Ryan M. McAdams

Concurrent erythropoietin and hypothermia treatment improve outcomes in a term nonhuman primate model of perinatal asphyxia.

Background: Up to 65% of untreated infants suffering from moderate to severe hypoxic-ischemic encephalopathy (HIE) are at risk of death or major disability. Therapeutic hypothermia (HT) reduces this risk to approximately 50% (number needed to treat: 7-9). Erythropoietin (Epo) is a neuroprotective treatment that is promising as an adjunctive therapy to decrease HIE-induced injury because Epo decreases apoptosis, inflammation, and oxidative injury and promotes glial cell survival and angiogenesis. We hypothesized that HT and concurrent Epo will be safe and effective, improve survival, and reduce moderate-severe cerebral palsy (CP) in a term nonhuman primate model of perinatal asphyxia. Methodology: Thirty-five Macacanemestrina were delivered after 15-18 min of umbilical cord occlusion (UCO) and randomized to saline (n = 14), HT only (n = 9), or HT+Epo (n = 12). There were 12 unasphyxiated controls. Epo (3,500 U/kg × 1 dose followed by 3 doses of 2,500 U/kg, or Epo 1,000 U/kg/day × 4 doses) was given on days 1, 2, 3, and 7. Timed blood samples were collected to measure plasma Epo concentrations. Animals underwent MRI/MRS and diffusion tensor imaging (DTI) at <72 h of age and again at 9 months. A battery of weekly developmental assessments was performed. Results: UCO resulted in death or moderate-severe CP in 43% of saline-, 44% of HT-, and 0% of HT+Epo-treated animals. Compared to non-UCO control animals, UCO animals exhibit poor weight gain, behavioral impairment, poor cerebellar growth, and abnormal brain DTI. Compared to UCO saline, UCO HT+Epo improved motor and cognitive responses, cerebellar growth, and DTI measures and produced a death/disability relative risk reduction of 0.911 (95% CI -0.429 to 0.994), an absolute risk reduction of 0.395 (95% CI 0.072-0.635), and a number needed to treat of 2 (95% CI 2-14). The effects of HT+Epo on DTI included an improved mode of anisotropy, fractional anisotropy, relative anisotropy, and volume ratio as compared to UCO saline-treated infants. No adverse drug reactions were noted in animals receiving Epo, and there were no hematology, liver, or kidney laboratory effects. Conclusions/Significance: HT+Epo treatment improved outcomes in nonhuman primates exposed to UCO. Adjunctive use of Epo combined with HT may improve the outcomes of term human infants with HIE, and clinical trials are warranted.

Outcomes of extremely low birth weight infants given early high-dose erythropoietin

Objective:To evaluate long-term outcomes of 60 extremely low birth weight (ELBW) infants treated with or without three injections of high-dose erythropoietin (Epo).Study Design:A retrospective analysis of anthropometric and neurodevelopmental outcome data comparing 30 ELBW infants enrolled in a phase I/II study examining the pharmacokinetics of high-dose Epo (500, 1000 and 2500 U/kg × 3 doses) administered to 30 concurrent controls.Result:Birth characteristics and growth from 4 to 36 months were similar for untreated and Epo-treated patients. Multiple linear regression analysis of neurodevelopmental follow-up scores from 17/25 Epo-treated and 18/26 control infants identified that Epo correlated with improvement of cognitive (R=0.22, P=0.044) and motor (R=0.15, P=0.026) scores. No negative long-term effects of Epo treatment were evident.Conclusion:Retrospective analysis of the only available long-term follow-up data from ELBW infants given high-dose Epo treatment suggests that Epo treatment is safe and correlates with modest improvement of neurodevelopmental outcomes.

The role of cytokines and inflammatory cells in perinatal brain injury.

Perinatal brain injury frequently complicates preterm birth and leads to significant long-term morbidity. Cytokines and inflammatory cells are mediators in the common pathways associated with perinatal brain injury induced by a variety of insults, such as hypoxic-ischemic injury, reperfusion injury, toxin-mediated injury, and infection. This paper examines our current knowledge regarding cytokine-related perinatal brain injury and specifically discusses strategies for attenuating cytokine-mediated brain damage.

