Joepe J. Kaandorp

Long-term neuroprotective effects of allopurinol after moderate perinatal asphyxia: follow-up of two randomised controlled trials

Objective Free-radical-induced reperfusion injury has been recognised as an important cause of brain tissue damage after birth asphyxia. Allopurinol reduces the formation of free radicals, thereby potentially limiting the amount of hypoxia–reperfusion damage. In this study the long-term outcome of neonatal allopurinol treatment after birth asphyxia was examined. Design Follow-up of 4 to 8 years of two earlier performed randomised controlled trials. Setting Leiden University Medical Center, University Medical Center Groningen and University Medical Center Utrecht, The Netherlands. Patients Fifty-four term infants were included when suffering from moderate-to-severe birth asphyxia in two previously performed trials. Intervention Infants either received 40 mg/kg allopurinol (with an interval of 12 h) starting within 4 h after birth or served as controls. Main outcome measures Children, who survived, were assessed with the Wechsler Preschool and Primary Scales of Intelligence test or Wechsler Intelligence Scale for Children and underwent a neurological examination. The effect of allopurinol on severe adverse outcome (defined as mortality or severe disability at the age of 4–8 years) was examined in the total group of asphyxiated infants and in a predefined subgroup of moderately asphyxiated infants (based on the amplitude integrated electroencephalogram). Results The mean age during follow-up (n=23) was 5 years and 5 months (SD 1 year and 2 months). There were no differences in long-term outcome between the allopurinol-treated infants and controls. However, subgroup analysis of the moderately asphyxiated group showed significantly less severe adverse outcome in the allopurinol-treated infants compared with controls (25% vs 65%; RR 0.40, 95%CI 0.17 to 0.94). Conclusions The reported data may suggest a (neuro)protective effect of neonatal allopurinol treatment in moderately asphyxiated infants.

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

Periods of impaired oxygenation or acute hypoxia in the fetus can be common during labour and how the fetus withstands these challenges is of interest. During hypoxia, the fetus shunts blood flow away from peripheral and towards essential vascular beds: the so called brain‐sparing effect. Part of the peripheral vasoconstriction is driven by reactive oxygen species (ROS) that inactivate nitric oxide (NO), thereby limiting its vasodilator action. Here, we investigate the source of ROS generation contributing to fetal peripheral vasoconstriction during hypoxia, and show that xanthine oxidase (XO) is fundamentally involved. Fetal exposure to the XO inhibitor allopurinol markedly diminished the peripheral vasoconstriction during hypoxia via NO‐dependent mechanisms. The data increase our understanding of the physiological control of fetal cardiovascular function during stress. The findings are also of significant clinical relevance as allopurinol is being administered to pregnant women in clinical obstetric trials.

Maternal allopurinol administration during suspected fetal hypoxia: a novel neuroprotective intervention? A multicentre randomised placebo controlled trial

Objective To determine whether maternal allopurinol treatment during suspected fetal hypoxia would reduce the release of biomarkers associated with neonatal brain damage. Design A randomised double-blind placebo controlled multicentre trial. Patients We studied women in labour at term with clinical indices of fetal hypoxia, prompting immediate delivery. Setting Delivery rooms of 11 Dutch hospitals. Intervention When immediate delivery was foreseen based on suspected fetal hypoxia, women were allocated to receive allopurinol 500 mg intravenous (ALLO) or placebo intravenous (CONT). Main outcome measures Primary endpoint was the difference in cord S100ß, a tissue-specific biomarker for brain damage. Results 222 women were randomised to receive allopurinol (ALLO, n=111) or placebo (CONT, n=111). Cord S100ß was not significantly different between the two groups: 44.5 pg/mL (IQR 20.2–71.4) in the ALLO group versus 54.9 pg/mL (IQR 26.8–94.7) in the CONT group (difference in median −7.69 (95% CI −24.9 to 9.52)). Post hoc subgroup analysis showed a potential treatment effect of allopurinol on the proportion of infants with a cord S100ß value above the 75th percentile in girls (ALLO n=5 (12%) vs CONT n=10 (31%); risk ratio (RR) 0.37 (95% CI 0.14 to 0.99)) but not in boys (ALLO n=18 (32%) vs CONT n=15 (25%); RR 1.4 (95% CI 0.84 to 2.3)). Also, cord neuroketal levels were significantly lower in girls treated with allopurinol as compared with placebo treated girls: 18.0 pg/mL (95% CI 12.1 to 26.9) in the ALLO group versus 32.2 pg/mL (95% CI 22.7 to 45.7) in the CONT group (geometric mean difference −16.4 (95% CI −24.6 to −1.64)). Conclusions Maternal treatment with allopurinol during fetal hypoxia did not significantly lower neuronal damage markers in cord blood. Post hoc analysis revealed a potential beneficial treatment effect in girls. Trial registration number NCT00189007, Dutch Trial Register NTR1383.

