SMC5 Plays Independent Roles in Congenital Heart Disease and Neurodevelopmental Disability.

Up to 50% of patients with severe congenital heart disease (CHD) develop life-altering neurodevelopmental disability (NDD). It has been presumed that NDD arises in CHD cases because of hypoxia before, during, or after cardiac surgery. Recent studies detected an enrichment in de novo mutations in CHD and NDD, as well as significant overlap between CHD and NDD candidate genes. However, there is limited evidence demonstrating that genes causing CHD can produce NDD independent of hypoxia. A patient with hypoplastic left heart syndrome and gross motor delay presented with a de novo mutation in SMC5. Modeling mutation of smc5 in Xenopus tropicalis embryos resulted in reduced heart size, decreased brain length, and disrupted pax6 patterning. To evaluate the cardiac development, we induced the conditional knockout (cKO) of Smc5 in mouse cardiomyocytes, which led to the depletion of mature cardiomyocytes and abnormal contractility. To test a role for Smc5 specifically in the brain, we induced cKO in the mouse central nervous system, which resulted in decreased brain volume, and diminished connectivity between areas related to motor function but did not affect vascular or brain ventricular volume. We propose that genetic factors, rather than hypoxia alone, can contribute when NDD and CHD cases occur concurrently.

De novo damaging variants associated with congenital heart diseases contribute to the connectome.

Congenital heart disease (CHD) survivors are at risk for neurodevelopmental disability (NDD), and recent studies identify genes associated with both disorders, suggesting that NDD in CHD survivors may be of genetic origin. Genes contributing to neurogenesis, dendritic development and synaptogenesis organize neural elements into networks known as the connectome. We hypothesized that NDD in CHD may be attributable to genes altering both neural connectivity and cardiac patterning. To assess the contribution of de novo variants (DNVs) in connectome genes, we annotated 229 published NDD genes for connectome status and analyzed data from 3,684 CHD subjects and 1,789 controls for connectome gene mutations. CHD cases had more protein truncating and deleterious missense DNVs among connectome genes compared to controls (OR = 5.08, 95%CI:2.81-9.20, Fisher's exact test P = 6.30E-11). When removing three known syndromic CHD genes, the findings remained significant (OR = 3.69, 95%CI:2.02-6.73, Fisher's exact test P = 1.06E-06). In CHD subjects, the top 12 NDD genes with damaging DNVs that met statistical significance after Bonferroni correction (PTPN11, CHD7, CHD4, KMT2A, NOTCH1, ADNP, SMAD2, KDM5B, NSD2, FOXP1, MED13L, DYRK1A; one-tailed binomial test P ≤ 4.08E-05) contributed to the connectome. These data suggest that NDD in CHD patients may be attributable to genes that alter both cardiac patterning and the connectome.

Blood Pressure in Young Adults Born at Very Low Birth Weight: Adults Born Preterm International Collaboration.

Adults born preterm at very low birth weight (VLBW; <1500 g) have higher blood pressure than those born at term. It is not known whether all VLBW adults are at risk or whether higher blood pressure could be attributed to some of the specific conditions underlying or accompanying preterm birth. To identify possible risk or protective factors, we combined individual-level data from 9 cohorts that measured blood pressure in young adults born at VLBW or with a more stringent birth weight criterion. In the absence of major heterogeneity, we performed linear regression analysis in our pooled sample of 1571 adults born at VLBW and 777 controls. Adults born at VLBW had 3.4 mm Hg (95% confidence interval, 2.2-4.6) higher systolic and 2.1 mm Hg (95% confidence interval, 1.3-3.0) higher diastolic pressure, with adjustment for age, sex, and cohort. The difference in systolic pressure was present in men (1.8 mm Hg; 95% confidence interval, 0.1-3.5) but was stronger in women (4.7 mm Hg; 95% confidence interval, 3.2-6.3). Among the VLBW group, blood pressure was unrelated to gestational age, maternal smoking, multiple pregnancy, retinopathy of prematurity, or bronchopulmonary dysplasia. Blood pressure was higher than that of controls among VLBW adults unexposed to maternal preeclampsia. Among those exposed, it was even higher, especially if born appropriate for gestational age. In conclusion, although female sex and maternal preeclampsia are additional risk factors, the risk of higher blood pressure is not limited to any etiologic subgroup of VLBW adults, arguing for vigilance in early detection of high blood pressure in all these individuals.

