Influence of Eat, Sleep, and Console on Infants Pharmacologically Treated for Opioid Withdrawal: A Post Hoc Subgroup Analysis of the ESC-NOW Randomized Clinical Trial.

Importance: The function-based eat, sleep, console (ESC) care approach substantially reduces the proportion of infants who receive pharmacologic treatment for neonatal opioid withdrawal syndrome (NOWS). This reduction has led to concerns for increased postnatal opioid exposure in infants who receive pharmacologic treatment. However, the effect of the ESC care approach on hospital outcomes for infants pharmacologically treated for NOWS is currently unknown. Objective: To evaluate differences in opioid exposure and total length of hospital stay (LOS) for pharmacologically treated infants managed with the ESC care approach vs usual care with the Finnegan tool. Design, Setting, and Participants: This post hoc subgroup analysis involved infants pharmacologically treated in ESC-NOW, a stepped-wedge cluster randomized clinical trial conducted at 26 US hospitals. Hospitals maintained pretrial practices for pharmacologic treatment, including opioid type, scheduled opioid dosing, and use of adjuvant medications. Infants were born at 36 weeks' gestation or later, had evidence of antenatal opioid exposure, and received opioid treatment for NOWS between September 2020 and March 2022. Data were analyzed from November 2022 to January 2024. Exposure: Opioid treatment for NOWS and the ESC care approach. Main Outcomes and Measures: For each outcome (total opioid exposure, peak opioid dose, time from birth to initiation of first opioid dose, length of opioid treatment, and LOS), we used generalized linear mixed models to adjust for the stepped-wedge design and maternal and infant characteristics. Results: In the ESC-NOW trial, 463 of 1305 infants were pharmacologically treated (143/603 [23.7%] in the ESC care approach group and 320/702 [45.6%] in the usual care group). Mean total opioid exposure was lower in the ESC care approach group with an absolute difference of 4.1 morphine milligram equivalents per kilogram (MME/kg) (95% CI, 1.3-7.0) when compared with usual care (4.8 MME/kg vs 8.9 MME/kg, respectively; P = .001). Mean time from birth to initiation of pharmacologic treatment was 22.4 hours (95% CI, 7.1-37.7) longer with the ESC care approach vs usual care (75.4 vs 53.0 hours, respectively; P = .002). No significant difference in mean peak opioid dose was observed between groups (ESC care approach, 0.147 MME/kg, vs usual care, 0.126 MME/kg). The mean length of treatment was 6.3 days shorter (95% CI, 3.0-9.6) in the ESC care approach group vs usual care group (11.8 vs 18.1 days, respectively; P < .001), and mean LOS was 6.2 days shorter (95% CI, 3.0-9.4) with the ESC care approach than with usual care (16.7 vs 22.9 days, respectively; P < .001). Conclusion and Relevance: When compared with usual care, the ESC care approach was associated with less opioid exposure and shorter LOS for infants pharmacologically treated for NOWS. The ESC care approach was not associated with a higher peak opioid dose, although pharmacologic treatment was typically initiated later. Trial Registration: ClinicalTrials.gov Identifier: NCT04057820.

Eat, Sleep, Console Approach or Usual Care for Neonatal Opioid Withdrawal.

BACKGROUND: Although clinicians have traditionally used the Finnegan Neonatal Abstinence Scoring Tool to assess the severity of neonatal opioid withdrawal, a newer function-based approach - the Eat, Sleep, Console care approach - is increasing in use. Whether the new approach can safely reduce the time until infants are medically ready for discharge when it is applied broadly across diverse sites is unknown. METHODS: In this cluster-randomized, controlled trial at 26 U.S. hospitals, we enrolled infants with neonatal opioid withdrawal syndrome who had been born at 36 weeks' gestation or more. At a randomly assigned time, hospitals transitioned from usual care that used the Finnegan tool to the Eat, Sleep, Console approach. During a 3-month transition period, staff members at each hospital were trained to use the new approach. The primary outcome was the time from birth until medical readiness for discharge as defined by the trial. Composite safety outcomes that were assessed during the first 3 months of postnatal age included in-hospital safety, unscheduled health care visits, and nonaccidental trauma or death. RESULTS: A total of 1305 infants were enrolled. In an intention-to-treat analysis that included 837 infants who met the trial definition for medical readiness for discharge, the number of days from birth until readiness for hospital discharge was 8.2 in the Eat, Sleep, Console group and 14.9 in the usual-care group (adjusted mean difference, 6.7 days; 95% confidence interval [CI], 4.7 to 8.8), for a rate ratio of 0.55 (95% CI, 0.46 to 0.65; P<0.001). The incidence of adverse outcomes was similar in the two groups. CONCLUSIONS: As compared with usual care, use of the Eat, Sleep, Console care approach significantly decreased the number of days until infants with neonatal opioid withdrawal syndrome were medically ready for discharge, without increasing specified adverse outcomes. (Funded by the Helping End Addiction Long-term (HEAL) Initiative of the National Institutes of Health; ESC-NOW ClinicalTrials.gov number, NCT04057820.).

