Macrophages, circulating in the blood or concatenated into different organs and tissues constitute the first barrier against any disease. They are foremost controllers of both innate and acquired immunity, healthy tissue homeostasis, vasculogenesis and congenital metabolism. Two hallmarks of macrophages are diversity and plasticity due to which they acquire a wobbling array of phenotypes. These phenotypes are appropriately synchronized responses to a variety of different stimuli from either the tissue microenvironment or - microbes or their products. Based on the phenotype, macrophages are classified into classically activated/(M1) and alternatively activated/(M2) which are further sub-categorized into M2a, M2b, M2c and M2d based upon gene expression profiles. Macrophage phenotype metamorphosis is the regulating factor in initiation, progression, and termination of numerous inflammatory diseases. Several transcriptional factors and other factors controlling gene expression such as miRNAs contribute to the transformation of macrophages at different points in different diseases. Understanding the mechanisms of macrophage polarization and modulation of their phenotypes to adjust to the micro environmental conditions might provide us a great prospective for designing novel therapeutic strategy. In view of the above, this review summarises the activation of macrophages, the factors intricated in activation along with benefaction of macrophage polarization in response to microbial infections, pulmonary toxicity, lung injury and other inflammatory diseases such as chronic obstructive pulmonary dysplasia (COPD), bronchopulmonary dysplasia (BPD), asthma and sepsis, along with the existing efforts to develop therapies targeting this facet of macrophage biology.

Hyperoxia-induced acute lung injury (HALI) is a key contributor to the pathogenesis of bronchopulmonary dysplasia (BPD) in neonates, for which no specific preventive or therapeutic agent is available. Here we show that lung micro-RNA (miR)-34a levels are significantly increased in lungs of neonatal mice exposed to hyperoxia. Deletion or inhibition of miR-34a improves the pulmonary phenotype and BPD-associated pulmonary arterial hypertension (PAH) in BPD mouse models, which, conversely, is worsened by miR-34a overexpression. Administration of angiopoietin-1, which is one of the downstream targets of miR34a, is able to ameliorate the BPD pulmonary and PAH phenotypes. Using three independent cohorts of human samples, we show that miR-34a expression is increased in type 2 alveolar epithelial cells in neonates with respiratory distress syndrome and BPD. Our data suggest that pharmacologic miR-34a inhibition may be a therapeutic option to prevent or ameliorate HALI/BPD in neonates.

Premature infants are at high risk for developing bronchopulmonary dysplasia (BPD), characterized by chronic inflammation and inhibition of lung development, which we have recently identified as being modulated by microRNAs (miRNAs) and alterations in the airway microbiome. Exosomes and exosomal miRNAs may regulate cell differentiation and tissue and organ development. We discovered that tracheal aspirates from infants with severe BPD had increased numbers of, but smaller, exosomes compared with term controls. Similarly, bronchoalveolar lavage fluid from hyperoxia-exposed mice (an animal model of BPD) and supernatants from hyperoxia-exposed human bronchial epithelial cells (in vitro model of BPD) had increased exosomes compared with air controls. Next, in a prospective cohort study of tracheal aspirates obtained at birth from extremely preterm infants, utilizing independent discovery and validation cohorts, we identified unbiased exosomal miRNA signatures predictive of severe BPD. The strongest signal of reduced miR-876-3p in BPD-susceptible compared with BPD-resistant infants was confirmed in the animal model and in vitro models of BPD. In addition, based on our recent discovery of increased Proteobacteria in the airway microbiome being associated with BPD, we developed potentially novel in vivo and in vitro models for BPD combining Proteobacterial LPS and hyperoxia exposure. Addition of LPS led to a larger reduction in exosomal miR 876-3p in both hyperoxia and normoxia compared with hyperoxia alone, thus indicating a potential mechanism by which alterations in microbiota can suppress miR 876-3p. Gain of function of miR 876-3p improved the alveolar architecture in the in vivo BPD model, demonstrating a causal link between miR 876-3p and BPD. In summary, we provide evidence for the strong predictive biomarker potential of miR 876-3p in severe BPD. We also provide insights on the pathogenesis of neonatal lung disease, as modulated by hyperoxia and microbial product-induced changes in exosomal miRNA 876-3p, which could be targeted for future therapeutic development.