Genes and environment in neonatal intraventricular hemorrhage.

Emerging data suggest intraventricular hemorrhage (IVH) of the preterm neonate is a complex disorder with contributions from both the environment and the genome. Environmental analyses suggest factors mediating both cerebral blood flow and angiogenesis contribute to IVH, while candidate gene studies report variants in angiogenesis, inflammation, and vascular pathways. Gene-by-environment interactions demonstrate the interaction between the environment and the genome, and a non-replicated genome-wide association study suggests that both environmental and genetic factors contribute to the risk for severe IVH in very low-birth weight preterm neonates.

Adolescents born prematurely with isolated grade 2 haemorrhage in the early 1990s face increased risks of learning challenges.

AIM: To compare the impact of low-grade haemorrhage on neurocognitive function in 16-year-old adolescents born preterm, by grade of intraventricular haemorrhage, and term controls. METHODS: We evaluated 338 preterm adolescents (birth weight 600-1250 g) for intelligence, executive function and memory tasks. Eleven had grade 3-4 haemorrhage, 44 had grade 2, 31 had grade 1, and 251 had no haemorrhage. Group comparisons were made with 102 term age-matched controls, and regression models used to identify the risk that low-grade haemorrhage posed for cognitive, executive function and memory deficits. RESULTS: Preterm adolescents with grade 2 haemorrhage had higher deficit rates of verbal intelligence, receptive vocabulary, phonemic fluency, cognitive flexibility and phonological fluency than preterm adolescents with grade 1 or no haemorrhage, compared with term controls. After excluding preterm adolescents with both grade 2 haemorrhage and cystic periventricular leukomalacia, those with isolated grade 2 haemorrhage remained at greater risk of cognitive and executive function deficits than term controls and of cognitive deficits than preterm adolescents with no haemorrhage. CONCLUSION: Our findings suggest that preterm adolescents born in the early 1990s with isolated grade 2 haemorrhage are at increased risk of learning challenges, including cognitive and executive function deficits.

The role of neuroimaging in predicting neurodevelopmental outcomes of preterm neonates.

Magnetic resonance imaging (MRI) is a safe and high-resolution neuroimaging modality that is increasingly used in the neonatal population to assess brain injury and its consequences on brain development. It is superior to cranial ultrasound for the definition of patterns of both white and gray matter maturation and injury and therefore has the potential to provide prognostic information on the neurodevelopmental outcomes of the preterm population. Furthermore, the development of sophisticated MRI strategies, including diffusion tensor imaging, resting state functional connectivity, and magnetic resonance spectroscopy, may increase the prognostic value, helping to guide parental counseling and allocate early intervention services.

Candidate gene analysis: severe intraventricular hemorrhage in inborn preterm neonates.

Intraventricular hemorrhage (IVH) is a disorder of complex etiology. We analyzed genotypes for 7 genes from 224 inborn preterm neonates treated with antenatal steroids and grade 3-4 IVH and 389 matched controls. Only methylenetetrahydrofolate reductase was more prevalent in cases of IVH, emphasizing the need for more comprehensive genetic strategies.

Systemic inflammation, intraventricular hemorrhage, and white matter injury.

To see if the systemic inflammation profile of 123 infants born before the 28th week of gestation who had intraventricular hemorrhage without white matter injury differed from that of 68 peers who had both lesions, we compared both groups to 677 peers who had neither. Cranial ultrasound scans were read independently by multiple readers until concordance. The concentrations of 25 proteins were measured with multiplex arrays using an electrochemiluminescence system. Infants who had both hemorrhage and white matter injury were more likely than others to have elevated concentrations of C-reactive protein and interleukin 8 on days 1, 7, and 14, and elevated concentrations of serum amyloid A and tumor necrosis factor-α on 2 of these days. Intraventricular hemorrhage should probably be viewed as 2 entities: hemorrhage alone and hemorrhage with white matter injury. Each entity is associated with inflammation, but the combination has a stronger inflammatory signal than hemorrhage alone.

Environmental enrichment increases the GFAP+ stem cell pool and reverses hypoxia-induced cognitive deficits in juvenile mice.

