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Background In newborns with hypoxic-ischemic encephalopathy (HIE), the correlation between neonatal neuroimaging and the degree of neurodevelopmental impairment (NDI) is unclear. Methods Infants with HIE enrolled in a randomized controlled trial underwent neonatal MRI/MR spectroscopy (MRS) using a harmonized protocol at 4–6 days of age. The severity of brain injury was measured with a validated scoring system. Using proportional odds regression, we calculated adjusted odds ratios (aOR) for the associations between MRI/MRS measures of injury and primary ordinal outcome (i.e., normal, mild NDI, moderate NDI, severe NDI, or death) at age 2 years. Results Of 451 infants with MRI/MRS at a median age of 5 days (IQR 4.5–5.8), outcomes were normal (51%); mild (12%), moderate (14%), severe NDI (13%); or death (9%). MRI injury score (aOR 1.06, 95% CI 1.05, 1.07), severe brain injury (aOR 39.6, 95% CI 16.4, 95.6), and MRS lactate/n-acetylaspartate (NAA) ratio (aOR 1.6, 95% CI 1.4,1.8) were associated with worse primary outcomes. Infants with mild/moderate MRI brain injury had similar BSID-III cognitive, language, and motor scores as infants with no injury. Conclusion In the absence of severe injury, brain MRI/MRS does not accurately discriminate the degree of NDI. Given diagnostic uncertainty, families need to be counseled regarding a range of possible neurodevelopmental outcomes. Impact Half of all infants with hypoxic-ischemic encephalopathy (HIE) enrolled in a large clinical trial either died or had neurodevelopmental impairment at age 2 years despite receiving therapeutic hypothermia. Severe brain injury and a global pattern of brain injury on MRI were both strongly associated with death or neurodevelopmental impairment. Infants with mild or moderate brain injury had similar mean BSID-III cognitive, language, and motor scores as infants with no brain injury on MRI. Given the prognostic uncertainty of brain MRI among infants with less severe degrees of brain injury, families should be counseled regarding a range of possible neurodevelopmental outcomes.

To examine the frequency of placental abnormalities in a multicenter cohort of newborn infants with hypoxic-ischemic encephalopathy (HIE) and to determine the association between acuity of placental abnormalities and clinical characteristics of HIE.Infants born at ≥36 weeks of gestation (n = 500) with moderate or severe HIE were enrolled in the High-dose Erythropoietin for Asphyxia and Encephalopathy Trial. A placental pathologist blinded to clinical information reviewed clinical pathology reports to determine the presence of acute and chronic placental abnormalities using a standard classification system.Complete placental pathologic examination was available for 321 of 500 (64%) trial participants. Placental abnormalities were identified in 273 of 321 (85%) and were more common in infants ≥40 weeks of gestation (93% vs 81%, P = .01). A combination of acute and chronic placental abnormalities (43%) was more common than either acute (20%) or chronic (21%) abnormalities alone. Acute abnormalities included meconium staining of the placenta (41%) and histologic chorioamnionitis (39%). Chronic abnormalities included maternal vascular malperfusion (25%), villitis of unknown etiology (8%), and fetal vascular malperfusion (6%). Infants with chronic placental abnormalities exhibited a greater mean base deficit at birth (-15.9 vs -14.3, P = .049) than those without such abnormalities. Patients with HIE and acute placental lesions had older mean gestational ages (39.1 vs 38.0, P .001) and greater rates of clinically diagnosed chorioamnionitis (25% vs 2%, P .001) than those without acute abnormalities.Combined acute and chronic placental abnormalities were common in this cohort of infants with HIE, underscoring the complex causal pathways of HIE.ClinicalTrials.gov: NCT02811263.

Neonatal hypoxic-ischemic encephalopathy is an important cause of death as well as long-term disability in survivors. Erythropoietin has been hypothesized to have neuroprotective effects in infants with hypoxic-ischemic encephalopathy, but its effects on neurodevelopmental outcomes when given in conjunction with therapeutic hypothermia are unknown.In a multicenter, double-blind, randomized, placebo-controlled trial, we assigned 501 infants born at 36 weeks or more of gestation with moderate or severe hypoxic-ischemic encephalopathy to receive erythropoietin or placebo, in conjunction with standard therapeutic hypothermia. Erythropoietin (1000 U per kilogram of body weight) or saline placebo was administered intravenously within 26 hours after birth, as well as at 2, 3, 4, and 7 days of age. The primary outcome was death or neurodevelopmental impairment at 22 to 36 months of age. Neurodevelopmental impairment was defined as cerebral palsy, a Gross Motor Function Classification System level of at least 1 (on a scale of 0 [normal] to 5 [most impaired]), or a cognitive score of less than 90 (which corresponds to 0.67 SD below the mean, with higher scores indicating better performance) on the Bayley Scales of Infant and Toddler Development, third edition.Of 500 infants in the modified intention-to-treat analysis, 257 received erythropoietin and 243 received placebo. The incidence of death or neurodevelopmental impairment was 52.5% in the erythropoietin group and 49.5% in the placebo group (relative risk, 1.03; 95% confidence interval [CI], 0.86 to 1.24; P = 0.74). The mean number of serious adverse events per child was higher in the erythropoietin group than in the placebo group (0.86 vs. 0.67; relative risk, 1.26; 95% CI, 1.01 to 1.57).The administration of erythropoietin to newborns undergoing therapeutic hypothermia for hypoxic-ischemic encephalopathy did not result in a lower risk of death or neurodevelopmental impairment than placebo and was associated with a higher rate of serious adverse events. (Funded by the National Institute of Neurological Disorders and Stroke; ClinicalTrials.gov number, NCT02811263.).

