Showing papers by "Simon G. Gregory published in 2022"

Lifetime marijuana use and epigenetic age acceleration: A 17-year prospective examination.

Abstract: This study was designed to assess links between lifetime levels of marijuana use and accelerated epigenetic aging.Prospective longitudinal study, following participants annually from age 13 to age 30.A community sample of 154 participants recruited from a small city in the Southeastern United States.Participants completed annual assessments of marijuana use from age 13 to age 29 and provided blood samples that yielded two indices of epigenetic aging (DNAmGrimAge and DunedinPoAm) at age 30. Additional covariates examined included history of cigarette smoking, anxiety and depressive symptoms, childhood illness, gender, adolescent-era family income, and racial/ethnic minority status.Lifetime marijuana use predicted accelerated epigenetic aging, with effects remaining even after covarying cell counts, demographic factors and chronological age (β's = 0.32 & 0.27, p's < 0.001, 95% CI's = 0.21-0.43 & 0.16-0.39 for DNAmGrimAge and DunedinPoAm, respectively). Predictions remained after accounting for cigarette smoking (β's = 0.25 & 0.21, respectively, p's < 0.001, 95% CI's = 0.14-0.37 & 0.09-0.32 for DNAmGrimAge and DunedinPoAm, respectively). A dose-response effect was observed and there was also evidence that effects were dependent upon recency of use. Effects of marijuana use appeared to be fully mediated by hypomethylation of a site linked to effects of hydrocarbon inhalation (cg05575921).Marijuana use predicted epigenetic changes linked to accelerated aging, with evidence suggesting that effects may be primarily due to hydrocarbon inhalation among marijuana smokers. Further research is warranted to explore mechanisms underlying this linkage.

Single Cell RNA-Seq Analysis of Human Red Cells

Abstract: Human red blood cells (RBCs), or erythrocytes, are the most abundant blood cells responsible for gas exchange. RBC diseases affect hundreds of millions of people and impose enormous financial and personal burdens. One well-recognized, but poorly understood feature of RBC populations within the same individual are their phenotypic heterogeneity. The granular characterization of phenotypic RBC variation in normative and disease states may allow us to identify the genetic determinants of red cell diseases and reveal novel therapeutic approaches for their treatment. Previously, we discovered diverse RNA transcripts in RBCs that has allowed us to dissect the phenotypic heterogeneity and malaria resistance of sickle red cells. However, these analyses failed to capture the heterogeneity found in RBC sub-populations. To overcome this limitation, we have performed single cell RNA-Seq to analyze the transcriptional heterogeneity of RBCs from three adult healthy donors which have been stored in the blood bank conditions and assayed at day 1 and day 15. The expression pattern clearly separated RBCs into seven distinct clusters that include one RBC cluster that expresses HBG2 and a small population of RBCs that express fetal hemoglobin (HbF) that we annotated as F cells. Almost all HBG2-expessing cells also express HBB, suggesting bi-allelic expression in single RBC from the HBG2/HBB loci, and we annotated another cluster as reticulocytes based on canonical gene expression. Additional RBC clusters were also annotated based on the enriched expression of NIX, ACVR2B and HEMGN, previously shown to be involved in erythropoiesis. Finally, we found the storage of RBC was associated with an increase in the ACVR2B and F-cell clusters. Collectively, these data indicate the power of single RBC RNA-Seq to capture and discover known and unexpected heterogeneity of RBC population.

Resistance of mitochondrial DNA to cadmium and Aflatoxin B1 damage-induced germline mutation accumulation in C. elegans

Abstract: Abstract Mitochondrial DNA (mtDNA) is prone to mutation in aging and over evolutionary time, yet the processes that regulate the accumulation of de novo mtDNA mutations and modulate mtDNA heteroplasmy are not fully elucidated. Mitochondria lack certain DNA repair processes, which could contribute to polymerase error-induced mutations and increase susceptibility to chemical-induced mtDNA mutagenesis. We conducted error-corrected, ultra-sensitive Duplex Sequencing to investigate the effects of two known nuclear genome mutagens, cadmium and Aflatoxin B1, on germline mtDNA mutagenesis in Caenorhabditis elegans. Detection of thousands of mtDNA mutations revealed pervasive heteroplasmy in C. elegans and that mtDNA mutagenesis is dominated by C:G → A:T mutations generally attributed to oxidative damage. However, there was no effect of either exposure on mtDNA mutation frequency, spectrum, or trinucleotide context signature despite a significant increase in nuclear mutation rate after aflatoxin B1 exposure. Mitophagy-deficient mutants pink-1 and dct-1 accumulated significantly higher levels of mtDNA damage compared to wild-type C. elegans after exposures. However, there were only small differences in mtDNA mutation frequency, spectrum, or trinucleotide context signature compared to wild-type after 3050 generations, across all treatments. These findings suggest mitochondria harbor additional previously uncharacterized mechanisms that regulate mtDNA mutational processes across generations.

