Ex Utero Extracorporeal Support as a Model for Fetal Hypoxia and Brain Dysmaturity.

BACKGROUND: Congenital heart disease (CHD) is associated with abnormal fetal brain development, a phenomenon that may be related to decreased cerebral oxygen delivery in utero. We used an artificial womb model to test the hypothesis that decreasing fetal oxygen delivery would reproduce physiologic changes identified in fetuses with CHD. METHODS: Experimental (hypoxemic) fetal lambs (mean gestational age, 111 ± 3 days; n = 4) and control animals (112 days; n = 5) were maintained in the artificial womb for a mean of 22 ± 6 days. Oxygen delivery was reduced to 15.6 ± 1.0 mL/kg/min in the hypoxemia animals versus 21.6 ± 2.0 mL/kg/min in the control animals. Blood chemistry analysis and sonographic evaluation were performed daily. An additional control group (n = 7) was maintained in utero and harvested for analysis at gestational age 134 ± 4 days. RESULTS: Physiologic variables were monitored continuously, and no statistical differences between the groups were identified. Fetal oxygen delivery and arterial partial pressure of oxygen were remarkably lower in the experimental group longitudinally. Increased umbilical artery and decreased middle cerebral artery resistance resulted in a lower cerebral to umbilical resistance ratio, similar to the brain sparing effect observed in human fetuses with CHD. Experimental brains were smaller than control brains in relation to the calvarium on magnetic resonance. CONCLUSIONS: Sustained hypoxemia in fetal sheep leads to altered cerebrovascular resistances and loss of brain mass, similar to human fetuses with CHD. This unique model provides opportunities to investigate the pathologic process underlying CHD-associated brain dysmaturity and to evaluate potential fetal neuroprotective therapies.

Chronic intrauterine hypoxia alters neurodevelopment in fetal sheep.

OBJECTIVE: We tested the hypothesis that chronic fetal hypoxia, at a severity present in many types of congenital heart disease, would lead to abnormal neurodevelopment. METHODS: Eight mid-gestation fetal sheep were cannulated onto a pumpless extracorporeal oxygenator via the umbilical vessels and supported in a fluid-filled environment for 22 ± 2 days under normoxic or hypoxic conditions. Total parenteral nutrition was provided. Control fetuses (n = 7) were harvested at gestational age 133 ± 4 days. At necropsy, brains were fixed for histopathology. Neurons were quantified in white matter tracts, and the thickness of the external granular layer of the cerebellum was measured to assess neuronal migration. Capillary density and myelination were quantified in white matter. Data were analyzed with unpaired Student t tests or 1-way analysis of variance, as appropriate. RESULTS: Oxygen delivery was reduced in hypoxic fetuses (15.6 ± 1.8 mL/kg/min vs 24.3 ± 2.3 mL/kg/min, P < .01), but umbilical blood flow and caloric delivery were not different between the 2 groups. Compared with normoxic and control animals, hypoxic fetuses had reduced neuronal density and increased external granular layer thickness. Compared with normoxic and control animals, hypoxic fetuses had increased capillary density in white matter. Cortical myelin integrity score was lower in the hypoxic group compared with normoxic and control animals. There was a significant negative correlation between myelin integrity and capillary density. CONCLUSIONS: Chronic fetal hypoxia leads to white matter hyper-vascularity, decreased neuronal density, and impaired myelination, similar to the neuropathologic findings observed in children with congenital heart disease. These findings support the hypothesis that fetal hypoxia, even in the setting of normal caloric delivery, impairs neurodevelopment.

Arterio-venous blood gas Δvalues for validation of umbilical cord blood samples at birth are not only biased by sample mix ups but also affected by clinical factors.

