Joanne Van der Velden

Crayfish Recognize the Faces of Fight Opponents

The capacity to associate stimuli underlies many cognitive abilities, including recognition, in humans and other animals. Vertebrates process different categories of information separately and then reassemble the distilled information for unique identification, storage and recall. Invertebrates have fewer neural networks and fewer neural processing options so study of their behavior may reveal underlying mechanisms still not fully understood for any animal. Some invertebrates form complex social colonies and are capable of visual memory–bees and wasps, for example. This ability would not be predicted in species that interact in random pairs without strong social cohesion; for example, crayfish. They have chemical memory but the extent to which they remember visual features is unknown. Here we demonstrate that the crayfish Cherax destructor is capable of visual recognition of individuals. The simplicity of their interactions allowed us to examine the behavior and some characteristics of the visual features involved. We showed that facial features are learned during face-to-face fights, that highly variable cues are used, that the type of variability is important, and that the learning is context-dependent. We also tested whether it is possible to engineer false identifications and for animals to distinguish between twin opponents.

TGF-β signaling in stromal cells acts upstream of FGF-10 to regulate epithelial stem cell growth in the adult lung

Tissue resident mesenchymal stromal cells (MSCs) contribute to tissue regeneration through various mechanisms, including the secretion of trophic factors that act directly on epithelial stem cells to promote epithelialization. However, MSCs in tissues constitute a heterogeneous population of stromal cells and different subtypes may have different functions. In this study we show that CD166(neg) and CD166(pos) lung stromal cells have different proliferative and differentiative potential. CD166(neg) lung stromal cells exhibit high proliferative potential with the capacity to differentiate along the lipofibroblastic and myofibroblastic lineages, whereas CD166(pos) lung stromal cells have limited proliferative potential and are committed to the myofibroblastic lineage. Moreover, we show that CD166(pos) lung stromal cells do not share the same epithelial-supportive capacity as their CD166(neg) counterparts, which support the growth of lung epithelial stem cell (EpiSPC) colonies in vitro. In addition, ex vivo expansion of lung stromal cells also resulted in the loss of epithelial-supportive capacity, which could be reinstated by inhibition of the TGF-β signaling pathway. We show that epithelial-supportive capacity correlated with the level of FGF-10 expression and the reactivation of several lung development-associated genes. In summary, these studies suggest that TGF-β signaling in stromal cells acts upstream of FGF-10 to regulate epithelial stem cell growth in the adult lung.

Airway disease: the use of large animal models for drug discovery.

Large animal models have contributed to our current understanding of respiratory pathophysiology and the effects of pulmonary disease modifying drugs. For drug development, the benefit of using large animals over smaller animal species is primarily due to the greater similarity between humans and equivalent sized animals in terms of gross anatomy, morphometry, structure and physiology of their respiratory systems. Thus, when appropriate lung structure and function are required for correctly assessing the efficacy of novel drugs, large animals can play an important role in the development of these drugs to combat respiratory disease. The most widely used and best characterised large animal for drug development has been the sheep model of asthma. Recently, large animal models for chronic obstructive pulmonary disease (COPD) and cystic fibrosis (CF) have been reported but thus far have not been used extensively for drug development. Some important limitations of using large animals are the large costs associated with this type of research, as well as the poorer understanding of disease mechanisms in these species relative to rodents. In this review we discuss the extent of correlations between preclinical testing performed in large animal models and the initial indication of clinical efficacy in ongoing clinical trials.

KCa3.1 Channel-Blockade Attenuates Airway Pathophysiology in a Sheep Model of Chronic Asthma

Background The Ca2+-activated K+ channel KCa3.1 is expressed in several structural and inflammatory airway cell types and is proposed to play an important role in the pathophysiology of asthma. The aim of the current study was to determine whether inhibition of KCa3.1 modifies experimental asthma in sheep. Methodology and Principal Findings Atopic sheep were administered either 30 mg/kg Senicapoc (ICA-17073), a selective inhibitor of the KCa3.1-channel, or vehicle alone (0.5% methylcellulose) twice daily (orally). Both groups received fortnightly aerosol challenges with house dust mite allergen for fourteen weeks. A separate sheep group received no allergen challenges or drug treatment. In the vehicle-control group, twelve weeks of allergen challenges resulted in a 60±19% increase in resting airway resistance, and this was completely attenuated by treatment with Senicapoc (0.25±12%; n = 10, P = 0.0147). The vehicle-control group had a peak-early phase increase in lung resistance of 82±21%, and this was reduced by 58% with Senicapoc treatment (24±14%; n = 10, P = 0.0288). Senicapoc-treated sheep also demonstrated reduced airway hyperresponsiveness, requiring a significantly higher dose of carbachol to increase resistance by 100% compared to allergen-challenged vehicle-control sheep (20±5 vs. 52±18 breath-units of carbachol; n = 10, P = 0.0340). Senicapoc also significantly reduced eosinophil numbers in bronchoalveolar lavage taken 48 hours post-allergen challenge, and reduced vascular remodelling. Conclusions These findings suggest that KCa3.1-activity contributes to allergen-induced airway responses, inflammation and vascular remodelling in a sheep model of asthma, and that inhibition of KCa3.1 may be an effective strategy for blocking allergen-induced airway inflammation and hyperresponsiveness in humans.

