Sex differences in neonatal outcomes following prenatal opioid exposure.

The impact of the opioid epidemic on pregnant people and children is a growing public health crisis. Understanding how opioids affect the developing brain during pregnancy and postnatally remains a critical area of investigation. Biological sex plays a crucial role in all physiologic processes, with the potential for a significant impact on neonatal outcomes, including those infants with opioid exposure. Here, we aim to explore current literature on the effect of sex on neonatal outcomes following prenatal opioid exposure. Sex differences in adults with opioid use disorder have been well studied, including increased mortality among males and higher rates of psychiatric comorbidities and likelihood of relapse in females. However, such differences are not yet well understood in neonates. Emerging clinical data suggest sex-specific effects in infants with prenatal opioid exposure on the expression of genes related to feeding regulation and reward signaling pathways. Increased susceptibility to white matter injury has also been noted in female infants following prenatal opioid exposure. Understanding the impact of sex as a biological variable on neonatal outcomes following prenatal opioid exposure is paramount to improving the health and well-being of infants, children, and adults impacted by the opioid epidemic.

Prenatal Opioid Exposure and Immune-Related Conditions in Children.

Importance: Prenatal opioid exposure (POE) may alter with fetal development of the immune system, which may influence long-term health and susceptibility to immune-related conditions. Objective: To compare the risk of hospitalization and emergency department presentation for immune-related conditions in children with and without POE. Design, Setting, and Participants: This retrospective, population-based cohort study used linked administrative health records of all children born in Western Australia between January 1, 2003, and December 31, 2018 (N = 401 462). Exposure: Prenatal exposure to prescription opioids (overall and by trimester), neonatal abstinence syndrome diagnosis, and opioid indication (pain or opioid use disorder [OUD]). Main Outcomes and Measures: The main outcome was hospital admissions and emergency department presentations during which a child was diagnosed with an immune-related condition, including infections, conditions associated with an overactive immune system (eg, asthma, eczema, and allergy and anaphylaxis), and autoimmune diseases diagnosed before age 5 years or June 30, 2020. Data were analyzed between August 30, 2022, and February 27, 2023. Results: Neonates with POE (1656 [0.4%]; mean [SD] gestational age, 37.7 [2.1] weeks; 836 females [50.5%]; 820 males [49.5%]) were more likely to be born preterm, have low birth weight for gestational age, and be coexposed to cigarette smoke compared with nonexposed neonates. Perinatal opioid exposure was associated with an increased risk of perinatal infection (adjusted odds ratio [AOR], 1.62; 95% CI, 1.38-1.90) and eczema and dermatitis (AOR, 11.91; 95% CI, 9.84-14.41) compared with nonexposure. Neonatal abstinence syndrome was also associated with both conditions (AOR, 2.91 [95% CI, 2.36-3.57] and 31.11 [95% CI, 24.64-39.28], respectively). Prenatal opioid exposure was also associated with an increased risk of childhood asthma (adjusted hazard ratio [AHR], 1.44; 95% CI, 1.16-1.79), but not allergies and anaphylaxis. It was also associated with an increased risk of childhood eczema and dermatitis, but only in children with POE from opioids used to treat OUD (AHR, 1.47; 95% CI, 1.08-1.99) rather than pain. In contrast, POE from opioids used for pain was associated with an increased risk of infection (AHR, 1.44; 95% CI, 1.32-1.58), but POE to opioids used to treat OUD was not. Autoimmune conditions were rare and were not observed to be associated with POE. Conclusions and Relevance: In this cohort study, POE was associated with an increased risk of infection, eczema and dermatitis, and asthma, but not allergies and anaphylaxis or autoimmune conditions. These findings highlight the importance of further study of opioid-induced immune changes during pregnancy, the potential impact on long-term health in exposed children, and the mechanisms of opioid-induced immune dysregulation.

Soluble PD-L1 reprograms blood monocytes to prevent cerebral edema and facilitate recovery after ischemic stroke.

