Custom-Designed Wells and Flow Chamber for Exposing Air–Liquid Interface Cultures to Wall Shear Stress

Abstract

The effects of mechanical stimuli such as wall shear stresses (WSS) on cellular processes have been studied in vitro in numerous cell types. In order to study WSS effects on cells cultured under air-liquid interface (ALI) conditions, we developed a custom-designed well that can be disassembled into sub-units that allow installation of the cultured cells in a flow chamber, and then, re-assembled for further incubation or biological tests. Human nasal epithelial cells were cultured in the new wells under ALI conditions, and some of their biological characteristics were compared with those cultured in commercial Millicells. The cultured cells from both types of wells secreted the same amount of mucin and had similar cytoskeletal structures. Preliminary WSS experiments demonstrated the advantage of the new wells and provided initial indications that WSS affects the performance of ALI cultured respiratory epithelial cells.

Wall Shear Stress Effects on Endothelial-Endothelial and Endothelial-Smooth Muscle Cell Interactions in Tissue Engineered Models of the Vascular Wall

“…The cell cultures for the in vitro models of the vascular wall in this work were grown in modified wells that were previously developed in our laboratory [36]. Briefly, these wells are constructed from a well bottom and a cylinder.…”

Role of mechanical stress in regulating airway surface hydration and mucus clearance rates

“…3A, accelerates/decelerates human airway epithelial cultures inside a highly humidified incubator to generate "thin-film" liquid flow over airway epithelial surfaces whilst avoiding airflow-induced dehydration of the ASL (Tarran et al, 2005). The change in relative velocities of ASL and the underlying epithelium caused by this oscillatory motion generates a shear stress over the apical surface with profiles similar to airflow-induced shear stress generated during tidal breathing (Tarran et al, 2005;Elad et al, 2006;Even-Tzur et al, 2006). The acceleration rate is controlled so that the magnitude of shear stress at the epithelial surface can be varied over a wide range of shear stress (0.01 to 10 dynes·cm −2 ) at 28 cycles per minute (CPM), a rate approximating the combined number of both inhalations and exhalations in one minute.…”

Air–Liquid Interface Culture of Nasal Epithelial Cells on Denuded Amniotic Membranes

“…12 Several studies investigated the biological response of cultured airway and NEC to airflow induced WSS, and specifically the changes in mucin secretion and the cytoskeleton due to such stimuli. 14,15,45 In the current study we examined the strength of cells attachment to the substrate by applying different levels of WSS on top of the different cultures. The application of a relatively high WSS of 20 dyn/cm 2 induced detachment of the cells from the synthetic membrane while they remained attached to the denuded .…”

“…The cells were then cultured on the denuded AM under ALI conditions as previously described. [14][15][16]37 The epithelial cells were dissociated from the excised turbinates after incubation in pronase solution and scraping of the cells from the tissues. The cell suspension was then centrifuged, washed and resuspended in a serum-free hormone culture bronchial epithelial growth medium (BEGM) supplemented with additives.…”

“…14,15 The chamber is an 18 cm long conduit with a rectangular cross-sectional area of 20 mm 9 10 mm that can be connected to inlet and outlet tubes. The chamber base can host up to 3 well-bottoms in carefully designed holes which are filled with culture medium that is in contact with the inferior plane of the membrane in the well-bottom throughout the experiment.…”