Early-life exposure to tobacco smoke alters airway signaling pathways and later mortality in D. melanogaster.

Early life environmental influences such as exposure to cigarette smoke (CS) can disturb molecular processes of lung development and thereby increase the risk for later development of chronic respiratory diseases. Among the latter, asthma and chronic obstructive pulmonary disease (COPD) are the most common. The airway epithelium plays a key role in their disease pathophysiology but how CS exposure in early life influences airway developmental pathways and epithelial stress responses or survival is poorly understood. Using Drosophila melanogaster larvae as a model for early life, we demonstrate that CS enters the entire larval airway system, where it activates cyp18a1 which is homologues to human CYP1A1 to metabolize CS-derived polycyclic aromatic hydrocarbons and further induces heat shock protein 70. RNASeq studies of isolated airways showed that CS dysregulates pathways involved in oxidative stress response, innate immune response, xenobiotic and glutathione metabolic processes as well as developmental processes (BMP, FGF signaling) in both sexes, while other pathways were exclusive to females or males. Glutathione S-transferase genes were further validated by qPCR showing upregulation of gstD4, gstD5 and gstD8 in respiratory tracts of females, while gstD8 was downregulated and gstD5 unchanged in males. ROS levels were increased in airways after CS. Exposure to CS further resulted in higher larval mortality, lower larval-pupal transition, and hatching rates in males only as compared to air-exposed controls. Taken together, early life CS induces airway epithelial stress responses and dysregulates pathways involved in the fly's branching morphogenesis as well as in mammalian lung development. CS further affected fitness and development in a highly sex-specific manner.

Different responses of the oral, nasal and lung microbiomes to cigarette smoke.

To examine the role of smoking on the bacterial community composition of the upper and the lower respiratory tract, a monocentric, controlled prospective study was performed, including healthy smokers, ex-smokers and never-smokers. Smokers were further grouped according to their smoking history. Bacterial diversity was analysed using a molecular barcoding approach based on directly extracted DNA. Our study shows for the first time distinct bacterial response patterns in the upper and lower respiratory tract to cigarette smoking leading to a higher abundance of opportunistic pathogens. The clinical significance of these dysbioses for health needs to be further explored.

Cigarette Smoke Affects Dendritic Cell Populations, Epithelial Barrier Function, and the Immune Response to Viral Infection With H1N1.

Smokers with apparently "healthy" lungs suffer from more severe and frequent viral respiratory infections, but the mechanisms underlying this observation are still unclear. Epithelial cells and dendritic cells (DC) form the first line of defense against inhaled noxes such as smoke or viruses. We therefore aimed to obtain insight into how cigarette smoke affects DCs and epithelial cells and how this influences the response to viral infection. Female C57BL/6J mice were exposed to cigarette smoke (CS) for 1 h daily for 24 days and then challenged i.n. with the viral mimic and Toll-like receptor 3 (TLR3) ligand poly (I:C) after the last exposure. DC subpopulations were analyzed 24 h later in whole lung homogenates by flow cytometry. Calu-3 cells or human precision-cut lung slices (PCLS) cultured at air-liquid interface were exposed to CS or air and subsequently inoculated with influenza H1N1. At 48 h post infection cytokines were analyzed by multiplex technology. Cytotoxic effects were measured by release of lactate dehydrogenase (LDH) and confocal imaging. In Calu-3 cells the trans-epithelial electrical resistance (TEER) was assessed. Smoke exposure of mice increased numbers of inflammatory and plasmacytoid DCs in lung tissue. Additional poly (I:C) challenge further increased the population of inflammatory DCs and conventional DCs, especially CD11b+ cDCs. Smoke exposure led to a loss of the barrier function in Calu-3 cells, which was further exaggerated by additional influenza H1N1 infection. Influenza H1N1-induced secretion of antiviral cytokines (IFN-α2a, IFN-λ, interferon-γ-induced protein 10 [IP-10]), pro-inflammatory cytokine IL-6, as well as T cell-associated cytokines (e.g., I-TAC) were completely suppressed in both Calu-3 cells and human PCLS after smoke exposure. In summary, cigarette smoke exposure increased the number of inflammatory DCs in the lung and disrupted epithelial barrier functions, both of which was further enhanced by viral stimulation. Additionally, the antiviral immune response to influenza H1N1 was strongly suppressed by smoke. These data suggest that smoke impairs protective innate mechanisms in the lung, which could be responsible for the increased susceptibility to viral infections in "healthy" smokers.