The effect of oxytocin nasal spray on social interaction in young children with autism: a randomized clinical trial

Abstract: Abstract Early supports to enhance social development in children with autism are widely promoted. While oxytocin has a crucial role in mammalian social development, its potential role as a medication to enhance social development in humans remains unclear. We investigated the efficacy, tolerability, and safety of intranasal oxytocin in young children with autism using a double-blind, randomized, placebo-controlled, clinical trial, following a placebo lead-in phase. A total of 87 children (aged between 3 and 12 years) with autism received 16 International Units (IU) of oxytocin ( n = 45) or placebo ( n = 42) nasal spray, morning and night (32 IU per day) for twelve weeks, following a 3-week placebo lead-in phase. Overall, there was no effect of oxytocin treatment over time on the caregiver-rated Social Responsiveness Scale (SRS-2) ( p = 0.686). However, a significant interaction with age ( p = 0.028) showed that for younger children, aged 3–5 years, there was some indication of a treatment effect. Younger children who received oxytocin showed improvement on caregiver-rated social responsiveness ( SRS-2). There was no other evidence of benefit in the sample as a whole, or in the younger age group, on the clinician-rated Clinical Global Improvement Scale (CGI-S), or any secondary measure. Importantly, placebo effects in the lead-in phase were evident and there was support for washout of the placebo response in the randomised phase. Oxytocin was well tolerated, with more adverse side effects reported in the placebo group. This study suggests the need for further clinical trials to test the benefits of oxytocin treatment in younger populations with autism. Trial registration www.anzctr.org.au (ACTRN12617000441314).

Genetic and epigenetic signatures associated with plasma oxytocin levels in children and adolescents with autism spectrum disorder

Abstract: Oxytocin (OT), the brain's most abundant neuropeptide, plays an important role in social salience and motivation. Clinical trials of the efficacy of OT in autism spectrum disorder (ASD) have reported mixed results due in part to ASD's complex etiology. We investigated whether genetic and epigenetic variation contribute to variable endogenous OT levels that modulate sensitivity to OT therapy. To carry out this analysis, we integrated genome‐wide profiles of DNA‐methylation, transcriptional activity, and genetic variation with plasma OT levels in 290 participants with ASD enrolled in a randomized controlled trial of OT. Our analysis identified genetic variants with novel association with plasma OT, several of which reside in known ASD risk genes. We also show subtle but statistically significant association of plasma OT levels with peripheral transcriptional activity and DNA‐methylation profiles across several annotated gene sets. These findings broaden our understanding of the effects of the peripheral oxytocin system and provide novel genetic candidates for future studies to decode the complex etiology of ASD and its interaction with OT signaling and OT‐based interventions.

Sex-dimorphic gene effects on survival outcomes in people with coronary artery disease

Abstract: Ischemic coronary heart disease (IHD) is the leading cause of death worldwide. Genetic variation is presumed to be a major factor underlying sex differences for IHD events, including mortality. The purpose of this study was to identify sex-specific candidate genes associated with all-cause mortality among people diagnosed with coronary artery disease (CAD). We performed a sex-stratified, exploratory genome-wide association (GWAS) screen using existing data from CAD-diagnosed males (n = 510) and females (n = 174) who reported European ancestry from the Duke Catheterization Genetics biorepository. Extant genotype data for 785,945 autosomal SNPs generated with the Human Omni1-Quad BeadChip (Illumina, CA, USA) were analyzed using an additive inheritance model. We estimated instantaneous risk of all-cause mortality by genotype groups across the 11-year follow-up using Cox multivariate regression, covarying for age and genomic ancestry. The top GWAS hits associated with all-cause mortality among people with CAD included 8 SNPs among males and 15 among females (p = 1 × 10−6 or 10−7), adjusted for covariates. Cross-sex comparisons revealed distinct candidate genes. Biologically relevant candidates included rs9932462 (EMP2/TEKT5) and rs2835913 (KCNJ6) among males and rs7217169 (RAP1GAP2), rs8021816 (PRKD1), rs8133010 (PDE9A), and rs12145981 (LPGAT1) among females. We report 20 sex-specific candidate genes having suggestive association with all-cause mortality among CAD-diagnosed subjects. Findings demonstrate proof of principle for identifying sex-associated genetic factors that may help explain differential mortality risk in people with CAD. Replication and meta-analyses in larger studies with more diverse samples will strengthen future work in this area.