Acute cerebral ischemia triggers a profound inflammatory response. While macrophages polarized to an M2-like phenotype clear debris and facilitate tissue repair, aberrant or prolonged macrophage activation is counterproductive to recovery. The inhibitory immune checkpoint Programmed Cell Death Protein 1 (PD-1) is upregulated on macrophage precursors (monocytes) in the blood after acute cerebrovascular injury. To investigate the therapeutic potential of PD-1 activation, we immunophenotyped circulating monocytes from patients and found that PD-1 expression was upregulated in the acute period after stroke. Murine studies using a temporary middle cerebral artery (MCA) occlusion (MCAO) model showed that intraperitoneal administration of soluble Programmed Death Ligand-1 (sPD-L1) significantly decreased brain edema and improved overall survival. Mice receiving sPD-L1 also had higher performance scores short-term, and more closely resembled sham animals on assessments of long-term functional recovery. These clinical and radiographic benefits were abrogated in global and myeloid-specific PD-1 knockout animals, confirming PD-1+ monocytes as the therapeutic target of sPD-L1. Single-cell RNA sequencing revealed that treatment skewed monocyte maturation to a non-classical Ly6Clo, CD43hi, PD-L1+ phenotype. These data support peripheral activation of PD-1 on inflammatory monocytes as a therapeutic strategy to treat neuroinflammation after acute ischemic stroke.

Chorioamnionitis disrupts erythropoietin and melatonin homeostasis through the placental-fetal-brain axis during critical developmental periods.

Introduction: Novel therapeutics are emerging to mitigate damage from perinatal brain injury (PBI). Few newborns with PBI suffer from a singular etiology. Most experience cumulative insults from prenatal inflammation, genetic and epigenetic vulnerability, toxins (opioids, other drug exposures, environmental exposure), hypoxia-ischemia, and postnatal stressors such as sepsis and seizures. Accordingly, tailoring of emerging therapeutic regimens with endogenous repair or neuro-immunomodulatory agents for individuals requires a more precise understanding of ligand, receptor-, and non-receptor-mediated regulation of essential developmental hormones. Given the recent clinical focus on neurorepair for PBI, we hypothesized that there would be injury-induced changes in erythropoietin (EPO), erythropoietin receptor (EPOR), melatonin receptor (MLTR), NAD-dependent deacetylase sirtuin-1 (SIRT1) signaling, and hypoxia inducible factors (HIF1α, HIF2α). Specifically, we predicted that EPO, EPOR, MLTR1, SIRT1, HIF1α and HIF2α alterations after chorioamnionitis (CHORIO) would reflect relative changes observed in human preterm infants. Similarly, we expected unique developmental regulation after injury that would reveal potential clues to mechanisms and timing of inflammatory and oxidative injury after CHORIO that could inform future therapeutic development to treat PBI. Methods: To induce CHORIO, a laparotomy was performed on embryonic day 18 (E18) in rats with transient uterine artery occlusion plus intra-amniotic injection of lipopolysaccharide (LPS). Placentae and fetal brains were collected at 24 h. Brains were also collected on postnatal day 2 (P2), P7, and P21. EPO, EPOR, MLTR1, SIRT1, HIF1α and HIF2α levels were quantified using a clinical electrochemiluminescent biomarker platform, qPCR, and/or RNAscope. MLT levels were quantified with liquid chromatography mass spectrometry. Results: Examination of EPO, EPOR, and MLTR1 at 24 h showed that while placental levels of EPO and MLTR1 mRNA were decreased acutely after CHORIO, cerebral levels of EPO, EPOR and MLTR1 mRNA were increased compared to control. Notably, CHORIO brains at P2 were SIRT1 mRNA deficient with increased HIF1α and HIF2α despite normalized levels of EPO, EPOR and MLTR1, and in the presence of elevated serum EPO levels. Uniquely, brain levels of EPO, EPOR and MLTR1 shifted at P7 and P21, with prominent CHORIO-induced changes in mRNA expression. Reductions at P21 were concomitant with increased serum EPO levels in CHORIO rats compared to controls and variable MLT levels. Discussion: These data reveal that commensurate with robust inflammation through the maternal placental-fetal axis, CHORIO impacts EPO, MLT, SIRT1, and HIF signal transduction defined by dynamic changes in EPO, EPOR, MLTR1, SIRT1, HIF1α and HIF2α mRNA, and EPO protein. Notably, ligand-receptor mismatch, tissue compartment differential regulation, and non-receptor-mediated signaling highlight the importance, complexity and nuance of neural and immune cell development and provide essential clues to mechanisms of injury in PBI. As the placenta, immune cells, and neural cells share many common, developmentally regulated signal transduction pathways, further studies are needed to clarify the perinatal dynamics of EPO and MLT signaling and to capitalize on therapies that target endogenous neurorepair mechanisms.

