Prospective epigenome and transcriptome analyses of cord and peripheral blood from preterm infants at risk of bronchopulmonary dysplasia.

Bronchopulmonary dysplasia (BPD) is a prevalent chronic lung disease of prematurity with limited treatment options. To uncover biomarkers of BPD risk, this study investigated epigenetic and transcriptomic signatures of prematurity at birth and during the neonatal period at day 14 and 28. Peripheral blood DNAs from preterm infants were applied to methylation arrays and cell-type composition was estimated by deconvolution. Covariate-adjusted robust linear regression elucidated BPD- and prolonged oxygen (≥ 14 days) exposure-associated CpGs. RNAs from cord and peripheral blood were sequenced, and differentially expressed genes (DEGs) for BPD or oxygen exposure were determined. Estimated neutrophil-lymphocyte ratios in peripheral blood at day 14 in BPD infants were significantly higher than nonBPD infants, suggesting an heightened inflammatory response in developing BPD. BPD-DEGs in cord blood indicated lymphopoiesis inhibition, altered Th1/Th2 responses, DNA damage, and organ degeneration. On day 14, BPD-associated CpGs were highly enriched in neutrophil activation, infection, and CD4 + T cell quantity, and BPD-DEGs were involved in DNA damage, cellular senescence, T cell homeostasis, and hyper-cytokinesis. On day 28, BPD-associated CpGs along with BPD-DEGs were enriched for phagocytosis, neurological disorder, and nucleotide metabolism. Oxygen supplementation markedly downregulated mitochondrial biogenesis genes and altered CpGs annotated to developmental genes. Prematurity-altered DNA methylation could cause abnormal lymphopoiesis, cellular assembly and cell cycle progression to increase BPD risk. Similar pathways between epigenome and transcriptome networks suggest coordination of the two in dysregulating leukopoiesis, adaptive immunity, and innate immunity. The results provide molecular insights into biomarkers for early detection and prevention of BPD.

Epigenomic profiling of isolated blood cell types reveals highly specific B cell smoking signatures and links to disease risk.

BACKGROUND: Tobacco smoking alters the DNA methylation profiles of immune cells which may underpin some of the pathogenesis of smoking-associated diseases. To link smoking-driven epigenetic effects in specific immune cell types with disease risk, we isolated six leukocyte subtypes, CD14+ monocytes, CD15+ granulocytes, CD19+ B cells, CD4+ T cells, CD8+ T cells, and CD56+ natural killer cells, from whole blood of 67 healthy adult smokers and 74 nonsmokers for epigenome-wide association study (EWAS) using Illumina 450k and EPIC methylation arrays. RESULTS: Numbers of smoking-associated differentially methylated sites (smCpGs) at genome-wide significance (p < 1.2 × 10-7) varied widely across cell types, from 5 smCpGs in CD8+ T cells to 111 smCpGs in CD19+ B cells. We found unique smoking effects in each cell type, some of which were not apparent in whole blood. Methylation-based deconvolution to estimate B cell subtypes revealed that smokers had 7.2% (p = 0.033) less naïve B cells. Adjusting for naïve and memory B cell proportions in EWAS and RNA-seq allowed the identification of genes enriched for B cell activation-related cytokine signaling pathways, Th1/Th2 responses, and hematopoietic cancers. Integrating with large-scale public datasets, 62 smCpGs were among CpGs associated with health-relevant EWASs. Furthermore, 74 smCpGs had reproducible methylation quantitative trait loci single nucleotide polymorphisms (SNPs) that were in complete linkage disequilibrium with genome-wide association study SNPs, associating with lung function, disease risks, and other traits. CONCLUSIONS: We observed blood cell-type-specific smCpGs, a naïve-to-memory shift among B cells, and by integrating genome-wide datasets, we identified their potential links to disease risks and health traits.

Impacts of the invasive Spartina anglica on C-S-Hg cycles and Hg(II) methylating microbial communities revealed by hgcA gene analysis in intertidal sediment of the Han River estuary, Yellow Sea.