INTRODUCTION: Traditional validation of umbilical cord blood samples with positive veno-arterial ΔpH and arterio-venous ΔpCO2 values confirms the source of samples, whereas negative Δvalues represent mix-up of samples. To investigate whether this is true, the distributions of V-A ΔpO2 and A-V Δlactate were also explored and related to clinical characteristics. In addition, different cord blood sampling techniques were evaluated. MATERIAL AND METHODS: Register study with cord blood acid-base and clinical data from 27 233 newborns. Clinical characteristics were related to positive, zero and negative Δvalues. Blood samplings from unclamped and double-clamped cords were compared. A two-sided P < 0.05 was considered significant. RESULTS: ΔpH and ΔpCO2 values distributed into positive, around zero, and negative sub-populations, with significant differences in pH and clinical characteristics between sub-populations. No such sub-populations were distinguished for ΔpO2 and Δlactate. The 2.5th and 5th ΔpH percentiles were 0.013 and 0.022, respectively, and for ΔpCO2 0.30 and 0.53 kPa. Applying 5th percentile criteria resulted in 3.5% of "approved" cases showing a ΔpO2  ≤ 0. Puncture and sampling of the unclamped cord resulted in significantly better sample quality. CONCLUSIONS: Unphysiological negative ΔpO2 values occurred despite correct validation with traditional criteria. Δlactate cannot be used for validation because both positive and negative values are physiological. Positive/around zero/negative ΔpH and ΔpCO2 sub-populations were associated with significant differences in pH and clinical characteristics, indicating that defective sampling and sample handling are not the sole explanations for negative Δvalues. Prompt puncture and sampling of the unclamped cord resulted in best sample quality.

Effects of inter-twin vascular anastomoses of monochorionic twins with selective intrauterine growth restriction on the contents of placental mitochondria DNA.

BACKGROUND: Placental mitochondrial DNA (mtDNA) has been proposed to be an indicator for placental hypoxia. This study was designed to evaluate the effect of vascular anastomoses between monochorionic (MC) twins on placental mtDNA. METHODS: In this study, twin-twin transfusion syndrome (TTTS) treated with laser therapy and MC twins without TTTS (without laser therapy) resulting in two live babies were included in this study. The placental mtDNA fold changes (FC) between the small and large twins were analyzed using real-time quantitative PCR. TTTS twins with selective intrauterine growth restriction (sIUGR) are categorized as group 1, TTTS without sIUGR as group 2, MC twins without TTTS but with sIUGR as group 3, and MC twins without both TTTS and sIUGR as group 4. RESULTS: There were seven cases in group 1, eight in group 2, 26 in group 3, and 24 in group 4 cases. The placental mtDNA FC were significantly higher in group 1 (1.57 ± 0.9) compared to that of the group 3 (0.86 ± 0.6). CONCLUSION: In MC twin pregnancies with sIUGR, the placental mtDNA FC between the small and large twins are different between cases with and without inter-twin anastomoses. These findings suggest that the inter-twin anastomoses in the MC twins with sIUGR may provide rescue perfusion from the appropriate-for-gestational-age twin to the sIUGR one.

Decreased neuroinflammation correlates to higher vagus nerve activity fluctuations in near-term ovine fetuses: a case for the afferent cholinergic anti-inflammatory pathway?

BACKGROUND: Neuroinflammation in utero may contribute to brain injury resulting in life-long neurological disabilities. The pivotal role of the efferent cholinergic anti-inflammatory pathway (CAP) in controlling inflammation, e.g., by inhibiting the HMGB1 release, via the macrophages' α7 nicotinic acetylcholine receptor (α7nAChR) has been described in adults, but its importance in the fetus is unknown. Moreover, it is unknown whether CAP may also exert anti-inflammatory effects on the brain via the anatomically predominant afferent component of the vagus nerve. METHODS: We measured microglial activation in the ovine fetal brain near term 24 h after the umbilical cord occlusions mimicking human labor versus controls (no occlusions) by quantifying HMGB1 nucleus-to-cytosol translocation in the Iba1+ and α7nAChR+ microglia. Based on multiple clinical studies in adults and our own work in fetal autonomic nervous system, we gauged the degree of CAP activity in vivo using heart rate variability measure RMSSD that reflects fluctuations in vagus nerve activity. RESULTS: RMSSD correlated to corresponding plasma IL-1ß levels at R = 0.57 (p = 0.02, n = 17) and to white matter microglia cell counts at R = -0.89 (p = 0.03). The insult increased the HMGB1 translocation in α7nAChR+ microglia in a brain region-dependent manner (p < 0.001). In parallel, RMSSD at 1 h post insult correlated with cytosolic HMGB1 of thalamic microglia (R = -0.94, p = 0.005), and RMSSD at pH nadir correlated with microglial α7nAChR in the white matter (R = 0.83, p = 0.04). Overall, higher RMSSD values correlated with lower HMGB1 translocation and higher α7nAChR intensity per area in a brain region-specific manner. CONCLUSIONS: Afferent fetal CAP may translate increased vagal cholinergic signaling into suppression of cerebral inflammation in response to near-term hypoxic acidemia as might occur during labor. Our findings suggest a new control mechanism of fetal neuroinflammation via the vagus nerve, providing novel possibilities for its non-invasive monitoring in utero and for targeted treatment.

