Gestational age dependence of 11 beta-hydroxysteroid dehydrogenase and its relationship to the enzymes of phosphatidylcholine synthesis in lung and liver of fetal rat.

Increase in fetal surfactant synthesis and lung maturity is caused by the glucocorticoidal induction of enzymes required for phosphatidylcholine (PC) synthesis towards the end of gestation. The regulation of gestational age-dependent induction of PC synthesis by glucocorticoids is still unclear. Since 11-beta-hydroxysteroid dehydrogenase (11 beta-HSD) activity and its metabolising capacity for glucocorticoids have been suggested to play a central role in this regulation, we measured the gestational age-dependent changes in 11 beta-HSD and PC synthesizing enzymes in lung and liver of fetal rat. The activity of cholinephosphate cytidyltransferase (CCT; key enzyme in PC synthesis), choline phosphotransferase (CPT) and lysolecithin acyltransferase (LAT) were found to increase gradually in the lung towards the end of gestation, reached peak values at term followed by a decrease of activity reaching finally adult levels. Only CK activity exhibited constant levels until term followed by a slight increase after the birth. In comparison with the lung, the liver enzymes followed a similar pattern, but at a higher rate of activity except for CCT which was higher in the lung. The activity of 11 beta-HSD in fetal lung microsomes was detectable from day 20 and increased towards the end of gestation in the lung and liver of the rat. Oxidase activity was always found to exceed the reductase activity. The activity of 11 beta-HSD continued to increase after delivery and reached peak levels in adult animals in both organs. In order to test the hypothesis, whether 11 beta-HSD activity and PC synthesis are induced by increasing endogenous glucocorticoidal levels, we examined on day 19 of gestation the effect of dexamethasone (DEXA) on enzymatic activities (11 beta-HSD, CCT) and on [14C]choline incorporation in phosphatidylcholine in fetal lung organoid cultures. Additionally, changes in CCT activity in fetal lungs after maternal administration of DEXA were measured. DEXA accelerated 11 beta-HSD and CCT activities as well as [14C]choline incorporation. We conclude, that endogenous glucocorticoids induce PC synthesis as well as 11 beta-HSD activity in lung and liver of the fetal rat. Fetal PC synthesis is not altered by increasing 11 beta-HSD levels, because the increase of free serum corticosterone levels apparently exceeds the metabolising capacity of 11 beta-HSD towards term.

Correlation of surfactant phosphatidylcholine synthesis and 11 beta-hydroxysteroid dehydrogenase in the fetal lung.

Glucocorticosteroids (GCS) are prerequisite for the induction of surfactant synthesis in the fetal lung. The 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) regulates the intracellular concentration of biologically active GCS. In this study we demonstrate the correlation of 11 beta-HSD activity and the GCS-induced surfactant phosphatidylcholine synthesis in organoid cultures of fetal rat lung. In the first series of experiments, [3H]corticosterone (CORT) or [3H]11-dehydrocorticosterone (11-DHC) were added to lung organoid cultures to test 11 beta-HSD activity. We found a low oxidative and a high reductive conversion indicating that in intact cells the equilibrium tends to biologically active GCS. However, in homogenized organoid cultures oxidative outweighed reductive activity. Secondly, the phosphatidylcholine synthesis of organoid cultures was enhanced by preincubation with GCS. CORT, as well as the hormonally inactive 11-DHC, increased the incorporation of [14C]choline into phosphatidylcholine. The effect of the latter was completely inhibited by glycyrrhetinic acid (inhibitor of 11 beta-HSD) indicating that it is caused by a previous conversion of 11-DHC into CORT via 11 beta-HSD. Thirdly, preincubation with GCS also altered 11 beta-HSD activity: dexamethasone or CORT both decreased the oxidative and increased the reductive activity in intact cells, indicating that glucocorticoids increase the rate of their own activation by positive feedback.

Inhibition of 11 beta-hydroxysteroid dehydrogenase activity enhances the antiproliferative effect of glucocorticosteroids on MCF-7 and ZR-75-1 breast cancer cells.