We investigated the impact of invasive vegetation on mercury cycles, and identified microorganisms directly related to Hg(II) methylation using hgcA gene in vegetated mud flats (VMF) inhabited by native Suaeda japonica (SJ) and invasive Spartina anglica (SA), and unvegetated mud flats (UMF) in Ganghwa intertidal sediments. Sulfate reduction rate (SRR) and rate constants of Hg(II) methylation (Km) and methyl-Hg demethylation (Kd) were consistently greater in VMF than in UMF, specifically 1.5, 2 and 11.7 times higher, respectively, for SA. Both Km and Kd were significantly correlated with SRR and the abundance of sulfate-reducing bacteria. These results indicate that the rhizosphere of invasive SA provides a hotspot for Hg dynamics coupled with sulfate reduction. HgcA gene analysis revealed that Hg(II)-methylators were dominated by Deltaproteobacteria, Chloroflexi and Euryarchaeota, comprising 37.9%, 35.8%, and 6.5% of total hgcA gene sequences, respectively, which implies that coastal sediments harbor diverse Hg(II)-methylating microorganisms that previously underrepresented.

Targeted thrombolysis by magnetoacoustic particles in photothrombotic stroke model.

BACKGROUND: Recombinant tissue plasminogen activator (rtPA) has a short half-life, and additional hemorrhagic transformation (HT) can occur when treatment is delayed. Here, we report the design and thrombolytic performance of 3 [Formula: see text]m discoidal polymeric particles loaded with rtPA and superparamagnetic iron oxide nanoparticles (SPIONs), referred to as rmDPPs, to address the HT issues of rtPA. METHODS: The rmDPPs consisted of a biodegradable polymeric matrix, rtPA, and SPIONs and were synthesized via a top-down fabrication. RESULTS: The rmDPPs could be concentrated at the target site with magnetic attraction, and then the rtPA could be released under acoustic stimulus. Therefore, we named that the particles had magnetoacoustic properties. For the in vitro blood clot lysis, the rmDPPs with magnetoacoustic stimuli could not enhance the lytic potential compared to the rmDPPs without stimulation. Furthermore, although the reduction of the infarcts in vivo was observed along with the magnetoacoustic stimuli in the rmDPPs, more enhancement was not achieved in comparison with the rtPA. A notable advantage of rmDPPs was shown in delayed administration of rmDPPs at poststroke. The late treatment of rmDPPs with magnetoacoustic stimuli could reduce the infarcts and lead to no additional HT issues, while rtPA alone could not show any favorable prognosis. CONCLUSION: The rmDPPs may be advantageous in delayed treatment of thrombotic patients.

Epigenome-wide association study of bronchopulmonary dysplasia in preterm infants: results from the discovery-BPD program.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is a lung disease in premature infants caused by therapeutic oxygen supplemental and characterized by impaired pulmonary development which persists into later life. While advances in neonatal care have improved survival rates of premature infants, cases of BPD have been increasing with limited therapeutic options for prevention and treatment. This study was designed to explore the relationship between gestational age (GA), birth weight, and estimated blood cell-type composition in premature infants and to elucidate early epigenetic biomarkers associated with BPD. METHODS: Cord blood DNA from preterm neonates that went on to develop BPD (n = 14) or not (non-BPD, n = 93) was applied to Illumina 450 K methylation arrays. Blood cell-type compositions were estimated using DNA methylation profiles. Multivariable robust regression analysis elucidated CpGs associated with BPD risk. cDNA microarray analysis of cord blood RNA identified differentially expressed genes in neonates who later developed BPD. RESULTS: The development of BPD and the need for oxygen supplementation were strongly associated with GA (BPD, p < 1.0E-04; O2 supplementation, p < 1.0E-09) and birth weight (BPD, p < 1.0E-02; O2 supplementation, p < 1.0E-07). The estimated nucleated red blood cell (NRBC) percent was negatively associated with birth weight and GA, positively associated with hypomethylation of the tobacco smoke exposure biomarker cg05575921, and high-NRBC blood samples displayed a hypomethylation profile. Epigenome-wide association study (EWAS) identified 38 (Bonferroni) and 275 (false discovery rate 1%) differentially methylated CpGs associated with BPD. BPD-associated CpGs in cord blood were enriched for lung maturation and hematopoiesis pathways. Stochastic epigenetic mutation burden at birth was significantly elevated among those who developed BPD (adjusted p = 0.02). Transcriptome changes in cord blood cells reflected cell cycle, development, and pulmonary disorder events in BPD. CONCLUSIONS: While results must be interpreted with caution because of the small size of this study, NRBC content strongly impacted DNA methylation profiles in preterm cord blood and EWAS analysis revealed potential insights into biological pathways involved in BPD pathogenesis.