Foetal scalp blood sampling during labour for pH and lactate measurements.

Second-line methods of foetal monitoring have been developed in an attempt to reduce unnecessary interventions due to continuous cardiotocography (CTG), and to better identify foetuses that are at risk of intrapartum asphyxia. Very few studies directly compared CTG with foetal scalp blood (FBS) and CTG only. Only one randomised controlled trial (RCT) was published in the 1970s and had limited power to assess neonatal outcome. Direct and indirect comparisons conclude that FBS could reduce the number of caesarean deliveries associated with the use of continuous CTG. The main drawbacks of FBS are its invasive and discontinuous nature and the need for a sufficient volume of foetal blood for analysis, especially for pH measurement, resulting in failure rates reaching 10%. FBS for lactate measurement became popular with the design of test-strip devices, requiring <0.5 mL of foetal blood. RCTs showed similar outcomes with the use of FBS for lactates compared with pH in terms of obstetrical interventions and neonatal outcomes. In conclusion, there is some evidence that FBS reduces the need for operative deliveries. However, the evidence is limited with regard to actual standards, and large RCTs, directly comparing CTG only with CTG with FBS, are still needed.

Did antepartum hypoxic insult caused by fetal vessel thrombosis influence the procalcitonin level in umbilical blood? A case report.

We report a case of marked elevation of the procalcitonin level in umbilical blood and neonatal blood at birth. The mother did not perceive fetal motion. Antepartum fetal heart rate monitoring showed a loss of variability and absence of acceleration. No fetal breathing movement, fetal movement, or fetal tone were observed by ultrasonography. The female neonate was delivered by cesarean section at 25 weeks of gestation, with birthweight 774 g. The umbilical arterial pH value at birth was 7.29. Mild elevation in interleukin-6 and tumor necrosis factor-α in umbilical blood were observed. Cytochrome c showed a high level in umbilical and neonatal blood at birth. Placental histopathology revealed multiple fetal vessel thrombosis in the large stem villi and chorionic vessels. The neonate showed no infectious signs throughout the neonatal period. Computed tomography at 3 months of age revealed atrophy in the cerebrum and cerebellum. At 1 year after birth, the infant showed spastic quadriplegia. In this case, antepartum asphyxia due to fetal vessel thrombosis may have influenced the elevation of procalcitonin level in umbilical blood and neonatal blood at birth.

[Birth hypoxia]. / Porodní hypoxie.

OBJECTIVE: Evaluation of the commonly used laboratory and clinical parameters of the newborn shortly after birth. Check thresholds acidemia, and in relation to the method of termination of pregnancy. DESIGN: Retrospective epidemiological study. SETTING: Department of Obstetrics and Gynecology, University Hospital, Olomouc. METHODS: Of the 26,869 children born in the years 2000 to 2013 Inclusion criteria (complete clinical and laboratory findings after birth) fulfill 23,471 (87.4%) neonates. Methods for evaluation of newborns included Apgar score calculation and arterial umbilical cord blood pH and lactate analysis. RESULTS: A total of 0.7% (157) of the neonates had severe acidosis pH below 7.00 arterial umbilical cord blood, its prevalence varies annually between 0.1 to 1.1%. Cutoff lactate in relation to pH < 7.00 was 6.3 mmol/l (n = 23 471, the sensitivity of 92.99%, specificity 92.15%, AUC = 0.972). For children of low weight < 2500 g the cutoff value is lower, 5.3 mmol/l (n = 2592, 89.66% sensitivity, specificity 91.10% AUC = 0.912). Suprathreshold lactate values was 8.4% (1977) newborns. Correlation of pH and lactate to Apgar evaluation is very low and in the range from 1 to 10 minutes gradually decreases. Worse Apgar evaluation in children of low birth weight do not correspond to laboratory findings acidosis, which is probably related to prematurity and lower energy reserves. Operating cesarean births in particular accounts for more than half of those with worse clinical findings Apgar and pH <7.00, but only 30% supratreshold lactate values. Also worse clinical evaluation after caesarean section is not in accordance with the laboratory findings. Vaginal surgery, especially forceps have a significant share of severe acidosis than cesarean, regardless of their frequency. Risk factor of forceps to pH less 7.00,OR = 9.28 (5.39 -15.77), P = 0.0000000, while caesarean to pH less 7,00 had OR = 1.52 (1.08 to 2.14), P = 0.01408156. CONCLUSION: The results obtained confirm that acidosis after birth is quite common, although they may not have response on the clinical condition of the newborn after birth. Evaluation of Apgar is little objective for the detection of hypoxia during birth and is influenced by the immaturity of newborn and method of delivery. Lactate levels may contribute to an objective assessment of hypoxia during birth. Values above 6.3 mmol/l can be considered an important indicator of newborn acidosis and birth hypoxia.