Effects of intrauterine growth restriction on embryonic hippocampal dentate gyrus neurogenesis and postnatal critical period of synaptic plasticity that govern learning and memory function.

Intrauterine growth restriction (IUGR) complicates up to 10% of human pregnancies and is the second leading cause of perinatal morbidity and mortality after prematurity. The most common etiology of IUGR in developed countries is uteroplacental insufficiency (UPI). For survivors of IUGR pregnancies, long-term studies consistently show a fivefold increased risk for impaired cognition including learning and memory deficits. Among these, only a few human studies have highlighted sex differences with males and females having differing susceptibilities to different impairments. Moreover, it is well established from brain magnetic resonance imaging that IUGR affects both white and gray matter. The hippocampus, composed of the dentate gyrus (DG) and cornu ammonis (CA) subregions, is an important gray matter structure critical to learning and memory, and is particularly vulnerable to the chronic hypoxic-ischemic effects of UPI. Decreased hippocampal volume is a strong predictor for learning and memory deficits. Decreased neuron number and attenuated dendritic and axonal morphologies in both the DG and CA are additionally seen in animal models. What is largely unexplored is the prenatal changes that predispose an IUGR offspring to postnatal learning and memory deficits. This lack of knowledge will continue to hinder the design of future therapy to improve learning and memory. In this review, we will first present the clinical susceptibilities and human epidemiology data regarding the neurological sequelae after IUGR. We will follow with data generated using our laboratory's mouse model of IUGR, that mimics the human IUGR phenotype, to dissect at the cellular and molecular alterations in embryonic hippocampal DG neurogenesis. We will lastly present a newer topic of postnatal neuron development, namely the critical period of synaptic plasticity that is crucial in achieving an excitatory/inhibitory balance in the developing brain. To our knowledge, these findings are the first to describe the prenatal changes that lead to an alteration in postnatal hippocampal excitatory/inhibitory imbalance, a mechanism that is now recognized to be a cause of neurocognitive/neuropsychiatric disorders in at-risk individuals. Studies are ongoing in our laboratory to elucidate additional mechanisms that underlie IUGR-induced learning and memory impairment and to design therapy aimed at ameliorating such impairment.

Hospital Outcomes of Infants with Neonatal Opioid Withdrawal Syndrome at a Tertiary Care Hospital with High Rates of Concurrent Nonopioid (Polysubstance) Exposure.

OBJECTIVE: Neonatal opioid withdrawal syndrome (NOWS) describes infants' withdrawal signs and symptoms after birth due to an interruption of prenatal opioid exposure. Many infants with NOWS are also exposed to nonopioids, however. This study was to determine hospital outcomes of infants exposed to opioids alone or coexposed with nonopioid substances (polysubstance). STUDY DESIGN: We reviewed infants of ≥34 weeks of gestation with prenatal opioid exposure from April 2015 to May 2018. We compared the median lengths of stay (LOS) and treatment (LOT) and the percentages of infants requiring pharmacologic and adjunctive treatment in infants exposed to opioids alone or polysubstance. We used Wilcoxon's test for continuous outcomes or Chi-squared test for categorical outcomes to determine statistical significance. We used multivariable regression model to calculate each drug category's estimates of adjusted mean ratios for LOS and LOT plus estimates of adjusted odds ratios for pharmacologic/adjunctive treatments. RESULTS: Of the 175 infants, 33 (19%) infants had opioid exposure alone. Opioid exposure included short- and/or long-acting opioids. A total of 142 (81%) had polysubstance exposure with 47% of mothers using nicotine products. We saw similar hospital outcomes between infants exposed to opioids alone or polysubstance; however, a higher percentage of infants with both short- and long-acting opioid exposure required pharmacologic treatment compared with either opioid alone. Focusing on individual drug categories, we detected differential hospital outcomes in which short-acting opioids decreased LOT, whereas long-acting opioids increased LOS, LOT, and need for pharmacologic and adjunctive treatment. Coexposure of opioids with stimulants decreased LOT and reduced need for adjunctive treatment. Coexposures with antidepressants increased LOT, while with antiepilepetics increased LOS. CONCLUSION: Because infants with NOWS often have coexposures to other nonopioid substances, appreciating the associated risks of individual or combination of drugs in modulating hospital outcomes may help counsel families on their infants' expected hospital course. KEY POINTS: · Hospital outcomes were similar between infants exposed to opioids alone or polysubstance including opioids.. · Infants with short- and long-acting opioids required pharmacologic treatment more often than either opioid alone.. · Differential hospital outcomes exist for various co-exposures of opioids with nonopioids..