Premature children born with very low birth weight (VLBW) can suffer chronic hypoxic injury as a consequence of abnormal lung development and cardiovascular abnormalities, often leading to grave neurological and behavioral consequences. Emerging evidence suggests that environmental enrichment improves outcome in animal models of adult brain injury and disease; however, little is known about the impact of environmental enrichment following developmental brain injury. Intriguingly, data on socio-demographic factors from longitudinal studies that examined a number of VLBW cohorts suggest that early environment has a substantial impact on neurological and behavioral outcomes. In the current study, we demonstrate that environmental enrichment significantly enhances behavioral and neurobiological recovery from perinatal hypoxic injury. Using a genetic fate-mapping model that allows us to trace the progeny of GFAP+ astroglial cells, we show that hypoxic injury increases the proportion of astroglial cells that attain a neuronal fate. In contrast, environmental enrichment increases the stem cell pool, both through increased stem cell proliferation and stem cell survival. In mice subjected to hypoxia and subsequent enrichment there is an additive effect of both conditions on hippocampal neurogenesis from astroglia, resulting in a robust increase in the number of neurons arising from GFAP+ cells by the time these mice reach full adulthood.

Trajectories of receptive language development from 3 to 12 years of age for very preterm children.

OBJECTIVES: The goal was to examine whether indomethacin use, gender, neonatal, and sociodemographic factors predict patterns of receptive language development from 3 to 12 years of age in preterm children. METHODS: A total of 355 children born in 1989-1992 with birth weights of 600 to 1250 g were evaluated at 3, 4.5, 6, 8, and 12 years with the Peabody Picture Vocabulary Test-Revised. Hierarchical growth modeling was used to explore differences in language trajectories. RESULTS: From 3 to 12 years, preterm children displayed catch-up gains on the Peabody Picture Vocabulary Test-Revised. Preterm children started with an average standardized score of 84.1 at 3 years and gained 1.2 points per year across the age period studied. Growth-curve analyses of Peabody Picture Vocabulary Test-Revised raw scores revealed an indomethacin-gender effect on initial scores at 3 years, with preterm boys assigned randomly to receive indomethacin scoring, on average, 4.2 points higher than placebo-treated boys. However, the velocity of receptive vocabulary development from 3 to 12 years did not differ for the treatment groups. Children with severe brain injury demonstrated slower gains in skills over time, compared with those who did not suffer severe brain injury. Significant differences in language trajectories were predicted by maternal education and minority status. CONCLUSION: Although indomethacin yielded an initial benefit for preterm boys, this intervention did not alter the developmental trajectory of receptive language scores. Severe brain injury leads to long-term sequelae in language development, whereas a socioeconomically advantaged environment supports better language development among preterm children.

Lasting effects of preterm birth and neonatal brain hemorrhage at 12 years of age.

OBJECTIVES: Our goals were to compare cognitive, language, behavioral, and educational outcomes of preterm children to term controls and to evaluate the impact of neonatal brain injury, indomethacin, and environmental risk factors on intellectual function at 12 years of age. METHODS: A total of 375 children born in 1989-1992 with birth weights of 600 to 1250 g enrolled in the Indomethacin Intraventricular Hemorrhage Prevention Trial and 111 controls were evaluated. Neuropsychometric testing, neurologic examination, and interviews on educational needs were completed. Severe brain injury was defined as the presence of grade 3 to 4 indomethacin intraventricular hemorrhage, periventricular leukomalacia, or severe ventriculomegaly on cranial ultrasound. RESULTS: On the Wechsler Scales of Intelligence for Children, the preterm cohort obtained a full-scale IQ of 87.9 +/- 18.3, verbal IQ of 90.8 +/- 18.9, and performance IQ of 86.8 +/- 17.9. Preterm children obtained scores 6 to 14 points lower than term controls on all psychometric tests after adjustment for sociodemographic factors. On the Clinical Evaluation of Language Fundamentals (test of basic language skills), 22% to 24% of preterm children scored in the abnormal ranges (<70) as opposed to 2% to 4% of controls. Preterm children with and without brain injury required more school services (76% and 44% vs 16%), and support in reading (44% and 28% vs 9%), writing (44% and 20% vs 4%), and mathematics (47% and 30% vs 6%) compared with controls. Preterm children also displayed more behavior problems than their term counterparts. Severe neonatal brain injury was the strongest predictor of poor intelligence. Antenatal steroids, higher maternal education, and 2-parent family were associated with better cognition, whereas minority status incurred a disadvantage. Indomethacin did not affect intellectual function among preterm children. CONCLUSIONS: Preterm children born in the early 1990s, especially those with severe brain injury, demonstrate serious deficits in their neuropsychological profile, which translates into increased use of school services at 12 years.