This in vitro study on MCF-7 and ZR-75-1 breast cancer cells showed that the antiproliferative action of glucocorticosteroids (GCS) on breast cancer cells is weakened by a high oxidative activity of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD; EC 1.1.1.146): both endogenic as well as synthetic GCS (dexamethasone, prednisolone) were metabolised to hormonally inactive 11-dehydro metabolites. This enzymatic shield protected the breast cancer cells from the antiproliferative action of GCS. Continuous exposure of breast cancer cells to GCS resulted in enhanced 11 beta-HSD activity. The intracellular GCS concentration was further reduced by this feedback and thus the antiproliferative effect was additionally weakened. These mechanisms of GCS deactivation could be influenced by inhibiting 11 beta-HSD with the liquorice compound glycyrrhetinic acid (GLY). In MCF-7 and ZR-75-1 cultures the antiproliferative effect of GCS was significantly increased by GLY.

Could a combined administration of dexamethasone and 3,5-dimethyl-3'-isopropyl-L-thyronine (DIMIT) be a more effective alternative to dexamethasone alone in the prevention of RDS?

OBJECTIVE: To test whether the combined application of dexamethasone (DEXA) and 3,5-dimethyl-3'-isopropyl-L-thyronine (DIMIT) induces the synthesis of surfactant protein A (SP-A) mRNA at a higher rate than both substances given alone? STUDY DESIGN: Organoid culture of fetal rat lungs (Wistar rats; day 19 of gestation) was prepared. After 48h of incubation we added DEXA (10(-5), 10(-7), 10(-8) and 10(-9)mol/l), DIMIT (10(-5), 10(-7) and 10(-9)mol/l) and the combination of DEXA in 10(-8)mol/l with various concentrations of DIMIT. After another 48h of incubation, northern blot and hybridization with a 32P-labeled SP-A cDNA probe was performed. One-way-variance-analysis with a Scheffé-test, Levene-test and one-sample-t-test were used for statistical analysis. RESULTS: DEXA alone above 10(-8)mol/l resulted in a significant increase, DIMIT resulted in a decrease of SP-A mRNA induction. Combined application of DIMIT and DEXA resulted in a significant increase compared to the controls. Compared to DEXA alone in 10(-8)mol/l, we found an increased induction, but the data were not significant. CONCLUSIONS: The combined application of DEXA and DIMIT shows a higher induction of SP-A mRNA than both drugs given alone.

Higher efficacy of letrozole in combination with trastuzumab compared to letrozole monotherapy as first-line treatment in patients with HER2-positive, hormone-receptor-positive metastatic breast cancer - results of the eLEcTRA trial.

The eLEcTRA trial compared efficacy and safety of letrozole combined with trastuzumab to letrozole alone in patients with HER2 and hormone receptor (HR) positive metastatic breast cancer (MBC). Patients were randomized to either letrozole alone (arm A, n = 31) or letrozole plus trastuzumab (arm B, n = 26) as first-line treatment. Additional 35 patients with HER2 negative and HR positive tumors received letrozole alone (arm C). Median time to progression in arm A was 3.3 months compared to 14.1 months in arm B (hazard ratio 0.67; p = 0.23) and 15.2 months in arm C (hazard ratio 0.71; p = 0.03). Clinical benefit rate was 39% for arm A compared to 65% in arm B (odds ratio 2.99, 95% CI 1.01-8.84) and 77% in arm C (odds ratio 5.34, 95% CI 1.83-15.58). The eLEcTRA trial showed that the combination of letrozole and trastuzumab is a safe and effective treatment option for patients with HER2 positive and HR positive MBC.

From biochemical to biophysical placental function tests in fetal surveillance.