Intrauterine Growth Restriction Causes Abnormal Embryonic Dentate Gyrus Neurogenesis in Mouse Offspring That Leads to Adult Learning and Memory Deficits.

Human infants who suffer from intrauterine growth restriction (IUGR), which is a failure to attain their genetically predetermined weight, are at increased risk for postnatal learning and memory deficits. Hippocampal dentate gyrus (DG) granule neurons play an important role in memory formation; however, it is unknown whether IUGR affects embryonic DG neurogenesis, which could provide a potential mechanism underlying abnormal postnatal learning and memory function. Using a mouse model of the most common cause of IUGR, induced by hypertensive disease of pregnancy, we first assessed adult learning and memory function. We quantified the percentages of embryonic hippocampal DG neural stem cells (NSCs) and progenitor cells and developing glutamatergic granule neurons, as well as hippocampal volumes and neuron cell count and morphology 18 and 40 d after delivery. We characterized the differential embryonic hippocampal transcriptomic pathways between appropriately grown and IUGR mouse offspring. We found that IUGR offspring of both sexes had short-term adult learning and memory deficits. Prenatally, we found that IUGR caused accelerated embryonic DG neurogenesis and Sox2+ neural stem cell depletion. IUGR mice were marked by decreased hippocampal volumes and decreased doublecortin+ neuronal progenitors with increased mean dendritic lengths at postnatal day 18. Consistent with its known molecular role in embryonic DG neurogenesis, we also found evidence for decreased Wnt pathway activity during IUGR. In conclusion, we have discovered that postnatal memory deficits are associated with accelerated NSC differentiation and maturation into glutamatergic granule neurons following IUGR, a phenotype that could be explained by decreased embryonic Wnt signaling.

Intrauterine growth restriction followed by oxygen support uniquely interferes with genetic regulators of myelination.

Intrauterine growth restriction (IUGR) and oxygen exposure in isolation and combination adversely affect the developing brain, putting infants at risk for neurodevelopmental disability including cerebral palsy. Rodent models of IUGR and postnatal hyperoxia have demonstrated oligodendroglial injury with subsequent white matter injury (WMI) and motor dysfunction. Here we investigate transcriptomic dysregulation in IUGR with and without hyperoxia exposure to account for the abnormal brain structure and function previously documented. We performed RNA sequencing and analysis using a mouse model of IUGR and found that IUGR, hyperoxia, and the combination of IUGR with hyperoxia (IUGR/hyperoxia) produced distinct changes in gene expression. IUGR in isolation demonstrated the fewest differentially expressed genes compared to control. In contrast, we detected several gene alterations in IUGR/hyperoxia; genes involved in myelination were strikingly downregulated. We also identified changes to specific regulators including TCF7L2, BDNF, SOX2, and DGCR8, through Ingenuity Pathway Analysis, that may contribute to impaired myelination in IUGR/hyperoxia. Our findings show that IUGR with hyperoxia induces unique transcriptional changes in the developing brain. These indicate mechanisms for increased risk for WMI in IUGR infants exposed to oxygen and suggest potential therapeutic targets to improve motor outcomes.Significance StatementThis study demonstrates that perinatal exposures of IUGR and/or postnatal hyperoxia result in distinct transcriptomic changes in the developing brain. In particular, we found that genes involved in normal developmental myelination, myelin maintenance, and remyelination were most dysregulated when IUGR was combined with hyperoxia. Understanding how multiple risk factors lead to WMI is the first step in developing future therapeutic interventions. Additionally, because oxygen exposure is often unavoidable after birth, an understanding of gene perturbations in this setting will increase our awareness of the need for tight control of oxygen use to minimize future motor disability.

Effects of excess thromboxane A2 on placental development and nutrient transporters in a Mus musculus model of fetal growth restriction.