Xanthine oxidase and the fetal cardiovascular defence to hypoxia in late gestation ovine pregnancy.

Hypoxia is a common challenge to the fetus, promoting a physiological defence to redistribute blood flow towards the brain and away from peripheral circulations. During acute hypoxia, reactive oxygen species (ROS) interact with nitric oxide (NO) to provide an oxidant tone. This contributes to the mechanisms redistributing the fetal cardiac output, although the source of ROS is unknown. Here, we investigated whether ROS derived from xanthine oxidase (XO) contribute to the fetal peripheral vasoconstrictor response to hypoxia via interaction with NO-dependent mechanisms. Pregnant ewes and their fetuses were surgically prepared for long-term recording at 118 days of gestation (term approximately 145 days). After 5 days of recovery, mothers were infused i.v. for 30 min with either vehicle (n = 11), low dose (30 mg kg(-1), n = 5) or high dose (150 mg kg(-1), n = 9) allopurinol, or high dose allopurinol with fetal NO blockade (n = 6). Following allopurinol treatment, fetal hypoxia was induced by reducing maternal inspired O2 such that fetal basal P aO 2 decreased approximately by 50% for 30 min. Allopurinol inhibited the increase in fetal plasma uric acid and suppressed the fetal femoral vasoconstrictor, glycaemic and lactate acidaemic responses during hypoxia (all P < 0.05), effects that were restored to control levels with fetal NO blockade. The data provide evidence for the activation of fetal XO in vivo during hypoxia and for XO-derived ROS in contributing to the fetal peripheral vasoconstriction, part of the fetal defence to hypoxia. The data are of significance to the understanding of the physiological control of the fetal cardiovascular system during hypoxic stress. The findings are also of clinical relevance in the context of obstetric trials in which allopurinol is being administered to pregnant women when the fetus shows signs of hypoxic distress.

Maternal and fetoplacental hypoxia do not alter circulating angiogenic growth effectors during human pregnancy.

One causal model of preeclampsia (PE) postulates that placental hypoxia alters the production of angiogenic growth effectors (AGEs), causing an imbalance leading to maternal endothelial cell dysfunction. We tested this model using the natural experiment of high-altitude (HA) residence. We hypothesized that in HA pregnancies 1) circulating soluble fms-like tyrosine kinase 1 (sFlt-1) is increased and placental growth factor (PlGF) decreased, and 2) AGE concentrations correlate with measures of hypoxia. A cross-sectional study of healthy pregnancies at low altitude (LA) (400 m) versus HA (3600 m) compared normal (n = 80 at HA, n = 90 at LA) and PE pregnancies (n = 20 PE at HA, n = 19 PE at LA). Blood was collected using standard serum separation and, in parallel, by a method designed to inhibit platelet activation. AGEs were measured by enzyme-linked immunosorbent assays. AGEs did not differ between altitudes in normal or PE pregnancies. AGE concentrations were unrelated to measures of maternal or fetal hypoxia. PlGF was lower and sFlt-1 higher in PE, but overlapped considerably with the range observed in normal samples. PlGF correlated with placental mass in both normal and PE pregnancies. The contribution of peripheral cells to the values measured for AGEs was similar at LA and HA, but was greater in PE than in normotensive women. Hypoxia, across a wide physiological range in pregnancy, does not alter levels of circulating AGEs in otherwise normal pregnancies. Peripheral cell release of AGEs with the hemostasis characteristic of standard blood collection is highly variable and contributes to a doubling of the amount of sFlt-1 measured in PE as compared to normal pregnancies.