Poor birth weight recovery among low birth weight/preterm infants following hospital discharge in Kampala, Uganda

BackgroundHealthy infants typically regain their birth weight by 21 days of age; however, failure to do so may be due to medical, nutritional or environmental factors. Globally, the incidence of low birth weight deliveries is high, but few studies have assessed the postnatal weight changes in this category of infants, especially in Africa. The aim was to determine what proportion of LBW infants had not regained their birth weight by 21 days of age after discharge from the Special Care Unit of Mulago hospital, Kampala.MethodsA cross sectional study was conducted assessing weight recovery of 235 LBW infants attending the Kangaroo Clinic in the Special Care Unit of Mulago Hospital between January and April 2010. Infants aged 21 days with a documented birth weight and whose mothers gave consent to participate were included in the study. Baseline information was collected on demographic characteristics, history on pregnancy, delivery and postnatal outcome through interviews. Pertinent infant information like gestation age, diagnosis and management was obtained from the medical records and summarized in the case report forms.ResultsOf the 235 LBW infants, 113 (48.1%) had not regained their birth weight by 21 days. Duration of hospitalization for more than 7 days (AOR: 4.2; 95% CI: 2.3 - 7.6; p value < 0.001) and initiation of the first feed after 48 hours (AOR: 1.9; 95% CI 1.1 - 3.4 p value 0.034) were independently associated with failure to regain birth weight. Maternal factors and the infants physical examination findings were not significantly associated with failure to regain birth weight by 21 days of age.ConclusionFailure to regain birth weight among LBW infants by 21 days of age is a common problem in Mulago Hospital occurring in almost half of the neonates attending the Kangaroo clinic. Currently, the burden of morbidity in this group of high-risk infants is undetected and unaddressed in many developing countries. Measures for consideration to improve care of these infants would include; discharge after regaining birth weight and use of total parenteral nutrition. However, due to the pressure of space, keeping the baby and mother is not feasible at the moment hence the need for a strong community system to boost care of the infant. Close networking with support groups within the childs environment could help alleviate this problem.

Choriodecidual Group B Streptococcal Inoculation Induces Fetal Lung Injury without Intra-Amniotic Infection and Preterm Labor in Macaca nemestrina

Background Early events leading to intrauterine infection and fetal lung injury remain poorly defined, but may hold the key to preventing neonatal and adult chronic lung disease. Our objective was to establish a nonhuman primate model of an early stage of chorioamnionitis in order to determine the time course and mechanisms of fetal lung injury in utero. Methodology/Principal Findings Ten chronically catheterized pregnant monkeys (Macaca nemestrina) at 118–125 days gestation (term = 172 days) received one of two treatments: 1) choriodecidual and intra-amniotic saline (n = 5), or 2) choriodecidual inoculation of Group B Streptococcus (GBS) 1×106 colony forming units (n = 5). Cesarean section was performed regardless of labor 4 days after GBS or 7 days after saline infusion to collect fetal and placental tissues. Only two GBS animals developed early labor with no cervical change in the remaining animals. Despite uterine quiescence in most cases, blinded review found histopathological evidence of fetal lung injury in four GBS animals characterized by intra-alveolar neutrophils and interstitial thickening, which was absent in controls. Significant elevations of cytokines in amniotic fluid (TNF-α, IL-8, IL-1β, IL-6) and fetal plasma (IL-8) were detected in GBS animals and correlated with lung injury (p<0.05). Lung injury was not directly caused by GBS, because GBS was undetectable in amniotic fluid (∼10 samples tested/animal), maternal and fetal blood by culture and polymerase chain reaction. In only two cases was GBS cultured from the inoculation site in low numbers. Chorioamnionitis occurred in two GBS animals with lung injury, but two others with lung injury had normal placental histology. Conclusions/Significance A transient choriodecidual infection can induce cytokine production, which is associated with fetal lung injury without overt infection of amniotic fluid, chorioamnionitis or preterm labor. Fetal lung injury may, thus, occur silently without symptoms and before the onset of the fetal systemic inflammatory response syndrome.

