Marie Geerlings

Female mice are more susceptible to the development of allergic airway inflammation than male mice.

Background In humans the prevalence of asthma is higher among females than among males after puberty. The reason for this phenomenon is not clear.

Macrophages Regulators of Sex Differences in Asthma

Females are more susceptible to development of asthma than are males. In a mouse model of ovalbumin-induced airway inflammation, with aggravated disease in females compared with males, we studied interactions between immune and resident lung cells during asthma development to elucidate which processes are affected by sex. We studied numbers of regulatory T cells (Tregs), effector T cells, myeloid dendritic cells (mDCs), and alternatively activated macrophages (AAMPhi), and their functional capabilities. Male and female mice had comparable Treg numbers in lung tissue and comparable Treg function, but effector T cells had expanded to a greater extent in lungs of females after ovalbumin exposure. This difference in T cell expansion was therefore not the result of lack of Treg control, but appeared to be driven by a greater number of inflammatory mDCs migrating from the lungs to lymph nodes in females. Resident lung cells can influence mDC migration, and AAMPhi in lung tissue were found to be involved. Artificially elevating the number of AAMPhi in lung tissue increased the migration of mDCs and airway inflammation. We found greater numbers of AAMPhi in female lungs than in males; we therefore postulate that AAMPhi are involved in increased airway inflammation found in female mice.

Maternal smoking during pregnancy induces airway remodelling in mice offspring

Children from smoking mothers have an increased risk of developing asthma for reasons largely unknown. The effects of maternal smoking during pregnancy on remodelling, allergic airway inflammation and hyperresponsiveness in offspring were investigated in an experimental asthma model. Mice were exposed to fresh air or cigarette smoke from 3 weeks prior to conception until birth. Offspring were exposed to house dust mite (HDM) or PBS intranasally four times per week from week 5 to week 10 after birth onwards. Maternal smoking increased airway smooth muscle layer, collagen III deposition and HDM-induced goblet cell numbers in offspring. It additionally increased methacholine responsiveness, which correlated significantly with increased airway smooth muscle layer and collagen deposition. Maternal smoking increased HDM-induced numbers of neutrophils and mast cells in lung tissue. No further effects were observed. Smoking during pregnancy induces airway remodelling in mice offspring, which may contribute to increased methacholine responsiveness. This takes place irrespective of allergen exposure but may worsen the outcome of the allergic stimulus, resulting in higher methacholine responsiveness in house dust mite-exposed offspring from smoking mothers when compared to nonsmoking mothers. The results provide a possible mechanism behind the association between maternal smoking and asthma.

Current smoking-specific gene expression signature in normal bronchial epithelium is enhanced in squamous cell lung cancer†

Cigarette smoking is the main risk factor for the development of squamous cell lung carcinoma (SCC). However, the smoking‐related molecular changes in SCC have not been studied. Gene expression studies in both histologically normal bronchial epithelium and SCC epithelial samples identified genes differentially expressed between current and ex‐smokers. Subsequently, expression levels of the smoking‐related genes in normal bronchial epithelium were compared with those in SCC cells, since we hypothesized that the smoking‐induced changes would be also deregulated in SCC. Gene expression profiles were generated using Agilent whole human genome microarrays on laser‐microdissected normal bronchial epithelium and SCC samples. Expression levels of 246 genes, mainly related to oxidative stress response, were significantly different between normal bronchial epithelium of current and ex‐smokers. Such a differential gene expression profile did not exist in SCC cells of smokers and ex‐smokers. Interestingly, when comparing SCC and normal bronchial epithelium from ex‐smokers, the vast majority of these 246 genes were also deregulated in SCC. When comparing SCC with normal epithelium from smokers, 22% of the up‐regulated genes showed a similar high expression in SCC whereas 79% of the down‐regulated genes were even further reduced in SCC as compared to current smokers. The down‐regulated genes included several tumour suppressor genes, such as C9orf9, INHBB, LRIG1, SCGB3A1, SERPINI2, STEAP3 and ZMYND10. Thus, our study shows that the majority of genes up‐regulated in normal bronchial epithelium of current smokers show similar high expression levels in SCC, while down‐regulated genes are even further repressed in SCC. Our data indicate that smoking‐related changes in normal bronchial epithelial cells persist in malignant transformed squamous cells. Copyright

Airway Inflammation and Remodeling in Two Mouse Models of Asthma: Comparison of Males and Females

Background: Asthma and especially severe asthma affect women more frequently than men. Since asthma severity correlates with remodeling changes in the lung, a female propensity to remodeling could be expected. We studied whether our previous observation that female mice have more pronounced airway inflammation than males is associated with more pronounced remodeling in two models of chronic allergic asthma. Methods: Male and female BALB/c mice were (1) sensitized and subsequently challenged with ovalbumin (OVA) for 4 weeks, or (2) exposed to house dust mite (HDM) for 5 weeks. In both models, allergic inflammation, remodeling, antigen-specific IgE and methacholine (MCh) responsiveness were assessed. Results: Females had higher antigen-specific serum IgE levels, higher numbers of eosinophils and were more responsive to MCh. In the OVA model, females also had higher levels of Th2 cytokines in lung tissue than males. Both sexes developed similar airway remodeling (smooth muscle layer thickness, collagen III deposition and goblet cell hyperplasia) in the two models. Conclusions: Combining results of an OVA- and a HDM-induced mouse model of allergic airway inflammation, we have shown that more severe allergic inflammation in females is not accompanied with more pronounced airway remodeling.

Effects of 4 months of smoking in mice with ovalbumin-induced airway inflammation.