The absolute nucleated red blood cell (aNRBC) count at birth is not an indicator for retinopathy of prematurity (ROP).

OBJECTIVE: To establish the reproducibility of a published observation by Lubetzky et al. that infants affected by retinopathy of prematurity (ROP) had higher absolute nucleated red blood cell (aNRBC) counts than those unaffected. The authors suggested that infants exposed to intrauterine hypoxia are at higher risk for ROP. We attempted to verify this reported relationship of ROP with the aNRBCs at birth and hypothesized that infants with ROP ≥ stage 2 have higher aNRBCs at birth. STUDY DESIGN: We report a retrospective 1:1 case matched analysis where cases had a diagnosis of grade II ROP or worse and matching infants had confirmed stage I or no ROP. Eligible infants had birth weights of 501 to 1500 g and were discharged alive from 1st January 2000 to 31st December 2008. Wilcoxon's signed rank test was performed for continuous comparisons. This study was approved by two local Institutional Review Boards. RESULT: In all, 66 matched pairs were analyzed. When comparing aNRBCs there was no statistically significant relationship (w=-0.265, P=0.791) between the ROP affected group (M=4550, s.d.=7342) and the unaffected group (M=5287, s.d.=6524). CONCLUSION: We are unable to support the previously reported relationship of aNRBCs with ROP. Our population was three times larger, had higher aNRBCs and less retinopathy than previously reported. A biological principle of cause and effect or predisposition to ROP as reflected by aNRBCs should have been easier for us to demonstrate, if it existed.

Effect of the umbilical cord blood acid-base status and gas values on the yield of mononuclear cells and CD34⁺ cells.

AIM: To investigate the influence of umbilical cord blood (CB) acid-base status and gas values on the yield of mononuclear cells and CD34⁺ cells, pH, pCO2, pO2, HCO3⁻ and base excess were measured in arterial CB samples obtained from normal full-term deliveries. The relationship of these values with the yield of mononuclear cells and CD34⁺ cells detected in venous CB was analyzed. MATERIAL AND METHODS: A total of 145 CB units were collected from full-term vaginal deliveries at a single hospital. Immediately after delivery, a segment of the umbilical cord was double clamped, and arterial CB was analyzed to determine the acid-base status and gases. Venous CB was collected in a sterile collection bag and processed for cell separation within 24 h of collection. The relationship between umbilical arterial acid-base status, each gas value, and the yield of mononuclear cells and CD34⁺ cells was analyzed. RESULTS: Statistically significant correlations were observed between the net weight of CB and the total mononuclear and CD34⁺ cell counts. In addition, there was a negative correlation between the mononuclear cell counts and pH, but a positive correlation between the mononuclear cell counts and pCO2. However, no significant differences were observed between the primipara and multipara groups in terms of the net weight of CB, total mononuclear cell counts and total CD34⁺ cell counts. CONCLUSION: The findings of the present study show that the mononuclear cell counts are correlated with arterial CB pH and pCO2, suggesting the involvement of fetal hypoxia on the yield of mononuclear cells.

Conduta obstétrica na centralização da circulação fetal / Obstetric management in fetal brain sparing effect

A centralização do fluxo sanguíneo fetal é um fenômeno de compensação vascular bastante estudado na atualidade. Trata-se de alterações na resistência da circulação fetal, caracterizada pela redistribuição hemodinâmica do fluxo sanguíneo, com perfusão preferencial para órgãos nobres (cérebro, coração e glândulas adrenais) em detrimento dos pulmões, rins, baço e esqueleto, o que pode ser diagnosticado pelo estudo dopplervelocimétrico. O momento ideal para intervenção obstétrica ainda não é consenso. Uma das grandes preocupações, em relação à avaliação da vitalidade fetal, diz respeito ao momento ideal para interrupção da gravidez, uma vez que alguns dos métodos utilizados apresentam uma alta frequência de resultado falso-positivos, podendo ocasionar um nascimento prematuro, por vezes, desnecessário. Em fetos muito prematuros a opção pela interrupção da gravidez pode trazer consequências irreversíveis. Na tentativa de minimizar os danos, optou-se pela realização de uma revisão, baseada nas melhores evidências sobre a conduta nos fetos centralizados.

