Human Preterm Neonates Research Articles

Ontogeny of CYP3A and UGT activity in preterm piglets: a translational model for drug metabolism in preterm newborns.

Despite considerable progress in understanding drug metabolism in the human pediatric population, data remains scarce in preterm neonates. Improving our knowledge of the ADME properties in this vulnerable age group is of utmost importance to avoid suboptimal dosing, which may lead to adverse drug reactions. The juvenile (mini)pig is a representative model for hepatic drug metabolism in human neonates and infants, especially phase I reactions. However, the effect of prematurity on the onset of hepatic phase I and phase II enzyme activity has yet to be investigated in this animal model. Therefore, the aim of this study was to assess the ontogeny of CYP3A and UGT enzyme activity in the liver of preterm (gestational day 105-107) and term-born (gestational day 115-117) domestic piglets. In addition, the ontogeny pattern between the preterm and term group was compared to examine whether postconceptional or postnatal age affects the onset of enzyme activity. The following age groups were included: preterm postnatal day (PND) 0 (n = 10), PND 5 (n = 10), PND 11 (n = 8), PND 26 (n = 10) and term PND 0 (n = 10), PND 5 (n = 10), PND 11 (n = 8), PND 19 (n = 18) and PND 26 (n = 10). Liver microsomes were extracted, and the metabolism of CYP3A and UGT-specific substrates assessed enzyme activity. Preterm CYP3A activity was only detectable at PND 26, whereas term CYP3A activity showed a gradual postnatal increase from PND 11 onwards. UGT activity gradually increased between PND 0 and PND 26 in preterm and term-born piglets, albeit, being systematically lower in the preterm group. Thus, postconceptional age is suggested as the main driver affecting porcine CYP3A and UGT enzyme ontogeny. These data are a valuable step forward in the characterization of the preterm piglet as a translational model for hepatic drug metabolism in the preterm human neonate.

Red cell infusion but not saline is effective for volume expansion in preterm piglets.

A common first-line treatment for supporting cardiovascular function in preterm infants is volume expansion using saline, but this does not improve outcomes. This study aimed to determine if volume expansion with saline increases blood volume, blood pressure and cerebral oxygenation; and if volume expansion with packed red blood cells (RBC) is more effective. We hypothesized that RBC infusion is more effective than saline for increasing blood volume and maintaining cardiovascular function and cerebral oxygenation. Five groups of preterm piglets (98/115d gestation) were infused with saline (10 or 20 mL/kg) or RBC (10 or 20 mL/kg) or no treatment. Blood volume, blood pressure, central venous pressure, heart rate, carotid flow, cerebral oxygenation, arterial pH, base excess, and lactate levels were assessed for 6 h after treatment started. Both RBC groups had significant increases in blood volume, and improved measures of cardiovascular function, cerebral oxygenation and metabolic acidosis. Saline infusion did not increase blood volume or measures of cardiovascular function, cerebral oxygenation or metabolic acidosis. The results suggest that the deteriorating cardiovascular function in the hours after birth in preterm piglets, and possibly in premature babies, may be reversed or halted by more effective support of blood volume. Blood volume decreases after birth in preterm piglets and this decrease is associated with deteriorating cardiovascular function and cerebral oxygenation. Infusion of saline does not increase blood volume nor prevent deterioration in cardiovascular function. Infusion of packed red blood cells results in an increase in blood volume and improvements in cardiovascular function and cerebral oxygenation. Deteriorating cardiovascular function in the hours after birth in preterm piglets, and possibly in human preterm neonates, may be reversed or halted by more effective support of blood volume.

Determining Intestinal Permeability using Lucifer Yellow in an Apical-Out Enteroid Model.

