Fluorescence anisotropy of tracheal and oropharyngeal aspirates as an indicator of lung disease in neonates.

Fluorescence anisotropy was assessed in tracheal and oropharyngeal aspirates to evaluate its use in the diagnosis of respiratory distress syndrome. Samples from 31 neonates at birth were purified by using Sephacryl S-300 column chromatography. After the addition of the molecular probe, 1,6-diphenyl 1,3,5 hexatriene fluorescence was measured and anisotropy calculated. Ten neonates with normal respiratory function had a mean anisotropy (+/- SD) of 0.226 +/- 0.023. Fourteen infants with a diagnosis of respiratory distress syndrome had a mean anisotropy of 0.260 +/- 0.014 (p < 0.001). Seven neonates with respiratory problems other than respiratory distress syndrome had a mean anisotropy of 0.211 +/- 0.027 (p < 0.001 vs respiratory distress syndrome). A diagnosis of respiratory distress syndrome was associated with higher tracheal fluid anisotropy, indicating a qualitative difference in the characteristics of the pulmonary surfactant. We conclude that this 30-minute test could help identify neonates who benefit from surfactant replacement therapy.

High glucose causes delayed fetal lung maturation as measured by fluorescence anisotropy.

Fluorescence anisotropy has been used to estimate the microviscosity of the surfactant phospholipid bilayer and no predict fetal lung maturity in human amniotic fluid; its usefulness in in vitro systems has been recently demonstrated. To investigate the effect of high glucose on lung development, anisotropy measurements were performed on 20-day fetal rat lung explant homogenates and culture media after culture for 48 hours in medium containing final concentrations of 10, 50, and 100mM glucose. Anisotropy of lung tissue cultured in 100mM glucose was significantly increased when compared to those cultured in 10mM glucose (p < .01). After 48 hours, the media from samples grown in 100mM glucose had significantly higher anisotropy (.2210 +/- .0031) than did media from explants grown in 50mM glucose (.2027 +/- .0079; p < .05), or in 10mM glucose (.1886 +/- .0046; p < .001). Relative fluorescence intensity of explants grown in 100mM glucose was 74.4 +/- 5.7% of those grown in 10mM glucose (p < .01). Fluorescence intensity of media was also decreased by 15-30% under higher glucose considerations (p < .05). These data suggest that surfactant synthesized and secreted under high glucose conditions, such as exist in the infant of the diabetic gestation, may have qualitative as well as quantitative changes.

Fluorescence anisotropy as an index of fetal lung maturation in vitro.

Fluorescence techniques have been used to assess the viscosity of surfactant-containing fluids in vivo and have been successfully employed clinically as indices of lung maturity. However, fluorescence measurements have not been previously used as indicators of fetal lung maturation in an in vitro system. Lung explants derived from 19-, 20-, and 21-day fetal rats were cultured in F-12 medium for 24-72 h. Tissue homogenates and culture medium were eluted on Sephacryl S-300 columns, a diphenylhexatriene (DPH) probe was added to each fraction, and fluorescence anisotropy and intensity were measured after excitation at 357 nm and emission at 435 nm. Elution fractions containing the major fluorescence peak were demonstrated to correspond to the phosphatidylcholine-containing fraction and were shown to contain lamellar bodies. Fluorescence anisotropy of tissue homogenates obtained from 19-day lung explants decreased after 72 h in culture, suggesting lower microviscosity of the surfactant-containing fractions. Assessment of culture media collected at 24-h intervals revealed significant decreases in anisotropy by 48 h for the 19-day explants, and by 24 h for the 20- and 21-day explants. Anisotropy of the final (48-72 h) culture media aliquots was significantly lower for 21-day explants (0.144 +/- 0.004, SE), than for 20-day (0.172 +/- 0.013) or 19-day explants (0.197 +/- 0.008), p < .005. Anisotropy of culture medium tended to be lower than anisotropy of corresponding tissue homogenates, suggesting that viscosity of recently secreted surfactant may be different from viscosity of surfactant within lamellar bodies in type II cells. Relative fluorescence intensity of tissue homogenates also increased with time in culture. These results indicate that fluorescence anisotropy can be used to assess the viscosity of surfactant in vitro and serve as another index of fetal lung maturation in in vitro systems. Estimation of the microviscosity of the surfactant phospholipid bilayer using anisotropy measurements may provide additional insight into such roles of surfactant function as adsorption and spreading.