Hepatocyte Smoothened Activity Controls Susceptibility to Insulin Resistance and Nonalcoholic Fatty Liver Disease

Abstract: Nonalcoholic steatohepatitis (NASH), a leading cause of cirrhosis, strongly associates with the metabolic syndrome, an insulin-resistant proinflammatory state that disrupts energy balance and promotes progressive liver degeneration. We aimed to define the role of Smoothened (Smo), an obligatory component of the Hedgehog signaling pathway, in controlling hepatocyte metabolic homeostasis and, thereby, susceptibility to NASH.We conditionally deleted Smo in hepatocytes of healthy chow-fed mice and performed metabolic phenotyping, coupled with single-cell RNA sequencing (RNA-seq), to characterize the role of hepatocyte Smo in regulating basal hepatic and systemic metabolic homeostasis. Liver RNA-seq datasets from 2 large human cohorts were also analyzed to define the relationship between Smo and NASH susceptibility in people.Hepatocyte Smo deletion inhibited the Hedgehog pathway and promoted fatty liver, hyperinsulinemia, and insulin resistance. We identified a plausible mechanism whereby inactivation of Smo stimulated the mTORC1-SREBP1c signaling axis, which promoted lipogenesis while inhibiting the hepatic insulin cascade. Transcriptomics of bulk and single Smo-deficient hepatocytes supported suppression of insulin signaling and also revealed molecular abnormalities associated with oxidative stress and mitochondrial dysfunction. Analysis of human bulk RNA-seq data revealed that Smo expression was (1) highest in healthy livers, (2) lower in livers with NASH than in those with simple steatosis, (3) negatively correlated with markers of insulin resistance and liver injury, and (4) declined progressively as fibrosis severity worsened.The Hedgehog pathway controls insulin sensitivity and energy homeostasis in adult livers. Loss of hepatocyte Hedgehog activity induces hepatic and systemic metabolic stress and enhances susceptibility to NASH by promoting hepatic lipoxicity and insulin resistance.

Abstract 646: Liquid biopsy transcriptomics identify pathways associated with poor outcomes and immune phenotypes in men with mCRPC