Enteroids are an emerging research tool in the study of inflammatory bowel diseases such as necrotizing enterocolitis (NEC). They are traditionally grown in the basolateral-out (BO) conformation, where the apical surface of the epithelial cell faces the inner lumen. In this model, access to the luminal surface of enteroids for treatment and experimentation is challenging, which limits the ability to study host-pathogen interactions. To circumvent this, a neonatal apical-out (AO) model for necrotizing enterocolitis was created. Since intestinal epithelial cell permeability changes are pathognomonic for NEC, this protocol outlines using lucifer yellow (LY) as a marker of paracellular permeability. LY traverses the intestinal epithelial barrier via all three major paracellular pathways: pore, leak, and unrestricted. Using LY in an AO model allows for a broader study of permeability in NEC. Following IRB approval and parental consent, surgical samples of intestinal tissue were collected from human preterm neonates. Intestinal stem cells were harvested via crypt isolation and used to grow enteroids. Enteroids were grown to maturity and then transformed AO or left in BO conformation. These were either not treated (control) or were treated with lipopolysaccharide (LPS) and subjected to hypoxic conditions for the induction of in vitro NEC. LY was used to assess for permeability. Immunofluorescent staining of the apical protein zonula occludens-1 and basolateral protein β-catenin confirmed AO conformation. Both AO and BO enteroids treated with LPS and hypoxia demonstrated significantly increased paracellular permeability compared to controls. Both AO and BO enteroids showed increased uptake of LY into the lumen of the treated enteroids compared to controls. The utilization of LY in an AO enteroid model allows for the investigation of all three major pathways of paracellular permeability. It additionally allows for the investigation of host-pathogen interactions and how this may affect permeability compared to the BO enteroid model.

Preterm infant meconium microbiota transplant induces growth failure, inflammatory activation, and metabolic disturbances in germ-free mice

SummaryPreterm birth may result in adverse health outcomes. Very preterm infants typically exhibit postnatal growth restriction, metabolic disturbances, and exaggerated inflammatory responses. We investigated the differences in the meconium microbiota composition between very preterm (<32 weeks), moderately preterm (32–37 weeks), and term (>37 weeks) human neonates by 16S rRNA gene sequencing. Human meconium microbiota transplants to germ-free mice were conducted to investigate whether the meconium microbiota is causally related to the preterm infant phenotype in an experimental model. Our results indicate that very preterm birth is associated with a distinct meconium microbiota composition. Fecal microbiota transplant of very preterm infant meconium results in impaired growth, altered intestinal immune function, and metabolic parameters as compared to term infant meconium transplants in germ-free mice. This finding suggests that measures aiming to minimize the long-term adverse consequences of very preterm birth should be commenced during pregnancy or directly after birth.

Lung ultrasound features and relationships with respiratory mechanics of evolving BPD in preterm rabbits and human neonates.

Evolving bronchopulmonary dysplasia (BPD) is characterized by impaired alveolarization leading to lung aeration inhomogeneities. Hyperoxia-exposed preterm rabbits have been proposed to mimic evolving BPD; therefore, we aimed to verify if this model has the same lung ultrasound and mechanical features of evolving BPD in human neonates. Semiquantitative lung ultrasound and lung mechanics measurement was performed in 25 preterm rabbits (28 days of gestation) and 25 neonates (mean gestational age ≈ 26 wk) with evolving BPD. A modified rabbit lung ultrasound score (rLUS) and a validated neonatal lung ultrasound score (LUS) were used. Lung ultrasound images were recorded and evaluated by two independent observers blinded to each other's evaluation. Lung ultrasound findings were equally heterogeneous both in rabbits as in human neonates and encompassed all the classical lung ultrasound semiology. Lung ultrasound and histology examination were also performed in 13 term rabbits kept under normoxia as further control and showed the absence of ultrasound and histology abnormalities compared with hyperoxia-exposed preterm rabbits. The interrater absolute agreement for the evaluation of lung ultrasound images in rabbits was very high [ICC: 0.989 (95%CI: 0.975-0.995); P < 0.0001], and there was no difference between the two observers. Lung mechanics parameters were similarly altered in both rabbits and human neonates. There were moderately significant correlations between airway resistances and lung ultrasound scores in rabbits (ρ = 0.519; P = 0.008) and in neonates (ρ = 0.409; P = 0.042). In conclusion, the preterm rabbit model fairly reproduces the lung ultrasound and mechanical characteristics of preterm neonates with evolving BPD.NEW & NOTEWORTHY We have reported that hyperoxia-exposed preterm rabbits and human preterm neonates with evolving BPD have the same lung ultrasound appearance, and that lung ultrasound can be fruitfully applied on this model with a brief training. The animal model and human neonates also presented the same relationship between semiquantitative ultrasound-assessed lung aeration and airway resistances. In conclusion, this animal model fairly reproduce evolving BPD as it is seen in clinical practice.

