间亦有较好的正性相关（r=0.788,P ＜0.01）.结论：血清和羊水IL-8水平、羊水TNF-α水平与胎膜早破合并宫内感染有明显的正相关性,其中羊水IL-8的相关性最好.孕妇羊水IL -8、TNF-α水平可作为早期诊断胎膜早破合并宫内感染的指标.     

Tetranectin in amniotic fluid, maternal serum and fetal fluids

The concentration of the newly discovered protein tetranectin has been measured in different fetal and maternal compartments. In amniotic fluid a significant, positive correlation between the tetranectin concentration and gestational age was found (a mean of 0.2 mg l-1 at week 14 to a mean of 0.5 mg l-1 at week 21). In maternal serum a slight negative correlation was found between tetranectin concentration and gestational week (a mean of 6.17 mg l-1 at week 14 to a mean of 5.79 mg l-1 at week 21). In-term cord blood collected at delivery a mean level of 6.0 mg l-1 was found, and no difference was found between arterial- and venous-blood tetranectin concentration. In fetal serum the overall mean level was 2.6 mg l-1 and a significant positive correlation between tetranectin concentration and gestational age was found. The mean level was 1.1 mg l-1 in fetal cerebrospinal fluid and no correlation to gestational age was found. Fetal tetranectin may, therefore, be correlated to fetal maturation.     

Unconjugated pteridines in amniotic fluid during gestation

Neopterin and biopterin concentrations were measured in amniotic fluid in 226 pregnancies from the 12th week of gestation to term. At mid-gestation, neopterin and biopterin levels were low and remained relatively constant between 12 and 26 weeks of gestation, whereas during the third trimester, a progressive increase was observed. Near term the values were greater than those in maternal serum and the higher neopterin to biopterin ratio suggested that pteridine concentration in amniotic fluid may reflect the maturation of pteridine metabolism in the fetus.     

Tobramycin: Maternal-Fetal Pharmacology

To investigate the maternal-fetal transfer of tobramycin (TBM) and its distribution in the fetus, a single dose of 2 mg/kg was administered intramuscularly to 35 pregnant patients (13 first trimester, 22 second trimester) 0.5 to 34 h before hysterectomy. TBM concentration was assayed microbiologically in maternal serum, fetal tissues (placenta, brain, lung, liver, and kidney), and fluids (amniotic, cerebrospinal fluid [CSF], urine, and serum). Mean maternal serum half-life (1.54 h) and mean peak serum concentration of TBM were within ranges reported for nonpregnant adults. In fetal serum, half-life was 5.2 h, and TBM levels did not exceed 0.58 μg/ml. For intervals up to 34 h, the mean TBM concentration in placental tissues was 1.4 μg/g. Concentration differences related to fetal maturation were found for fetal CSF, amniotic fluid, and fetal kidney. No antimicrobial activity was found in the fetal CSF of >16 weeks' gestation. TBM was present predominantly in the second trimester amniotic fluid specimens. Fetal kidney concentrations reached 7.2 μg/g at 34 h after maternal drug administration. Higher TBM concentrations were related to advanced maturation of the fetal kidney. Second trimester fetal urine concentrations for TBM ranged from 0.1 to 3.4 μg/ml, and the fetal urinary half-life was 3.7 h. Knowledge of fetal pharmacology is essential for weighing the fetal benefits or risks of antimicrobial therapy for the infected gravid patient.     

Evaluation of a commercial immunoenzymometric assay for alpha-fetoprotein

A two-site immunoenzymometric assay (Abbott Diagnostics) for alpha-fetoprotein (AFP) in maternal serum and amniotic fluid has been evaluated for its suitability as a screening test for open neural tube defects. In a retrospective study based on 190 pregnancies of known outcome, performance of the kit in measuring both serum and amniotic fluid AFP correlated well with that of an in-house radioimmunoassay. Of 39 pregnancies associated with open neural tube defects, only four would not have been detected by the use of sequential measurement of serum and amniotic fluid AFP (also essentially in agreement with results obtained by the RIA). We conclude that this immunoassay could form the basis for a screening program for antenatal detection of open neural tube defects.     