Pilot Translational Precision Biobehavioral Assays for Early Detection of Motor Impairments in a Rat Model of Cerebral Palsy.

BACKGROUND: Cutting-edge neonatal programs diagnose cerebral palsy (CP) or "high risk of CP" using validated neurobehavioral exams in combination with risk history and neuroimaging. In rat models, digital gait analyses are the gold standard adult assessment, but tools in infant rats are limited. Refinement of infant rat neurobehavioral correlates of CP will establish translational behavioral biomarkers to delineate early mechanisms of CP in both humans and rodent models of CP. OBJECTIVE: To facilitate precision medicine approaches of neurodevelopmental health and integrate basic and clinical research approaches for CP, we developed and piloted a new assay of neonatal rat neurobehavior to mimic human neonate exams. METHODS: Our established rat model of CP secondary to chorioamnionitis (CHORIO) that induces bilateral motor impairment reminiscent of spastic CP was used. On postnatal day 10 (P10), 5 min videos were recorded of 26 (6 sham and 20 CHORIO) animals moving freely in a cage were reviewed by an evaluator trained in the human General Movements Assessment (GMA). Non-blinded observation revealed two behaviors that differed between rat pups in each group (time spent rearing; multi-dimensional nose sweeping; and sniffing). Each video was re-coded for these criteria by an evaluator blind to group status. Differences between sham and CP groups were analyzed using a Mann-Whitney U-test or Student's t-test (p < 0.05 level of significance). RESULTS: Neonatal rats with CP exhibited sensorimotor impairment and decreased spatial exploration. CP rats spent significantly less time rearing (17.85 ± 1.60 s vs. 34.8 ± 2.89 s, p = 0.007) and engaged in multi-dimensional nose sweeping and sniffing (2.2 ± 0.58 episodes vs. 5.5 ± 0.96 episodes, p = 0.03) than sham controls. CONCLUSIONS: These pilot findings of harmonized translational and precision biobehavioral assays provide an opportunity for increased expediency of clinical trials at the earliest stages of brain development.

Sexual Dimorphism in the Closure of the Hippocampal Postnatal Critical Period of Synaptic Plasticity after Intrauterine Growth Restriction: Link to Oligodendrocyte and Glial Dysregulation.

Intrauterine growth restriction (IUGR) resulting from hypertensive disease of pregnancy (HDP) leads to sexually dimorphic hippocampal-dependent cognitive and memory impairment in humans. In our translationally relevant mouse model of IUGR incited by HDP, we have previously shown that the synaptic development in the dorsal hippocampus including GABAergic development, NPTX2+ excitatory synaptic formation, axonal myelination, and perineural net (PNN) formation were perturbed by IUGR at adolescent equivalence in humans (P40). The persistence of these disturbances through early adulthood and the potential upstream mechanisms are currently unknown. Thus, we hypothesized that NPTX2+ expression, PNN formation, axonal myelination, all events closing synaptic development in the hippocampus, will be persistently perturbed, particularly affecting IUGR female mice through P60 given the fact that they had worse short-term recognition memory in this model. We additionally hypothesized that such sexual dimorphism is linked to persistent glial dysregulation. We induced IUGR by a micro-osmotic pump infusion of a potent vasoconstrictor U-46619, a thromboxane A2-analog, in the last week of the C57BL/6 mouse gestation to precipitate HDP. Sham-operated mice were used as controls. At P60, we assessed hippocampal and hemispheric volumes, NPTX2 expression, PNN formation, as well as myelin basic protein (MBP), Olig2, APC/CC1, and M-NF expression. We also evaluated P60 astrocytic (GFAP) reactivity and microglial (Iba1 and TMEM119) activation using immunofluorescent-immunohistochemistry and Imaris morphological analysis plus cytokine profiling using Meso Scale Discovery platform. IUGR offspring continued to have smaller hippocampal volumes at P60 not related to changes in hemisphere volume. NPTX2+ puncta counts and volumes were decreased in IUGR hippocampal CA subregions of female mice compared to sex-matched shams. Intriguingly, NPTX2+ counts and volumes were concurrently increased in the dentate gyrus (DG) subregion. PNN volumes were smaller in CA1 and CA3 of IUGR female mice along with PNN intensity in CA3 but they had larger volumes in the CA3 of IUGR male mice. The myelinated axon (MBP+) areas, volumes, and lengths were all decreased in the CA1 of IUGR female mice compared to sex-matched shams, which correlated with a decrease in Olig2 nuclear expression. No decrease in the number of APC/CC1+ mature oligodendrocytes was identified. We noted an increase in M-NF expression in the mossy fibers connecting DG to CA3 only in IUGR female mice. Reactive astrocytes denoted by GFAP areas, volumes, lengths, and numbers of branching were increased in IUGR female CA1 but not in IUGR male CA3 compared to sex-matched shams. Lastly, activated microglia were only detected in IUGR female CA1 and CA3 subregions. We detected no difference in the cytokine profile between sham and IUGR adult mice of either sex. Collectively, our data support a sexually dimorphic impaired closure of postnatal critical period of synaptic plasticity in the hippocampus of young adult IUGR mice with greater effects on females. A potential mechanism supporting such dimorphism may include oligodendrocyte dysfunction in IUGR females limiting myelination, allowing axonal overgrowth followed by a reactive glial-mediated synaptic pruning.