Background Neonatal stroke affects 1 in 2800 live births and is a major cause of neurological injury. The Sonic hedgehog (Shh) signaling pathway is critical for central nervous system (CNS) development and has neuroprotective and reparative effects in different CNS injury models. Previous studies have demonstrated beneficial effects of small molecule Shh-Smoothened agonist (SAG) against neonatal cerebellar injury and it improves Down syndrome-related brain structural deficits in mice. Here we investigated SAG neuroprotection in rat models of neonatal ischemia–reperfusion (stroke) and adult focal white matter injury. Methods We used transient middle cerebral artery occlusion at P10 and ethidium bromide (EB) injection in adult rats to induce damage. Following surgery and SAG or vehicle treatment, we analyzed tissue loss, cell proliferation and fate, and behavioral outcome. Results We report that a single dose of SAG administered following neonatal stroke preserved brain volume, reduced gliosis, enhanced oligodendrocyte progenitor cell (OPC) and EC proliferation, and resulted in long-term cognitive improvement. Single-dose SAG also promoted proliferation of OPCs following focal demyelination in the adult rat. Conclusions These findings indicate benefit of one-time SAG treatment post insult in reducing brain injury and improving behavioral outcome after experimental neonatal stroke. Impact A one-time dose of small molecule Sonic hedgehog agonist protected against neonatal stroke and improved long-term behavioral outcomes in a rat model. This study extends the use of Sonic hedgehog in treating developing brain injury, previously shown in animal models of Down syndrome and cerebellar injury. Sonic hedgehog agonist is one of the most promising therapies in treating neonatal stroke thanks to its safety profile and low dosage.

The COVID-19 pandemic will leave an indelible mark on the careers of current medical trainees. Given the disruptions to medical education, economic impact on institutions, and the uncertainties around future job prospects, trainees are facing unprecedented challenges. This situation is especially concerning for futures of pediatric physician-scientist trainees, where concerns regarding maintaining the pipeline were well documented prior to the emergence of COVID-19. In this Perspectives article, we leverage the unique expertise of our workgroup to address concerns of physician-scientist trainees and to provide suggestions on how to navigate career trajectories in the post-COVID-19 era. We identified and addressed four major areas of concern: lack of in-person conferences and the associated decrease access to mentors and networking activities, decreased academic productivity, diminished job prospects, and mental health challenges. We also suggest actions for trainees, mentors and educational leaders, and institutions to help support trainees during the pandemic, with a goal of maintaining the pediatric physician-scientist pipeline., Impact The advice in this article will provide guidance to pediatric physician-scientist trainees on sustaining and developing their career during the continued challenges presented by COVID-19.We also provide suggestions for mentors, educational leaders, institutions, and professional societies on ways to help support the pediatric physician-scientist pipeline.

Background and Purpose: Perinatal stroke is a common cause of life-long neurobehavioral compromise. Mesenchymal stromal cells (MSCs) and EPO (erythropoietin) have each demonstrated short-term benefit with delayed administration after stroke, and combination therapy may provide the most benefit. The purpose of this study is to determine the long-term histological and functional efficacy of enhanced, intranasal stem cell therapy (MSC preexposed to EPO) compared with standard MSC or multidose systemic EPO. Methods: Transient middle cerebral artery occlusion or sham surgery was performed in postnatal day (P) 10 Sprague-Dawley rats, who were treated with single-dose intranasal MSC, MSC preexposed to EPO (MSC/EPO), multidose systemic EPO (EPO3; 1000 u/kg per dose×3 every 72 hours), or cell-conditioned media on P13 (day 3 [P13–P19] for EPO), or on P17 (day 7 [P17–P23] for EPO). At 2 months of age, animals underwent novel object recognition, cylinder rearing, and open field testing to assess recognition memory, sensorimotor function, and anxiety in adulthood. Results: MSC, MSC/EPO, and EPO3 improved brain volume when administered at 3 or 7 days after middle cerebral artery occlusion. MSC/EPO also enhanced long-term recognition memory with either day 3 or day 7 treatment, but EPO3 had the most long-term benefit, improving recognition memory and exploratory behavior and reducing anxiety. Conclusions: These data suggest that single-dose MSC/EPO and multidose systemic EPO improve long-term neurobehavioral outcomes even when administration is delayed, although EPO was the most effective treatment overall. It is possible that EPO represents a final common pathway for improved long-term repair, although the specific mechanisms remain to be determined.