Background The effects of smoking on asthma pathogenesis are complex and not well studied. We have shown recently that 3 weeks of smoking attenuates ovalbumin (OVA)‐induced airway inflammation in mice and that 4–6 months of smoking induces emphysema in mice without airway inflammation. Effects of combined long‐term smoking and OVA exposure have not been investigated so far.

Maternal smoking during pregnancy decreases Wnt signalling in neonatal mice

Epidemiological studies have shown that maternal smoking during pregnancy is a risk factor for the development of asthma. However, the mechanisms underlying the increased risk of developing asthma are largely unknown. We have shown that maternal smoking during pregnancy increases smooth muscle layer and collagen III deposition around the airways in mouse offspring in association with increased airway hyper-responsiveness.1 Other factors also appear to contribute to the development of increased airway hyper-responsiveness. We hypothesise that lung development is such a factor. Since genes in the Wnt-β-catenin pathway are essential for lung development and epithelial stem cell differentiation/expansion, we investigated the effects of maternal smoking during pregnancy on Wnt gene expression in lung tissue from neonatal offspring. Balb/c mice were exposed …

Heme oxygenase-1 prevents smoke induced B-cell infiltrates: a role for regulatory T cells?

BackgroundSmoking is the most important cause for the development of COPD. Since not all smokers develop COPD, it is obvious that other factors must be involved in disease development. We hypothesize that heme oxygenase-1 (HO-1), a protective enzyme against oxidative stress and inflammation, is insufficiently upregulated in COPD.The effects of HO-1 modulation on cigarette smoke induced inflammation and emphysema were tested in a smoking mouse model.MethodsMice were either exposed or sham exposed to cigarette smoke exposure for 20 weeks. Cobalt protoporphyrin or tin protoporphyrin was injected during this period to induce or inhibit HO-1 activity, respectively. Afterwards, emphysema development, levels of inflammatory cells and cytokines, and the presence of B-cell infiltrates in lung tissue were analyzed.ResultsSmoke exposure induced emphysema and increased the numbers of inflammatory cells and numbers of B-cell infiltrates, as well as the levels of inflammatory cytokines in lung tissue. HO-1 modulation had no effects on smoke induced emphysema development, or the increases in neutrophils and macrophages and inflammatory cytokines. Interestingly, HO-1 induction prevented the development of smoke induced B-cell infiltrates and increased the levels of CD4+CD25+ T cells and Foxp3 positive cells in the lungs. Additionally, the CD4+CD25+ T cells correlated positively with the number of Foxp3 positive cells in lung tissue, indicating that these cells were regulatory T cells.ConclusionThese results support the concept that HO-1 expression influences regulatory T cells and indicates that this mechanism is involved in the suppression of smoke induced B-cell infiltrates. The translation of this interaction to human COPD should now be pursued.

Increased levels of (class switched) memory B cells in peripheral blood of current smokers

There is increasing evidence that a specific immune response contributes to the pathogenesis of COPD. B-cell follicles are present in lung tissue and increased anti-elastin titers have been found in plasma of COPD patients. Additionally, regulatory T cells (Tregs) have been implicated in its pathogenesis as they control immunological reactions. We hypothesize that the specific immune response in COPD is smoke induced, either by a direct effect of smoking or as a result of smoke-induced lung tissue destruction (i.e. formation of neo-epitopes or auto antigens). Furthermore, we propose that Tregs are involved in the suppression of this smoke-induced specific immune response.The presence of B cells, memory B cells and Tregs was assessed by flow cytometry in peripheral blood of 20 COPD patients and 29 healthy individuals and related to their current smoking status.COPD patients had lower (memory) B-cell percentages and higher Treg percentages in peripheral blood than healthy individuals, with a significant negative correlation between these cells. Interestingly, current smokers had higher percentages of (class-switched) memory B cells than ex-smokers and never smokers, irrespective of COPD.This increase in (class-switched) memory B cells in current smokers is intriguing and suggests that smoke-induced neo-antigens may be constantly induced in the lung. The negative correlation between B cells and Tregs in blood is in line with previously published observations that Tregs can suppress B cells. Future studies focusing on the presence of these (class switched) memory B cells in the lung, their antigen specificity and their interaction with Tregs are necessary to further elucidate the specific B-cell response in COPD.

Localization and enhanced mRNA expression of the orphan chemokine receptor L-CCR in the lung in a murine model of ovalbumin-induced airway inflammation

Various CC chemokine receptors are expressed on effector cells in allergic inflammation and their distinct expression pattern may dictate, to a large extent, the migration of inflammatory cells to sites of airway inflammation. The lipopolysaccharide (LPS)-inducible CC chemokine receptor (L-CCR) is an orphan chemokine receptor that has previously been identified in the murine macrophage cell line RAW 264.7 and in murine brain glial cells. In this study we investigated the induction and localization of L-CCR mRNA expression in mouse lung after ovalbumin (OVA)-induced airway inflammation. Both RT-PCR experiments and in situ hybridization (ISH) experiments in whole lung sections revealed a rapid upregulation of L-CCR mRNA expression as early as 1 hr and 3 hr after OVA challenge. Expression was found predominantly in MAC3+ macrophages and in bronchial epithelium, as shown by ISH and immunohistochemistry (IHC). We demonstrated that L-CCR mRNA expression is strongly upregulated in mouse lung after OVA challenge and is localized in macrophages and bronchial epithelium. Regarding the likely role of L-CCR as a chemokine receptor with the putative ligand monocyte chemotactic protein-1 (MCP-1, CCL2), this receptor may have an important function in the early phase of airway inflammation.