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Trieu D.,University of Toronto | Waddell T.K.,University of Toronto | Waddell T.K.,Latner Thoracic Surgery Research Laboratories | McGuigan A.P.,University of Toronto
Biomicrofluidics | Year: 2014

Organization of airway epithelium determines ciliary beat direction and coordination for proper mucociliary clearance. Fluidic shear stresses have the potential to influence ciliary organization. Here, an in vitro fluidic flow system was developed for inducing long-term airflow shear stresses on airway epithelium with a view to influencing epithelial organization. Our system consists of a fluidic device for cell culture, integrated into a humidified airflow circuit. The fluidic device has a modular design and is made from a combination of polystyrene and adhesive components incorporated into a 6-well filter membrane insert. We demonstrate the system operates within physiologically relevant shear and pressure ranges and estimate the shear stress exerted on the epithelial cell layer as a result of air flow using a computational model. For both the bronchial epithelial cell line BEAS2B and primary human tracheal airway epithelial cells, we demonstrate that cells remain viable within the device when exposed to airflow for 24?h and that normal differentiation and cilia formation occurs. Furthermore, we demonstrate the utility of our device for exploring the impact of exposing cells to airflow: our tool enables quantification of cytoskeletal organization, and is compatible with in situ bead assays to assess the orientation of cilia beating. © 2014 AIP Publishing LLC. Source


Oliver J.R.,University of Toronto | Kushwah R.,University of Toronto | Wu J.,University of Toronto | Cutz E.,University of Toronto | And 3 more authors.
Laboratory Investigation | Year: 2011

E74-like transcription factor-3 (Elf3), a member of the E26 transformation-specific transcription factor family, is strongly expressed in epithelial-rich tissues, such as small intestine, fetal lung, and various lung cancers. Although previous studies have shown a defect in terminal differentiation of the small intestinal epithelium of Elf3-deficient (Elf3/) mice during embryonic development, very little is known about the role Elf3 may play in repair of the airway epithelium after injury. In order to investigate whether Elf3 is involved in regeneration of the bronchiolar epithelium after Clara cell-specific injury, we administered naphthalene to both wild-type (Elf3/) and Elf3/mice. Histopathological analysis revealed no significant difference in the extent of naphthalene-induced Clara cell necrosis between Elf3/mice and Elf3/mice. In the bronchiolar epithelium of Elf3/mice, there was a substantial delay in the kinetics of cell proliferation and mitosis along with Clara cell renewal, whereas in the peribronchiolar interstitium, there was a significantly greater level of cell proliferation and mitosis in Elf3/mice than in Elf3/mice. Last, the intensity of immunopositive signal for transforming growth factor-Β type II receptor, which is a well-known transcriptional target gene of Elf3 and involved in the induction of epithelial cell differentiation, was significantly lower in the bronchiolar epithelium of Elf3/mice when compared with Elf3/mice. Taken together, our results suggest that Elf3 plays an important role in the regulation of lung cell proliferation and differentiation during repair of the injured bronchiolar airway epithelium. © 2011 USCAP, Inc All rights reserved. Source


Soleas J.P.,University of Toronto | Soleas J.P.,Latner Thoracic Surgery Research Laboratories | Waddell T.K.,University of Toronto | Waddell T.K.,Latner Thoracic Surgery Research Laboratories | McGuigan A.P.,University of Toronto
Biomaterials Science | Year: 2015

Epithelial tissues are a critical component of all tubular organs. Engineering artificial epithelium requires an understanding of the polarization of epithelia: both apicobasal and in a planar fashion. Air liquid interface (ALI) culture is typically used to generate apicobasal polarized airway epithelium in vitro; however, this approach does not provide any signalling cues to induce morphological planar polarization of the generated epithelial layer. Here we describe a microgrooved gelatin hydrogel insert that can induce alignment of confluent epithelial cell sheets under ALI conditions to induce both apicobasal and morphologically planar polarized epithelium. Microgrooves are imprinted into the surface of the gelatin insert using elastomeric stamps moulded from a diffraction grating film and gels are stabilized by crosslinking with glutaraldehyde. We show that microgrooved gelatin inserts produce alignment of 3T3 fibroblasts and a number of epithelial cell lines (ARPE-19, BEAS2B and IMCD3 cells). Furthermore, we show that BEAS2B apicobasally polarize and form a similar density of cilia on both gelatin inserts and standard transwell filters used for ALI culture but that as apicobasal polarization progresses cell alignment on the grooves is lost. Our method provides a simple strategy that can easily be adopted by labs without microfabrication expertise for manipulating epithelial organization in transwell culture and studying the interplay of various polarization forces. This journal is © The Royal Society of Chemistry 2015. Source


Soleas J.P.,Latner Thoracic Surgery Research Laboratories | Soleas J.P.,University of Toronto | Paz A.,University of Toronto | Marcus P.,Latner Thoracic Surgery Research Laboratories | And 4 more authors.
Journal of Biomedicine and Biotechnology | Year: 2012

Airway epithelium is constantly presented with injurious signals, yet under healthy circumstances, the epithelium maintains its innate immune barrier and mucociliary elevator function. This suggests that airway epithelium has regenerative potential (I. R. Telford and C. F. Bridgman, 1990). In practice, however, airway regeneration is problematic because of slow turnover and dedifferentiation of epithelium thereby hindering regeneration and increasing time necessary for full maturation and function. Based on the anatomy and biology of the airway epithelium, a variety of tissue engineering tools available could be utilized to overcome the barriers currently seen in airway epithelial generation. This paper describes the structure, function, and repair mechanisms in native epithelium and highlights specific and manipulatable tissue engineering signals that could be of great use in the creation of artificial airway epithelium. © 2012 John P. Soleas et al. Source


Paz A.C.,University of Toronto | Soleas J.,Latner Thoracic Surgery Research Laboratories | Soleas J.,University of Toronto | Poon J.C.H.,University of Toronto | And 6 more authors.
Tissue Engineering - Part B: Reviews | Year: 2014

The epithelium is one of the most important tissue types in the body and the specific organization of the epithelial cells in these tissues is important for achieving appropriate function. Since many tissues contain an epithelial component, engineering functional epithelium and understanding the factors that control epithelial maturation and organization are important for generating whole artificial organ replacements. Furthermore, disruption of the cellular organization leads to tissue malfunction and disease; therefore, engineered epithelium could provide a valuable in vitro model to study disease phenotypes. Despite the importance of epithelial tissues, a surprisingly limited amount of effort has been focused on organizing epithelial cells into artificial polarized epithelium with an appropriate structure that resembles that seen in vivo. In this review, we provide an overview of epithelial tissue organization and highlight the importance of cell polarization to achieve appropriate epithelium function. We next describe the in vitro models that exist to create polarized epithelium and summarize attempts to engineer artificial epithelium for clinical use. Finally, we highlight the opportunities that exist to translate strategies from tissue engineering other tissues to generate polarized epithelium with a functional structure. © 2014 Mary Ann Liebert, Inc. Source

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