The relationship of birth weight and intrauterine diagnostic ultrasound exposure.

Imaging with ultrasound is common in obstetric practice. Several laboratory animal studies have shown retardation in fetal growth after experimental ultrasound exposure. This investigation was conducted to determine whether human fetuses exposed to diagnostic ultrasound (sonography) have a greater risk of growth retardation than fetuses not so exposed. This retrospective cohort study compares the birth weights of 1598 exposed and 944 unexposed single live births at the Johns Hopkins Hospital in Baltimore, Maryland during calendar year 1981. Confounding variables, defined as those associated with both exposure status and birth weight outcome, were included in multivariable analysis. Both exposure to more than one ultrasound procedure and first exposure during the third trimester were associated with a reduction in birth weight. However, the most consistent effect associated with birth weight appeared to be the indication for an ultrasound examination. The relationship of ultrasound exposure and reduced birth weight appeared to be due to shared common risk factors, which lead to both exposure and a reduction in birth weight.

Fetal umbilical artery flow velocimetry in postdate pregnancies.

This study prospectively examined the use of umbilical artery flow velocimetry for monitoring fetal health in postdate pregnancies. Forty-six patients with well-established dates were evaluated with semiweekly biophysical profiles and umbilical artery flow velocimetry (characterized by the ratio of the peak systolic to end-diastolic velocity). Their labor records were reviewed, and neonates were examined for signs of postmaturity. Twenty neonates had an abnormal test result or outcome (identified as an abnormal nonstress test, oligohydramnios, meconium, intrapartum fetal distress, or a 5-minute Apgar score less than 7). Nine neonates had a physical examination consistent with the postmaturity syndrome. Twenty-one neonates were entirely normal. Comparisons of the mean systolic/diastolic ratios for neonates with and without the complications associated with postdatism showed no significant differences. In addition, all systolic/diastolic ratios were within the normal range. Therefore, umbilical artery flow velocimetry is unlikely to be useful for the routine antenatal assessment of the postdate fetus.

Temporal response of immunoreactive erythropoietin to acute hypoxemia in fetal sheep.

Acute hypoxemia was produced in chronically catheterized sheep fetuses to determine the response time necessary to increase plasma immunoreactive erythropoietin (Ep) concentration. Sodium nitrite (0.2 mM) was infused via a fetal vein to induce fetal hypoxemia. The resultant fetal methemoglobinemia was associated with a predictable, incremental decrease in arterial oxygen content. Twelve nitrite infusions were performed in eight fetal sheep preparations (gestational ages 115-146 days). Mean methemoglobin level increased to 33% of total Hb after 1-2 h of NaNO2 infusion. These results were compared to those obtained in nine control studies in eight fetuses in which no change was observed for plasma Ep, arterial oxygen content, PaO2, pHa, or whole blood lactate. In the nitrite infused group, however, a significant and progressive increase in mean plasma Ep level over baseline levels was observed during the 4th and 5th h of hypoxemia (p less than 0.01). This change in Ep was significantly greater compared to the control group. These results, however, were confounded by the concomitant development of a lactic acidemia secondary to the fetal hypoxemia. To examine the theoretic possibility that lactic acidemia may primarily affect fetal Ep levels, an additional group of five fetuses was infused with L-lactic acid for the same time period. Although the decrements in pHa and whole blood lactate levels achieved in these fetuses were in excess of those observed during the nitrite infusions, this possibility was ruled out since no change in fetal plasma Ep levels occurred. We conclude that during the 4th h of acute fetal hypoxemia a predictable, progressive increase in plasma Ep level is observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Correlation of the interpretation of fetal heart rate records with cord plasma erythropoietin levels.