Divergence of mechanistic pathways mediating cardiovascular aging and developmental programming of cardiovascular disease

Aging and developmental programming are both associated with oxidative stress and endothelial dysfunction, suggesting common mechanistic origins. However, their interrelationship has been little explored. In a rodent model of programmed cardiovascular dysfunction we determined endothelial function and vascular telomere length in young (4 mo) and aged (15 mo) adult offspring of normoxic or hypoxic pregnancy with or without maternal antioxidant treatment. We show loss of endothelial function [maximal arterial relaxation to acetylcholine (71 ± 3 vs. 55 ± 3%) and increased vascular short telomere abundance (4.2–1.3 kb) 43.0 ± 1.5 vs. 55.1 ± 3.8%) in aged vs. young offspring of normoxic pregnancy (P < 0.05). Hypoxic pregnancy in young offspring accelerated endothelial dysfunction (maximal arterial relaxation to acetylcholine: 42 ± 1%, P < 0.05) but this was dissociated from increased vascular short telomere length abundance. Maternal allopurinol rescued maximal arterial relaxation to acetylcholine in aged offspring of normoxic or hypoxic pregnancy but not in young offspring of hypoxic pregnancy. Aged offspring of hypoxic allopurinol pregnancy compared with aged offspring of untreated hypoxic pregnancy had lower levels of short telomeres (vascular short telomere length abundance 35.1 ± 2.5 vs. 48.2 ± 2.6%) and of plasma proinflammatory chemokine (24.6 ± 2.8 vs. 36.8 ± 5.5 pg/ml, P < 0.05). These data provide evidence for divergence of mechanistic pathways mediating cardiovascular aging and developmental programming of cardiovascular disease, and aging being decelerated by antioxidants even prior to birth.—Allison, B. J., Kaandorp, J. J., Kane, A. D., Camm, E. J., Lusby, C., Cross, C. M., Nevin‐Dolan, R., Thakor, A. S., Derks, J. B., Tarry‐Adkins, J. L., Ozanne, S. E., Giussani, D. A Divergence of mechanistic pathways mediating cardiovascular aging and developmental programming of cardiovascular disease. FASEB J. 30, 1968–1975 (2016). www.fasebj.org

Antenatal Allopurinol Reduces Hippocampal Brain Damage After Acute Birth Asphyxia in Late Gestation Fetal Sheep

Free radical–induced reperfusion injury is a recognized cause of brain damage in the newborn after birth asphyxia. The xanthine oxidase inhibitor allopurinol reduces free radical synthesis and crosses the placenta easily. Therefore, allopurinol is a promising therapeutic candidate. This study tested the hypothesis that maternal treatment with allopurinol during fetal asphyxia limits ischemia–reperfusion (I/R) damage to the fetal brain in ovine pregnancy. The I/R challenge was induced by 5 repeated measured compressions of the umbilical cord, each lasting 10 minutes, in chronically instrumented fetal sheep at 0.8 of gestation. Relative to control fetal brains, the I/R challenge induced significant neuronal damage in the fetal hippocampal cornu ammonis zones 3 and 4. Maternal treatment with allopurinol during the I/R challenge restored the fetal neuronal damage toward control scores. Maternal treatment with allopurinol offers potential neuroprotection to the fetal brain in the clinical management of perinatal asphyxia.

Rapid target allopurinol concentrations in the hypoxic fetus after maternal administration during labour