Fetal brain-sparing effect is a vascular compensation phenomenon widely studied today. Diagnosed by Doppler study it consists of changes on resistance in the fetal circulation characterized by hemodynamic redistribution of blood flow, with preferential perfusion to brain, heart and adrenal glands compared to the lungs, kidneys, spleen and skeleton. There is no consensus over ideal time for obstetric intervention. Ideal time for pregnancy termination is of major concern when assessing fetal vitality since methods used today have high false positives rate, leading to unnecessary prematurity. In extreme prematurity the decision to terminate pregnancy can lead to irreversible consequences. In an attempt to minimize damage, it was decided to carry out a review, based on the best evidence regarding conduct in fetal brain sparing effect.

Application of comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry method to identify potential biomarkers of perinatal asphyxia in a non-human primate model.

Perinatal asphyxia is a leading cause of brain injury in infants, occurring in 2-4 per 1000 live births. The clinical response to asphyxia is variable and difficult to predict with current diagnostic tests. Reliable biomarkers are needed to help predict the timing and severity of asphyxia, as well as response to treatment. Two-dimensional gas chromatography-time-of-flight-mass spectrometry (GC×GC-TOFMS) was used herein, in conjunction with chemometric data analysis approaches for metabolomic analysis in order to identify significant metabolites affected by birth asphyxia. Blood was drawn before and after 15 or 18 min of cord occlusion in a Macaca nemestrina model of perinatal asphyxia. Postnatal samples were drawn at 5 min of age (n=20 subjects). Metabolomic profiles of asphyxiated animals were compared to four controls delivered at comparable gestational age. Fifty metabolites with the greatest change pre- to post-asphyxia were identified and quantified. The metabolic profile of post-asphyxia samples showed marked variability compared to the pre-asphyxia samples. Fifteen of the 50 metabolites showed significant elevation in response to asphyxia, ten of which remained significant upon comparison to the control animals. This metabolomic analysis confirmed lactate and creatinine as markers of asphyxia and discovered new metabolites including succinic acid and malate (intermediates in the Krebs cycle) and arachidonic acid (a brain fatty acid and inflammatory marker) as potential biomarkers. GC×GC-TOFMS coupled with chemometric data analysis are useful tools to identify acute biomarkers of brain injury. Further study is needed to correlate these metabolites with severity of disease, and response to treatment.

Antenatal allopurinol for reduction of birth asphyxia induced brain damage (ALLO-Trial); a randomized double blind placebo controlled multicenter study.

BACKGROUND: Hypoxic-ischaemic encephalopathy is associated with development of cerebral palsy and cognitive disability later in life and is therefore one of the fundamental problems in perinatal medicine. The xanthine-oxidase inhibitor allopurinol reduces the formation of free radicals, thereby limiting the amount of hypoxia-reperfusion damage. In case of suspected intra-uterine hypoxia, both animal and human studies suggest that maternal administration of allopurinol immediately prior to delivery reduces hypoxic-ischaemic encephalopathy. METHODS/DESIGN: The proposed trial is a randomized double blind placebo controlled multicenter study in pregnant women at term in whom the foetus is suspected of intra-uterine hypoxia.Allopurinol 500 mg IV or placebo will be administered antenatally to the pregnant woman when foetal hypoxia is suspected. Foetal distress is being diagnosed by the clinician as an abnormal or non-reassuring foetal heart rate trace, preferably accompanied by either significant ST-wave abnormalities (as detected by the STAN-monitor) or an abnormal foetal blood scalp sampling (pH < 7.20).Primary outcome measures are the amount of S100B (a marker for brain tissue damage) and the severity of oxidative stress (measured by isoprostane, neuroprostane, non protein bound iron and hypoxanthine), both measured in umbilical cord blood. Secondary outcome measures are neonatal mortality, serious composite neonatal morbidity and long-term neurological outcome. Furthermore pharmacokinetics and pharmacodynamics will be investigated.We expect an inclusion of 220 patients (110 per group) to be feasible in an inclusion period of two years. Given a suspected mean value of S100B of 1.05 ug/L (SD 0.37 ug/L) in the placebo group this trial has a power of 90% (alpha 0.05) to detect a mean value of S100B of 0.89 ug/L (SD 0.37 ug/L) in the 'allopurinol-treated' group (z-test2-sided). Analysis will be by intention to treat and it allows for one interim analysis. DISCUSSION: In this trial we aim to answer the question whether antenatal allopurinol administration reduces hypoxic-ischaemic encephalopathy in neonates exposed to foetal hypoxia. TRIAL REGISTRATION NUMBER: Clinical Trials, protocol registration system: NCT00189007.