Maternal Thyroid Function is the Major Determinant of Amniotic Fluid 3,3′,5′-Triiodothyronine in the Rat

3,3′,5′-triiodothyronine, (rT₃), is easily measured in human amniotic fluid (AF) during the second and third trimesters. To determine if AF rT₃ levels are maintained by either maternal or fetal thyroid function, or both, models of fetal hypothyroidism (FH), maternal hypothyroidism (MH), and combined maternal and fetal hypothyroidism (MFH) were developed in pregnant rats. Hormone analyses of maternal and fetal serum and AF were performed at term. Thyroxine (T₄) and 3,3′,5-triiodothyronine (T₃) were not detectable in the sera and AF of term fetuses in all groups. MFH rats were prepared by administration of methimazole to the dams, and in some experiments, by maternal thyroidectomy and a low iodine diet as well. In the MFH groups from the three experiments serum thyrotropin (TSH) was markedly elevated in the dams and in the fetuses. FH rats were prepared by administering T₄ by various routes to dams treated according to the MFH protocols and serum TSH was elevated in fetal serum. Analysis of FH maternal serum T₄, T₃, and TSH concentrations suggested mild maternal hyperthyroidism or hypothyroidism depending upon the schedule of T₄ administration. The MH groups were prepared by maternal thyroidectomy and in all experiments the fetuses had normal serum TSH concentrations. The degree of maternal hypothyroidism in the MH and MFH groups was equivalent. The mean concentration of AF rT₃ in normal rats in three experiments was 28.4±2.5 ng/dl (±SEM). In the three experiments, AF rT₃ was undetectable or markedly reduced in the MH and MFH rats and was normal in the FH rats. These results in the amniotic fluid could not be explained by transfer of rT₃ from fetal serum to the AF because fetal serum rT₃ concentrations in these various models did not correlate with AF rT₃ concentration. Furthermore, infusion of large doses of rT₃ in MFH dams resulted in a 35-fold elevation in maternal serum rT₃ concentration, a twofold elevation in fetal serum rT₃ concentration, and only a minimal increase in AF rT₃. These studies demonstrated that, in the rat, the maternal thyroid has the dominant role in maintaining AF rT₃, whereas little effect of fetal thyroid status on AF rT₃ could be demonstrated. Transfer of maternal rT₃ or of fetal rT₃ derived from maternal T₄ to the AF do not appear to be the mechanisms whereby the maternal thyroid maintains AF rT₃.     

Studies on serum apolipoproteins and lipids in amniotic fluid and neonatal urine

Amniotic fluid and neonatal urine were examined for the presence of lipids and serum apolipoproteins. Human apolipoproteins A-I, A-II, and ApoD found principally in serum high density lipoproteins were identified in both neonatal urine and amniotic fluid. A lecithin/sphingomyelin ratio of greater than 5 associated with fetal lung maturity was accompanied by the disappearance of A-II from amniotic fluid. Dissimilarities of total fatty acid composition of amniotic fluid when compared to cord serum or neonatal urine indicate other tissue sources for fatty acids found in amniotic fluid. In addition, the presence of serum apolipoproteins and lipids in both amniotic fluid and neonatal urine suggests that a least a portion of these constituents could be derived from fetal urine.     

Longitudinal evaluation of azithromycin and cytokine concentrations in amniotic fluid following one‐time oral dosing in pregnancy

To utilize noninvasive collection of amniotic fluid in the setting of preterm premature rupture of membranes (PPROMs) to report the time concentration profile of azithromycin in amniotic fluid over 7 days from a single dose, and evaluate the correlation between azithromycin concentration and inflammatory markers in amniotic fluid. Prospective cohort study of five pregnant patients admitted with PPROMs and treated with a single 1 g oral azithromycin dose. Amniotic fluid was collected from pads and used to quantify azithromycin concentration as well as TNFa, IL‐1a, IL‐1b, IL‐6, IL‐8, and IL‐10 concentrations. Primary outcome was time/concentration profile of azithromycin in amniotic fluid. Secondary outcome included correlation between azithromycin concentration and cytokine concentrations. Five patients were enrolled. Mean gestational age on admission with PPROM was 27.5 ± 2.3 weeks with a median latency of 7 days (interquartile range [IQR] = 4–13). A median of two samples/day (IQR = 1–3) were collected per participant. Azithromycin was quantified in duplicate; intra‐assay coefficient of variation was 17%. Azithromycin concentration was less than 60 ng/ml after day 3. Azithromycin concentration was positively correlated with IL‐8 (r = 0.38, p = 0.03), IL1a (r = 0.39, p = 0.03), and IL‐1b (r = 0.36, p = 0.04) in amniotic fluid. Azithromycin is detectable in amniotic fluid over 7 days from a single 1 g maternal dose, however, it is not sustained over the range of minimum inhibitory concentration for common genitourinary flora. Based on correlation with specific cytokines, azithromycin penetration in amniotic fluid may relate to maternal monocyte concentration in amniotic fluid in the setting of PPROM.