Influence of probiotic supplementation on the developing microbiota in human preterm neonates

ABSTRACT Background Oral administration of probiotic bacteria to preterm neonates has been recommended to prevent the development of necrotizing enterocolitis (NEC). The influence of probiotics on the endogenous microbiome, however, has remained incompletely understood. Study design & methods Here, we performed an observational study including 80 preterm neonates born at a gestational age <32-weeks to characterize the persistence of probiotic bacteria after no treatment or oral administration of two different probiotic formula and their influence on the microbial ecosystem during and after the intervention and their association with the development of NEC. Weekly fecal samples were profiled by 16S rRNA sequencing and monitored for the presence of the probiotic bacteria by quantitative PCR. Results Microbiota profiles differed significantly between the control group and both probiotic groups. Probiotic supplementation was associated with lower temporal variation as well as higher relative abundance of Bifidobacterium and Enterobacter combined with reduced abundance of Escherichia, Enterococcus, and Klebsiella. Colonization by probiotic bifidobacteria was observed in approximately 50% of infants although it remained transient in the majority of cases. A significantly reduced monthly incidence of NEC was observed in neonates supplemented with probiotics. Conclusion Our results demonstrate successful transient colonization by probiotic bacteria and a significant influence on the endogenous microbiota with a reduced abundance of bacterial taxa associated with the development of NEC. These results emphasize that probiotic supplementation may allow targeted manipulation of the enteric microbiota and confer a clinical benefit. (Clinical Trial Registry accession number: DRKS/GCTR 00021034)

Lung transcriptional unresponsiveness and loss of early influenza virus control in infected neonates is prevented by intranasal Lactobacillus rhamnosus GG.

Respiratory viral infections contribute substantially to global infant losses and disproportionately affect preterm neonates. Using our previously established neonatal murine model of influenza infection, we demonstrate that three-day old mice are exceptionally sensitive to influenza virus infection and exhibit high mortality and viral load. Intranasal pre- and post-treatment of neonatal mice with Lactobacillus rhamnosus GG (LGG), an immune modulator in respiratory viral infection of adult mice and human preterm neonates, considerably improves neonatal mice survival after influenza virus infection. We determine that both live and heat-killed intranasal LGG are equally efficacious in protection of neonates. Early in influenza infection, neonatal transcriptional responses in the lung are delayed compared to adults. These responses increase by 24 hours post-infection, demonstrating a delay in the kinetics of the neonatal anti-viral response. LGG pretreatment improves immune gene transcriptional responses during early infection and specifically upregulates type I IFN pathways. This is critical for protection, as neonatal mice intranasally pre-treated with IFNβ before influenza virus infection are also protected. Using transgenic mice, we demonstrate that the protective effect of LGG is mediated through a MyD88-dependent mechanism, specifically via TLR4. LGG can improve both early control of virus and transcriptional responsiveness and could serve as a simple and safe intervention to protect neonates.