Brain volume reductions within multiple cognitive systems in male preterm children at age twelve.

OBJECTIVES: To more precisely examine regional and subregional microstructural brain changes associated with preterm birth. STUDY DESIGN: We obtained brain volumes from 29 preterm children, age 12 years, with no ultrasound scanning evidence of intraventricular hemorrhage or cystic periventricular leukomalacia in the newborn period, and 22 age- and sex-matched term control subjects. RESULTS: Preterm male subjects demonstrated significantly lower white matter volumes in bilateral cingulum, corpus callosum, corticospinal tract, prefrontal cortex, superior and inferior longitudinal fasciculi compared with term male subjects. Gray matter volumes in prefrontal cortex, basal ganglia, and temporal lobe also were significantly reduced in preterm male subjects. Brain volumes of preterm female subjects were not significantly different from those of term female control subjects. Voxel-based morphometry results were not correlated with perinatal variables or cognitive outcome. Higher maternal education was associated with higher cognitive performance in preterm male subjects. CONCLUSIONS: Preterm male children continue to demonstrate abnormal neurodevelopment at 12 years of age. However, brain morphology in preterm female children may no longer differ from that of term female children. The neurodevelopmental abnormalities we detected in preterm male subjects appear to be relatively diffuse, involving multiple neural systems. The relationship between aberrant neurodevelopment and perinatal variables may be mediated by genetic factors, environmental factors, or both reflected in maternal education level.

Sickle cell disease complicated by post-streptococcal glomerulonephritis, cerebral hemorrhage and reversible posterior leucoencephalopathy syndrome.

A patient with homozygous hemoglobin SS disease presented with an intracerebral hemorrhage complicating reversible posterior leucoencephalopathy syndrome (RPLS), secondary to hypertension associated with acute post-streptococcal glomerulonephritis (APSGN). Distinguishing potentially reversible causes of central nervous system events from primary cerebral infarction or hemorrhage in patients with sickle cell disease is important because the management and prognosis of these complications is very different. Similarly, because of the difference in prognosis between APSGN and other forms of sickle cell nephropathy, it is also important to differentiate these conditions.

Genetic susceptibility to retinopathy of prematurity.

OBJECTIVES: The goals were to isolate and to estimate the genetic susceptibility to retinopathy of prematurity. METHODS: A retrospective study (1994-2004) from 3 centers was performed with zygosity data for premature twins who were born at a gestational age of < or = 32 weeks and survived beyond a postmenstrual age of 36 weeks. Retinopathy of prematurity was diagnosed and staged by pediatric ophthalmologists at each center. Data analyses were performed with mixed-effects logistic regression analysis and latent variable probit modeling. RESULTS: A total of 63 monozygotic and 137 dizygotic twin pairs were identified and analyzed. Data on gestational age, birth weight, gender, respiratory distress syndrome, retinopathy of prematurity, bronchopulmonary dysplasia, duration of ventilation and supplemental oxygen use, and length of stay were comparable between monozygotic and dizygotic twins. In the mixed-effects logistic regression analysis for retinopathy of prematurity, gestational age and duration of supplemental oxygen use were significant covariates. After controlling for known and unknown nongenetic factors, genetic factors accounted for 70.1% of the variance in liability for retinopathy of prematurity. CONCLUSION: In addition to prematurity and environmental factors, there is a strong genetic predisposition to retinopathy of prematurity.

Familial and genetic susceptibility to major neonatal morbidities in preterm twins.