Unbiased Quantitative Single-Cell Morphometric Analysis to Identify Microglia Reactivity in Developmental Brain Injury.

Microglia morphological studies have been limited to the process of reviewing the most common characteristics of a group of cells to conclude the likelihood of a "pathological" milieu. We have developed an Imaris-software-based analytical pipeline to address selection and operator biases, enabling use of highly reproducible machine-learning algorithms to quantify at single-cell resolution differences between groups. We hypothesized that this analytical pipeline improved our ability to detect subtle yet important differences between groups. Thus, we studied the temporal changes in Iba1+ microglia-like cell (MCL) populations in the CA1 between P10-P11 and P18-P19 in response to intrauterine growth restriction (IUGR) at E12.5 in mice, chorioamnionitis (chorio) at E18 in rats and neonatal hypoxia-ischemia (HI) at P10 in mice. Sholl and convex hull analyses differentiate stages of maturation of Iba1+ MLCs. At P10-P11, IUGR or HI MLCs were more prominently 'ameboid', while chorio MLCs were hyper-ramified compared to sham. At P18-P19, HI MLCs remained persistently 'ameboid' to 'transitional'. Thus, we conclude that this unbiased analytical pipeline, which can be adjusted to other brain cells (i.e., astrocytes), improves sensitivity to detect previously elusive morphological changes known to promote specific inflammatory milieu and lead to worse outcomes and therapeutic responses.

In utero methadone exposure permanently alters anatomical and functional connectivity: A preclinical evaluation.

The opioid epidemic is an ongoing public health crisis, and children born following prenatal opioid exposure (POE) have increased risk of long-term cognitive and behavioral sequelae. Clinical studies have identified reduced gray matter volume and abnormal white matter microstructure in children with POE but impacts on whole-brain functional brain connectivity (FC) have not been reported. To define effects of POE on whole brain FC and white matter injury in adult animals, we performed quantitative whole-brain structural and functional MRI. We used an established rat model of POE in which we have previously reported impaired executive function in adult rats analogous to persistent neurocognitive symptoms described in humans with POE. Pregnant Sprague-Dawley rat dams received continuous methadone (12 mg/kg/day) vs. saline infusion for 28 days via osmotic mini-pumps, exposing rats to pre- and postnatal opioid until weaning. At young adult age (P60), POE and saline exposed offspring underwent in vivo MRI included diffusion tensor imaging and functional MRI (fMRI). Results indicate that fractional anisotropy (FA) was decreased in adult animals with POE [n = 11] compared to animals that received saline [n = 9] in major white matter tracts, including the corpus callosum (p < 0.001) and external capsule (p < 0.01). This change in FA was concomitant with reduced axial diffusivity in the external capsule (p < 0.01) and increased radial diffusivity in the corpus callosum (p < 0.01). fMRI analyses reveal brainwide FC was diffusely lower in POE (p < 10-6; 10% of variance explained by group). Decreased connectivity in cortical-cortical and cortico-basal ganglia circuitry was particularly prominent with large effect sizes (Glass's Δ > 1). Taken together, these data confirm POE reduces brainwide functional connectivity as well as microstructural integrity of major white matter tracts. Altered neural circuitry, dysregulated network refinement, and diffuse network dysfunction have been implicated in executive function deficits that are common in children with POE. FC may serve as a translatable biomarker in children with POE.