Newborns with hypoxic–ischemic encephalopathy (HIE) may exhibit abnormalities on placental histology. In this phase II clinical trial ancillary study, we hypothesized that placental abnormalities correlate with MRI brain injury and with response to treatment. Fifty newborns with moderate/severe encephalopathy who received hypothermia were enrolled in a double-blind, placebo-controlled trial of erythropoietin for HIE. A study pathologist reviewed all available clinical pathology reports to determine the presence of chronic abnormalities and acute chorioamnionitis. Neonatal brain MRIs were scored using a validated HIE scoring system. Placental abnormalities in 19 of the 35 (54%) patients with available pathology reports included chronic changes (N = 13), acute chorioamnionitis (N = 9), or both (N = 3). MRI subcortical brain injury was less common in infants with a placental abnormality (26 vs. 69%, P = 0.02). Erythropoietin treatment was associated with a lower global brain injury score (median 2.0 vs. 11.5, P = 0.003) and lower rate of subcortical brain injury (33 vs. 90%, P = 0.01) among patients with no chronic placental abnormality but not in patients whose placentas harbored a chronic abnormality. Erythropoietin treatment was associated with less brain injury only in patients whose placentas exhibited no chronic histologic changes. Placentas may provide clues to treatment response in HIE.

Objective Stroke is a major cause of death and disability in the United States. Current acute stroke therapy consists of clot-dissolving drugs, catheter-based interventions and physical rehabilitation. To date, there are no therapies that directly enhance neuronal survival after a stroke. Previous work from our lab demonstrated that Interleukin-15 (IL-15) peptide could rescue cardiomyocytes subjected to hypoxia. We sought to extend these findings to cortical neurons since IL-15 has been implicated to have an important role in neuronal homeostasis. Methods We have evaluated the effect of IL-15 peptide on primary cortical neurons derived from embryonic rats in vitro under conditions of anoxia and glucose deprivation, and in vivo following middle cerebral artery occlusion. Results IL-15 administration rescued neuronal cells subjected to anoxia coupled with glucose deprivation (AGD), as well as with reoxygenation. A hallmark of stroke is the ischemic microenvironment and associated oxidative stress, which results in DNA damage and ER stress, both of which contribute to neuronal cell damage and death. The expression of anoxia, ER stress, and DNA damage factors/markers was evaluated via western blot and correlated with the cellular survival effects of IL-15 in vitro. In addition, IL-15 effect of alleviating ER stress and increasing cell survival was also observed in vivo. Interpretation Our data indicate, for the first time, that administration of the pleiotropic factor IL-15 reduces neuronal cell death during AGD, which correlates with modulation of multiple cellular stress pathways.

ObjectiveNeonatal stroke affects 1 in 2800 live births and is a major cause of neurological injury. The Sonic Hedgehog (Shh) signaling pathway is critical for central nervous system (CNS) development and has neuroprotective and reparative effects in different CNS injury models. Previous studies have demonstrated beneficial effects of small molecule Shh-Smoothened-agonist (SAG) against neonatal cerebellar injury and it improves Down syndrome-related brain structural deficits in mice. Here, we investigated SAG neuroprotection in rat models of neonatal ischemia-reperfusion (stroke) and adult focal white matter injury.MethodsWe used transient middle cerebral artery occlusion at P10 and ethidium bromide injection in adult rats to induce damage. Following surgery and SAG or vehicle treatment we analyzed tissue loss, cell proliferation and fate, and behavioral outcome.ResultsWe report that a single dose of SAG administered following neonatal stroke preserved brain volume, reduced inflammation, enhanced oligodendrocyte progenitor cell (OPC) and EC proliferation, and resulted in long-term cognitive improvement. Single-dose SAG also promoted proliferation of OPCs following focal demyelination in the adult rat.ConclusionThese findings indicate benefit of one-time SAG treatment post-insult in reducing brain injury and improving behavioral outcome after experimental neonatal stroke.

As the resident macrophages of the brain and spinal cord, microglia are crucial for the phagocytosis of infectious agents, apoptotic cells and synapses. During brain injury or infection, bone-marrow derived macrophages invade neural tissue, making it difficult to distinguish between invading macrophages and resident microglia. In addition to circulation-derived monocytes, other non-microglial central nervous system (CNS) macrophage subtypes include border-associated meningeal, perivascular and choroid plexus macrophages. Using immunofluorescent labeling, flow cytometry and Cre-dependent ribosomal immunoprecipitations, we describe P2ry12-CreER, a new tool for the genetic targeting of microglia. We use this new tool to track microglia during embryonic development and in the context of ischemic injury and neuroinflammation. Because of the specificity and robustness of microglial recombination with P2ry12-CreER, we believe that this new mouse line will be particularly useful for future studies of microglial function in development and disease.

