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Asthma is a chronic disease related to airway hyperresponsiveness and airway remodeling. Airway remodeling is the important reason of refractory asthma and is associated with differentiation of airway epithelia into myofibroblasts via epithelial-mesenchymal transition (EMT) to increase the process of subepithelial fibrosis. There is growing evidence that autophagy modulates remodeling. However, the underlying molecular mechanisms of these effects are still unclear. In this study, we hypothesized that Follistatin-like 1 (FSTL1) promotes EMT and airway remodeling by intensifying autophagy. With the use of transmission electron microscopy (TEM), double-membrane autophagosomes were detected in the airways of patients and mice. More autophagosomes were in patients with asthma and OVA-challenged mice compared with healthy controls. The expression of FSTL1 and beclin-1 was upregulated in the airways of patients with asthma and OVA-challenged mice, accompanied by airway EMT and remodeling. In OVA-challenged Fstl1+/- mice, the degree of airway remodeling and autophagy was decreased compared with control mice. The effects of FSTL1 on autophagy and EMT were also tested in 16HBE cells in vitro. Additionally, inhibition of autophagy by using LY-294002 and siRNA-ATG5 reduced the FSTL1-induced EMT in 16HBE cells, as measured by E-cadherin, N-cadherin, and vimentin expression. In line herewith, administration of LY-294002 reduced the expression of autophagy, EMT, and airway remodeling markers in FSTL1-challenged WT mice. Taken together, our study suggests that FSTL1 may induce EMT and airway remodeling by activating autophagy. These findings may provide novel avenues for therapeutic research targeting the autophagy and FSTL1 pathway, which may be beneficial to patients with refractory asthma. [ABSTRACT FROM AUTHOR]

MicroRNAs are a family of small noncoding RNAs that regulate the expression of their target proteins at the posttranscriptional level. Their functions cover almost every aspect of cell physiology. However, the roles of microRNAs in fetal lung development are not completely understood. The objective of this study is to investigate the regulation and molecular mechanisms of alveolar epithelial cell maturation during fetal lung development by miR-124. We discovered that miR-124 was downregulated during rat fetal lung development and predominantly expressed in the epithelial cells at late stage of the lung development. Overexpression of miR-124 with an adenovirus vector led to the inhibition of epithelial maturation in rat fetal lung organ cultures and fetal alveolar epithelial type II cells, as demonstrated by a decrease in the type II cell marker expression and an increase in glycogen content. We further demonstrated by luciferase reporter assays that miR-124 inhibited the NF-αB, cAMP/PKA, and MAPK/ERK pathways. In addition, nuclear factor I/B (NFIB), a critical protein in fetal lung maturation, was validated as a direct target of miR-124. Furthermore, miR-124 expression was induced by the Wnt/β-catenin signaling pathway through a direct interaction of LEF1 and the miR-124 promoter region. We concluded that miR-124 downregulation is critical to fetal lung epithelial maturation and miR-124 inhibits this maturation process at least partially through the inhibition of NFIB. [ABSTRACT FROM AUTHOR]

Cigarette smoke (CS) exposure increases the frequency and severity of respiratory tract infections. Despite this association, the mechanisms underlying the increased susceptibility to respiratory virus infection are poorly understood. Retinoic acid-inducible gene I (RIG-I) is an important regulator of influenza virus-induced expression of antiviral cytokines, mainly interferons (IFNs), which are necessary to clear viral infections. In this study, we compared the innate cytokine responses of two mouse CS exposure models following a challenge with influenza A virus (IAV): 1) exposure of the mice to cigarette smoke extract (CSE) intratracheally and 2) exposure of the mice to CS in a whole body exposure chamber. Both intratracheal CSE treatment and whole body CS exposure caused antiviral immunosuppression in these mice, and both CS exposure methods inhibited RIG-I induction. CS attenuated influenza-induced antiviral IFNs and IP-10 expression in vivo. However, we did not find that CS inhibited induction of the proinflammatory cytokines IL-6 and TNF-α, whose expression was induced by IAV. Interestingly, IAV infection also increased Toll-like receptor 3 (TLR3) expression in mouse lung, but CS exposure did not impact TLR3 induction in these mice. Together, the results support our previous finding in a human lung organ culture model that the suppression of RIG-I induction and antiviral cytokine responses by CS are likely important in the enhanced susceptibility of smokers to influenza infection in the lung. [ABSTRACT FROM AUTHOR]