BACKGROUND: Intraventricular hemorrhage, necrotizing enterocolitis, and bronchopulmonary dysplasia remain significant causes of morbidity and mortality in preterm newborns. OBJECTIVES: Our goal was to assess the familial and genetic susceptibility to intraventricular hemorrhage, necrotizing enterocolitis, and bronchopulmonary dysplasia. METHODS: Mixed-effects logistic-regression and latent variable probit model analysis were used to assess the contribution of several covariates in a multicenter retrospective study of 450 twin pairs born at < or =32 weeks of gestation. To determine the genetic contribution, concordance rates in a subset of 252 monozygotic and dizygotic twin pairs were compared. RESULTS: The study population had a mean gestational age of 29 weeks and birth weight of 1286 g. After controlling for effects of covariates, the twin data showed that 41.3%, 51.9%, and 65.2%, respectively, of the variances in liability for intraventricular hemorrhage, necrotizing enterocolitis, and bronchopulmonary dysplasia could be accounted for by genetic and shared environmental factors. Among the 63 monozygotic twin pairs, the observed concordance for bronchopulmonary dysplasia was significantly higher than the expected concordance; 12 of 18 monozygotic twin pairs with > or =1 affected member had both members affected versus 3.69 expected. After controlling for covariates, genetic factors accounted for 53% of the variance in liability for bronchopulmonary dysplasia. CONCLUSIONS: Twin analyses show that intraventricular hemorrhage, necrotizing enterocolitis, and bronchopulmonary dysplasia are familial in origin. These data demonstrate, for the first time, the significant genetic susceptibility for bronchopulmonary dysplasia in preterm infants.

Cortical neurogenesis enhanced by chronic perinatal hypoxia.

Most regions of the mature mammalian brain, including the cerebral cortex, appear to be unable to support the genesis of new neurons. Here, we report that a low level of neurogenesis occurs in the cerebral cortex of the infant mouse brain and is enhanced by chronic perinatal hypoxia. When mice were reared in a low-oxygen environment from postnatal days 3 to 11, approximately 30% of the cortical neurons were lost after the insult; yet this damage was transient. The loss of cortical neuron number, cortical volume, and brain weight were all reversed during the recovery period. At P18, 7 days after the cessation of hypoxia, there was a marked increase in astroglial cell proliferation within the SVZ, as assessed by 5-bromodeoxyuridine (BrdU) incorporation in S-phase cells. One month after BrdU incorporation, 40% more BrdU-positive cells were found in the cerebral cortex of hypoxic-reared as compared to normoxic control mice. Among these newly generated cortical cells, approximately 45% were oligodendrocytes, 35% were astrocytes, and 10% were neurons in both hypoxic and normoxic mice. However, twice as many BrdU-labeled cells expressed neuronal markers in the neocortex in mice recovering from hypoxia as compared to controls. In both hypoxic-reared and normoxic infant/juvenile mice, putative neuroblasts could be seen detaching from the forebrain subventricular zone, migrating through the subcortical white matter and entering the lower cortical layers, 5 to 11 days after their last mitotic division. We suggest that cortical neurogenesis may play a significant role in repairing neuronal losses after neonatal injury.

An evidence-based approach to predicting low IQ in very preterm infants from the neurological examination: outcome data from the indomethacin Indomethacin Intraventricular Hemorrhage Prevention Trial.

We evaluated whether the degree of cerebral palsy (CP) at age 3 in very preterm children is predictive of full-scale intelligence quotient (FSIQ) <70 at age 8 by calculating likelihood ratios (LRs) for findings on the neurologic examination. Data from the follow-up phase of the Indomethacin Intraventricular Hemorrhage Prevention Trial, which includes periodic neurologic examination and neuropsychometric testing, were used. Information was available on 366 of 440 (83%) children with birth weight of 600 to 1250 g who survived. Neurologic examination at age 3 was grouped by presence and type of CP, and the Weschler Intelligence Scale for Children-Third Edition FSIQ at age 8 was grouped dichotomously (<70 or > or =70). CP was identified in 35 of 366 3-year-olds (9.5%). An FSIQ <70 was identified in 47 of 366 children at 8 years old (12.8%). FSIQ <70 occurred in 14 of 17 children with tri- or quadriplegia (82%), 8 of 18 children with di- or hemiplegia (44%), and 25 of 331 children without CP (7.5%). Useful LRs were calculated for tri- or quadriplegia (30), di- or hemiplegia (5.7), and children without CP (0.55). These LRs have greater impact on posttest odds for FSIQ <70 than those for birth weight <1000 g, history of bronchopulmonary dysplasia, and Stanford-Binet Intelligence Score <70 at age 3. We conclude that the neurologic examination at 3 years old predicts FSIQ <70 at age 8 with LRs that allow evidence-based parental counseling and intervention planning.