Author(s): Gonzalez, Fernando F; Heyman, Melvin B | Abstract: Pediatric physician-scientists play a critical role in translating research findings from basic and clinical research into new paradigms and approaches pertinent to our care for children. The United States has long been a leader in biomedical research; however, this workforce has been diminishing over many years secondary to decreases in recruitment, limited funding and protected time, and attrition during training.1 This pipeline of pediatric physician-scientists is critical not only for discovery but also for contextualizing specific findings and extrapolating those findings to understanding development and treatment strategies that can reduce disease burden and improve quality of life.

In human preterm newborns, caffeine increases brain activity and improves neurodevelopmental outcomes. In animal models of hypoxic ischemic brain injury, caffeine pretreatment reduces infarct volume. We studied the relationship between tissue neuroprotection and brain activity after injury to further understand caffeine neuroprotection. Rat dams received caffeine prior to birth or on postnatal day 3 (P3) through P16. Caffeine-treated and -untreated pups underwent the Vannucci procedure (unilateral carotid ligation, global hypoxia) on P2. A subset had EEG recordings. Brain hemispheric infarct volume was measured on P16. P2 hypoxic ischemia (HI) results in histologic brain injury (mean ± standard deviation infarct volume 10.3 ± 4.6%) and transient suppression of EEG activity. Caffeine pretreatment reduces brain injury (mean ± standard deviation infarct volume 1.6 ± 4.5%, p < 0.001) and improves amplitude-integrated EEG (aEEG) and EEG burst duration and amplitude. Caffeine treatment after HI does not reduce infarct volume (mean ± standard deviation 8.3 ± 4.1%, p = 1.0). However, caffeine posttreatment was equally effective at restoring aEEG amplitude and EEG burst duration and amplitude. Thus, caffeine supports brain background electrical activity independent of tissue neuroprotection.

Therapeutic hypothermia is now well established to significantly improve survival without disability after neonatal encephalopathy (NE). To further improve outcomes, we need to better understand the mechanisms of brain injury. The central finding, which offers the potential for neuroprotective and neurorestorative interventions, is that brain damage after perinatal hypoxia-ischemia evolves slowly over time. Although brain cells may die during profound hypoxia-ischemia, even after surprisingly severe insults many cells show transient recovery of oxidative metabolism during a “latent” phase characterized by actively suppressed neural metabolism and activity. Critically, after moderate to severe hypoxia-ischemia, this transient recovery is followed after ~6 h by a phase of secondary deterioration, with delayed seizures, failure of mitochondrial function, cytotoxic edema, and cell death over ~72 h. This is followed by a tertiary phase of remodeling and recovery. This review discusses the mechanisms of injury that occur during the primary, latent, secondary and tertiary phases of injury and potential treatments that target one or more of these phases. By analogy with therapeutic hypothermia, treatment as early as possible in the latent phase is likely to have the greatest potential to prevent injury (“neuroprotection”). In the secondary phase of injury, anticonvulsants can attenuate seizures, but show limited neuroprotection. Encouragingly, there is now increasing preclinical evidence that late, neurorestorative interventions have potential to improve long-term outcomes.

Newborns with hypoxic-ischemic encephalopathy (HIE) may exhibit abnormalities on placental histology. In this phase II clinical trial ancillary study, we hypothesized that placental abnormalities correlate with MRI brain injury and with response to treatment.Fifty newborns with moderate/severe encephalopathy who received hypothermia were enrolled in a double-blind, placebo-controlled trial of erythropoietin for HIE. A study pathologist reviewed all available clinical pathology reports to determine the presence of chronic abnormalities and acute chorioamnionitis. Neonatal brain MRIs were scored using a validated HIE scoring system.Placental abnormalities in 19 of the 35 (54%) patients with available pathology reports included chronic changes (N = 13), acute chorioamnionitis (N = 9), or both (N = 3). MRI subcortical brain injury was less common in infants with a placental abnormality (26 vs. 69%, P = 0.02). Erythropoietin treatment was associated with a lower global brain injury score (median 2.0 vs. 11.5, P = 0.003) and lower rate of subcortical brain injury (33 vs. 90%, P = 0.01) among patients with no chronic placental abnormality but not in patients whose placentas harbored a chronic abnormality.Erythropoietin treatment was associated with less brain injury only in patients whose placentas exhibited no chronic histologic changes. Placentas may provide clues to treatment response in HIE.