Radioimmunoassays of human placental lactogen and estriol levels in the maternal plasma, ultrasound biometry of the abdominal diameter (AD), pulsed Doppler measurements of uteroplacental arteries, the common carotid artery (CCA), and the umbilical artery (UA) and fetal heart rate monitoring were simultaneously performed in 219 risk pregnancies from 26 weeks' onward at regular intervals. The prognostic value of all tests to predict small for gestational age infants (SGA) and fetal distress was evaluated simultaneously. Receiver operator characteristic allowed the simultaneous comparison of all methods: The AD was most predictive for early detection of SGA, even more than uteroplacental blood flow. Fetal blood flow redistribution expressed as a ratio of the resistance index of CCA/UA was the most significant test for detection of fetal distress later in pregnancy, even more than antenatal cardiotocography. Considering cutoff levels used in clinical routine, the sensitivity and specificity of the fetal AD in detecting intrauterine growth retardation more than 28 days before birth, were 71 and 81%, respectively. Pulsed Doppler measurements of CCA/UA less than 7 days before the delivery had a sensitivity and specificity of 83 and 90%, respectively. Our results demonstrate the historical change in fetal surveillance within one study group: If accurate routine ultrasound is available, the use of biochemical placental function tests is not justified, a procedure that is already accepted in nearly all perinatal centers. Fetal cerebral versus umbilical blood flow measurements should be applied as a tool to measure fetal blood flow redistribution in small fetuses to predict most precisely the risk for poor outcome and perinatal hypoxia.

[Current methods of prenatal determination of lung maturity of the infant from amniotic fluid]. / Aktuelle Methoden zur antepartalen Lungenreifebestimmung des Kindes aus dem Fruchtwasser.

Two currently used methods of determining fetal pulmonary maturity were verified on the basis of 148 amniotic fluid samples: one dimensional sequential thin-layer chromatographic determination of the lecithin: sphingomyelin ratio and of phosphatidyl glycerol, and an immunologic slide detection of phosphatidyl glycerol (Amniostat-FLM). With both methods, which have the advantage that they can be performed quickly and can sometimes even be used at the bedside (Amniostat-FLM), a respiratory distress syndrome in the newborn can be ruled out with a high degree of confidence if the threshold values are exceeded (L/S greater than 2; phosphatidyl glycerol present). As with all historical methods, the positive correctness of these two modern methods (no RDS if the child is born within 72 hours despite a negative phosphatidyl glycerol test in the amniotic fluid) is low. Clearly, apart from the quantitative existence of certain surfactant phospholipids (lecithin, phosphatidyl glycerol), there are other perinatal events or measures which determine the extent and severity of RDS in the immature newborn.

Actual state in the diagnosis of fetal lung maturity.

Four methods of phospholipid analysis in amniotic fluid were compared: miniaturized version of a one-dimensional thin-layer chromatographic determination of the lecithin/sphingomyelin ratio; A one-dimensional thin-layer chromatographic separation of all phospholipid components in amniotic fluid; A completely enzymatic determination of amniotic fluid lecithin concentration; An immunological method for semiquantitative measurement of phosphatidylglycerol. The various phospholipid parameters (L/S ratio, lecithin concentration, phosphatidylglycerol detection either by thin-layer chromatography or immunological methods) show a strong correlation to advancing gestational age. While we were not able to prove a statistically significant correlation between enzymatic lecithin values or L/S ratio values and the occurrence of neonatal RDS cases in this study due to the relatively small number of RDS cases, the L/S ratio values obtained by one-dimensional thin-layer chromatography and the phosphatidylglycerol values, determined either chromatographically or immunologically, showed a clear correlation to the expected development of RDS in the newborn. Thus we can conclude that under the actual methods of amniotic fluid phospholipid evaluation immunological phosphatidylglycerol determination as well as the one-dimensional separation of amniotic fluid phospholipids, which is easier to run than the "lung profile" determination of Kulovich and Gluck provide good information about the exclusion of RDS.

[Effect of maternal age on the course of labor--analysis of women over 40 years of age]. / Einfluss des mütterlichen Alters auf den Geburtsverlauf--Analyse bei Frauen über 40 Jahre.

In order to determine the birth risk for pregnant women in the age group > or = 40, the delivery data of 143 pregnant women in this age group were retrospectively evaluated over a 3-year period. Here, 37 of these 143 women (25.9%) were primiparae. The following was examined: The number of prenatal examinations (including ultrasound examinations), the incidence of genetic examinations, the delivery methods with the percentage of surgical deliveries, complications at the time of delivery as well as the fetal outcome with APGAR values, umbilical artery pH, birth weights, neonatal morbidity and mortality. The delivery results were compared with representative populations of women between 20-29 years (n = 2252) and 30-39 years (n = 1980). Pregnancy in older women still ends significantly more often with cesarean section than in younger women. Here, the rate of cesarean sections was 32.7% compared to 21.9% in 30-39-year-olds and 15.8% in 20-29-year-olds. However, parity has an even greater influence on the mode of delivery than age. Only 30.1% of multi-parae over 40 years underwent surgical delivery (vaginal and abdominal) compared to 77.3% of primiparae. It was also found that multiparae more rarely had surgical delivery than younger primiparae (30-39 years 53.3%, 20-29 years 39.3%). Despite the high surgical delivery rate being in the group of primiparae over 40 years, the fetal outcome was comparatively poor, so that the less restrictive indication for surgical delivery seems justified.