On the basis that fetal levels of plasma erythropoietin (Ep) may reflect fetal oxygenation the primary purpose of the present study was to assess the relation between Ep measured in cord plasma at delivery and the intrapartum fetal heart rate (FHR) record. A scoring system for interpreting FHR recordings blindly was prospectively utilized in 41 selected human pregnancies during the 4 h immediately preceding birth. The correlation of the overall mean FHR score for each individual patient with cord plasma Ep was significant such that the highest Ep levels were observed in those infants with the most abnormal FHR scores. Furthermore, when the birthweights of the infants were adjusted for gestational age, sex, and birth order, birthweight centile was negatively correlated with cord plasma Ep. When both FHR score and birthweight were simultaneously correlated with cord plasma Ep using multiple regression, the combined effect of these two factors improved the association of either alone with both contributing approximately equally.

Purification of lamellar bodies from human amniotic fluid.

Lamellar bodies were purified from human amniotic fluid obtained from pregnancies at 17, 31, and 40 weeks' gestation by means of Sephacryl S-300 column chromatography. Fractions were labeled with diphenylhexatriene and two peaks of phospholipid were identified at each gestational age. The size of peak I increased relative to that of peak II with advancing gestational age. Further study of the two peaks from 40-week pregnancies showed that peak I contained lamellar bodies that could be identified by electron microscopy. The ratio of the concentration of protein to phospholipid for peak I varied from 0.2 to 1.1. There were four proteins which could be identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (130K, 80K, 50K, and 20K), and there was 90-degree scattering of 400 nm light. Peak II also contained phospholipid. However, bilayer structures could not be visualized by electron microscopy, and there was no 90-degree scattering of 400 nm light. Peak II had a much higher ratio of protein to phospholipid (approximately 100) and two broad bands of protein on sodium dodecyl sulfate electrophoresis (80K, 50K). The anisotropy of diphenylhexatriene in both peaks decreased with advancing gestational age. The anisotropy in peak I was always lower than that in peak II, indicating that the microenvironment in peak I was more fluid. Peak I may represent mature surfactant and peak II, precursors of surfactant.

Nanosecond time-resolved emission anisotropy of the fluorescent probe 1,6-diphenyl-1,3,5-hexatriene in human amniotic fluid.

The fluorescence emission anisotropy of 1,6-diphenyl-1,3,5-hexatriene in human amniotic fluid has been studied using nanosecond time-resolved emission techniques. These studies demonstrate that the previously reported decrease in the steady state emission anisotropy, [r], with gestational age is due to a change in the rate of rotational motion of the probe. The emission anisotropy decays to a limiting value (r infinity) greater than zero, suggesting a hindered rotation of the probe, and this is independent of gestational age. The decay function for the emission anisotropy of amniotic fluids from 17, 29, 40 and 41 weeks in gestational age can be best expressed as a single exponential plus a constant term, with rotational correlation times varying from 17 ns to 2.2 ns, respectively. The zero time emission anisotropy remains approx. 0.30 for both early and late gestational times.

Acute bacterial endocarditis with postpartum aortic valve replacement.

A case of acute bacterial endocarditis with aortic valve abscess, aortic insufficiency, and congestive heart failure at 32 weeks' gestation is described. Prompt valve replacement is indicated due to the risks of embolism to the coronary arteries and brain, and to the high mortality of such patients with medical management only. The infant was delivered prematurely to avoid the intraoperative risks to the fetus of cardiac surgery. General rather than regional anesthesia was chosen because venous pooling from a regional block would necessitate preoperative fluid loading and vasopressor therapy, which would be stressful for an already failing heart. In the presence of severe congestive heart failure, the patient underwent cesarean section and delivered a health 2020-g male infant; 36 hours later the aortic valve was successfully replaced with a no. 21 Byork-Shiley prosthesis. The infecting organism was Streptococcus viridans.