Background: Hypoxic-ischemic encephalopathy (HIE) remains an important cause of neonatal death and frequently leads to significant long-term disability in survivors. Therapeutic hypothermia, while beneficial, still leaves many treated infants with lifelong disabilities. Adjunctive therapies are needed, and erythropoietin (Epo) has the potential to provide additional neuroprotection. Objectives: The aim of this study was to review the current incidence, mechanism of injury, and sequelae of HIE, and to describe a new phase III randomized, placebo-controlled trial of Epo neuroprotection in term and near-term infants with moderate to severe HIE treated with therapeutic hypothermia. Methods: This article presents an overview of HIE, neuroprotective functions of Epo, and the design of a double-blind, placebo-controlled, multicenter trial of high-dose Epo administration, enrolling 500 neonates ≥36 weeks of gestation with moderate or severe HIE diagnosed by clinical criteria. Results and Conclusions: Epo has robust neuroprotective effects in preclinical studies, and phase I/II trials suggest that multiple high doses of Epo may provide neuroprotection against brain injury in term infants. The High Dose Erythropoietin for Asphyxia and Encephalopathy (HEAL) Trial will evaluate whether high-dose Epo reduces the combined outcome of death or neurodevelopmental disability when given in conjunction with hypothermia to newborns with moderate/severe HIE.

In recent years, growth factor therapy has emerged as a potential treatment for ischemic brain injury. The efficacy of therapies that either directly introduce or stimulate local production of growth factors and their receptors in damaged brain tissue has been tested in a multitude of models for different Central Nervous System (CNS) diseases. These growth factors include erythropoietin (EPO), vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), and insulin-like growth factor (IGF-1), among others. Despite the promise shown in animal models, the particular growth factors that should be used to maximize both brain protection and repair, and the therapeutic critical period, are not well defined. We will review current pre-clinical and clinical evidence for growth factor therapies in treating different causes of brain injury, as well as issues to be addressed prior to application in humans.

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature and mature brain. Stroke occurs commonly in the perinatal period in humans, and transient ischemia-reperfusion is the most common form of stroke in neonates. The reperfusion phase is a critical component of injury progression, which occurs over a period of days to weeks, and of the endogenous response to injury. This postnatal day 10 (p10) rat model of transient middle cerebral artery occlusion (tMCAO) creates a unilateral, non-hemorrhagic focal ischemia-reperfusion injury that can be utilized to study the mechanisms of focal injury and repair in the full-term-equivalent brain. The injury pattern that is produced by tMCAO is consistent and highly reproducible and can be confirmed with MRI or histological analyses. The severity of injury can be manipulated through changes in occlusion time and other methods that will be discussed.

Brain injury in the full-term and near-term neonates is a significant cause of mortality and long-term morbidity, resulting in injury patterns distinct from that seen in premature infants and older patients. Therapeutic hypothermia improves long-term outcomes for many of these infants, but there is a continued search for therapies to enhance the plasticity of the newborn brain, resulting in long-term repair. It is likely that a combination strategy utilizing both early and late interventions may have the most benefit, capitalizing on endogenous mechanisms triggered by hypoxia or ischemia. Optimizing care of these critically ill newborns in the acute setting is also vital for improving both short- and long-term outcomes.

Background and Purpose— Stroke is a common cause of neonatal brain injury. The subventricular zone is a lifelong source of newly generated cells in rodents, and erythropoietin (EPO) treatment has shown benefit in different animal models of brain injury. The purpose of this study is to investigate the specific role of exogenous EPO on subventricular zone progenitor cell populations in response to neonatal stroke. Methods— Intraventricular injections of green fluorescent protein (GFP)–expressing lentivirus to label subventricular zone precursor cells were made in postnatal day 1 (P1) Long-Evans rats, which then underwent transient middle cerebral artery occlusion on P7. Middle cerebral artery occlusion and sham rats were treated with either vehicle or EPO (1000 U/kg) at reperfusion, 24 hours, and 7 days later. The density of double-labeled DCx+/GFP+, NeuN+/GFP+, O4+/GFP+, GFAP+/GFP+, as well as single-labeled GFP+ and Ki67+ cells, was calculated to determine cell fate outcome in the striatum at 72 hours and 2 weeks after stroke. Results— There was a significant increase in DCx+/GFP+ and NeuN+/GFP+ neurons and O4+/GFP+ oligodendrocyte precursors, with decreased GFAP+/GFP+ astrocytes at both time points in EPO-middle cerebral artery occlusion animals. There was also a significant increase in GFP+ cells and Ki67+ proliferating cells in EPO compared with vehicle-middle cerebral artery occlusion animals. Conclusions— These data suggest that subventricular zone neural progenitor cells proliferate and migrate to the site of injury after neonatal stroke and multiple doses of EPO, with a shift in cell fate toward neurogenesis and oligodendrogliosis at both early and late time points. The contribution of local cell proliferation and neurogenesis remains to be determined.