Effect of epidermal growth factor on enzymes of phospholipid biosynthesis in lung and liver of fetal rat in vivo and in vitro.

Epidermal growth factor (EGF), a mitogenic polypeptide that binds to cell surface receptors, is an important regulator of cell differentiation and fetal lung surfactant synthesis, and may be used as a potential novel therapeutic agent in prematurity. Nevertheless, the distinct role in lung development and its mechanisms of action are not well understood. We investigated in vivo the systemic effect of intrafetally administered EGF (200 ng/g fetal body weight) and maternally administered dexamethasone (DEXA; 0.2 and 2.0mg/kg maternal body weight) on the activity of important enzymes of the phospholipid synthesis in the fetal rat lung and liver: choline kinase (EC 2.7.1.32), cholinephosphate cytidyltransferase (EC 2.7.7.15), choline phosphotransferase (EC 2.7.8.2), lysolecithin acyltransferase (EC 2.3.1.23) and glycerolphosphate phosphatidyltransferase (EC 2.7.8.5). Additionally, in vivo and in vitro effects of DEXA on EGF receptor synthesis, and the effects of EGF on protein content and morphogenesis of the fetal rat lung organoid culture, were evaluated. Whereas DEXA induced the activity of all investigated enzymes of phospholipid synthesis and increased EGF receptor synthesis, EGF has no effects on the enzymes, either in vivo or in vitro. EGF enhanced protein synthesis and morphogenesis in vitro. With respect to our data and the literature, we hypothesize that DEXA and EGF may act on different cellular sides. Whereas glucocorticoids induce surfactant phospholipid synthesis, EGF should be more involved in cell proliferation and morphogenesis.

[Prenatal determination of lung maturity from amniotic fluid--indications and new methods]. / Antepartale Lungenreifebestimmung aus dem Fruchtwasser--Indikationen und neue Methoden.

Although fetal lung maturity determination is carried out more and more rarely in the German-speaking area, a reliable information about the degree of lung maturity is still very important in the care of high-risk pregnancies. The side effects and costs of a postpartal surfactant administration can be avoided if lung maturity is proved. Indications for determination of fetal lung maturity are the threatening preterm delivery and the premature rupture of membranes before the 34th week of gestation and uncertain gestational age. Furthermore, in preeclampsia resp. in diabetes mellitus, which is difficult to control, premature delivery may be necessary. To improve lung maturity testing we introduce a new "sequence scheme" containing three lung maturity tests which are easy to carry out (in the following sequence: Amniostat-FLM ultrasensitive, counting of the lamellar bodies in amniotic fluid, surfactant/albumin ratio with TDx-FLM). The principle of this scheme is, that if any of these three tests indicates lung maturity, the sequence is terminated and no further test is performed. Only if all three tests indicated immaturity, the child was at risk for RDS. In 87 amniotic fluid samples with 7 RDS-cases, we achieved high predictive values for lung maturity (specificity 90%, sensitivity 100%, predictive value of positive test 47%, predictive value of negative test 100%). In 62% only one test was needed for lung maturity determination. It is possible to use other combinations in such a scheme (e.g. the L/S ratio). This might lead to equal or perhaps better results. An advantage of this suggested "sequence scheme" is that it can be performed in any clinic.

[Prevention of neonatal risk by general screening for diabetes in pregnancy, intensive diagnosis and subsequent therapy]. / Vermeidung kindlicher Risiken durch ein generelles Gestationsdiabetes-Screening, intensivierte Diagnostik und konsequente Therapie.