Objective Perinatal hypoxia-induced free radical formation is an important cause of hypoxic-ischaemic encephalopathy and subsequent neurodevelopmental disabilities. Allopurinol reduces the formation of free radicals, which potentially limits hypoxia-induced brain damage. We investigated placental transfer and safety of allopurinol after maternal allopurinol treatment during labour to evaluate its potential role as a neuroprotective agent in suspected fetal hypoxia. Design We used data from a randomised, double-blind multicentre trial comparing maternal allopurinol versus placebo in case of imminent fetal hypoxia (NCT00189007). Patients We studied 58 women in labour at term, with suspected fetal hypoxia prompting immediate delivery, in the intervention arm of the study. Setting Delivery rooms of 11 Dutch hospitals. Intervention 500 mg allopurinol, intravenously to the mother, immediately prior to delivery. Main outcome measures Drug disposition (maternal plasma concentrations, cord blood concentrations) and drug safety (maternal and fetal adverse events). Results Within 5 min after the end of maternal allopurinol infusion, target plasma concentrations of allopurinol of ≥2 mg/L were present in cord blood. Of all analysed cord blood samples, 95% (52/55) had a target allopurinol plasma concentration at the moment of delivery. No adverse events were observed in the neonates. Two mothers had a red and/or painful arm during infusion. Conclusions A dose of 500 mg intravenous allopurinol rapidly crosses the placenta and provides target concentrations in 95% of the fetuses at the moment of delivery, which makes it potentially useful as a neuroprotective agent in perinatology with very little side effects. Trial registration The study is registered in the Dutch Trial Register (NTR1383) and the Clinical Trials protocol registration system (NCT00189007).

Maternal-to-fetal allopurinol transfer and xanthine oxidase suppression in the late gestation pregnant rat

Fetal brain hypoxic injury remains a concern in high‐risk delivery. There is significant clinical interest in agents that may diminish neuronal damage during birth asphyxia, such as in allopurinol, an inhibitor of the prooxidant enzyme xanthine oxidase. Here, we established in a rodent model the capacity of allopurinol to be taken up by the mother, cross the placenta, rise to therapeutic levels, and suppress xanthine oxidase activity in the fetus. On day 20 of pregnancy, Wistar dams were given 30 or 100 mg kg−1 allopurinol orally. Maternal and fetal plasma allopurinol and oxypurinol concentrations were measured, and xanthine oxidase activity in the placenta and maternal and fetal tissues determined. There were significant strong positive correlations between maternal and fetal plasma allopurinol (r = 0.97, P < 0.05) and oxypurinol (r = 0.88, P < 0.05) levels. Under baseline conditions, maternal heart (2.18 ± 0.62 mU mg−1), maternal liver (0.29 ± 0.08 mU mg−1), placenta (1.36 ± 0.42 mU mg−1), fetal heart (1.64 ± 0.59 mU mg−1), and fetal liver (0.14 ± 0.08 mU mg−1) samples all showed significant xanthine oxidase activity. This activity was suppressed in all tissues 2 h after allopurinol administration and remained suppressed 24 h later (P < 0.05), despite allopurinol and oxypurinol levels returning toward baseline. The data establish a mammalian model of xanthine oxidase inhibition in the mother, placenta, and fetus, allowing investigation of the role of xanthine oxidase–derived reactive oxygen species in the maternal, placental, and fetal physiology during healthy and complicated pregnancy.

Behavioral and neurodevelopmental outcome of children after maternal allopurinol administration during suspected fetal hypoxia: 5-year follow up of the ALLO-trial

Objective To evaluate the long-term neurodevelopmental and behavioral outcome of antenatal allopurinol treatment during suspected fetal hypoxia. Study design We studied children born from women who participated in a randomized double-blind placebo controlled multicenter study (ALLO-trial). Labouring women in whom the fetus was suspected to have fetal hypoxia were randomly allocated to receive allopurinol or placebo. At 5 years of age, the children were assessed with 2 parent reported questionnaires, the Ages and Stages Questionnaire (ASQ) and the Child Behavior Checklist (CBCL). A child was marked abnormal for ASQ if it scored below 2 standard deviation under the normative mean of a reference population in at least one domain. For CBCL, a score above the cut-off value (95th percentile for narrowband scale, 85th percentile for broadband scale) in at least one scale was marked as abnormal. Results We obtained data from 138 out of the original 222 mildly asphyxiated children included in the ALLO-trial (response rate 62%, allopurinol n = 73, placebo n = 65). At 5 years of age, the number of children that scored abnormal on the ASQ were 11 (15.1%) in the allopurinol group versus 11 (9.2%) in the placebo group (relative risk (RR) 1.64, 95% confidence interval (CI): 0.64 to 4.17, p = 0.30). On CBCL 21 children (30.4%) scored abnormal in de allopurinol group versus 12 children (20.0%) in the placebo group (RR 1.52, 95% CI: 0.82 to 2.83, p = 0.18). Conclusion We found no proof that allopurinol administered to labouring women with suspected fetal hypoxia improved long-term developmental and behavioral outcome. These findings are limited due to the fact that the study was potentially underpowered. Trial registration NCT00189007 Dutch Trial Register NTR1383