Abstract: Androgen receptor signaling inhibitors (ARSi) and taxanes are mainstays for patients with metastatic castration-resistant prostate cancer (mCRPC). However, patient response is heterogeneous, and the molecular underpinnings of treatment resistance are not well elucidated. To identify clinically meaningful mechanisms of treatment resistance, we performed transcriptome analysis of circulating tumor cells (CTCs) isolated from mCRPC patients enrolled in two independent prospective clinical trials: PROPHECY, a clinical study of patients (n=118) treated with abiraterone or enzalutamide followed by docetaxel; and TAXYNERGY where patients were randomized to docetaxel or cabazitaxel treatment. CTCs were obtained at baseline (before treatment), on treatment and at progression and their comprehensive transcriptomic analysis was correlated with clinical outcomes. To uncover potential involvement of the circulating immune macroenvironment (CIME) in treatment resistance, we performed transcriptomic analysis of matching peripheral blood mononuclear cells (PBMCs) using an established, rigorous, blood-derived transcriptional modular framework. In PROPHECY, CTC RNA-seq identified that RB loss concurrently with enhanced E2F signaling networks were associated with intrinsic ARSi resistance. Using single sample GSEA (ssGSEA) score, we identified that the RB/E2F common signature at baseline was associated with short PFS (median PFS=6.5 months) and OS (median OS=24.5 months) (hazard ratio (HR) = 3.5; 95% CI 1.5-8.2) in men with mCRPC. We further developed a BRCA-loss transcriptional signature, and validated it in the SU2C mCRPC patient cohort, expanding the identification of patients with BRCA-loss phenotypes beyond genomic loss. Applying this signature to PROPHECY baseline samples, we showed that men with high BRCA-loss scores experienced shorter OS (HR=2.42; 95% CI=1-5.9). Through the comparison of CTC transcriptomic profiles at progression with baseline, we identified an inflammatory response signature in CTCs which was significantly associated with acquired ARSi resistance. Transcriptomic PBMC analysis further identified enrichment of inflammasome gene signatures at progression, with concurrent downregulation of CD8+ T and NK cells. Furthermore, preliminary data from both clinical trials, showed a significant upregulation of TGF-β1 and corresponding TGFβ-Receptor signaling pathway in CTCs from patients at progression following taxane treatment, suggesting a role for TGFβ pathway in clinical response to taxane chemotherapy. Taken together, these data demonstrate that liquid biopsy transcriptomics of both tumor cells and immune cells can identify molecular pathways associated with treatment resistance paving the way for treatment optimization and the development of novel precision therapies in patients with mCRPC. Citation Format: Jiaren Zhang, Bob Zimmermann, Giuseppe Galletti, Susan Halabi, Ada Gjyrezi, Qian Yang, Santosh Gupta, Akanksha Verma, Andrea Sboner, Monika Anand, Daniel J. George, Simon G. Gregory, Prerna Mahtani, Seunghee Hong, Virginia Pascual, Clio P. Mavragani, Emmanuel S. Antonarakis, David M. Nanus, Scott T. Tagawa, Olivier Elemento, Andrew J. Armstrong, Paraskevi Giannakakou. Liquid biopsy transcriptomics identify pathways associated with poor outcomes and immune phenotypes in men with mCRPC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 646.

Association of circulating tumor cell RB1 loss RNA signature with outcomes and immune phenotypes in men with mCRPC.

Abstract: 139 Background: Androgen receptor signaling inhibitors (ARSi) are a mainstay for patients with metastatic castration-resistant prostate cancer (mCRPC). However, patient response is heterogeneous and the molecular underpinnings of ARSi resistance are not well elucidated. Methods: We performed transcriptome analysis of circulating tumor cells (CTCs) and peripheral blood mononuclear cells (PBMC) in the context of PROPHECY, a prospective clinical trial of men (n = 118) with mCRPC treated with abiraterone (Abi) or enzalutamide (Enza). We obtained CTCs at baseline (before treatment) and at the time of progression on Abi/Enza, performed a comprehensive transcriptomic analysis of CTC patient samples (n = 40) and correlated with clinical outcomes to identify mechanisms of ARSi resistance. In addition, we also performed a transcriptomic analysis of matching peripheral blood mononuclear cells (PBMCs) in order to uncover potential involvement of the circulating immune macroenvironment (CIME) in ARSi resistance. The proportional hazard model was used to determine the prognostic significance of these signatures in predicting overall survival (OS) and progression-free survival (PFS). Results: CTC RNA-sequencing identified that RB loss concurrently with enhanced E2F signaling transcriptional networks were associated with intrinsic ARSi resistance. Using single sample GSEA (ssGSEA) score, we identified that the RB/E2F common signature at baseline was associated with short PFS (median PFS = 6.5 months) and OS (median OS = 24.5 months) (hazard ratio (HR) = 3.5; 95% CI 1.5-8.2) in men with mCRPC. We further developed a BRCA loss transcriptional signature which we validated in the SU2C mCRPC patient cohort, by showing that BRCA loss transcriptional network reflected BRCA genomic alterations as it was significantly enriched in the SU2C BRCA-altered patients vs unaltered patients. Generating BRCA loss ssGSEA scores in the PROPHECY cohort we observed that patients with high BRCA loss scores at baseline experienced shorter OS (HR = 2.42; 95% CI = 1-5.9). Through the comparison of CTC transcriptomic profiles at progression with baseline, we identified an inflammatory response signature in CTCs which was significantly associated with acquired ARSi resistance. Transcriptomic analysis of matching PBMCs identified enrichment of inflammasome gene signatures indicative of activated innate immunity at progression, with concurrent downregulation of CD8 T and NK cells. Importantly, CTC gene signatures of RB loss/E2F signaling had a significant positive association with this CIME signatures. Conclusions: Taken together, these data demonstrate that liquid biopsy transcriptomics of both tumor cells and immune cells can identify molecular pathways associated with clinical ARSi resistance paving the way for treatment optimization and the development of novel precision therapies in patients with mCRPC.