SUN-132 PRETERM BIRTH: LONG-TERM EFFECTS ON RENAL MORPHOLOGY AND GLOMERULAR CAPILLARISATION IN ADULT SHEEP

Preterm birth (delivery < 37 weeks of gestation) is an important risk factor for the later development of hypertension and chronic kidney disease. The exact cause of this association is unknown, but likely relates to impaired kidney development in early life. We have previously shown that preterm birth can result in the formation of glomeruli with shrunken capillary tufts in the kidneys of human preterm neonates. We have also shown, in lambs, that preterm birth and/or postnatal mechanical ventilation lead to substantial reductions in glomerular capillary length and filtration surface area at 3 days after birth. If this persists, it has the potential to cause glomerulosclerosis and renal dysfunction. Therefore, in this study we aimed to determine the long-term impact of preterm birth on glomerular morphology in adult sheep. Singleton male sheep were delivered moderately preterm at 132 days (~0.9) of gestation or at term (146 days of gestation). The ewes assigned to deliver preterm were administered 11.4mg betamethasone at 48 and 24 hours prior to birth, and the preterm sheep did not require mechanical ventilation postnatally. After weaning, the Preterm (n=6) and Term (n=6) sheep were maintained in natural grass paddocks and were euthanised at 14.5 months of age (early adulthood). The right kidney from each sheep was perfusion-fixed, and stereological methods were used to determine mean renal corpuscle and glomerular volumes, and glomerular capillary length and surface area in the outer, mid, and inner regions of the renal cortex. Glomerulosclerosis, and interstitial collagen levels in the medulla and cortex, were assessed histologically. Sheep born preterm were on average 2.5 kg lighter at birth than those born at term (p=0.0001), but at 14.5 months of age there was no difference between the Term and Preterm sheep in body or kidney weight (Term: 128 ± 8 g, Preterm: 121 ± 5 g, p=0.45). Renal corpuscle volume was significantly (p=0.03) larger in the Preterm sheep than the Term sheep (e.g. mean for inner cortex – Term: 0.0018 ± 0.0001 mm3, Preterm: 0.0024 ± 0.0003 mm3) with the Preterm sheep exhibiting enlarged Bowman’s spaces; however, there was no difference in glomerular volume between groups. Similarly, preterm birth did not impact mean glomerular capillary length or surface area per glomerulus in adult sheep. There was also no difference between groups in interstitial collagen levels, or glomerulosclerosis index. Although there is some evidence of morphological changes in the kidneys following preterm birth, findings suggest that moderate preterm birth per se does not adversely affect total renal filtration surface area or result in renal disease in early adulthood. It is now important to determine whether glomerular capillarisation is similarly normalised in preterm-born adults that were exposed to mechanical ventilation in the neonatal period.

Microvascular circulatory dysregulation driven in part by cystathionine gamma-lyase: A new paradigm for cardiovascular compromise in the preterm newborn.

H2 S may explain the dysregulation of microvascular tone associated with poor outcome following preterm birth. In adult vasculature, H2 S is predominantly produced by CSE. We hypothesized that vascular CSE activity contributes to microvascular tone regulation during circulatory transition. Preterm (GA62) and full-term (GA69) guinea pig fetuses and neonates were studied. Microvascular blood flow was assessed by laser Doppler flowmetry. Thiosulfate, primary urinary metabolite of H2 S, was determined by high-performance liquid chromatography. Real-time H2 S production was assessed using a microrespiration system in fetal and postnatal (10, 24hours) skin and heart samples. CSE contribution was investigated by inhibition via propargylglycine. In preterm animals, postnatal H2 S production capacity in peripheral vasculature increased significantly and was significantly reduced by the inhibition of CSE. Urinary thiosulfate correlated with both microvascular blood flow and capacity of the vasculature to produce H2 S. H2 S produced via CSE did not correlate directly with microvascular blood flow. In preterm neonates, H2 S production increases during fetal-to-neonatal transition and CSE contribution to total H2 S increases postnatally. CSE-dependent mechanisms may therefore underpin the increase in H2 S production over the first 72hours of life in preterm human neonates, associated with both central and peripheral cardiovascular instability.

Magnetic Resonance Imaging Findings of Term and Preterm Hypoxic-Ischemic Encephalopathy: A Review of Relevant Animal Models and Correlation to Human Imaging.

Background:Neonatal hypoxic-ischemic encephalopathy is brain injury caused by decreased perfusion and oxygen delivery that most commonly occurs in the context of delivery complications such as umbilical cord compression or placental abruption. Imaging is a key component for guiding treatment and prediction of prognosis, and the most sensitive clinical imaging modality for the brain injury patterns seen in hypoxic-ischemic encephalopathy is magnetic resonance imaging.Objective:The goal of this review is to compare magnetic resonance imaging findings demonstrated in the available animal models of hypoxic-ischemic encephalopathy to those found in preterm (≤ 36 weeks) and term (>36 weeks) human neonates with hypoxic-ischemic encephalopathy, with special attention to the strengths and weaknesses of each model.Methods:A structured literature search was performed independently by two authors and the results of the searches were compiled. Animal model, human brain age equivalency, mechanism of injury, and area of brain injury were recorded for comparison to imaging findings in preterm and term human neonates with hypoxic-ischemic encephalopathy.Conclusion:Numerous animal models have been developed to better elicit the expected findings that occur after HIE by allowing investigators to control many of the clinical variables that result in injury. Although modeling the same disease process, magnetic resonance imaging findings in the animal models vary with the species and methods used to induce hypoxia and ischemia. The further development of animal models of HIE should include a focus on comparing imaging findings, and not just pathologic findings, to human studies.