Long-term effects of betamethasone on fetal development.

In previous studies, we noted that treatment of pregnant rhesus monkeys with betamethasone resulted in a marked increase in fetal lung distensibility. The purpose of the present study was to determine whether these changes persisted during subsequent in utero development. Pregnant rhesus monkeys were treated with 2 mg of betamethasone intramuscularly from day 120 to day 133 and underwent delivery by cesarean section one month later. The treated fetuses were found to have smaller lungs (-31%; p less than 0.005), and lower alveolar stability (-14%; p less than 0.025) than the control fetuses. Additional findings included smaller weights for the brain (p less than 0.01), liver, pancreas, and heart (p less than 0.05). Smaller adrenal (p less than 0.025) and larger pituitary weights (p less than 0.05) and lower plasma corticoid concentrations (p less than 0.001) indicated long-standing adrenal insufficiency in the treated fetuses. These persistent sequelae caution the indiscriminate and prolonged use of these potent glucocorticoids during pregnancy.

The relationship of amniotic fluid fluorescence polarization to neonatal lung function.

Steady-state fluorescence polarization (FP) of 1,6-diphenyl-1,3,5-hexatriene (DPH) in monkey and human amniotic fluid was studied over a wide range of gestational ages. In both of these systems, the fluorescence polarization decreased with advancing gestational age. In the monkey, these measurements were correlated with both biochemical and physiologic parameters of lung function, including maximal lung volume, alveolar stability, percentage of disaturated phosphatidylcholine in lung homogenate, and lecithin/sphingomyelin ratio of amniotic fluid. Fluorescence polarization values correlated well with the lung disaturated phosphatidylcholine content expressed as a percentage of phosphatidylcholine, thus suggesting that the fluorescent probe interacts with a fraction of the amniotic fluid which is closely related to development of the pulmonary surfactant. system. Comparison of monkey and human amniotic fluid fluorescence polarizations showed a greater anisotropy of DPH in the monkey fluid at all stages of gestation, thereby indicating a greater microviscosity in monkey pulmonary surfactant.

Induction and decay of human fibroblast interferon mRNA.

Polyadenylylated interferon mRNA, obtained from induced human fibroblasts, was quantitatively assayed by synthesis of biologically active human interferon in Xenopus laevis oocytes. The assay for interferon mRNA was used to distinguish between various hypotheses relating to interferon induction and biosynthesis. The data demonstrate that on induction with poly(I-poly(C) human fibroblasts accumulate interferon mRNA for 1-1.5 hr, after which time the mRNA is rapidly degraded with a half-life (t 1/2) of 18 min. Treatment of cells with cycloheximide prolongs the period of accumulation to 3 hr and decreases the rate of mRNA inactivation (t 1/2 = 49 min). Treatment with actinomycin D decreases the rate of inactivation still further (t 1/2 = 68 min). A comparison of cellular interferon synthesis with the relative amounts of interferon m RNA after simple induction or inductionin the presence of the inhibitors (superinduction) indicated a general correlation. Thus, on induction, the genes for interferon are activated to produce a transcript for a short time. The superinducing treatments prolong the period of accumulation and decrease the rate of degradation of this transcript.

Synthesis of human interferon by Xenopus laevis oocytes: two structural genes for interferons in human cells.

Human fibroblasts and leukocytes produce interferons which may be distinguished by their antigenic and species specificity as well as by their molecular weight distributions. To elucidate the basis for these differences, we isolated mRNA from induced human fibroblasts and lymphoblastoid (Namalva) cells and studied the products of translation in Xenopus laevis oocytes. The mRNA from the respective cells yielded translation products, in oocytes, that were characteristic of the cells from which the mRNA was derived. We conclude that human cells contain at least two structural genes for interferon, coding for polypeptides differing in primary sequence. Fibroblasts synthesize a single species of interferon; lymphoblastoid cells synthesize two species, the fibroblast and leukocyte types.