NRF2 Alters Mitochondrial Gene Expression in Neonate Mice Exposed to Hyperoxia.

Approximately 1 in 10 newborns are born preterm and require supplemental oxygen (O2) in an extrauterine environment following birth. Supplemental O2 can induce oxidative stress that can impair mitochondrial function, resulting in lung injury and increased risk in early life pulmonary diseases. The nuclear factor-erythroid 2 related factor 2 (NRF2) protects the cells from oxidative stress by regulating the expression of genes containing antioxidant response elements and many mitochondrial-associated genes. In this study, we compared Nrf2-deficient (Nrf2-/-) and wild-type (Nrf2+/+) mice to define the role of NRF2 in lung mitochondrial genomic features in late embryonic development in mice (embryonic days, E13.5 and E18.5) versus birth (postnatal day 0, PND0). We also determined whether NRF2 protects lung mitochondrial genome parameters in postnatal mice exposed to a 72 h hyperoxia environment. We found Nrf2-/- embryonic lungs were characterized by decreases in mtDNA copies from E13.5 to E18.5. Interestingly, Nrf2-/- heteroplasmy frequency was significantly higher than Nrf2+/+ at E18.5, though this effect reversed at PND0. In postnatal mice exposed to hyperoxia, we identified three- to four-fold increases in mitochondria-encoded mitochondrial genes, which regulate oxidative phosphorylation. Overall, our findings demonstrate a potentially critical role of NRF2 in mediating long-term effects of hyperoxia on mitochondrial function.

Shifts in benthic bacterial communities associated with farming stages and a microbiological proxy for assessing sulfidic sediment conditions at fish farms.

To assess the aquaculture-induced sediment conditions associated with sulfur cycles, shifts in bacterial communities across farming stages were investigated. The sulfate reduction rate (SRR), and concentrations of acid volatile sulfide (AVS) and H2S were significantly higher at the mid- and post-farming stages than at the early stage, indicating that the aquaculture effects persist even after harvest. Incomplete organic carbon-oxidizing sulfate-reducing bacteria (IO-SRB) affiliated with Desulfobulbaceae, and gammaproteobacterial sulfur oxidizing bacteria (SOB) (Thiohalobacter, Thioprofundum, and Thiohalomonas) were dominant during the early stage, whereas fermenting bacteria (Bacteroidetes and Firmicutes) and complete oxidizing SRB (CO-SRB) belonging to Desulfobacteraceae, and epsilonproteobacterial SOB (Sulfurovum) dominated during the mid- and post-stages. The shift in SRB and SOB communities well reflected the anoxic and sulfidic conditions of farm sediment. Especially, the Sulfurovum-like SOB correlated highly and positively with H2S, AVS, and SRR, suggesting that they could be relevant microbiological proxies to assess sulfidic conditions in farm sediment.

Murine Neonatal Oxidant Lung Injury: NRF2-Dependent Predisposition to Adulthood Respiratory Viral Infection and Protection by Maternal Antioxidant.

NRF2 protects against oxidant-associated airway disorders via cytoprotective gene induction. To examine if NRF2 is an important determinant of respiratory syncytial virus (RSV) susceptibility after neonate lung injury, Nrf2-deficient (Nrf2-/-) and wild-type (Nrf2+/+) mice neonatally exposed to hyperoxia were infected with RSV. To investigate the prenatal antioxidant effect on neonatal oxidative lung injury, time-pregnant Nrf2-/- and Nrf2+/+ mice were given an oral NRF2 agonist (sulforaphane) on embryonic days 11.5-17.5, and offspring were exposed to hyperoxia. Bronchoalveolar lavage and histopathologic analyses determined lung injury. cDNA microarray analyses were performed on placenta and neonatal lungs. RSV-induced pulmonary inflammation, injury, oxidation, and virus load were heightened in hyperoxia-exposed mice, and injury was more severe in hyperoxia-susceptible Nrf2-/- mice than in Nrf2+/+ mice. Maternal sulforaphane significantly alleviated hyperoxic lung injury in both neonate genotypes with more marked attenuation of severe neutrophilia, edema, oxidation, and alveolarization arrest in Nrf2-/- mice. Prenatal sulforaphane altered different genes with similar defensive functions (e.g., inhibition of cell/perinatal death and inflammation, potentiation of angiogenesis/organ development) in both strains, indicating compensatory transcriptome changes in Nrf2-/- mice. Conclusively, oxidative injury in underdeveloped lungs NRF2-dependently predisposed RSV susceptibility. In utero sulforaphane intervention suggested NRF2-dependent and -independent pulmonary protection mechanisms against early-life oxidant injury.