OBJECTIVE: To determine if multiple doses of erythropoietin (Epo) administered with hypothermia improve neuroradiographic and short-term outcomes of newborns with hypoxic-ischemic encephalopathy. METHODS: In a phase II double-blinded, placebo-controlled trial, we randomized newborns to receive Epo (1000 U/kg intravenously; n = 24) or placebo (n = 26) at 1, 2, 3, 5, and 7 days of age. All infants had moderate/severe encephalopathy; perinatal depression (10 minute Apgar RESULTS: The mean age at first study drug was 16.5 hours (SD, 5.9). Neonatal deaths did not significantly differ between Epo and placebo groups (8% vs 19%, P = .42). Brain MRI at mean 5.1 days (SD, 2.3) showed a lower global brain injury score in Epo-treated infants (median, 2 vs 11, P = .01). Moderate/severe brain injury (4% vs 44%, P = .002), subcortical (30% vs 68%, P = .02), and cerebellar injury (0% vs 20%, P = .05) were less frequent in the Epo than placebo group. At mean age 12.7 months (SD, 0.9), motor performance in Epo-treated (n = 21) versus placebo-treated (n = 20) infants were as follows: Alberta Infant Motor Scale (53.2 vs 42.8, P = .03); Warner Initial Developmental Evaluation (28.6 vs 23.8, P = .05). CONCLUSIONS: High doses of Epo given with hypothermia for hypoxic-ischemic encephalopathy may result in less MRI brain injury and improved 1-year motor function.

Preterm birth is a major cause of neonatal mortality and morbidity. While advances in medical care have improved the survival of preterm infants, neurodevelopmental problems persist in this population. This article aims to review factors associated with their neurodevelopmental outcomes. English language studies of neurodevelopmental outcomes in preterm infants were retrieved from PubMed. A total of 100 related publications were included. Early gestational age and birth weight are the most significant predictors of poor long-term neurological outcome. Structural changes of the brain, infection, male gender and neonatal intensive care unit course are also important factors affecting eventual outcome. Other complex biological and socio-economic factors, which extend from prenatal through postnatal periods, up through and including adulthood, also affect the trajectory of brain development in preterm infants. Neurodevelopmental problems continue to affect the preterm population. There is a critical need for collaboration among geneticists, obstetricians, pediatricians, and neuroimaging and rehabilitation experts to determine early predictive factors and neuroprotective therapies to properly treat or prevent poor neurodevelopmental outcomes in these infants.

Aim: To analyse the bacterial pathogens and drug sensitivities for neonatal community-acquired pneumonia. Methods: Seven hundred sixty sputum samples from newborns with community-acquired pneumonia were cultured to determine microbial organisms present and their drug sensitivities. Results: Of the 760 specimens, 425 grew pathogens for a 55.9% positive rate. Among the 425 positive cultures, 278 grew gram-negative organisms (65.4%), 142 grew gram-positive organisms (33.3%), while 5 grew fungus (1.3%). The most common gram-negative organisms were Escherichia coli, Klebsiella pneumoniae and Hemophilus influenzae, while the most common gram-positive organisms were Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus. To the gram-negative organisms, the most sensitive drugs were meropenem, imipenem and amikacin, while to the gram-positive ones were vancomycin, teicoplanin and quinupristin/dalfopristin. Conclusions: The most common causative bacteria were gram-negative organisms, which were highly sensitive to Meropenem, Imipenem and Amikacin, yet often treatable with more focused antibiotic coverage, which depended on the bacterium identified.

Objective: To investigate the effects of neonatal stroke on progenitor cells lining the lateral ventricles. Methods: Intraventricular injection of replication-incompetent green fluorescent protein (GFP)-expressing lentivirus was performed in postnatal day 1 (P1) rats to specifically label radial glia/type B neural stem cells and ependymal cells of the lateral ventricle. A subset of animals was exposed to transient middle cerebral artery occlusion (MCAO) at P7, with mild or moderate injury confirmed by diffusion-weighted MRI and histology. Newborn cells were identified by GFP expression, location and expression of cell type-specific markers in the striatum, cortex and olfactory bulb using confocal microscopy and systematic random sampling. Results: Three weeks lentiviral GFP transduction of cells in the lateral ventricle, abundant GFP-expressing neurons and glia were identified in the rostral migratory stream, olfactory bulb and striatum as expected from labeling the subventricular zone (SVZ) type B neural stem cell lineage. Two weeks following mild or severe focal stroke at P7, no GFP-expressing neurons were detected in striatum or cortex although some single-labeled doublecortin+ immature neurons were detected in the penumbra. The densities of GFP+/ glial fibrillary acidic protein (GFAP)+ astrocytes and GFP+/O4+ oligodendrocytes were reduced in the striatum following MCAO (4.8 ± 1.02 vs. 2.5 ± 0.4 cells/high-power field, HPF; p = 0.005; 2.8± 1 vs. 0.5 ± 0.2 cells/HPF, p = 0.008). Furthermore, there was a reduction of GFP+ cells in the olfactory bulb following MCAO (58.8 ± 14.9 vs. 19.6 ± 5.4 cells/HPF, p = 0.025). Finally, there was an increased percentage of GFP+/GFAP+ cells (70 vs. 50%), with a decreased proportion of GFP+/O4+ cells (14 vs. 30%) in injured animals. Conclusion: Neurogenesis originating from cells of the lateral ventricle, including SVZ type B cells, is significantly reduced following neonatal stroke. Furthermore, neonatal stroke disrupts gliogenesis in the striatum, decreasing overall numbers of new glia and shifting the population towards astrocytes.

