Circadian patterns of heart rate variability in fetal sheep after hypoxia-ischaemia: A biomarker of evolving brain injury.

Hypoxia-ischaemia (HI) before birth is a key risk factor for stillbirth and severe neurodevelopmental disability in survivors, including cerebral palsy, although there are no reliable biomarkers to detect at risk fetuses that may have suffered a transient period of severe HI. We investigated time and frequency domain measures of fetal heart rate variability (FHRV) for 3 weeks after HI in preterm fetal sheep at 0.7 gestation (equivalent to preterm humans) until 0.8 gestation (equivalent to term humans). We have previously shown that this is associated with delayed development of severe white and grey matter injury, including cystic white matter injury (WMI) resembling that observed in human preterm infants. HI was associated with suppression of time and frequency domain measures of FHRV and reduced their circadian rhythmicity during the first 3 days of recovery. By contrast, circadian rhythms of multiple measures of FHRV were exaggerated over the final 2 weeks of recovery, mediated by a greater reduction in FHRV during the morning nadir, but no change in the evening peak. These data suggest that the time of day at which FHRV measurements are taken affects their diagnostic utility. We further propose that circadian changes in FHRV may be a low-cost, easily applied biomarker of antenatal HI and evolving brain injury. KEY POINTS: Hypoxia-ischaemia (HI) before birth is a key risk factor for stillbirth and probably for disability in survivors, although there are no reliable biomarkers for antenatal brain injury. In preterm fetal sheep, acute HI that is known to lead to delayed development of severe white and grey matter injury over 3 weeks, was associated with early suppression of multiple time and frequency domain measures of fetal heart rate variability (FHRV) and loss of their circadian rhythms during the first 3 days after HI. Over the final 2 weeks of recovery after HI, exaggerated circadian rhythms of frequency domain FHRV measures were observed. The morning nadirs were lower with no change in the evening peak of FHRV. Circadian changes in FHRV may be a low-cost, easily applied biomarker of antenatal HI and evolving brain injury.

Effects of antenatal dexamethasone and hyperglycemia on cardiovascular adaptation to asphyxia in preterm fetal sheep.

Antenatal glucocorticoids improve outcomes among premature infants but are associated with hyperglycemia, which can exacerbate hypoxic-ischemic injury. It is still unclear how antenatal glucocorticoids or hyperglycemia modulate fetal cardiovascular adaptations to severe asphyxia. In this study, preterm fetal sheep received either saline or 12 mg im maternal dexamethasone, followed 4 h later by complete umbilical cord occlusion (UCO) for 25 min. An additional cohort of fetuses received titrated glucose infusions followed 4 h later by UCO to control for the possibility that hyperglycemia contributed to the cardiovascular effects of dexamethasone. Fetuses were studied for 7 days after UCO. Maternal dexamethasone was associated with fetal hyperglycemia (P < 0.001), increased arterial pressure (P < 0.001), and reduced femoral (P < 0.005) and carotid (P < 0.05) vascular conductance before UCO. UCO was associated with bradycardia, femoral vasoconstriction, and transient hypertension. For the first 5 min of UCO, fetal blood pressure in the dexamethasone-asphyxia group was greater than saline-asphyxia (P < 0.001). However, the relative increase in arterial pressure was not different from saline-asphyxia. Fetal heart rate and femoral vascular conductance fell to similar nadirs in both saline and dexamethasone-asphyxia groups. Dexamethasone did not affect the progressive decline in femoral vascular tone or arterial pressure during continuing UCO. By contrast, there were no effects of glucose infusions on the response to UCO. In summary, maternal dexamethasone but not fetal hyperglycemia increased fetal arterial pressure before and for the first 5 min of prolonged UCO but did not augment the cardiovascular adaptations to acute asphyxia.

Magnetic Resonance Imaging Correlates of White Matter Gliosis and Injury in Preterm Fetal Sheep Exposed to Progressive Systemic Inflammation.

Progressive fetal infection/inflammation is strongly associated with neural injury after preterm birth. We aimed to test the hypotheses that progressively developing fetal inflammation leads to neuroinflammation and impaired white matter development and that the histopathological changes can be detected using high-field diffusion tensor magnetic resonance imaging (MRI). Chronically instrumented preterm fetal sheep at 0.7 of gestation were randomly assigned to receive intravenous saline (control; n = 6) or a progressive infusion of lipopolysaccharide (LPS, 200 ng intravenous over 24 h then doubled every 24 h for 5 days to induce fetal inflammation, n = 7). Sheep were killed 10 days after starting the infusions, for histology and high-field diffusion tensor MRI. Progressive LPS infusion was associated with increased circulating interleukin (IL)-6 concentrations and moderate increases in carotid artery perfusion and the frequency of electroencephalogram (EEG) activity (p < 0.05 vs. control). In the periventricular white matter, fractional anisotropy (FA) was increased, and orientation dispersion index (ODI) was reduced (p < 0.05 vs. control for both). Histologically, in the same brain region, LPS infusion increased microglial activation and astrocyte numbers and reduced the total number of oligodendrocytes with no change in myelination or numbers of immature/mature oligodendrocytes. Numbers of astrocytes in the periventricular white matter were correlated with increased FA and reduced ODI signal intensities. Astrocyte coherence was associated with increased FA. Moderate astrogliosis, but not loss of total oligodendrocytes, after progressive fetal inflammation can be detected with high-field diffusion tensor MRI.