Aging and Obesity Prime the Methylome and Transcriptome of Adipose Stem Cells for Disease and Dysfunction

Abstract: The epigenome of stem cells occupies a critical interface between genes and environment, serving to regulate expression through modification by intrinsic and extrinsic factors. We hypothesized that aging and obesity, which represent major risk factors for a variety of diseases, synergistically modify the epigenome of adult adipose stem cells (ASCs). Using integrated RNA- and targeted bisulfite-sequencing in murine ASCs from lean and obese mice at 5- and 12- months of age, we identified global DNA hypomethylation with either aging or obesity, and a synergistic effect of aging combined with obesity. The transcriptome of ASCs in lean mice was relatively stable to the effects of age, but this was not true in obese mice. Functional pathway analyses identified a subset of genes with critical roles in progenitors and in diseases of obesity and aging. Specifically, Mapt, Nr3c2, App, and Ctnnb1 emerged as potential hypomethylated upstream regulators in both aging and obesity (AL vs YL and AO vs YO), and App, Ctnnb1, Hipk2, Id2, and Tp53 exhibited additional effects of aging in obese animals. Further, Foxo3 and Ccnd1 were potential hypermethylated upstream regulators of healthy aging (AL vs YL), and of the effects of obesity in young animals (YO vs YL), suggesting that these factors could play a role in accelerated aging with obesity. Finally, we identified candidate driver genes that appeared recurrently in all analyses and comparisons undertaken. Further mechanistic studies are needed to validate the roles of these genes capable of priming ASCs for dysfunction in aging- and obesity-associated pathologies.

Adolescent peer struggles predict accelerated epigenetic aging in midlife.

Abstract: This study examined struggles to establish autonomy and relatedness with peers in adolescence and early adulthood as predictors of advanced epigenetic aging assessed at age 30. Participants (N = 154; 67 male and 87 female) were observed repeatedly, along with close friends and romantic partners, from ages 13 through 29. Observed difficulty establishing close friendships characterized by mutual autonomy and relatedness from ages 13 to 18, an interview-assessed attachment state of mind lacking autonomy and valuing of attachment at 24, and self-reported difficulties in social integration across adolescence and adulthood were all linked to greater epigenetic age at 30, after accounting for chronological age, gender, race, and income. Analyses assessing the unique and combined effects of these factors, along with lifetime history of cigarette smoking, indicated that each of these factors, except for adult social integration, contributed uniquely to explaining epigenetic age acceleration. Results are interpreted as evidence that the adolescent preoccupation with peer relationships may be highly functional given the relevance of such relationships to long-term physical outcomes.

Single-cell genome-wide association reveals that a nonsynonymous variant in ERAP1 confers increased susceptibility to influenza virus

Abstract: During pandemics, individuals exhibit differences in risk and clinical outcomes. Here, we developed single-cell high-throughput human in vitro susceptibility testing (scHi-HOST), a method for rapidly identifying genetic variants that confer resistance and susceptibility. We applied this method to influenza A virus (IAV), the cause of four pandemics since the start of the 20th century. scHi-HOST leverages single-cell RNA sequencing (scRNA-seq) to simultaneously assign genetic identity to cells in mixed infections of cell lines of European, African, and Asian origin, reveal associated genetic variants for viral burden, and identify expression quantitative trait loci. Integration of scHi-HOST with human challenge and experimental validation demonstrated that a missense variant in endoplasmic reticulum aminopeptidase 1 (ERAP1; rs27895) increased IAV burden in cells and human volunteers. rs27895 exhibits population differentiation, likely contributing to greater permissivity of cells from African populations to IAV. scHi-HOST is a broadly applicable method and resource for decoding infectious-disease genetics.