Only 10% of all gestational diabetic mothers in Germany are diagnosed with the current risk-screening. The elevated perinatal risks in case of an unrecognized or insufficiently treated gestational diabetes remains controversial. The purpose of our study was to determine if the number of recognized cases could be increased by a general screening method, and with intensive medical diagnostics the complication rate reduced. Routine blood glucose samplings during the outpatient care were performed throughout the pregnancy. In case of values over 100 mg/dl a 75 g OGTT was done for an exclusion of gestational diabetes. In case of gestational diabetes the patients were asked to follow a special exercise and diet programme as well as self-blood glucose determinations throughout the day. The amniotic fluid insulin level was of substantial value for the indication of insulin therapy. In 6% of the screened patients a gestational diabetes was diagnosed. There was a significant increase (p < 0.001) of fetal macrosomia and diabetic fetopathy in the group without amniocentesis (n = 22) in comparison to the group with invasive (n = 81). We demand the introduction of a general screening for every pregnant patient. By an intensification of the diagnostic methods as well as by a strictly appropriate therapy it should be possible to reduce the fetal and neonatal complications.

Evaluation of the fetal assessment score in pregnancies at risk for intrauterine hypoxia.

OBJECTIVE: Our purpose was to define the diagnostic value of a new fetal assessment score that is based on each of the components of the Apgar score. STUDY DESIGN: A fetal assessment score was established to study the main fetal vital functions: (1) cardiovascular (heart rate, color of the skin in the Apgar score), now based on fetal heart rate patterns and Doppler assessment of fetal blood flow redistribution, (2) fetal respiratory (quality of breathing in the Apgar score), now based on Doppler assessment of uteroplacental perfusion, and (3) neuromuscular function (tone and reflexes in the Apgar score), now based on fetal tone and response to external stimuli. The fetal assessment score was used in the study of 110 postdate pregnancies and 103 small-for-gestational-age infants and was compared with the traditional biophysical profile score in the prediction of perinatal outcome. RESULTS: There were significant associations between both the fetal assessment score and the biophysical profile score with fetal distress that necessitated operative delivery, low Apgar scores, and low umbilical cord arterial blood pH. However, receiver-operator characteristic plots demonstrated that the fetal assessment score was superior to the biophysical profile score in predicting fetal distress and low Apgar values particularly in the small-for-gestational-age infants. The best single parameters in predicting fetal distress were the amniotic fluid volume in the biophysical profile score and fetal heart rate patterns and pulsed Doppler measurements in the new score. CONCLUSION: A fetal Apgar score in which respiration is assessed by placental perfusion rather than chest movements and in which skin color is assessed by centralization of fetal blood flow may be better than the traditional biophysical profile score in predicting fetal hypoxic compromise.

Prediction of RDS by amniotic fluid analysis: a comparison of the prognostic value of traditional and recent methods.

The determination of lecithin or even more the lecithin/sphingomyelin (L/S) ratio in amniotic fluid are both well established in the prediction of neonatal RDS. The immunological measurement of phosphatidylglycerol (PG) and the determination of the surfactant/albumin (S/A) ratio by fluorescence polarization (TDx FLMR) have recently been introduced for the detection of fetal lung maturity. In order to compare traditional versus recent methods L/S ratio and PG determination by one dimensional thin-layer chromatography, enzymatic analysis of lecithin, immunological determination of PG by Amniostat-FLMR and the fluorescence polarization of S/A-ratio by the TDx FLMR were all performed in 141 amniotic fluid samples of 122 patients. Only one out of 72 samples was false negative in the enzymatic lecithin determination (sensitivity 88%). All other methods have a sensitivity and a negative predictive value of 100%. The positive predictive values and the specificity varied between 22%-50% and 58%-87% respectively. The false positive rate, which is high for all methods, is lowest for the L/S ratio. This study demonstrates, that the recent methods are reasonable alternatives in all cases with a positive test. In clinical practice they have the advantage, that the result can be obtained in 15 minutes. If the test is predictive for lung immaturity the L/S ratio should be performed in addition to decrease the false positive rate before any clinical decision is made.

[Accuracy of ultrasound weight assessment--comparison of vertex vs. breech presentation]. / Genauigkeit der sonographischen Gewichtsschätzung--Vergleich Schädellage vs. Beckenendlage.