UTP is known to regulate alveolar fluid clearance. However, the relative contribution of alveolar type I cells and type 11 cells to this process is unknown. In this study, we investigated the effects of UTP on ion transport in type I-like cell (AEC I) and type Il-like cell (AEC II) monolayers. Luminal treatment of cell monolayers with UTP increased short-circuit current (Isc) of AEC II but decreased Isc of AEC 1. The Cl- channel blockers NPPB and DIDS inhibited the UTP-induced changes in Isc (Δ/sc) in both types of cells. Amiloride, an inhibitor of epithelial Na+ channels (ENaC), abolished the UTP-induced Δ/sc in AEC I, but not in AEC II. The general blocker of K+ channels, BaCI2, eliminated the UTP-induced Δ/sc in AEC II, but not in AEC I. The intermediate conductance (lKca,) blocker, clofilium, also blocked the UTP effect in AEC II. The signal transduction pathways mediated by UTP were the same in AEC I and AEC II. Furthermore, UTP increased Cl- secretion in AEC II and Cl- absorption in AEC I. Our results suggest that UTP induces opposite changes in Isc in AEC I and AEC II, likely due to the reversed Cl- flux and different contributions of ENaC and lKCa. Our results further imply a new concept that type II cells contribute to UTP-induced fluid secretion and type I cells contribute to UTP-induced fluid absorption in alveoli. [ABSTRACT FROM AUTHOR]

Fetal lung development, is a complex biological process that involves temporal and spatial regulations of many genes. To understand the molecular mechanisms of this process, we investigated gene expression profiles of fetal lungs on gestational days 18, 19, 20, and 21, as well as newborn and adult rat lungs. For this analysis, we used an in-house rat DNA microarray containing 6,000 known genes and 4,000 expressed sequence tags (ESTs). Of these, 1,512 genes passed the statistical significance analysis of microarray (SAM) test; an at least twofold change was shown for 583 genes (402 known genes and 181 ESTs) between at least two time points. K-means cluster analysis revealed seven major expression patterns. In one of the clusters, gene expression increased from day 18 to day 20 and then decreased. In this cluster, which contained 10 known genes and 5 ESTs, 8 genes are associated with development. These genes can be integrated into regulatory pathways, including growth factors, plasma membrane receptors, adhesion molecules, intracellular signaling molecules, and transcription factors. Real-time PCR analysis of these 10 genes showed an 88% consistency with the microarray data. The mRNA of LIM homeodomain protein 3a (Lhx3), a transcription factor, was enriched in fetal type II cells. In contrast, pleiotrophin, a growth factor, had a much higher expression in fetal lung tissues than in fetal type II cells. Immunohistochemistry revealed that Lhx3 was localized in fetal lung epithelial cells and pleiotrophin in the mesenchymal cells adjacent to the developing epithelium and blood vessel. Using GenMAPP, we identified four regulatory pathways: transforming growth factor-β signaling, inflammatory response, cell cycle, and G protein signaling. We also identified two metabolic pathways: glycolysis-gluconeogenesis and proteasome degradation. Our results may provide new insights into the complex regulatory pathways that control fetal lung development. [ABSTRACT FROM AUTHOR]

Several studies have suggested that exposure to hyperoxia causes lung injury through increased generation of reactive oxygen and nitrogen species. The present study was aimed to investigate the effects of hyperoxia exposure on protein nitration in lungs. Rats were exposed to hyperoxia (>95%) for 48, 60, and 72 h. Histopathological analysis showed a dramatic change in the severity of lung injury in terms of edema and hemorrhage between 48- and 60-h exposure times. Western blot for nitrotyrosine showed that several proteins with molecular masses of 29-66 kDa were nitrated in hyperoxic lung tissues. Immunohistochemical analyses indicate nitrotyrosine staining of alveolar epithelial and interstitial regions. Furthermore, immunoprecipitation followed by Western blot revealed the nitration of surfactant protein A and t1α, proteins specific for alveolar epithelial type II and type I cells, respectively. The increased myeloperoxidase (MPO) activity and total nitrite levels in bronchoalveolar lavage and lung tissue homogenates were observed in hyperoxic lungs. Neutrophils and macrophages isolated from the hyperoxia-exposed rats, when cocultured with a rat lung epithelial L2 cell line, caused a significant protein nitration in L2 cells. Inclusion of nitrite further increased the protein nitration. These studies suggest that protein nitration during hyperoxia may be mediated in part by MPO generated from activated phagocytic cells, and such protein modifications may contribute to hyperoxia-mediated lung injury. [ABSTRACT FROM AUTHOR]