Increased vascular density is a persistent feature of airway remodeling in a sheep model of chronic asthma

ABSTRACT Background: Increases in blood vessel density and vascular area are now recognized as important features of remodeled airways in asthma. However, the time sequence for these vascular changes and whether they resolve in the absence of continued antigenic exposure is not well elucidated. The aim of the present study was to correlate progressive changes in airway vascularity with changes in functional airway responses in sheep chronically challenged with house dust mite (HDM) allergen, and to examine the resolution of vascular remodeling following allergen withdrawal. Methods: Progressive changes in vascular indices were examined in four spatially separate lung segments that received weekly challenges with HDM allergen for 0, 8, 16, or 24 weeks. Reversibility of these changes was assessed in a separate experiment in which two lung segments received 24 weeks of HDM challenges and either no rest or 12 weeks rest. Lung tissue was collected from each segment 7 days following the final challenge and vascular changes assessed by a morphometric analysis of airways immunohistochemically stained with an antibody against type IV collagen. Results: Blood vessel density and percent airway vascularity were significantly increased in bronchi following 24 weeks of HDM challenges compared to untreated controls (P < .05), but not at any of the other time-points. There was no significant correlation between vascular indices and airway responses to allergic or nonspecific stimuli. The increase in blood vessel density induced by repeated allergen exposures did not return to baseline levels following a 12-week withdrawal period from allergen. Conclusions: Our results show for the first time that the airways of sheep chronically exposed to HDM allergen undergo vascular remodeling. These findings show the potential of this large animal model for investigating airway angiogenesis in asthma.

Measurement and impact of remodeling in the lung: airway neovascularization in asthma.

Expansion of the airway wall vascular compartment has recently been established as a feature of asthma involving both enlargement of existing vascular structures and the formation of new vessels (angiogenesis). Both processes are likely to occur together and are fundamental for supporting the many aspects of tissue inflammation and remodeling manifest in the clinical symptoms of airway disease. Multiple growth factors are implicated in airway angiogenesis, with vascular endothelial growth factor among the most important. Other asthma-associated stimuli, including ADAM33, environmental tobacco smoke, and rhinovirus infection, are emerging as proangiogenic regulators. Increasing attention is also focused on the complex interplay of airway wall inflammatory and structural cells, including airway smooth muscle in driving expansion of the bronchial submucosal vascular plexus in asthma. Here, we provide a brief update on recent developments in this emerging area and highlight the potential role played by airway smooth muscle.

LysoTracker is a marker of differentiated alveolar type II cells

BackgroundLysoTracker Green DND-26 is a fluorescent dye that stains acidic compartments in live cells and has been shown to selectively accumulate in lamellar bodies in alveolar type II (AT2) cells in the lung. The aim of this study was to determine whether the accumulation of LysoTracker in lamellar bodies can be used to isolate viable AT2 cells by flow cytometry and track their differentiation in live-cell culture by microscopy.MethodsMouse lung cells were sorted on the basis of CD45negCD31negEpCAMposLysoTrackerpos expression and characterized by immunostaining for SP-C and cultured in a three-dimensional epithelial colony-forming unit (CFU-Epi) assay. To track AT2 cell differentiation, lung epithelial stem and progenitor cells were cultured in a CFU-Epi assay with LysoTracker-supplemented media.ResultsThe purity of sorted AT2 cells as determined by SP-C staining was 97.4% and viability was 85.3%. LysoTrackerpos AT2 cells generated SP-Cpos alveolar epithelial cell colonies in culture, and when added to the CFU-Epi culture medium, LysoTracker marked the differentiation of stem/progenitor-derived AT2 cells.ConclusionsThis study describes a novel method for isolating AT2 cells from mouse lungs. The high purity and viability of cells attained by this method, makes them suitable for functional analysis in vitro. The application of LysoTracker to live cell cultures will allow better assessment of the cellular and molecular mechanisms that regulate AT2 cell differentiation.