AIM: To determine the quality of prenatal sonographic weight estimation, comparing breech and vertex presentations. METHOD: In 147 breech presentations (BP) and 149 vertex presentations (VP), the biparietal head diameter (BPD), the fronto-occipital head diameter (FOD) and the transverse abdominal diameter (ATD) were measured. From these data, the weight was estimated, using Shepard's formula before 28 weeks and Hansmann's thereafter, and compared to the delivery weight. Both formula were modified to incorporate the virtual BPD (vBPD), derived from the BPD, FOD and the calculated head circumference. In the BP group, the role of examiner skills was evaluated, comparing the accuracy of experienced (DEGUM II qualifications) and average individuals. RESULTS: The accuracy for BP was nearly identical to that for VP (= 0.915 vs. = 0.929). The examiners' qualifications had a detectable but not significant influence on the results (= 0.942 vs. = 0.892). CONCLUSION: Ultrasound measurements yield comparable results in both BP and VP, if the vBPD is used. In our opinion, ultrasonic weight estimation is a useful adjunct when determining the manner of delivery in BP.

[Management of premature rupture of fetal membranes in obstetrical departments in Germany]. / Management des frühen vorzeitigen Blasensprungs an geburtshilflichen Abteilungen in Deutschland.

Since the treatment of premature ruptures of membranes is not only controversial in the German but also in the international literature, we performed a survey of all obstetrics departments in Germany. From a total of 843 hospitals, 444 questionnaires were returned for evaluation (52.7%). The purpose was to determine which diagnostic and therapeutic regimes are used and how these agree with the literature. In addition to questions on the type of hospital, birth rates with a percentage of premature births and applied diagnostic parameters, our special interest focused on therapy, particularly with regard to prophylactic antibiotic application, tocolytic treatment and lung maturity induction. Prophylactic antibiotics are used in 36.7% and prophylactic tocolytic therapy in 41.7% of the departments. Interestingly, lung maturity induction was performed in 93.5%, in part even before the 28th week of pregnancy, although the effect of this therapy has not yet been proven at a very early stage of gestation. Due to the different views in the literature and, in part, a lack of basic scientific data, it seems there is a preference for the procedure, in which the best personal experience has been made. Because premature ruptures of the membranes is responsible for 30-40% of premature births, it is urgently necessary to clarify this controversial problem by large multicenter studies so that the treatment of early premature ruptures of the amnion can be founded on a rational basis.

[Insulin-like-growth-factor-binding-protein 1 (IGFBP-1) and fetal fibronectin in diagnosis of premature rupture of fetal membranes]. / Insulin like growth factor binding protein 1 (IGFBP-1) und fetales Fibronectin in der Diagnostik eines vorzeitigen Blasensprunges.

The diagnosis of a premature rupture of membranes presents no problem in the vast majority of cases. However, a reliable diagnosis is clinically not possible in about 10%. Most methods available lack the necessary sensitivity and specificity. Since the clinical consequences of a false diagnosis are considerable (overtreatment for false-positive and risk of infection for false-negative results), it is essential to clinically establish new, minimally invasive methods with higher predictive powers. In the present study we compared: the AMNI Check for detection of insulin-like growth-factor binding protein 1 (IGFBP-1); a membrane immunoassay for detection of fetal fibronectin (fFn); pH indicator paper; and, to verify a rupture of membranes in unclear cases, amniocentesis with installation of indigo carmine. The examination was performed in a group of 75 patients, 35 with and 40 without rupture of the membranes. The best results were obtained for the AMNI Check (sensitivity and negative correctness 100%, specificity and positive correctness 83%). With the same sensitivity and negative correctness, the membrane immunoassay for fFn achieved a specificity of 70% and a positive correctness of 74%. The pH indicator paper had the lowest predictive value (sensitivity 94%, negative correctness 93%, specificity 63%, positive correctness 69%). Both the AMNI Check and the test for detection of fetal fibronectin can be recommended for reliable exclusion of premature rupture of membranes. Amniocentesis should however be performed in uncertain cases with a positive test result. Nevertheless, considerable reduction of this invasive method is possible.