Hypertensive disease of pregnancy (HDP) with placental insufficiency is the most common cause of fetal growth restriction (FGR) in the developed world. Despite the known negative consequences of HDP both to the mother and fetus, little is known about the longitudinal placental changes that occur as HDP progresses in pregnancy. This is because longitudinal sampling of human placentae during each gestation is impossible. Therefore, using a mouse model of thromboxane A2-analog infusion to mimic human HDP in the last trimester, we calculated placental efficiencies based on fetal and placental weights; quantified spongiotrophoblast and labyrinth thicknesses and vascular density within these layers; examined whether hypoxia signaling pathway involving vascular endothelial growth factor A (VEGFA) and its receptors (VEGFR1, VEGFR2) and matrix metalloproteinases (MMPs) contributed to vascular change; and examined nutrient transporter abundance including glucose transporters 1 and 3 (GLUT1, GLUT3), neutral amino acid transporters 1, 2, and 4 (SNAT1, SNAT2, and SNAT4), fatty acid transporters 2 and 4 (FATP2, FATP4), and fatty acid translocase (CD36) from embryonic day 15.5 to 19 in a 20-day C57Bl/6J mouse gestation. We conclude that early-to-mid gestation hypertensive placentae show compensatory mechanisms to preserve fetal growth by increasing placental efficiencies and maintaining abundance of important nutrient transporters. As placental vascular network diminishes over late hypertension, placental efficiency diminishes and fetal growth fails. Neither hypoxia signaling pathway nor MMPs mediated the vascular diminution in this model. Hypertensive placentae surprisingly exhibit a sex-differential expression of nutrient transporters in late gestation despite showing fetal growth failure in both sexes.

Lef1-dependent hypothalamic neurogenesis inhibits anxiety.

While innate behaviors are conserved throughout the animal kingdom, it is unknown whether common signaling pathways regulate the development of neuronal populations mediating these behaviors in diverse organisms. Here, we demonstrate that the Wnt/ß-catenin effector Lef1 is required for the differentiation of anxiolytic hypothalamic neurons in zebrafish and mice, although the identity of Lef1-dependent genes and neurons differ between these 2 species. We further show that zebrafish and Drosophila have common Lef1-dependent gene expression in their respective neuroendocrine organs, consistent with a conserved pathway that has diverged in the mouse. Finally, orthologs of Lef1-dependent genes from both zebrafish and mouse show highly correlated hypothalamic expression in marmosets and humans, suggesting co-regulation of 2 parallel anxiolytic pathways in primates. These findings demonstrate that during evolution, a transcription factor can act through multiple mechanisms to generate a common behavioral output, and that Lef1 regulates circuit development that is fundamentally important for mediating anxiety in a wide variety of animal species.

Intrauterine growth restriction inhibits expression of eukaryotic elongation factor 2 kinase, a regulator of protein translation.

Nutrient deprivation suppresses protein synthesis by blocking peptide elongation. Transcriptional upregulation and activation of eukaryotic elongation factor 2 kinase (eEF2K) blocks peptide elongation by phosphorylating eukaryotic elongation factor 2. Previous studies examining placentas from intrauterine growth restricted (IUGR) newborn infants show decreased eEF2K expression and activity despite chronic nutrient deprivation. However, the effect of IUGR on hepatic eEF2K expression in the fetus is unknown. We, therefore, examined the transcriptional regulation of hepatic eEF2K gene expression in a Sprague-Dawley rat model of IUGR. We found decreased hepatic eEF2K mRNA and protein levels in IUGR offspring at birth compared with control, consistent with previous placental observations. Furthermore, the CpG island within the eEF2K promoter demonstrated increased methylation at a critical USF 1/2 transcription factor binding site. In vitro methylation of this binding site caused near complete loss of eEF2K promoter activity, designating this promoter as methylation sensitive. The eEF2K promotor in IUGR offspring also lost the protective histone covalent modifications associated with unmethylated CGIs. In addition, the +1 nucleosome was displaced 3' and RNA polymerase loading was reduced at the IUGR eEF2K promoter. Our findings provide evidence to explain why IUGR-induced chronic nutrient deprivation does not result in the upregulation of eEF2K gene transcription.

Intrauterine Growth Restriction Alters Mouse Intestinal Architecture during Development.