Study Highlights

• WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC?

Azithromycin is used to prevent infection in setting of preterm premature rupture of membranes (PPROMs), but there is no established dose. One cross‐sectional study with one patient found amniotic fluid azithromycin concentration was below minimum inhibitory concentration (MIC)50 of common genitourinary (GU) flora 7 days after a single 1 g maternal dose. There is no reported correlation between maternal azithromycin treatment and the concentration of drug and inflammatory cytokines in amniotic fluid.

• WHAT QUESTIONS DID THIS STUDY ADDRESS?

Our objective was to utilize noninvasive collection of amniotic fluid in pregnant patients admitted with PPROM to address whether one‐time maternal dosing of azithromycin can produce a sustained amniotic fluid concentration greater than the MIC of common GU flora over 7 days, and secondarily, whether amniotic fluid azithromycin concentration was correlated with inflammatory cytokine concentrations.

• WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE?

Azithromycin is detectable in amniotic fluid over 7 days from a single 1 g maternal dose, however, it is not sustained over the range of MIC for common GU flora. Azithromycin concentration in amniotic fluid is correlated with specific cytokines that suggest that azithromycin penetration in amniotic fluid may relate to maternal monocyte concentration in amniotic fluid in the setting of PPROM.

• HOW MIGHT THIS CHANGE CLINICAL PHARMACOLOGY OR TRANSLATIONAL SCIENCE?

Amniotic fluid may be collected noninvasively from patients with PPROM and used to evaluate the fetal environment, including medication penetrance and inflammatory markers. Given the benefit of maternal and fetal exposure to azithromycin for prevention of adverse perinatal outcomes, it is important to study how maternal dosing leads to azithromycin concentration within the maternal fetal unit in order to optimize therapy.     

Variations of the concentrations of certain glycoproteins in maternal serum, fetal serum and amniotic fluid during pregnancy (author's transl)]

The authors investigated systematically the variations during normal pregnancies of the concentrations of alpha-1-antitrypsin, orosomucoid, transferrin and alpha-fetoprotein simultaneously in maternal serum, fetal serum and amniotic fluid. The role of certain factors such as the gestational age birth weight, placental weight and pairty were studied with regard to variations in the concentrations of each of these proteins. This research permitted the definition during pregnancy of the normal concentrations for these four proteins and allowed us to learn more about protein exchanges between fetal blood, maternal blood and amniotic fluid. There exists a difference between the concentrations of alpha-1-antitrypsin and of orosomucoid found for primigravidae and for multigravidae. The role of these glycoproteins in preventing the mother from rejecting the fetus (insofar as the fetus may be considered as an allograft) is discussed.     

Cocaine and metabolite concentrations in the fetal guinea pig after chronic maternal cocaine administration.

To determine the disposition of cocaine (COC) and metabolites after chronic COC exposure in the late gestation guinea pig, six time-bred Dunkin-Hartley guinea pigs were given 10 daily 6 mg/kg COC s.c. injections from day 50 of gestation. Maternal blood and urine, fetal cord blood, and brain and amniotic fluid were collected 1 hr after the last injection. There was no difference between maternal and fetal plasma COC concentrations. This may be due to the combined effect of lower protein binding and ion trapping of COC in the fetus. Benzoylecgonine was higher in maternal plasma, but benzoylnorecgonine was higher in fetal plasma. COC brain-to-plasma ratios were similar in the dam and fetus. Benzoylecgonine was the only metabolite that could be detected in the brain, but levels were too low to quantitate. COC accumulated 3 to 4 times plasma concentrations in the amniotic fluid and was directly proportional to fetal plasma COC concentrations. Benzoylnorecgonine in amniotic fluid accumulated to 2 times fetal plasma levels. The in vitro half-life of COC in amniotic fluid was 30 times longer than plasma elimination half-life in vivo. The high level and long duration of COC in amniotic fluid serve as a reservoir for prolonged fetal COC exposure.