A review of the genomics of neonatal abstinence syndrome.

Neonatal abstinence syndrome (NAS) is a constellation of signs of withdrawal occurring after birth following in utero exposure to licit or illicit opioids. Despite significant research and public health efforts, NAS remains challenging to diagnose, predict, and manage due to highly variable expression. Biomarker discovery in the field of NAS is crucial for stratifying risk, allocating resources, monitoring longitudinal outcomes, and identifying novel therapeutics. There is considerable interest in identifying important genetic and epigenetic markers of NAS severity and outcome that can guide medical decision making, research efforts, and public policy. A number of recent studies have suggested that genetic and epigenetic changes are associated with NAS severity, including evidence of neurodevelopmental instability. This review will provide an overview of the role of genetics and epigenetics in short and longer-term NAS outcomes. We will also describe novel research efforts using polygenic risk scores for NAS risk stratification and salivary gene expression to understand neurobehavioral modulation. Finally, emerging research focused on neuroinflammation from prenatal opioid exposure may elucidate novel mechanisms that could lead to development of future novel therapeutics.

Outcomes of the 2019 hydrocephalus association workshop, "Driving common pathways: extending insights from posthemorrhagic hydrocephalus".

The Hydrocephalus Association (HA) workshop, Driving Common Pathways: Extending Insights from Posthemorrhagic Hydrocephalus, was held on November 4 and 5, 2019 at Washington University in St. Louis. The workshop brought together a diverse group of basic, translational, and clinical scientists conducting research on multiple hydrocephalus etiologies with select outside researchers. The main goals of the workshop were to explore areas of potential overlap between hydrocephalus etiologies and identify drug targets that could positively impact various forms of hydrocephalus. This report details the major themes of the workshop and the research presented on three cell types that are targets for new hydrocephalus interventions: choroid plexus epithelial cells, ventricular ependymal cells, and immune cells (macrophages and microglia).

Omics approaches: interactions at the maternal-fetal interface and origins of child health and disease.

Immunoperinatology is an emerging field. Transdisciplinary efforts by physicians, physician-scientists, basic science researchers, and computational biologists have made substantial advancements by identifying unique immunologic signatures of specific diseases, discovering innovative preventative or treatment strategies, and establishing foundations for individualized neonatal intensive care of the most vulnerable neonates. In this review, we summarize the immunobiology and immunopathology of pregnancy, highlight omics approaches to study the maternal-fetal interface, and their contributions to pregnancy health. We examined the importance of transdisciplinary, multiomic (such as genomics, transcriptomics, proteomics, metabolomics, and immunomics) and machine-learning strategies in unraveling the mechanisms of adverse pregnancy, neonatal, and childhood outcomes and how they can guide the development of novel therapies to improve maternal and neonatal health. IMPACT: Discuss immunoperinatology research from the lens of omics and machine-learning approaches. Identify opportunities for omics-based approaches to delineate infection/inflammation-associated maternal, neonatal, and later life adverse outcomes (e.g., histologic chorioamnionitis [HCA]).

Corrigendum: Methadone alters the peripheral inflammatory and central immune landscape following prenatal exposure in rats.

[This corrects the article DOI: 10.3389/adar.2022.10792.].

A second dimension of somatosensory system injury? Thalamic volume loss and neuropathic pain in adults with cerebral palsy and periventricular white matter injury.

Objectives: Lemniscal (motor-related) and spinothalamic (neuropathic pain-related) somatosensory abnormalities affect different subsets of adults with cerebral palsy (CP). Lemniscal/motor abnormalities are associated with posterior thalamic radiation white matter disruption in individuals with CP and white matter injury. We tested the hypothesis that neuropathic pain symptoms in this population are rather associated with injury of the somatosensory (posterior group nuclei) thalamus. Methods: In this cross-sectional study, communicative adults with CP and bilateral white matter injury and neurotypical control participants volunteered to self-report pain symptoms and undergo research MRI. Posterior group thalamic nuclei volume was computed and correlated against neuropathic pain scores. Results: Participants with CP (n=6) had, on average, 24% smaller posterior group thalamic volumes (95% CI [10-39%]; corrected p=0.01) than control participants. More severe volume loss was correlated with more severe neuropathic pain scores (ρ=-0.87 [-0.99,-0.20]; p=0.02). Discussion: Association with thalamic volume loss suggests that neuropathic pain in adults with CP may frequently be central neuropathic pain. Complementing assessments of white matter microstructure, regional brain volumes hold promise as diagnostic biomarkers for central neuropathic pain in individuals with structural brain disorders.