Systemic and cerebral inflammatory response to umbilical cord occlusions with worsening acidosis in the ovine fetus.

OBJECTIVE: We hypothesized that repetitive umbilical cord occlusions (UCOs) with worsening acidosis will lead to a fetal inflammatory response. STUDY DESIGN: Chronically instrumented fetal sheep underwent a series of UCOs until fetal arterial pH decreased to <7.00. Maternal and fetal blood samples were taken for blood gases/pH and plasma interleukin (IL)-1B and IL-6 levels. Animals were euthanized at 24 hours of recovery with brain tissue processed for subsequent measurement of microglia and mast cell counts. RESULTS: Repetitive UCOs resulted in a severe degree of fetal acidemia. Fetal plasma IL-1B values were increased approximately 2-fold when measured at maximal fetal acidosis and again at 1-2 hours of recovery. Fetal microglia cells were increased approximately 2-fold in the white matter and hippocampus, while mast cells were increased approximately 2-fold in the choroid plexus and now evident in the thalamus when analyzed at 24 hours recovery. CONCLUSION: Repetitive UCOs leading to severe acidemia in the ovine fetus near term will result in an inflammatory response both systemically and locally within the brain.

Increased fetal plasma and amniotic fluid erythropoietin concentrations: markers of intrauterine hypoxia.

Tissue hypoxia is the major stimulus of erythropoietin (EPO) synthesis in fetuses and adults. Since EPO does not cross the placenta and is not stored, fetal plasma and amniotic fluid levels indicate EPO synthesis and elimination. Acutely, the rate and magnitude of the increase in plasma EPO levels correlate with the intensity of hypoxia. Amniotic fluid EPO levels correlate with cord plasma levels in normal and abnormal pregnancies, with fetal plasma EPO levels in humans averaging 2.6 times higher than the corresponding amniotic fluid EPO levels. Recent experimental and clinical studies demonstrate that EPO has neuroprotective effects related to its anti-apoptotic and vascular growth-promoting properties. Although under basal conditions the fetal kidneys are the main site of EPO production, during hypoxia recent experimental data indicate an important role of the placenta. Amniotic fluid EPO levels have been shown to increase exponentially during fetal hypoxia in preeclamptic, diabetic and Rh-immunized pregnancies, to correlate inversely with cord blood pH, pO(2) and base excess and to predict neonatal morbidities and NICU admission. As an indicator of chronic intrauterine hypoxia, fetal EPO measurements have increased our knowledge about the pathogenesis and importance of intrauterine growth restriction, macrosomia, diabetic pregnancy, prolonged pregnancy, meconium staining, fetal hemorrhage, fetal anemia, maternal smoking and alcohol consumption, abnormal fetal heart rate and abnormal Doppler flow patterns. While the clinical utility of fetal amniotic fluid and plasma EPO measurements in the management of high-risk pregnancies and their offspring is promising, adequately powered clinical trials are urgently needed.

Relationships of maternal serum levels of vascular endothelial growth factor and tensile strength properties of the cervix in a rat model of chronic hypoxia.