Necrotizing enterocolitis attenuates developmental heart rate variability increases in newborn rats.

We have shown previously that a decreased high-frequency spectrum of heart rate variability (HF-HRV), indicative of reduced vagal tone, shows promise in predicting neonates likely to develop necrotizing enterocolitis (NEC) before its clinical onset. We hypothesized that NEC induction in rat pups decreases HF-HRV power; subdiaphragmatic vagotomy worsens the severity of the NEC phenotype, increases levels of pro-inflammatory cytokines, and alters the myenteric phenotype. Newborn Sprague-Dawley rats, representative of preterm human neonates, were subjected to 7-8days of brief periods of cold stress and hypoxia to induce NEC with or without unilateral subdiaphragmatic vagotomy. HRV was measured at postnatal days one and five, pups were sacrificed at day 8/9, and gastrointestinal tissues and blood were collected for immunohistochemical, corticosterone, and cytokine analysis. Compared to control, NEC-induced rats showed the following: (a) typical histological signs of grade 2 NEC, which were more severe in rats that underwent vagotomy; (b) reduced developmental increases in time (RMSSD) and frequency (HF) HRV spectra when combined with the stress of laparotomy/vagotomy; (c) increases in nitric oxide synthase-immunoreactivity in the myenteric plexus of jejunum and ileum; furthermore, compared to mild NEC and controls, vagotomized NEC rats had increased plasma values of pro-inflammatory cytokines IL-1β and IL-6. Our data suggest that in rodents, similar to neonatal observations, NEC induction attenuated developmental HF-HRV increases, furthermore, subdiaphragmatic vagotomy worsened the histological severity, increased pro-inflammatory cytokines, and altered the nitrergic myenteric phenotype, suggesting a role of the vagus in the development of NEC pathology.

Significant Hypo-Responsiveness to GPVI and CLEC-2 Agonists in Pre-Term and Full-Term Neonatal Platelets and following Immune Thrombocytopenia.

Neonatal platelets are hypo-reactive to the tyrosine kinase-linked receptor agonist collagen. Here, we have investigated whether the hypo-responsiveness is related to altered levels of glycoprotein VI (GPVI) and integrin α2β1, or to defects in downstream signalling events by comparison to platelet activation by C-type lectin-like receptor 2 (CLEC-2). GPVI and CLEC-2 activate a Src- and Syk-dependent signalling pathway upstream of phospholipase C (PLC) γ2. Phosphorylation of a conserved YxxL sequence known as a (hemi) immunotyrosine-based-activation-motif (ITAM) in both receptors is critical for Syk activation. Platelets from human pre-term and full-term neonates display mildly reduced expression of GPVI and CLEC-2, as well as integrin αIIbβ3, accounted for at the transcriptional level. They are also hypo-responsive to the two ITAM receptors, as shown by measurement of integrin αIIbβ3 activation, P-selectin expression and Syk and PLCγ2 phosphorylation. Mouse platelets are also hypo-responsive to GPVI and CLEC-2 from late gestation to 2 weeks of age, as determined by measurement of integrin αIIbβ3 activation. In contrast, the response to G protein-coupled receptor agonists was only mildly reduced and in some cases not altered in neonatal platelets of both species. A reduction in response to GPVI and CLEC-2, but not protease-activated receptor 4 (PAR-4) peptide, was also observed in adult mouse platelets following immune thrombocytopenia, whereas receptor expression was not impaired. Our results demonstrate developmental differences in platelet responsiveness to GPVI and CLEC-2, and also following immune platelet depletion leading to reduced Syk activation. The rapid generation of platelets during development or following platelet depletion is achieved at the expense of signalling by ITAM-coupled receptors.

A TLR/AKT/FoxO3 immune tolerance-like pathway disrupts the repair capacity of oligodendrocyte progenitors.