Administration of supplemental oxygen remains a critical clinical intervention for survival of preterm infants with respiratory failure. However, prolonged exposure to hyperoxia can augment pulmonary damage, resulting in developmental lung diseases embodied as hyperoxia-induced acute lung injury and bronchopulmonary dysplasia (BPD). We sought to investigate the role of autophagy in hyperoxia-induced apoptotic cell death in developing lungs. We identified increased autophagy signaling in hyperoxia-exposed mouse lung epithelial-12 cells, freshly isolated fetal type II alveolar epithelial cells, lungs of newborn wild-type mice, and human newborns with respiratory distress syndrome and evolving and established BPD. We found that hyperoxia exposure induces autophagy in a Trp53-dependent manner in mouse lung epithelial-12 cells and in neonatal mouse lungs. Using pharmacological inhibitors and gene silencing techniques, we found that the activation of autophagy, upon hyperoxia exposure, demonstrated a protective role with an antiapoptotic response. Specifically, inhibiting regulatory-associated protein of mechanistic target of rapamycin (RPTOR) in hyperoxia settings, as evidenced by wild-type mice treated with torin2 or mice administered (Rptor) silencing RNA via intranasal delivery or Rptor 1/2 , limited lung injury by increased autophagy, decreased apoptosis, improved lung architecture, and increased survival. Furthermore, we identified increased protein expression of phospho-beclin1, light chain-3-II and lysosomalassociated membrane protein 1, suggesting altered autophagic flux in the lungs of human neonates with established BPD. Collectively, our study unveils a novel demonstration of enhancing autophagy and antiapoptotic effects, specifically through the inhibition of RPTOR as a potentially useful therapeutic target for the treatment of hyperoxia-induced acute lung injury and BPD in developing lungs.Keywords: cell death; oxygen; newborn; pulmonary; bronchopulmonary dysplasia Premature infants provided with supplemental oxygen are at higher risk for developing a chronic respiratory disease, bronchopulmonary dysplasia (BPD) (1). BPD occurs in developing lungs resulting in impaired alveolarization and dysregulated vascularization-the pathologic hallmarks (2). Premature infants diagnosed with BPD have diminished pulmonary function and reduced lung capacity as they grow older and up to adulthood (3). This disease represents a common yet complicated clinical issue associated with significant long-term morbidity and mortality (2, 3). Pulmonary-specific oxygen toxicity is This work was supported in part by National Heart, Lung, and Blood Institute (NHLBI)/National Institutes of Health (NIH) grants HL63039 (G.P.) and HL116632 (A.R.), by the Sigrid Jusélius Foundation (S.A.), and by NHLBI/NIH grant HL85103 (V.B.).Author Contributions: Concept and design-A.S. and V.B.; acquisition of data-A.S., M.S., P.D., C.J., G.P., A.R., S.A., R.J.H., and V.B.; data analysis and interpretation-A.S., M.S., P.D., C.J., G.P., A.R., S...

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In Gram-negative bacterial sepsis, production of excess pro-inflammatory cytokines results in hyperinflammation and tissue injury. Anti-inflammatory cytokines such as IL-10 inhibit inflammation and enhance tissue healing. Here, we report a novel approach to treat septicemia associated with intra-abdominal infection in a murine model by delicately balancing pro- and anti-inflammatory cytokines. A novel oligosaccharide compound AVR-25 selectively binds to the TLR4 protein (IC 50 = 0.15 µM) in human peripheral blood monocytes and stimulates IL-10 production. Following the cecal ligation and puncture (CLP) procedure, intravenous dosing of AVR-25 (10 mg/kg, 6–12 h post-CLP) alone and in combination with antibiotic imipenem protected both young adult (10–12 week old) and aged (16–18 month old) mice against polymicrobial infection, organ dysfunction, and death. Proinflammatory cytokines (TNF-α, MIP-1, i-NOS) were decreased significantly and restoration of tissue damage was observed in all organs. A decrease in serum C-reactive protein (CRP) and bacterial colony forming unit (CFU) confirmed improved bacterial clearance. Together, these findings demonstrate the therapeutic ability of AVR-25 to mitigate the storm of inflammation and minimize tissue injury with high potential for adjunctive therapy in intra-abdominal sepsis.

SUMMARYBmp4 expression is tightly regulated during embryonic tooth development, with early expression in the dental epithelial placode leading to later expression in the dental mesenchyme. Msx1 is among several transcription factors that are induced by epithelial Bmp4 and that, in turn, are necessary for the induction and maintenance of dental mesenchymal Bmp4 expression. Thus, Msx1 −/− teeth arrest at early bud stage and show loss of Bmp4 expression in the mesenchyme. Ectopic expression of Bmp4 rescues this bud stage arrest. We have identified Tbx2 expression in the dental mesenchyme at bud stage and show that this can be induced by epithelial Bmp4. We also show that endogenous Tbx2 and Msx1 can physically interact in mouse C3H10T1/2 cells. In order to ascertain a functional relationship between Msx1 and Tbx2 in tooth development, we crossed Tbx2 and Msx1 mutant mice. Our data show that the bud stage tooth arrest in Msx1 −/− mice is partially rescued in Msx1;Tbx2 +/− compound mutants. This rescue is accompanied by formation of the enamel knot (EK) and by restoration of mesenchymal Bmp4 expression. Finally, knockdown of Tbx2 in C3H10T1/2 cells results in an increase in Bmp4 expression. Together, these data identify a novel role for Tbx2 in tooth development and suggest that, following their induction by epithelial Bmp4, Msx1 and Tbx2 in turn antagonistically regulate odontogenic activity that leads to EK formation and to mesenchymal Bmp4 expression at the key bud-to-cap stage transition.KEY WORDS: Bmp4, Msx1, Tbx2, Protein-protein interaction, Tooth

Glial-derived neurotrophic factor (Gdnf) is required for morphogenesis of the enteric nervous system (ENS) and it has been shown to regulate proliferation, differentiation, and survival of cultured enteric neural crest-derived cells (ENCCs). The goal of this study was to investigate its in vivo role in the colon, the site most commonly affected by intestinal neuropathies such as Hirschsprung's disease. Gdnf activity was modulated in ovo in the distal gut of avian embryos using targeted retrovirus-mediated gene overexpression and retroviral vector-based gene silencing. We find that Gdnf has a pleiotropic effect on colonic ENCCs, promoting proliferation, inducing neuronal differentiation, and acting as a chemoattractant. Down-regulating Gdnf similarly induces premature neuronal differentiation, but also inhibits ENCC proliferation, leading to distal colorectal aganglionosis with severe proximal hypoganglionosis. These results indicate an important role for Gdnf signaling in colonic ENS formation and emphasize the critical balance between proliferation and differentiation in the developing ENS.