School-age outcomes of very low birth weight infants in the indomethacin intraventricular hemorrhage prevention trial.

OBJECTIVE: The cohort consisted of 328 very low birth weight infants (600-1250 g birth weight) who were enrolled in the low-dose prophylactic indomethacin prevention trial and were intraventricular hemorrhage (IVH) negative at 6 postnatal hours. The objective was to determine the effects of both IVH and indomethacin on cognitive, language, and achievement performance at 8 years of age. METHODS: The cohort was divided into 4 subgroups for analysis: indomethacin plus IVH, indomethacin no IVH, saline plus IVH, and saline with no IVH. The children were evaluated prospectively at 8 years of age with a neurologic assessment, history of school performance, and a battery of cognitive, academic, behavioral, and functional assessments. RESULTS: Children in both IVH groups had more cerebral palsy; more hearing impairment; lower daily living skills scores; lower IQ, vocabulary, and reading and mathematics achievement test scores; and greater educational resource needs. With logistic regression analyses grade 3 to 4 IVH, periventricular leukomalacia and/or ventriculomegaly, male gender, maternal education, and language spoken in the home contributed to outcomes. No effects of indomethacin or gestational age were identified. CONCLUSIONS: Although biological factors including IVH, ventriculomegaly, and periventricular leukomalacia contribute significantly to school age outcomes among very low birth weight survivors at 8 years of age, social and environmental factors including maternal level of education and primary language spoken in the home are also important contributors to outcome.

Disrupted synaptic development in the hypoxic newborn brain.

Infants born prematurely risk significant life-long cognitive disability, representing a major pediatric health crisis. The neuropathology of this cohort is accurately modeled in mice subjected to sublethal postnatal hypoxia. Massively parallel transcriptome analysis using cDNA microchips (9,262 genes), combined with immunohistochemical and protein assays, reveals that sublethal hypoxia accentuates genes subserving presynaptic function, and it suppresses genes involved with synaptic maturation, postsynaptic function, and neurotransmission. Other significantly affected pathways include those involved with glial maturation, vasculogenesis, and components of the cortical and microtubular cytoskeleton. These patterns reveal a global dysynchrony in the maturation programs of the hypoxic developing brain, and offer insights into the vulnerabilities of processes that guide early postnatal cerebral maturation.

Paracrine and autocrine functions of neuronal vascular endothelial growth factor (VEGF) in the central nervous system.

Recent data have demonstrated that vascular endothelial growth factor (VEGF) is expressed by subsets of neurons, coincident with angiogenesis within the developing cerebral cortex. Here we investigate the characteristics of VEGF expression by neurons and test the hypothesis that VEGF may serve both paracrine and autocrine functions in the developing central nervous system. To begin to address these questions, we assayed expression of VEGF and one of its potential receptors, Flk-1 (VEGFR-2), in the embryonic mouse forebrain and embryonic cortical neurons grown in vitro. Both VEGF and Flk-1 are present in subsets of post-mitotic neurons in vivo and in vitro. Moreover, VEGF levels are up-regulated in neuronal cultures subjected to hypoxia, consistent with our previous results in vivo. While the abundance of Flk-1 is unaffected by hypoxia, the receptor exhibits a higher level of tyrosine phosphorylation, as do downstream signaling kinases, including extracellular signal-regulated protein kinase, p90RSK and STAT3a, demonstrating activation of the VEGF pathway. These same signaling components also exhibited higher tyrosine phosphorylation levels in response to exogenous addition of rVEGFA(165). This activation was diminished in the presence of specific inhibitors of Flk-1 function and agents that sequester VEGF, resulting in a dose-dependent increase in apoptosis in these neuronal cultures. Further, inhibition of MEK resulted in increased apoptosis, while inhibition of phosphatidylinositol 3-kinase had no appreciable affect. In addition to the novel function for VEGF that we describe in neuronal survival, neuronal VEGF also affected the organization and differentiation of brain endothelial cells in a three-dimensional culture paradigm, consistent with its more traditional role as a vascular agent. Thus, our in vitro data support a role for neuronal VEGF in both paracrine and autocrine signaling in the maintenance of neurons and endothelia in the central nervous system.