Neonatal brain injury is an important cause of death and neurodevelopmental delay. Multiple pathways of oxidant stress, inflammation, and excitotoxicity lead to both early and late phases of cell damage and death. Therapies targeting these different pathways have shown potential in protecting the brain from ongoing injury. More recent therapies, such as growth factors, have demonstrated an ability to increase cell proliferation and repair over longer periods of time. Even though hypothermia, which decreases cerebral metabolism and possibly affects other mechanisms, may show some benefit in particular cases, no widely effective therapeutic interventions for human neonates exist. In this review, we summarize recent findings in neuroprotection and neurogenesis for the immature brain, including combination therapy to optimize repair.

Perinatal hypoxic-ischaemic encephalopathy (HIE) occurs in 1 to 3 per 1000 term births. HIE is not preventable in most cases, and therapies are limited. Hypothermia improves outcomes and is the current standard of care. Yet, clinical trials suggest that 44-53% of infants who receive hypothermia will die or suffer moderate to severe neurological disability. In this article, we review the preclinical and clinical evidence for erythropoietin (EPO) as a potential novel neuroprotective agent for the treatment of HIE. EPO is a novel neuroprotective agent, with remarkable neuroprotective and neuroregenerative effects in animals. Rodent and primate models of neonatal brain injury support the safety and efficacy of multiple EPO doses for improving histological and functional outcomes after hypoxia-ischaemia. Small clinical trials of EPO in neonates with HIE have also provided evidence supporting safety and preliminary efficacy in humans. There is currently insufficient evidence to support the use of high-dose EPO in newborns with HIE. However, several on-going trials will provide much needed data regarding the safety and efficacy of this potential new therapy when given in conjunction with hypothermia for HIE. Novel neuroprotective therapies are needed to further reduce the rate and severity of neurodevelopmental disabilities resulting from HIE. High-dose EPO is a promising therapy that can be administered in conjunction with hypothermia. However, additional data are needed to determine the safety and efficacy of this adjuvant therapy for HIE.

STEM CELLS In recent years stem cell therapy has emerged as a potential treatment for neonatal ischemic brain injury. The efficacy of cellbased therapies in restoring damaged brain tissue has been tested in a multitude of models for different CNS diseases. Several different stem and progenitor cell populations have been utilized as cell-based therapy, including neural stem cells, embryonic stem cells, human umbilical cord blood cells (HUBCs), hematopoietic stem and progenitor cells, and mesenchymal stem cells (MSCs). Most stem cell types appear to enhance recovery to some extent (Pimentel-Coelho and Mendez-Otero, 2010). However, because of their low immunogenicity, availability and positive results obtained from preclinical studies, MSCs are a particularly promising candidate to repair the devastating effects that are associated with neonatal stroke. MSCs were first isolated and identified in bone marrow, but can now be isolated from many tissues, including adipose tissue, muscle, skin and extraembryonic tissues like the placenta, umbilical cord and Wharton’s jelly. The latter sources are of particular interest for neonates that experience an ischemic event around the time of birth, at which time cells can be harvested and transplanted from an autologous source. MSCs derived from different sources have slightly different characteristics, but as of yet it is unknown whether this influences their therapeutic potential. Our group and others have shown that administration of MSCs reduces lesion volume, provides positive effects on the white matter and improves motor function (van Velthoven et al., 2012). Numerous studies have been done under the premise that transplanted stem cells contribute to brain repair by directly replacing damaged or lost tissue. While there is evidence that transplanted cells undergo differentiation toward neuronal lineages, improved outcomes have been observed even when survival of transplanted cells is low and engrafted cells are absent. This suggests that rather than replacing damaged cells, transplanted cells may improve outcome via indirect mechanisms. For example, MSCs have been shown to secrete many factors that can influence important processes like apoptosis, neurogenesis, angiogenesis and synaptogenesis.