Group B Streptococcal Infection of the Choriodecidua Induces Dysfunction of the Cytokeratin Network in Amniotic Epithelium: A Pathway to Membrane Weakening

Early events leading to intrauterine infection remain poorly defined, but may hold the key to preventing preterm delivery. To determine molecular pathways within fetal membranes (chorioamnion) associated with early choriodecidual infection that may progress to preterm premature rupture of membranes (PPROM), we examined the effects of a Group B Streptococcus (GBS) choriodecidual infection on chorioamnion in a nonhuman primate model. Ten chronically catheterized pregnant monkeys (Macaca nemestrina) at 118–125 days gestation (term = 172 days) received choriodecidual inoculation of either GBS (n = 5) or saline (n = 5). Cesarean section was performed in the first week after GBS or saline inoculation. RNA extracted from chorioamnion (inoculation site) was profiled by microarray. Single gene, Gene Set, and Ingenuity Pathway Analysis results were validated using qRT-PCR (chorioamnion), Luminex (amniotic fluid, AF), immunohistochemistry, and transmission electron microscopy (TEM). Despite uterine quiescence in most cases, significant elevations of AF cytokines (TNF-α, IL-8, IL-1β, IL-6) were detected in GBS versus controls (p<0.05). Choriodecidual infection resolved by the time of cesarean section in 3 of 5 cases and GBS was undetectable by culture and PCR in the AF. A total of 331 genes were differentially expressed (>2-fold change, p<0.05). Remarkably, GBS exposure was associated with significantly downregulated expression of multiple cytokeratin (CK) and other cytoskeletal genes critical for maintenance of tissue tensile strength. Immunofluorescence revealed highly significant changes in the CK network within amniocytes with dense CK aggregates and retraction from the cell periphery (all p = 0.006). In human pregnancies affected by PPROM, there was further evidence of CK network retraction with significantly shorter amniocyte foot processes (p = 0.002). These results suggest early choriodecidual infection results in decreased cellular membrane integrity and tensile strength via dysfunction of CK networks. Downregulation of CK expression and perturbations in the amniotic epithelial cell intermediate filament network occur after GBS choriodecidual infection, which may contribute to PPROM.

Choriodecidual Infection Downregulates Angiogenesis and Morphogenesis Pathways in Fetal Lungs from Macaca Nemestrina

Background Intrauterine exposure to amniotic fluid (AF) cytokines is thought to predispose to bronchopulmonary dysplasia (BPD). We evaluated the effects of GBS exposure on RNA expression in fetal lung tissue to determine early molecular pathways associated with fetal lung injury that may progress to BPD. Methods Ten chronically catheterized pregnant monkeys (Macaca nemestrina) at 118–125 days gestation (term = 172 days) received choriodecidual inoculation of either: 1) Group B Streptococcus (n = 5) or 2) saline (n = 5). Cesarean section and fetal necropsy was performed in the first week after GBS or saline inoculation regardless of labor. RNA was extracted from fetal lungs and profiled by microarray. Results were analyzed using single gene, Gene Set, and Ingenuity Pathway Analysis. Validation was by RT-PCR and immunohistochemistry. Results Despite uterine quiescence in most cases, fetal lung injury occurred in four GBS cases (intra-alveolar neutrophils, interstitial thickening) and one control (peri-mortem hemorrhage). Significant elevations of AF cytokines (TNF-α, IL-8, IL-1β, IL-6) were detected in GBS versus controls (p<0.05). Lung injury was not directly caused by GBS, because GBS was undetectable by culture and PCR in the AF and fetal lungs. A total of 335 genes were differentially expressed greater than 1.5 fold (p<0.05) with GBS exposure associated with a striking upregulation of genes in innate and adaptive immunity and downregulation of pathways for angiogenesis, morphogenesis, and cellular growth and development. Conclusions A transient choriodecidual infection may induce fetal lung injury with profound alterations in the genetic program of the fetal lung before signs of preterm labor. Our results provide a window for the first time into early molecular pathways disrupting fetal lung angiogenesis and morphogenesis before preterm labor occurs, which may set the stage for BPD. A strategy to prevent BPD should target the fetus in utero to attenuate alterations in the fetal lung genetic program.

Time to implement delayed cord clamping.

Immediate umbilical cord clamping after delivery is routine in the United States despite little evidence to support this practice. Numerous trials in both term and preterm neonates have demonstrated the safety and benefit of delayed cord clamping. In premature neonates, delayed cord clamping has been shown to stabilize transitional circulation, lessening needs for inotropic medications and reducing blood transfusions, necrotizing enterocolitis, and intraventricular hemorrhage. In term neonates, delayed cord clamping has been associated with decreased iron-deficient anemia and increased iron stores with potential valuable effects that extend beyond the newborn period, including improvements in long-term neurodevelopment. The failure to more broadly implement delayed cord clamping in neonates ignores published benefits of increased placental blood transfusion at birth and may represent an unnecessary harm for vulnerable neonates.