Infants with intrauterine growth restriction (IUGR) are at increased risk for neonatal and lifelong morbidities affecting multiple organ systems including the intestinal tract. The underlying mechanisms for the risk to the intestine remain poorly understood. In this study, we tested the hypothesis that IUGR affects the development of goblet and Paneth cell lineages, thus compromising the innate immunity and barrier functions of the epithelium. Using a mouse model of maternal thromboxane A2-analog infusion to elicit maternal hypertension and resultant IUGR, we tested whether IUGR alters ileal maturation and specifically disrupts mucus-producing goblet and antimicrobial-secreting Paneth cell development. We measured body weights, ileal weights and ileal lengths from birth to postnatal day (P) 56. We also determined the abundance of goblet and Paneth cells and their mRNA products, localization of cellular tight junctions, cell proliferation, and apoptosis to interrogate cellular homeostasis. Comparison of the murine findings with human IUGR ileum allowed us to verify observed changes in the mouse were relevant to clinical IUGR. At P14 IUGR mice had decreased ileal lengths, fewer goblet and Paneth cells, reductions in Paneth cell specific mRNAs, and decreased cell proliferation. These findings positively correlated with severity of IUGR. Furthermore, the decrease in murine Paneth cells was also seen in human IUGR ileum. IUGR disrupts the normal trajectory of ileal development, particularly affecting the composition and secretory products of the epithelial surface of the intestine. We speculate that this abnormal intestinal development may constitute an inherent "first hit", rendering IUGR intestine susceptible to further injury, infection, or inflammation.

Intrauterine growth restriction perturbs nucleosome depletion at a growth hormone-responsive element in the mouse IGF-1 gene.

Intrauterine growth restriction (IUGR) is a common human pregnancy complication. IUGR offspring carry significant postnatal risk for early-onset metabolic syndrome, which is associated with persistent reduction in IGF-1 protein expression. We have previously shown that preadolescent IUGR male mice have decreased hepatic IGF-1 mRNA and circulating IGF-1 protein at postnatal day 21, the age when growth hormone (GH) normally upregulates hepatic IGF-1 expression. Here we studied nucleosome occupancy and CpG methylation at a putative growth hormone-responsive element in intron 2 (in2GHRE) of the hepatic IGF-1 gene in normal, sham-operated, and IUGR mice. Nucleosome occupancy and CpG methylation were determined in embryonic stem cells (ESCs) and in liver at postnatal days 14, 21, and 42. For CpG methylation, additional time points out to 2 yr were analyzed. We confirmed the putative mouse in2GHRE was GH-responsive, and in normal mice, a single nucleosome was displaced from the hepatic in2GHRE by postnatal day 21, which exposed two STAT5b DNA binding sites. Nucleosome displacement correlated with developmentally programmed CpG demethylation. Finally, IUGR significantly altered the nucleosome-depleted region (NDR) at the in2GHRE of IGF-1 on postnatal day 21, with either complete absence of the NDR or with a shifted NDR exposing only one of two STAT5b DNA binding sites. An NDR shift was also seen in offspring of sham-operated mothers. We conclude that prenatal insult such as IUGR or anesthesia/surgery could perturb the proper formation of a well-positioned NDR at the mouse hepatic IGF-1 in2GHRE necessary for transitioning to an open chromatin state.

IUGR prevents IGF-1 upregulation in juvenile male mice by perturbing postnatal IGF-1 chromatin remodeling.

BACKGROUND: Intrauterine growth restriction (IUGR) offspring with rapid catch-up growth are at increased risk for early obesity especially in males. Persistent insulin-like growth factor-1 (IGF-1) reduction is an important risk factor. Using a mouse model of maternal hypertension-induced IUGR, we examined IGF-1 levels, promoter DNA methylation, and histone H3 covalent modifications at birth (D1). We additionally investigated whether prenatal perturbations could reset at preadolescence (D21). METHODS: IUGR was induced via maternal thromboxane A2-analog infusion in mice. RESULTS: IUGR uniformly decreased D1 IGF-1 mRNA and protein levels with reduced promoter 1 (P1) transcription and increased P1 DNA methylation. IUGR males also had increased H3K4ac at exon 5 and 3' distal UTR. At D21, IUGR males continued to have decreased IGF-1 levels, originating from both P1 and P2 with reduced 1A variant. IUGR males also had decreased activation mark of H3K4me3 at P1 compared with sham males. In contrast, D21 IUGR females normalized their IGF-1 levels, in association with an increased activation mark of H3K4me3 at P1 compared with sham females. CONCLUSION: IUGR uniformly affected D1 hepatic IGF-1 epigenetic modifications in both sexes. However, at preadolescence, IUGR males are unable to correct for the prenatal reduction possibly due to a more perturbed IGF-1 chromatin structure.