Breast milk-associated oxysterol reverses neonatal white matter injury through Gli2-dependent oligodendrogenesis Proceedings of the 13th International Newborn Brain Conference: Fetal and/or neonatal brain development, both normal and abnormal

Abstract: brain injury to neurologic in are novel for is by appropriate for safety. Here, we identi fi ed oxysterols in human maternal breast milk and explored their therapeutic potential in directing neural stem cells (NSCs) into the oligodendrocyte (OL) lineage via the Sonic Hedgehog (Shh) pathway in vitro. Further, we investigated whether breast milk-associated oxysterols can rescue neonatal WMI in vivo. were from the subventricular (SVZ) of and treated with oxysterols. were analyzed by immunohistochemistry, western blot, fl ow cytometry, and scRNAseq, looking for markers of the OL lineage. Shh pathway activation was established by quantifying upregulation of target gene, Gli1, by western blot and RT-PCR. Gli-dependence was explored using pharmacological and genetic approaches. Neonatal sepsis leading to WMI was induced in mice on postnatal day 5, using an intraperitoneal stool injection. then received either breast milk-associated 20-(cid:31)hydroxycholesterol (20HC) or vehicle. Stereology determined OL numbers in the periventricular white matter regions. Myelination was evaluated by performing g-ratios to determine myelin thickness. Motor function in mice was analyzed using the CatWalk gait analysis system. To lineage trace postnatal nestin+ SVZ NSCs in vivo, neonatal sepsis was induced in Nestin-CreERT2;R26r-TdTomato pups as above, and then treated with either vehicle or 20HC. RESULTS: Multiple oxysterols were identi fi ed in human maternal breast milk that induced oligodendrocyte production from NSCs in vitro. We found that Gli2 is functionally required for oxysterol-induced oligodendrogenesis. Following neonatal WMI in vivo, 20HC treatment increased numbers of mature OLs, improved myelination and rescued motor de fi cits in mice. Lineage tracing experiments showed that 20HC-mediated recovery of OL de fi cit is mediated in part through 20HC-induced SVZ-derived oligodendrogenesis in vivo. Additional recovery may be due to the impact of 20HC on oligodendrocyte progenitor cell maturation. BACKGROUND: Preterm infants are born with low cir-culating insulin-like growth factor 1 (IGF-1) levels and in cohort studies, low IGF-1 levels have been associated with morbidities, including poor neurodevelopment (1). It remains unclear if supplemental IGF-1 promotes neurodevelopment in preterm infants (2). Using preterm pigs as a model for preterm infants (3,4), we investigated how supplemental IGF-1, in complex with the binding protein IGFBP3, affects brain maturation in the early postnatal period. METHODS: Preterm-born pigs were treated with 2.25 OBJECTIVE: High rates of neurodevelopmental impairments are associated with sequalae of extreme premature birth, especially necrotizing enterocolitis and bowel perforations. The mechanisms of brain injury in this population are poorly understood. In this study we investigated the impact of a novel injury to the postnatal subventricular zone following modeled bowel perforation. Altered gliogenic and neurogenic functionality of the subventricular zone may contribute to neurologic de fi cits observed in preterm infants. METHODS: Using a clinically relevant rodent model of neonatal bowel perforation leading to polymicrobial sepsis we replicated many features of brain injuries seen among preterm-born infants. These injuries include diffuse white matter injury as well as motor de fi cits. Our experiments focused on neuroimmune responses and histological changes at the subventricular zone (SVZ) neural stem cell niche that resides in the germinal matrix of human preterm infants. qPCR was utilized to measure localized in fl ammatory cytokine outcomes in BACKGROUND AND PURPOSE: Alterations in neurotransmitter systems, particularly changes in GABAergic and glutamatergic systems, during frontal lobe development have been hypothesized to play a key role in neurodevelopmental disorders seen in children born very preterm (<32 weeks gestation) and at very low birth weight [1]. Prematurity frequently results in neurodevelopmental disorders such as autism, attention de fi cit or epilepsy in the absence of radiologically visible lesions, suggesting that long-term physiological alterations might underlie these male and 9 female term infants. Gene expression levels were measured for 47 genes related to GABAergic development and this gene expression data was used to calculate a maturation index. To evaluate the impact of premature birth on the GABAergic system development, samples from one-month-old term (n=9 male, 4 female) and one-month corrected-age (n=8 male, 6 female) very preterm infants, were compared using the same gene list and methodology. RESULTS: In term infants, the maturation index correlated with the age of the donor and was more dynamic in male than in female infants, suggesting a link to the increased male infant susceptibility to impaired neurodevelopment. When comparing