Choriodecidual Group B Streptococcal Infection Induces miR-155-5p in the Fetal Lung in Macaca nemestrina

ABSTRACT The mechanisms underlying fetal lung injury remain poorly defined. MicroRNAs (miRNAs) are small noncoding, endogenous RNAs that regulate gene expression and have been implicated in the pathogenesis of lung disease. Using a nonhuman primate model of choriodecidual infection, we sought to determine if differentially expressed miRNAs were associated with acute fetal lung injury. After inoculating 10 chronically catheterized pregnant monkeys (Macaca nemestrina) with either group B streptococcus (GBS) at 1 × 106 CFU (n = 5) or saline (n = 5) in the choriodecidual space, we extracted fetal lung mRNA and miRNA and profiled the changes in expression by microarray analysis. We identified 9 differentially expressed miRNAs in GBS-exposed fetal lungs, but of these, only miR-155-5p was validated by quantitative reverse transcription-PCR (P = 0.02). Significantly elevated miR-155-5p expression was also observed when immortalized human fetal airway epithelial (FeAE) cells were exposed to proinflammatory cytokines (interleukin-6 [IL-6] and tumor necrosis factor alpha [TNF-α]). Overexpression of miR-155-5p in FeAE cells in turn increased the production of IL-6 and CXCL10/gamma interferon-induced protein 10, which are implicated in leukocyte recruitment but also in protection from lung injury. Interestingly, while miR-155-5p decreased fibroblast growth factor 9 (FGF9) expression in a luciferase reporter assay, FGF9 levels were actually increased in GBS-exposed fetal lungs in vivo. FGF9 overexpression is associated with abnormal lung development. Thus, upregulation of miR-155-5p may serve as a compensatory mechanism to lessen the increase in FGF9 and prevent aberrant lung development. Understanding the complicated networks regulating lung development in the setting of infection is a key step in identifying how to prevent fetal lung injury leading to bronchopulmonary dysplasia.

Implementation of Bubble CPAP in a Rural Ugandan Neonatal ICU

BACKGROUND: Respiratory distress is a leading cause of neonatal death in low-income and middle-income countries. CPAP is a simple and effective respiratory support modality used to support neonates with respiratory failure and can be used in low-income and middle-income countries. The goal of this study was to describe implementation of the Silverman-Andersen respiratory severity score (RSS) and bubble CPAP in a rural Ugandan neonatal NICU. We sought to determine whether physicians and nurses in a low-income/middle-income setting would assign similar RSS in neonates after an initial training period and over time. METHODS: We describe the process of training NICU staff to use the RSS to assist in decision making regarding initiation, titration, and termination of bubble CPAP for neonates with respiratory distress. Characteristics of all neonates with respiratory failure treated with bubble CPAP in a rural Ugandan NICU from January to June 2012 are provided. RESULTS: Nineteen NICU staff members (4 doctors and 15 nurses) received RSS training. After this, the Spearman correlation coefficient for respiratory severity scoring between doctor and nurse was 0.73. Twenty-one infants, all < 3 d of age, were treated with CPAP, with 17 infants starting on the day of birth. The majority of infants (16/21, 76%) were preterm, 10 (48%) were <1,500 g (birthweight), and 13 (62%) were outborn. The most common diagnoses were respiratory distress syndrome (16/21, 76%) and birth asphyxia (5/21, 24%). The average RSS was 7.4 ± 1.3 before starting CPAP, 5.2 ± 2.3 after 2–4 h of CPAP, 4.9 ± 2.7 after 12–24 h of CPAP, and 3.5 ± 1.9 before CPAP was discontinued. Duration of treatment with CPAP averaged 79 ± 43 h. Approximately half (11/21, 52%) of infants treated with CPAP survived to discharge. CONCLUSIONS: Implementing bubble CPAP in a low-income/middle-income setting is feasible. The RSS may be a simple and useful tool for monitoring a neonates respiratory status and for guiding CPAP management.