Transcriptomics Underlying Pulmonary Ozone Pathogenesis Regulated by Inflammatory Mediators in Mice.

Ozone (O3) is the predominant oxidant air pollutant associated with airway inflammation, lung dysfunction, and the worsening of preexisting respiratory diseases. We previously demonstrated the injurious roles of pulmonary immune receptors, tumor necrosis factor receptor (TNFR), and toll-like receptor 4, as well as a transcription factor NF-κB, in response to O3 in mice. In the current study, we profiled time-dependent and TNFR- and NF-κB-regulated lung transcriptome changes by subacute O3 to illuminate the underlying molecular events and downstream targets. Mice lacking Tnfr1/Tnfr2 (Tnfr-/-) or Nfkb1 (Nfkb1-/-) were exposed to air or O3. Lung RNAs were prepared for cDNA microarray analyses, and downstream and upstream mechanisms were predicted by pathway analyses of the enriched genes. O3 significantly altered the genes involved in inflammation and redox (24 h), cholesterol biosynthesis and vaso-occlusion (48 h), and cell cycle and DNA repair (48-72 h). Transforming growth factor-ß1 was a predicted upstream regulator. Lack of Tnfr suppressed the immune cell proliferation and lipid-related processes and heightened epithelial cell integrity, and Nfkb1 deficiency markedly suppressed lung cell cycle progress during O3 exposure. Common differentially regulated genes by TNFR and NF-κB1 (e.g., Casp8, Il6, and Edn1) were predicted to protect the lungs from cell death, connective tissue injury, and inflammation. Il6-deficient mice were susceptible to O3-induced protein hyperpermeability, indicating its defensive role, while Tnf-deficient mice were resistant to overall lung injury caused by O3. The results elucidated transcriptome dynamics and provided new insights into the molecular mechanisms regulated by TNFR and NF-κB1 in pulmonary subacute O3 pathogenesis.

Role for Mucin-5AC in Upper and Lower Airway Pathogenesis in Mice.

Mucin-5AC (MUC5AC) is a major secreted mucin in pathogenic airways. To determine its role in mucus-related airway disorders, Muc5ac-deficient (Muc5ac-/-) and wild-type (Muc5ac+/+) mice were compared in bleomycin-induced pulmonary fibrosis, respiratory syncytial virus (RSV) disease, and ozone toxicity. Significantly greater inflammation and fibrosis by bleomycin were developed in Muc5ac-/- lungs compared to Muc5ac+/+ lungs. More severe mucous cell metaplasia in fibrotic Muc5ac-/- lungs coincided with bronchial Muc2, Muc4, and Muc5b overexpression. Airway RSV replication was higher in Muc5ac-/- than in Muc5ac+/+ during early infection. RSV-caused pulmonary epithelial death, bronchial smooth muscle thickening, and syncytia formation were more severe in Muc5ac-/- compared to Muc5ac+/+. Nasal septal damage and subepithelial mucoserous gland enrichment by RSV were greater in Muc5ac-/- than in Muc5ac+/+. Ozone exposure developed more severe nasal airway injury accompanying submucosal gland hyperplasia and pulmonary proliferation in Muc5ac-/- than in Muc5ac+/+. Ozone caused periodic acid-Schiff-positive secretion only in Muc5ac-/- nasal airways. Lung E-cadherin level was relatively lower in Muc5ac-/- than in Muc5ac+/+ basally and after bleomycin, RSV, and ozone exposure. Results indicate that MUC5AC is an essential mucosal component in acute phase airway injury protection. Subepithelial gland hyperplasia and adaptive increase of other epithelial mucins may compensate airway defense in Muc5ac-/- mice.