Cerebral white matter injury (WMI) persistently disrupts myelin regeneration by oligodendrocyte progenitor cells (OPCs). We identified a specific bioactive hyaluronan fragment (bHAf) that downregulates myelin gene expression and chronically blocks OPC maturation and myelination via a tolerance-like mechanism that dysregulates pro-myelination signaling via AKT. Desensitization of AKT occurs via TLR4 but not TLR2 or CD44. OPC differentiation was selectively blocked by bHAf in a maturation-dependent fashion at the late OPC (preOL) stage by a noncanonical TLR4/TRIF pathway that induced persistent activation of the FoxO3 transcription factor downstream of AKT. Activated FoxO3 selectively localized to oligodendrocyte lineage cells in white matter lesions from human preterm neonates and adults with multiple sclerosis. FoxO3 constraint of OPC maturation was bHAf dependent, and involved interactions at the FoxO3 and MBP promoters with the chromatin remodeling factor Brg1 and the transcription factor Olig2, which regulate OPC differentiation. WMI has adapted an immune tolerance-like mechanism whereby persistent engagement of TLR4 by bHAf promotes an OPC niche at the expense of myelination by engaging a FoxO3 signaling pathway that chronically constrains OPC differentiation.

Early Procedural Pain Is Associated with Regionally-Specific Alterations in Thalamic Development in Preterm Neonates.

Very preterm human neonates are exposed to numerous invasive procedures as part of life-saving care. Evidence suggests that repetitive neonatal procedural pain precedes long-term alterations in brain development. However, to date the link between pain and brain development has limited temporal and anatomic specificity. We hypothesized that early exposure to painful stimuli during a period of rapid brain development, before pain modulatory systems reach maturity, will predict pronounced changes in thalamic development, and thereby cognitive and motor function. In a prospective cohort study, 155 very preterm neonates (82 males, 73 females) born 24-32 weeks' gestation underwent two MRIs at median postmenstrual ages 32 and 40 weeks that included structural, metabolic, and diffusion imaging. Detailed day-by-day clinical data were collected. Cognitive and motor abilities were assessed at 3 years, corrected age. The association of early (skin breaks, birth-Scan 1) and late pain (skin breaks, Scans 1-2) with thalamic volumes and N-acetylaspartate (NAA)/choline (Cho), and fractional anisotropy of white-matter pathways was assessed. Early pain was associated with slower thalamic macrostructural growth, most pronounced in extremely premature neonates. Deformation-based morphometry analyses confirmed early pain-related volume losses were localized to somatosensory regions. In extremely preterm neonates early pain was associated with decreased thalamic NAA/Cho and microstructural alterations in thalamocortical pathways. Thalamic growth was in turn related to cognitive and motor outcomes. We observed regionally-specific alterations in the lateral thalamus and thalamocortical pathways in extremely preterm neonates exposed to more procedural pain. Findings suggest a sensitive period leading to lasting alterations in somatosensory-system development.SIGNIFICANCE STATEMENT Early exposure to repetitive procedural pain in very preterm neonates may disrupt the development of regions involved in somatosensory processing, leading to poor functional outcomes. We demonstrate that early pain is associated with thalamic volume loss in the territory of the somatosensory thalamus and is accompanied by disruptions thalamic metabolic growth and thalamocortical pathway maturation, particularly in extremely preterm neonates. Thalamic growth was associated with cognitive and motor outcome at 3 years corrected age. Findings provide evidence for a developmentally sensitive period whereby subcortical structures in young neonates may be most vulnerable to procedural pain. Furthermore, results suggest that the thalamus may play a key role underlying the association between neonatal pain and poor neurodevelopmental outcomes in these high-risk neonates.

Hyperoxia reduces insulin release and induces mitochondrial dysfunction with possible implications for hyperoxic treatment of neonates.