Neuroinflammation represents a common theme amongst genetic and environmental risk factors for Alzheimer and Parkinson diseases.

Multifactorial diseases are characterized by inter-individual variation in etiology, age of onset, and penetrance. These diseases tend to be relatively common and arise from the combined action of genetic and environmental factors; however, parsing the convoluted mechanisms underlying these gene-by-environment interactions presents a significant challenge to their study and management. For neurodegenerative disorders, resolving this challenge is imperative, given the enormous health and societal burdens they impose. The mechanisms by which genetic and environmental effects may act in concert to destabilize homeostasis and elevate risk has become a major research focus in the study of common disease. Emphasis is further being placed on determining the extent to which a unifying biological principle may account for the progressively diminishing capacity of a system to buffer disease phenotypes, as risk for disease increases. Data emerging from studies of common, neurodegenerative diseases are providing insights to pragmatically connect mechanisms of genetic and environmental risk that previously seemed disparate. In this review, we discuss evidence positing inflammation as a unifying biological principle of homeostatic destabilization affecting the risk, onset, and progression of neurodegenerative diseases. Specifically, we discuss how genetic variation associated with Alzheimer disease and Parkinson disease may contribute to pro-inflammatory responses, how such underlying predisposition may be exacerbated by environmental insults, and how this common theme is being leveraged in the ongoing search for effective therapeutic interventions.

Prenatal and postnatal insults differentially contribute to executive function and cognition: Utilizing touchscreen technology for perinatal brain injury research.

The use of touchscreen technology to evaluate cognitive deficits in animal models has grown tremendously over the past 20 years. The touchscreen apparatus encompasses many advantages, namely a high level of standardization and translational capability. Improvements in technology in recent years have expanded the versatility of the touchscreen platform, as it is able to test distinct cognitive modalities including working memory, attention, discrimination, and association. Importantly, touchscreen technology has allowed researchers to explore deficits in multiple pillars of cognition in a wide variety of perinatal disorders with neurological sequelae across critical developmental windows. The touchscreen platform has been used to dissect deficits in antenatal CNS injury including fetal alcohol syndrome, prenatal opioid exposure, and chorioamnionitis, to peripartum insults such as term hypoxic-ischemic encephalopathy, to early postnatal insults including infantile traumatic brain injury. Most importantly, touchscreen technology offers the sensitivity necessary to detect subtle injury and treatment-induced changes in cognition and executive function beyond those offered by more rudimentary tests of rodent cognition. Understanding the pathophysiology of these disorders in rodents is paramount to addressing these deficits in human infants and dissecting the neural circuitry essential to perinatal brain injury pathophysiology and responsiveness to novel therapeutics. Touchscreen testing provides an effective, facile, sophisticated technique to accelerate the goal of improving cognitive and behavioral outcomes of children who suffer perinatal brain injury.

Pathogenesis of posthemorrhagic hydrocephalus of prematurity: New horizons.

Posthemorrhagic hydrocephalus of prematurity (PHHP) remains a vexing problem for patients, their families, and the healthcare system. The complexity of the pathogenesis of PHHP also presents a unique challenge within the fields of neonatology, neurology and neurosurgery. Here we focus on pathogenesis of PHHP and its impact on the development of CSF dynamics including choroid plexus, ependymal motile cilia and glymphatic system. PHHP is contrasted with infantile hydrocephalus from other etiologies, and with other types of posthemorrhagic hydrocephalus that occur later in life. The important concept that distinguishing ventricular volume from brain health and function is highlighted. The influence of the pathogenesis of PHHP on current interventions is reviewed, with particular emphasis on how the unique pathogenesis of PHHP contributes to the high rate of failure of current existing interventions. Finally, we discuss emerging interventions. A thorough understanding of the pathogenesis of PHHP is essential to developing effective non-surgical therapeutics to prevent the transformation from severe IVH to PHHP.