Dysmaturation of sleep state and electroencephalographic activity after hypoxia-ischaemia in preterm fetal sheep.

Antenatal hypoxia-ischaemia (HI) in preterm fetal sheep can trigger delayed evolution of severe, cystic white matter injury (WMI), in a similar timecourse to WMI in preterm infants. We therefore examined how severe hypoxia-ischaemia affects recovery of electroencephalographic (EEG) activity. Chronically instrumented preterm fetal sheep (0.7 gestation) received 25 min of complete umbilical cord occlusion (UCO, n = 9) or sham occlusion (controls, n = 9), and recovered for 21 days. HI was associated with a shift to lower frequency EEG activity for the first 5 days with persisting loss of EEG power in the delta and theta bands, and initial loss of power in the alpha and beta bands in the first 14 days of recovery. In the final 3 days of recovery, there was a marked rhythmic shift towards higher frequency EEG activity after UCO. The UCO group spent less time in high-voltage sleep, and in the early evening (7:02 pm ± 47 min) abruptly stopped cycling between sleep states, with a shift to a high frequency state for 2 h 48 min ± 40 min, with tonic electromyographic activity. These findings demonstrate persisting EEG and sleep state dysmaturation after severe hypoxia-ischaemia. Loss of fetal or neonatal sleep state cycling in the early evening may be a useful biomarker for evolving cystic WMI.

Fetal growth restriction and stillbirth: Biomarkers for identifying at risk fetuses.

Fetal growth restriction (FGR) is a major cause of stillbirth, prematurity and impaired neurodevelopment. Its etiology is multifactorial, but many cases are related to impaired placental development and dysfunction, with reduced nutrient and oxygen supply. The fetus has a remarkable ability to respond to hypoxic challenges and mounts protective adaptations to match growth to reduced nutrient availability. However, with progressive placental dysfunction, chronic hypoxia may progress to a level where fetus can no longer adapt, or there may be superimposed acute hypoxic events. Improving detection and effective monitoring of progression is critical for the management of complicated pregnancies to balance the risk of worsening fetal oxygen deprivation in utero, against the consequences of iatrogenic preterm birth. Current surveillance modalities include frequent fetal Doppler ultrasound, and fetal heart rate monitoring. However, nearly half of FGR cases are not detected in utero, and conventional surveillance does not prevent a high proportion of stillbirths. We review diagnostic challenges and limitations in current screening and monitoring practices and discuss potential ways to better identify FGR, and, critically, to identify the "tipping point" when a chronically hypoxic fetus is at risk of progressive acidosis and stillbirth.

Fetal heart rate variability is a biomarker of rapid but not progressive exacerbation of inflammation in preterm fetal sheep.

Perinatal infection/inflammation can trigger preterm birth and contribute to neurodevelopmental disability. There are currently no sensitive, specific methods to identify perinatal infection. We investigated the utility of time, frequency and non-linear measures of fetal heart rate (FHR) variability (FHRV) to identify either progressive or more rapid inflammation. Chronically instrumented preterm fetal sheep were randomly assigned to one of three different 5d continuous i.v. infusions: 1) control (saline infusions; n = 10), 2) progressive lipopolysaccharide (LPS; 200 ng/kg over 24 h, doubled every 24 h for 5d, n = 8), or 3) acute-on-chronic LPS (100 ng/kg over 24 h then 250 ng/kg/24 h for 4d plus 1 µg boluses at 48, 72, and 96 h, n = 9). Both LPS protocols triggered transient increases in multiple measures of FHRV at the onset of infusions. No FHRV or physiological changes occurred from 12 h after starting progressive LPS infusions. LPS boluses during the acute-on-chronic protocol triggered transient hypotension, tachycardia and an initial increase in multiple time and frequency domain measures of FHRV, with an asymmetric FHR pattern of predominant decelerations. Following resolution of hypotension after the second and third LPS boluses, all frequencies of FHRV became suppressed. These data suggest that FHRV may be a useful biomarker of rapid but not progressive preterm infection/inflammation.

Cerebral Oxygenation and Metabolism After Hypoxia-Ischemia.

Perinatal hypoxia-ischemia (HI) is still a significant contributor to mortality and adverse neurodevelopmental outcomes in term and preterm infants. HI brain injury evolves over hours to days, and involves complex interactions between the endogenous protective and pathological processes. Understanding the timing of evolution of injury is vital to guide treatment. Post-HI recovery is associated with a typical neurophysiological profile, with stereotypic changes in cerebral perfusion and oxygenation. After the initial recovery, there is a delayed, prolonged reduction in cerebral perfusion related to metabolic suppression, followed by secondary deterioration with hyperperfusion and increased cerebral oxygenation, associated with altered neurovascular coupling and impaired cerebral autoregulation. These changes in cerebral perfusion are associated with the stages of evolution of injury and injury severity. Further, iatrogenic factors can also affect cerebral oxygenation during the early period of deranged metabolism, and improving clinical management may improve neuroprotection. We will review recent evidence that changes in cerebral oxygenation and metabolism after HI may be useful biomarkers of prognosis.