We previously showed that hyperoxia in vitro negatively affects beta cells of the rat. Here, we tested for possible clinical significance as well as mitochondrial interactions by hyperoxia, using human islets (function and viability), INS‐1 832/13 cells (mitochondrial metabolism), and mouse neonates (effects in vivo). Lastly, we assessed relevant parameters in a cohort of individuals born preterm and then exposed to hyperoxia. Human islets and INS‐1 832/13 cells were exposed to 24 h of hyperoxia (90–92% oxygen). Mouse neonates were subjected to 5 days of continuous hyperoxia. Individuals born preterm were evaluated in terms of glucose homeostasis and beta cell function by HbA1c and the HOMA2 formula. In human islets, hyperoxia significantly reduced glucose‐stimulated insulin secretion by 42.2 ± 5.3% and viability assessed by MTT by 22.5 ± 5.4%. Hyperoxia down‐regulated mitochondrial complex II by 21 ± 5% and upregulated complex III by 26 ± 10.1% and complex IV by 37 ± 10.6%. Partly similar effects on mitochondrial complexes were found in hyperoxia‐exposed INS‐1 832/13 cells. Exposure to hyperoxia swiftly reduced oxygen consumption in these cells and increased mitochondrial uncoupling. Hyperoxia transiently but significantly reduced insulin release in mouse neonates. Individuals born preterm displayed higher HbA1c versus controls, as well as insulin resistance. Thus, hyperoxia exerts negative effects in vitro on human beta cells and results indicate inhibitory effects on insulin secretion in vivo in mouse neonates. Negative effects may be lessened by the demonstrated swift and profound mitochondrial adaptability. Our findings open the possibility that hyperoxia could negatively affect beta cells of preterm human neonates.

Neutrophil chemotaxis and transcriptomics in term and preterm neonates

Neutrophils play a crucial role in combating life-threatening bacterial infections in neonates. Previous studies investigating neonatal cell function have been limited because of restricted volume sampling. Here, using novel microfluidic approaches, we provide the first description of neutrophil chemotaxis and transcriptomics fromwhole blood of human term and preterm neonates, as well as youngadults.Exvivo percent cell migration, neutrophil velocity, and directionality to N-formylmethionyl-leucyl-phenylalanine were measured from whole blood using time-lapse imaging of microfluidic chemotaxis. Genome-wide expression was also evaluated in CD66b+ cells using microfluidic capture devices. Neutrophils from preterm neonates migrated in fewer numbers compared to term neonates (preterm 12.3%, term 30.5%, P=0.008) and at a reduced velocity compared to young adults (preterm 10.1μm/min, adult 12.7μm/min, P=0.003). Despite fewer neutrophils migrating at slower velocities, neutrophil directionality from preterm neonates was comparable to adults and term neonates. 3607 genes were differentially expressed among the 3 groups (P<0.001). Differences in gene expression between neutrophils from preterm and term neonates were consistent with reduced pathogen recognition and antimicrobial activity but not neutrophil migration, by preterm neonates. In summary, preterm neonates have significant disturbances in neutrophil chemotaxis compared to term neonates and adults, and these differences in phenotype appear at the transcriptional level to target inflammatory pathways in general, rather than in neutrophil migration and chemotaxis.

Electrophysiological and hemodynamic mismatch responses in rats listening to human speech syllables.

Speech is a complex auditory stimulus which is processed according to several time-scales. Whereas consonant discrimination is required to resolve rapid acoustic events, voice perception relies on slower cues. Humans, right from preterm ages, are particularly efficient to encode temporal cues. To compare the capacities of preterms to those observed in other mammals, we tested anesthetized adult rats by using exactly the same paradigm as that used in preterm neonates. We simultaneously recorded neural (using ECoG) and hemodynamic responses (using fNIRS) to series of human speech syllables and investigated the brain response to a change of consonant (ba vs. ga) and to a change of voice (male vs. female). Both methods revealed concordant results, although ECoG measures were more sensitive than fNIRS. Responses to syllables were bilateral, but with marked right-hemispheric lateralization. Responses to voice changes were observed with both methods, while only ECoG was sensitive to consonant changes. These results suggest that rats more effectively processed the speech envelope than fine temporal cues in contrast with human preterm neonates, in whom the opposite effects were observed. Cross-species comparisons constitute a very valuable tool to define the singularities of the human brain and species-specific bias that may help human infants to learn their native language.

Functional Maps at the Onset of Auditory Inputs in Very Early Preterm Human Neonates.