Assessment of Peripheral Airway Function following Chronic Allergen Challenge in a Sheep Model of Asthma

Background There is increasing evidence that the small airways contribute significantly to the pathophysiology of asthma. However, due to the difficulty in accessing distal lung regions in clinical settings, functional changes in the peripheral airways are often overlooked in studies of asthmatic patients. The aim of the current study was to characterize progressive changes in small airway function in sheep repeatedly challenged with house dust mite (HDM) allergen. Methodology/Principal Findings Four spatially separate lung segments were utilized for HDM challenges. The right apical, right medial, right caudal and left caudal lung segments received 0, 8, 16 and 24 weekly challenges with HDM respectively. A wedged-bronchoscope technique was used to assess changes in peripheral resistance (Rp) at rest, and in response to specific and non-specific stimuli throughout the trial. Allergen induced inflammatory cell infiltration into bronchoalveolar lavage and increases in Rp in response to HDM and methacholine were localized to treated lung segments, with no changes observed in adjacent lung segments. The acute response to HDM was variable between sheep, and was significantly correlated to airway responsiveness to methacholine (rs = 0.095, P<0.01). There was no correlation between resting Rp and the number of weeks of HDM exposure. Nor was there a correlation between the magnitude of early-phase airway response and the number of HDM-challenges. Conclusions Our findings indicate that airway responses to allergic and non-allergic stimuli are localized to specific treated areas of the lung. Furthermore, while there was a decline in peripheral airway function with HDM exposure, this decrease was not correlated with the length of allergen challenge.

Nebulized perflubron and carbon dioxide rapidly dilate constricted airways in an ovine model of allergic asthma

BackgroundThe low toxicity of perfluorocarbons (PFCs), their high affinity for respiratory gases and their compatibility with lung surfactant have made them useful candidates for treating respiratory diseases such as adult respiratory distress syndrome. We report results for treating acute allergic and non-allergic bronchoconstriction in sheep using S-1226 (a gas mixture containing carbon dioxide and small volumes of nebulized perflubron). The carbon dioxide, which is highly soluble in perflubron, was used to relax airway smooth muscle.MethodsSheep previously sensitized to house dust mite (HDM) were challenged with HDM aerosols to induce early asthmatic responses. At the maximal responses (characterised by an increase in lung resistance), the sheep were either not treated or treated with one of the following; nebulized S-1226 (perflubron + 12% CO2), nebulized perflubron + medical air, 12% CO2, salbutamol or medical air. Lung resistance was monitored for up to 20 minutes after cessation of treatment.In additional naïve sheep, a segmental bronchus was pre-contracted with methacholine (MCh) and treated with nebulized S-1226 administered via a bronchoscope catheter. Subsequent bronchodilatation was monitored by real time digital video recording.ResultsTreatment with S-1226 for 2 minutes following HDM challenge resulted in a more rapid, more profound and more prolonged decline in lung resistance compared with the other treatment interventions. Video bronchoscopy showed an immediate and complete (within 5 seconds) re-opening of MCh-constricted airways following treatment with S-1226.ConclusionsS-1226 is a potent and rapid formulation for re-opening constricted airways. Its mechanism(s) of action are unknown. The formulation has potential as a rescue treatment for acute severe asthma.

The Effects of Tumstatin on Vascularity, Airway Inflammation and Lung Function in an Experimental Sheep Model of Chronic Asthma

Tumstatin, a protein fragment of the alpha-3 chain of Collagen IV, is known to be significantly reduced in the airways of asthmatics. Further, there is evidence that suggests a link between the relatively low level of tumstatin and the induction of angiogenesis and inflammation in allergic airway disease. Here, we show that the intra-segmental administration of tumstatin can impede the development of vascular remodelling and allergic inflammatory responses that are induced in a segmental challenge model of experimental asthma in sheep. In particular, the administration of tumstatin to lung segments chronically exposed to house dust mite (HDM) resulted in a significant reduction of airway small blood vessels in the diameter range 10+–20 μm compared to controls. In tumstatin treated lung segments after HDM challenge, the number of eosinophils was significantly reduced in parenchymal and airway wall tissues, as well as in the bronchoalveolar lavage fluid. The expression of VEGF in airway smooth muscle was also significantly reduced in tumstatin-treated segments compared to control saline-treated segments. Allergic lung function responses were not attenuated by tumstatin administration in this model. The data are consistent with the concept that tumstatin can act to suppress vascular remodelling and inflammation in allergic airway disease.