Glutathione reductase deficiency alters lung development and hyperoxic responses in neonatal mice.

Cellular antioxidants protect against hyperoxic lung injury. The role of the glutathione (GSH) system in lung development and bronchopulmonary dysplasia (BPD) pathogenesis has not been systematically investigated. The current study utilized GSH reductase-deficient (Gsr-KO) neonatal mice to test the hypothesis that early disruption of the GSH system negatively impacts lung development and hyperoxic responses. Lungs from wild-type (Gsr-WT) and Gsr-KO mice were analyzed for histopathology, developmental markers, redox indices, and transcriptome profiling at different developmental stages following exposure to room air or hyperoxia (85% O2) for up to 14 d. Lungs from Gsr-KO mice exhibited alveolar epithelial dysplasia in the embryonic and neonatal periods with relatively normal lung architecture in adulthood. GSH and its oxidized form (GSSG) were 50-70% lower at E19-PND14 in Gsr-KO lungs than in age-matched Gsr-WT. Differential gene expression between Gsr-WT and Gsr-KO lungs was analyzed at discrete developmental stages. Gsr-KO lungs exhibited downregulated cell cycle and DNA damage checkpoint genes at E19, as well as lung lipid metabolism and surfactant genes at PND5. In addition to abnormal baseline lung morphometry, Gsr-KO mice displayed a blunted response to hyperoxia. Hyperoxia caused a more robust upregulation of the lung thioredoxin system in Gsr-KO compared to Gsr-WT. Gsr-dependent, hyperoxia-responsive genes were highly associated with abnormal cytoskeleton, skeletal-muscular function, and tissue morphology at PND5. Overall, our data in Gsr-KO mice implicate the GSH system as a key regulator of lung development, cellular differentiation, and hyperoxic responses in neonatal mice.

Mitochondrial biology in airway pathogenesis and the role of NRF2.

A constant improvement in understanding of mitochondrial biology has provided new insights into mitochondrial dysfunction in human disease pathogenesis. Impaired mitochondrial dynamics caused by various stressors are characterized by structural abnormalities and leakage, compromised turnover, and reactive oxygen species overproduction in mitochondria as well as increased mitochondrial DNA mutation frequency, which leads to modified energy production and mitochondria-derived cell signaling. The mitochondrial dysfunction in airway epithelial, smooth muscle, and endothelial cells has been implicated in diseases including chronic obstructive lung diseases and acute lung injury. Increasing evidence indicates that the NRF2-antioxidant response element (ARE) pathway not only enhances redox defense but also facilitates mitochondrial homeostasis and bioenergetics. Identification of functional or potential AREs further supports the role for Nrf2 in mitochondrial dysfunction-associated airway disorders. While clinical reports indicate mixed efficacy, NRF2 agonists acting on respiratory mitochondrial dynamics are potentially beneficial. In lung cancer, growth advantage provided by sustained NRF2 activation is suggested to be through increased cellular antioxidant defense as well as mitochondria reinforcement and metabolic reprogramming to the preferred pathways to meet the increased energy demands of uncontrolled cell proliferation. Further studies are warranted to better understand NRF2 regulation of mitochondrial functions as therapeutic targets in airway disorders.

Toll-like receptor 4-mediated respiratory syncytial virus disease and lung transcriptomics in differentially susceptible inbred mouse strains.