During the last trimester of human gestation, neurons reach their final destination and establish long- and short-distance connections. Due to the difficulties obtaining functional data at this age, the characteristics of the functional architecture at the onset of sensory thalamocortical connectivity in humans remain largely unknown. In particular, it is unknown to what extent responses evoked by an external stimulus are general or already sensitive to certain stimuli. In the present study, we recorded high-density event-related potentials (ERPs) in 19 neonates, tested ten weeks before term (28-32 weeks gestational age (wGA), that is, at an average age of 30 wGA) by means of a syllable discrimination task (i.e., a phonetic change: ba vs. ga; and a voice change: male vs. female voice). We first observed that the syllables elicited 4 peaks with distinct topographies implying a progression of the sensory input along a processing hierarchy; second, repetition induced a decrease in the amplitude (repetition suppression) of these peaks, but their latencies and topographies remained stable; and third, a change of stimulus generated mismatch responses, which were more precisely time-locked to event onset in the case of a phonetic change than in the case of a voice change. A hierarchical and parallel functional architecture is therefore able to process environmental sounds in a timely precise fashion, well before term birth. This elaborate functional architecture at the onset of extrinsic neural activity suggests that specialized areas weakly dependent on the environment are present in the perisylvian region as part of the genetic endowment of the human species.

Expression of genes of the cardiac and renal renin–angiotensin systems in preterm piglets: is this system a suitable target for therapeutic intervention?

The newborn circulating, cardiac and renal renin-angiotensin systems (RASs) are essential for blood pressure control, and for cardiac and renal development. If cardiac and renal RASs are immature this may contribute to cardiovascular compromise in preterm infants. This study measured mRNA expression of cardiac and renal RAS components in preterm, glucocorticoid (GC) exposed preterm, and term piglets. Renal and cardiac RAS mRNA levels were measured using real-time polymerase chain reaction (PCR). Genes studied were: (pro)renin receptor, renin, angiotensinogen, angiotensin converting enzyme (ACE), ACE2, angiotensin type 1 receptor (AT1R) and angiotensin type 2 receptor (AT2R). All the genes studied were expressed in the kidney; neither renin nor AT2R mRNA were detected in the heart. There were no gestational changes in (pro)renin receptor, renin, ACE or AT1R mRNA levels. Right ventricular angiotensinogen mRNA levels in females were lower in preterm animals than at term, and GC exposure increased levels in male piglets. Renal angiotensinogen mRNA levels in female term piglets were lower than females from both preterm groups, and lower than male term piglets. Left ventricular ACE2 mRNA expression was lower in GC treated preterm piglets. Renal AT2R mRNA abundance was highest in GC treated preterm piglets, and the AT1R/AT2R ratio was increased at term. Preterm cardiac and renal RAS mRNA levels were similar to term piglets, suggesting that immaturity of these RASs does not contribute to preterm cardiovascular compromise. Since preterm expression of both renal and cardiac angiotensin II-AT1R is similar to term animals, cardiovascular dysfunction in the sick preterm human neonate might be effectively treated by agents acting on their RASs.

Early microvascular changes in the preterm neonate: a comparative study of the human and guinea pig.

Dysfunction of the transition from fetal to neonatal circulatory systems may be a major contributor to poor outcome following preterm birth. Evidence exists in the human for both a period of low flow between 5 and 11 h and a later period of increased flow, suggesting a hypoperfusion–reperfusion cycle over the first 24 h following birth. Little is known about the regulation of peripheral blood flow during this time. The aim of this study was to conduct a comparative study between the human and guinea pig to characterize peripheral microvascular behavior during circulatory transition. Very preterm (≤28 weeks GA), preterm (29–36 weeks GA), and term (≥37 weeks GA) human neonates underwent laser Doppler analysis of skin microvascular blood flow at 6 and 24 h from birth. Guinea pig neonates were delivered prematurely (62 day GA) or at term (68–71 day GA) and laser Doppler analysis of skin microvascular blood flow was assessed every 2 h from birth. In human preterm neonates, there is a period of high microvascular flow at 24 h after birth. No period of low flow was observed at 6 h. In preterm animals, microvascular flow increased after birth, reaching a peak at 10 h postnatal age. Blood flow then steadily decreased, returning to delivery levels by 24 h. Preterm birth was associated with higher baseline microvascular flow throughout the study period in both human and guinea pig neonates. The findings do not support a hypoperfusion–reperfusion cycle in the microcirculation during circulatory transition. The guinea pig model of preterm birth will allow further investigation of the mechanisms underlying microvascular function and dysfunction during the initial extrauterine period.