BackgroundHypoxic-ischemic (HI) injury to the developing brain remains a major cause of morbidity. Hypothermia is effective but does not provide complete neuroprotection, prompting a search for adjunctive therapies. Erythropoietin (Epo) has been shown to be beneficial in several models of neonatal HI. This study examines combination hypothermia and treatment with erythropoietin in neonatal rat HI.MethodsRats at postnatal day 7 were subjected to HI (Vannucci model) and randomized into four groups: no treatment, hypothermia alone, Epo alone, or hypothermia and Epo. Epo (1,000 U/kg) was administered in three doses: immediately following HI, and 24 h and 1 wk later. Hypothermia consisted of whole-body cooling for 8 h. At 2 and 6 wk following HI, sensorimotor function was assessed via cylinder-rearing test and brain damage by injury scoring. Sham-treated animals not subjected to HI were also studied.ResultsDifferences between experimental groups, except for Epo treatment on histopathological outcome in males, were not statistically significant, and combined therapy had no adverse effects.ConclusionNo significant benefit was observed from treatment with either hypothermia or combination therapy. Future studies may require older animals, a wider range of functional assays, and postinsult assessment of injury severity to identify only moderately damaged animals for targeted therapy.

OBJECTIVE: To determine the safety and pharmacokinetics of erythropoietin (Epo) given in conjunction with hypothermia for hypoxic-ischemic encephalopathy (HIE). We hypothesized that high dose Epo would produce plasma concentrations that are neuroprotective in animal studies (ie, maximum concentration = 6000–10 000 U/L; area under the curve = 117 000–140 000 U*h/L). METHODS: In this multicenter, open-label, dose-escalation, phase I study, we enrolled 24 newborns undergoing hypothermia for HIE. All patients had decreased consciousness and acidosis (pH < 7.00 or base deficit ≥ 12), 10-minute Apgar score ≤ 5, or ongoing resuscitation at 10 minutes. Patients received 1 of 4 Epo doses intravenously: 250 ( N = 3), 500 ( N = 6), 1000 ( N = 7), or 2500 U/kg per dose ( N = 8). We gave up to 6 doses every 48 hours starting at

Neonatal brain injury is an important cause of death and disability, with pathways of oxidant stress, inflammation, and excitotoxicity that lead to damage that progresses over a long period of time. Therapies have classically targeted individual pathways during early phases of injury, but more recent therapies such as growth factors may also enhance cell proliferation, differentiation, and migration over time. More recent evidence suggests combined therapy may optimize repair, decreasing cell injury while increasing newly born cells.

Neonatal stroke leads to mortality and severe morbidity, but there currently is no effective treatment. Erythropoietin (EPO) promotes cytoprotection and neurogenesis in the short term following brain injury; however, long-term cognitive outcomes and optimal dosing regimens have not been clarified. We performed middle cerebral artery occlusion in postnatal day 10 rats, which were treated with either a single dose of EPO (5 U/g, i.p.) immediately upon reperfusion, or 3 doses of EPO (1 U/g, i.p. each) at 0 h, 24 h, and 7 days after injury. At 3 months after injury, rats treated with 3 doses of EPO did not differ from shams in the Morris water maze, and generally performed better than either rats treated with a single dose or vehicle-treated injured rats. These multiple-dose-treated rats also had increases in hemispheric volume and its subregions. These results suggest that additional, later doses of EPO may be required for cell repair, proliferation, and long-term incorporation into neural networks after neonatal brain injury.

Some studies on neurodevelopmental outcomes after neonatal encephalopathy have suggested that cognitive deficits do not occur in the absence of cerebral palsy. It is increasingly apparent that childhood survivors of overt neonatal encephalopathy may have cognitive impairments, even in the absence of functional motor deficits. The risk of cognitive deficits is related to the severity of neonatal encephalopathy and the pattern of brain injury on neuroimaging, particularly the watershed pattern of injury. A better understanding of the risk factors for cognitive abnormalities after neonatal encephalopathy will ultimately lead to interventions to prevent these deficits. Identifying the full spectrum of neurodevelopmental outcomes after neonatal encephalopathy will also allow care givers to identify children requiring early intervention to maximise their potential for independent function throughout development.

Neonatal stroke leads to mortality and severe morbidity, but there is no effective treatment currently available. Erythropoietin (EPO) has been shown to promote cytoprotection and neurogenesis and decrease subventricular zone morphologic changes following brain injury. The long-term cellular response to EPO has not been defined, and local changes in cell fate decision may play a role in functional improvement. We performed middle cerebral artery occlusion in P10 rats. EPO treatment (5 U/g IP) significantly preserved hemispheric brain volume 6 weeks after injury. Furthermore, EPO increased the percentage of newly generated neurons while decreasing newly generated astrocytes following brain injury, without demonstrating long-term differences in the subventricular zone. These results suggest that EPO may neuroprotect and direct cell fate toward neurogenesis and away from gliogenesis in neonatal stroke.