Respiratory syncytial virus (RSV) causes severe lower respiratory tract disease in infants, young children, and susceptible adults. The pathogenesis of RSV disease is not fully understood, although toll-like receptor 4 (TLR4)-related innate immune response is known to play a role. The present study was designed to determine TLR4-mediated disease phenotypes and lung transcriptomics and to elucidate transcriptional mechanisms underlying differential RSV susceptibility in inbred strains of mice. Dominant negative Tlr4 mutant (C3H/HeJ, HeJ, Tlr4Lps-d) and its wild-type (C3H/HeOuJ, OuJ, Tlr4Lps-n) mice and five genetically diverse, differentially responsive strains bearing the wild-type Tlr4Lps-n allele were infected with RSV. Bronchoalveolar lavage, histopathology, and genome-wide transcriptomics were used to characterize the pulmonary response to RSV. RSV-induced lung neutrophilia [1 day postinfection (pi)], epithelial proliferation (1 day pi), and lymphocytic infiltration (5 days pi) were significantly lower in HeJ compared with OuJ mice. Pulmonary RSV expression was also significantly suppressed in HeJ than in OuJ. Upregulation of immune/inflammatory (Cxcl3, Saa1) and heat shock protein (Hspa1a, Hsph1) genes was characteristic of OuJ mice, while cell cycle and cell death/survival genes were modulated in HeJ mice following RSV infection. Strain-specific transcriptomics suggested virus-responsive (Oasl1, Irg1, Mx1) and epidermal differentiation complex (Krt4, Lce3a) genes may contribute to TLR4-independent defense against RSV in resistant strains including C57BL/6J. The data indicate that TLR4 contributes to pulmonary RSV pathogenesis and activation of cellular immunity, the inflammasome complex, and vascular damage underlies it. Distinct transcriptomics in differentially responsive Tlr4-wild-type strains provide new insights into the mechanism of RSV disease and potential therapeutic targets.

Muc5ac null mice are predisposed to spontaneous gastric antro-pyloric hyperplasia and adenomas coupled with attenuated H. pylori-induced corpus mucous metaplasia.

Gastric cancer (GC) is the third leading cause of cancer-related deaths worldwide and is strongly associated with chronic Helicobacter pylori (Hp) infection. The ability of Hp to closely adhere to the gastric surface protective mucous layer containing mucins (MUC in humans and Muc in animals), primarily Muc5ac, is integral in the stepwise pathogenesis from gastritis to cancer. To probe the role of Muc5ac in Hp-induced gastric pathology, Muc5ac-/- and Muc5ac+/+ (WT) mice were experimentally infected with Hp Sydney strain (SS1). At 16 weeks and 32 weeks post infection (wpi), groups of mice were euthanized and evaluated for the following: gastric histopathological parameters, immunohistochemical expression of mucins (Muc5ac, Muc1, Muc2), Trefoil factor family proteins (Tff1 and Tff2), Griffonia (Bandeiraea) simplicifolia lectin II (GSL II) (mucous metaplasia marker) and Clusterin (Spasmolytic Polypeptide Expressing Metaplasia (SPEM) marker), Hp colonization density by qPCR and gastric cytokine mRNA levels. Our results demonstrate that Muc5ac-/- mice developed spontaneous antro-pyloric proliferation, adenomas and in one case with neuroendocrine differentiation; these findings were independent of Hp infection along with strong expression levels of Tff1, Tff2 and Muc1. Hp-infected Muc5ac-/- mice had significantly lowered gastric corpus mucous metaplasia at 16 wpi and 32 wpi (P = 0.0057 and P = 0.0016, respectively), with a slight reduction in overall gastric corpus pathology. GSII-positive mucous neck cells were decreased in Hp-infected Muc5ac-/- mice compared to WT mice and clusterin positivity was noted within metaplastic glands in both genotypes following Hp infection. Additionally, Hp colonization densities were significantly higher in Muc5ac-/- mice compared to WT at 16 wpi in both sexes (P = 0.05) along with a significant reduction in gastric Tnfα (16 wpi-males and females, P = 0.017 and P = 0.036, respectively and 32 wpi-males only, P = 0.025) and Il-17a (16 wpi-males) (P = 0.025). Taken together, our findings suggest a protective role for MUC5AC/Muc5ac in maintaining gastric antral equilibrium and inhibiting Hp colonization and associated inflammatory pathology.

The discovery BPD (D-BPD) program: study protocol of a prospective translational multicenter collaborative study to investigate determinants of chronic lung disease in very low birth weight infants.