term and preterm infants, the maturation index for the GABAergic system was signi fi cantly lower (–50% p<0.05) in male preterm infants, with major alterations in genes linked to GABAergic function in astrocytes. Conclusion/Impact: Taken together, this study suggests that the development of GABAergic system is more dynamic in male infants, potentially contributing to their increased susceptibility to perinatal neurological insults. We have identi fi ed a potential key mechanism in the alteration of astrocytic GABAergic development that we are now investigating. BACKGROUND/PURPOSE: in infants in fl uences INTRODUCTION: Prematurity remains a real public health concern in the world and in Africa, especially in the Democratic Republic of Congo (DRC) due to its high morbidity and mortality rate. Growth and psychomotor outcomes of preterm infant remain worrying. Although several researches have shown the bene fi cial effects of Kangaroo care (KC), few data exist in low-income countries such as the DRC on short-term effects of KC in growth and psychomotor outcomes on premature infants. OBJECTIVES: To assess the bene fi cial effects of KC on growth and psychomotor outcomes in preterm infants in a low-income country. METHODOLOGY: Sixty- fi ve preterm infants who were born in three public hospitals in Kinshasa were included in this study. We included those preterm infants weighted below 2000g, who had ability to have oral alimentation and have cardiovascular stability. We excluded those with congenital abnormalities that may in fl uence the neurodevelopmental outcomes and the growth. Neurodevelopmental outcomes were assessed by the Bayley scale II. Thirty-one preterm infants underwent KC and thirty-four the traditional method (incubators). KC was performed in two of the three public hospitals and traditional care was done in one hospital. The age and weight of the infant, the education and socioeconomic status of the family were matched into the two groups. RESULTS: Among sixty- fi ve preterm infants included in this study, 31 underwent KC and 34 traditional method. The growth and psychomotor outcomes were better in the KC group than in the traditional method group, especially in mental development index (MDI: 91.6±12.1 vs 81.2±12.6, p 0.001). Birth weight was found as a determinant of the growth. Sex, birth age, APGAR at 5th minute, socioeconomic status and nutritional status were found as determinants of a good psychomotor outcome. CONCLUSION: In our setting KC offers more bene fi cial effects to the preterm infant according to growth and psychomotor outcomes than the traditional method. BACKGROUND AND PURPOSE: It is well accepted that insulin-like growth factor 1 (IGF-1) plays a crucial role in brain development. However, previous studies have inferred that the transfer of systemically administered IGF-1 over the blood-brain-barrier is restricted, which would present a limitation for bene fi cial treatment effects within the brain (1). Extracellular vesicles (EVs) are small, cell-derived phospholipid membrane enclosed vesicles that constitute important cell-to-cell messengers, thereby regu-lating diverse cellular functions of recipient cells (2). EVs released from the choroid plexus (CP) to intraventricular cerebrospinal fl uid (CSF) have been shown to cross the ependyme and exert a regulatory function in periventricular brain tissue (3). We hypothesized that exposure of the CP to blood-born IGF-1 induces release of EVs into intraventricular CSF destined to affect the surrounding brain parenchyma. We evaluated this in a primary culture of CP epithelial (CPE) cells using a Transwell in vitro model. METHODOLOGY: CPE cells were collected from P3-P9 mice pups and plated on Transwell membranes. Cells were stimulated with 40 ng/ml IGF-1 at the basal (blood) side for 24 hours, and subsequently the apical supernatant was collected, and cells were fi xed. BACKGROUND AND PURPOSE: Healthy development of the central nervous system (CNS) that controls respiration is crucial for the transition from a fetus to a neonate, especially for preterm neonates.1 Fetal alcohol spectrum disorder (FASD) has been linked to numerous poor outcomes of the CNS, such as microcephaly, severe learning disorders and sudden infant death syndrome (SIDS).2 SIDS is thought to be caused by abnormal respiratory control.3 In this study, we investigated whether ethanol exposure disrupts various aspects of respiratory neural control. Speci fi cally, using a rodent model of preterm neonates, we hypothesized that ethanol exposure would alter the hypoxic and/or the hypercapnic drive in 5-day-old (P5) rats, compared to saline treated animals. METHODOLOGY: P5 Sprague-Dawley rat pups were treated with 0.8 or 4.4 mg/g of ethanol delivered via IP injection, a validated model of FASD4,5,6. Pups’ hypoxic and hypercapnic ventilatory responses were measured using plethysmography. Individual pups were allowed to acclimatize in the plethysmography chamber for 1 h, while normoxic air fl owed through the chamber. A baseline minute ventilation was recorded at 1 h, pups were then subjected to hypoxia (10% O2) fo