OBJECTIVE: It has been shown that hypoxia leads to alterations in maternal serum levels of vascular endothelial growth factor (VEGF). In this study, we sought to test the hypothesis that chronic hypoxia increases maternal serum levels of VEGF, which in turn cause measurable changes in the viscoelastic properties of the rat uterine cervix. STUDY DESIGN: Timed-pregnant adult Sprague-Dawley rats were exposed to hypoxia beginning on day 17 of gestation (term = day 22). The following groups of animals were studied: (1) nonpregnant controls (NP, n = 6); (2) normoxia 21% fraction of inspired oxygen (FiO2) (NMX, n = 6); and (3) severe hypoxia 10% FiO2 (HPX; n = 5). A hypoxic chamber was used to assure consistent hypoxic environment. Animals were killed on day 21 of gestation (before labor). Maternal blood was collected immediately following anesthesia and prior to euthanasia. Free serum levels of VEGF were measured by highly specific immunoassays. Tensile strength properties of the cervix were assessed using a stretching regimen designed to mimic labor. Physical parameters measured were: indicators of viscoelasticity (slope; measure of stiffness), plasticity (yield point [YP]; moment the tissue changes its properties from elastic to plastic), strength (break point [BP]; moment of tissue disruption), and displacement at YP (marks the duration of the viscoelastic phase of the stretching) and BP (a measure of the strength of the material). Data were normalized to the dry weight of the cervix. RESULTS: Hypoxia is associated with increased serum levels of VEGF compared to NP or NMX groups (P = .001). Cervical stiffness was lower in NMX, compared with NP animals (P = .004), and was not significantly influenced by hypoxia (P > .05). Overall there was a significant inverse correlation between slope and maternal serum levels of VEGF (r = -0.85, P < .001). The force required to reach YP was significantly higher for the NP, compared with NMX and HPX groups (P = .004). Hypoxia did not alter the force required to reach the YP (NMX vs HPX, P > .05). Conversely, hypoxia significantly decreased the displacement at YP, indicating a shortening of the elastic phase (NMX vs HPX, P = .021). There was a significant inverse correlation between maternal serum levels of VEGF and the displacement at YP (r = -0.68, P = .002). In vivo, hypoxia decreased the force required to reached the BP (NMX vs HPX, P = .025), but there was no correlation between the levels of maternal serum VEGF and this indicator (r = -0.35, P = .170). CONCLUSION: Chronic hypoxia induces measurable changes in maternal serum levels of VEGF and tensile properties of the rat cervix, specifically a shortening of the elastic phase. Hypoxia decreases the cervical strength to stretch and predisposes to rupture, but this effect seems to be unrelated to maternal serum levels of VEGF.

Chronic hypoxia differentially increases glutathione content and gamma-glutamyl cysteine synthetase expression in fetal guinea pig organs.

OBJECTIVE: Glutathione is a natural antioxidant in the fetus and adult. We sought to determine whether maternal hypoxia alters glutathione levels in fetal organs as an adaptive response to the reduced oxygenation. STUDY DESIGN: Timed pregnant guinea pigs were housed in either a Plexiglas chamber containing 10.5% O(2) from 46 to 60 days gestation (HPX, n=6) or in room air, as the normoxic control (NMX, n=5). Pregnant guinea pigs were anesthetized at near term ( approximately 60 days, term=65 days) and liver, lungand kidney were excised from anesthetized fetuses and stored frozen (-80 degrees C) prior to sample processing. Using the hypoxia marker, pimonidazole, we measured a hypoxia-induced increase in stained cells of fetal liver compared to no change in either the lung or kidney. To measure the effect of hypoxia among different organs, total glutathione (GSH) content and protein levels of gamma-glutamyl cysteine synthetase (gamma-GCS) were measured from the same organs. RESULTS: Maternal hypoxia increased (P<0.05) total glutathione levels by 121% in the fetal liver but had no effect in either fetal lung or kidney. Chronic hypoxia increased (P<0.05) gamma-GCS protein levels in all three fetal organs studied. CONCLUSION: These results demonstrate that the fetal response to maternal hypoxia may be organ specific. The increase in fetal liver glutathione via upregulation of gamma-GCS may be an important adaptive response to prolonged hypoxic stress.

Blood gas analysis of bovine fetal capillary blood during stage II labor.

The aim of this study was to determine whether blood gas variables in fetal capillary blood during the last 30 min of stage II labor can be used to diagnose fetal asphyxia. Twenty-five newborn calves were used to investigate the correlation between capillary blood gas values obtained from the dorsolateral aspect of the distal pastern and those in arterial and venous blood. The pH, partial pressure of oxygen, partial pressure of carbon dioxide, concentration of bicarbonate, base excess and oxygen saturation were determined. The bicarbonate concentration (arterial, r=0.759; venous, r=0.766; both P<0.0001) and base excess (arterial, r=0.730; venous, r=0.807; both P<0.0001) had the highest correlations. Fetal capillary blood was collected during the last 30 min of stage II labor and the results of blood gas analysis were compared with those of arterial and venous blood collected immediately after birth in 38 calves. The pH (arterial, r=0.806; venous, r=0.885; both P<0.0001) and base excess (arterial, r=0.822; venous, r=0.871; both P<0.0001) had the highest correlations. The pH and base excess were significantly lower after birth than during the last 30 min of stage II labor. The severity of fetal acidosis during stage II labor can be easily and reliably determined using the pH or base excess of fetal capillary blood.