BACKGROUND: Premature birth is a growing and serious public health problem affecting more than one of every ten infants worldwide. Bronchopulmonary dysplasia (BPD) is the most common neonatal morbidity associated with prematurity and infants with BPD suffer from increased incidence of respiratory infections, asthma, other forms of chronic lung illness, and death (Day and Ryan, Pediatr Res 81: 210-213, 2017; Isayama et la., JAMA Pediatr 171:271-279, 2017). BPD is now understood as a longitudinal disease process influenced by the intrauterine environment during gestation and modulated by gene-environment interactions throughout the neonatal and early childhood periods. Despite of this concept, there remains a paucity of multidisciplinary team-based approaches dedicated to the comprehensive study of this complex disease. METHODS: The Discovery BPD (D-BPD) Program involves a cohort of infants < 1,250 g at birth prospectively followed until 6 years of age. The program integrates analysis of detailed clinical data by machine learning, genetic susceptibility and molecular translation studies. DISCUSSION: The current gap in understanding BPD as a complex multi-trait spectrum of different disease endotypes will be addressed by a bedside-to-bench and bench-to-bedside approach in the D-BPD program. The D-BPD will provide enhanced understanding of mechanisms, evolution and consequences of lung diseases in preterm infants. The D-BPD program represents a unique opportunity to combine the expertise of biologists, neonatologists, pulmonologists, geneticists and biostatisticians to examine the disease process from multiple perspectives with a singular goal of improving outcomes of premature infants. TRIAL REGISTRATION: Does not apply for this study.

Impacts of typhoon-induced heavy rainfalls and resultant freshwater runoff on the partitioning of organic carbon oxidation and nutrient dynamics in the intertidal sediments of the Han River estuary, Yellow Sea.

Occurrence of typhoons accompanied by heavy precipitation has increased for the past 40 years in northeast Asia. To elucidate the impact of three consecutive typhoon-induced heavy rainfall events and resultant freshwater runoff on the partitioning of organic carbon (Corg) oxidation and nutrient dynamics, we investigated the geochemical constituents, the rate of anaerobic Corg oxidation, sulfate reduction (SR), iron reduction (FeR) and P speciation in the intertidal mud flat of the Han River estuary, Yellow Sea. Corg oxidation by SR and FeR and their metabolic products (∑CO2, NH4+, H2S, Fe2+) decreased significantly (P < 0.05) during and immediately after the heavy rainfall. Additional mesocosm experiments demonstrated that potential N2 production rates increased up to 2.4 times with increased nitrate concentrations during freshwater runoff. The results suggest that denitrification becomes a significant Corg oxidation pathway substituting for SR during high-nitrate freshwater runoff, which may remove substantial portion of the N introduced into the estuary. P speciation analysis further revealed that the concentrations of iron bound P decreased by 2.2 fold during the heavy rainfall compared to that measured before the rainfall. The results suggest that an excess supply of riverine Si keeps P from binding to Fe, thereby stimulating P release. Taking projections of enhanced rainfall events in the future into account, our results suggest that the intensified storm events and resultant riverine runoff induces a shift of Corg oxidation pathways in the sediments, which ultimately alters C-N-P-S-Fe dynamics and may deepen N-limiting conditions in coastal ecosystems of the Yellow Sea.

Sulforaphane enriched transcriptome of lung mitochondrial energy metabolism and provided pulmonary injury protection via Nrf2 in mice.

Nrf2 is essential to antioxidant response element (ARE)-mediated host defense. Sulforaphane (SFN) is a phytochemical antioxidant known to affect multiple cellular targets including Nrf2-ARE pathway in chemoprevention. However, the role of SFN in non-malignant airway disorders remain unclear. To test if pre-activation of Nrf2-ARE signaling protects lungs from oxidant-induced acute injury, wild-type (Nrf2+/+) and Nrf2-deficient (Nrf2-/-) mice were given SFN orally or as standardized broccoli sprout extract diet (SBE) before hyperoxia or air exposure. Hyperoxia-induced pulmonary injury and oxidation indices were significantly reduced by SFN or SBE in Nrf2+/+ mice but not in Nrf2-/- mice. SFN upregulated a large cluster of basal lung genes that are involved in mitochondrial oxidative phosphorylation, energy metabolism, and cardiovascular protection only in Nrf2+/+ mice. Bioinformatic analysis elucidated ARE-like motifs on these genes. Transcript abundance of the mitochondrial machinery genes remained significantly higher after hyperoxia exposure in SFN-treated Nrf2+/+ mice than in SFN-treated Nrf2-/- mice. Nuclear factor-κB was suggested to be a central molecule in transcriptome networks affected by SFN. Minor improvement of hyperoxia-caused lung histopathology and neutrophilia by SFN in Nrf2-/- mice implies Nrf2-independent or alternate effector mechanisms. In conclusion, SFN is suggested to be as a preventive intervention in a preclinical model of acute lung injury by linking mitochondria and Nrf2. Administration of SFN alleviated acute lung injury-like pathogenesis in a Nrf2-dependent manner. Potential AREs in the SFN-inducible transcriptome for mitochondria bioenergetics provided a new insight into the downstream mechanisms of Nrf2-mediated pulmonary protection.

Determinants of host susceptibility to murine respiratory syncytial virus (RSV) disease identify a role for the innate immunity scavenger receptor MARCO gene in human infants.

BACKGROUND: Respiratory syncytial virus (RSV) is the global leading cause of lower respiratory tract infection in infants. Nearly 30% of all infected infants develop severe disease including bronchiolitis, but susceptibility mechanisms remain unclear. METHODS: We infected a panel of 30 inbred strains of mice with RSV and measured changes in lung disease parameters 1 and 5days post-infection and they were used in genome-wide association (GWA) studies to identify quantitative trait loci (QTL) and susceptibility gene candidates. FINDINGS: GWA identified QTLs for RSV disease phenotypes, and the innate immunity scavenger receptor Marco was a candidate susceptibility gene; targeted deletion of Marco worsened murine RSV disease. We characterized a human MARCO promoter SNP that caused loss of gene expression, increased in vitro cellular response to RSV infection, and associated with increased risk of disease severity in two independent populations of children infected with RSV. INTERPRETATION: Translational integration of a genetic animal model and in vitro human studies identified a role for MARCO in human RSV disease severity. Because no RSV vaccines are approved for clinical use, genetic studies have implications for diagnosing individuals who are at risk for severe RSV disease, and disease prevention strategies (e.g. RSV antibodies).

A Polymorphic Antioxidant Response Element Links NRF2/sMAF Binding to Enhanced MAPT Expression and Reduced Risk of Parkinsonian Disorders.

The NRF2/sMAF protein complex regulates the oxidative stress response by occupying cis-acting enhancers containing an antioxidant response element (ARE). Integrating genome-wide maps of NRF2/sMAF occupancy with disease-susceptibility loci, we discovered eight polymorphic AREs linked to 14 highly ranked disease-risk SNPs in individuals of European ancestry. Among these SNPs was rs242561, located within a regulatory region of the MAPT gene (encoding microtubule-associated protein Tau). It was consistently occupied by NRF2/sMAF in multiple experiments and its strong-binding allele associated with higher mRNA levels in cell lines and human brain tissue. Induction of MAPT transcription by NRF2 was confirmed using a human neuroblastoma cell line and a Nrf2-deficient mouse model. Most importantly, rs242561 displayed complete linkage disequilibrium with a highly protective allele identified in multiple GWASs of progressive supranuclear palsy, Parkinson's disease, and corticobasal degeneration. These observations suggest a potential role for NRF2/sMAF in tauopathies and a possible role for NRF2 pathway activators in disease prevention.

Potential therapeutic targets in Nrf2-dependent protection against neonatal respiratory distress disease predicted by cDNA microarray analysis and bioinformatics tools.

Hyperoxia exposure of newborn rodents has served as a model for bronchopulmonary dysplasia (BPD) phenotypes found in a sub-population of human premature infants. We previously demonstrated that Nrf2 modulates molecular events during saccular-to-alveolar lung maturation and also has a protective role in the pathogenesis of hyperoxia-induced acute lung injury, mortality, arrest of saccular-to-alveolar transition, and lung injury, using Nrf2-deficient and wild-type neonate mice. In this review, we describe how whole-genome transcriptome analyses can identify the means through which Nrf2 transcriptionally modulates organ injury and morphology, cellular growth/proliferation, vasculature development, and immune response during BPD-like pathogenesis. We illustrate how recently developed bioinformatics tools can be used to identify sets of Nrf2-dependently modulated genes in the BPD model, and elucidate direct Nrf2 downstream targets and chemicals/drugs that may act on them. These approaches will provide significant insights into promising therapeutic agents for Nrf2-dependent treatments of complications of preterm birth like BPD.