Horizontal and vertical microchamber platforms for evaluation of the paracellular permeability of an epithelial cell monolayer

Abstract

Epithelial cells serve as a barrier by tightly adhering to each other and contribute to the homeostasis of living organisms by controlling substance permeation. Therefore, evaluation of the barrier function is important in pharmaceutical development processes. However, the widely used Transwell-based assays require the development of the defect-free epithelial cell monolayer above several tens of mm², often resulting in low reproducibility and requiring a long incubation time. In addition, the culture surface of cells is far from the bottom of the well plate, making it difficult to observe the cell morphology using an optical microscope. Herein, we propose simple polydimethylsiloxane microfluidic devices for evaluating the barrier function of an epithelial monolayer using a microchamber array. After the formation of the epithelial monolayer over microchambers, the permeation of the marker molecules introduced above resulted in increased fluorescence intensity in microchambers, which was monitored using confocal laser scanning microscopy. We show that using this technique, alteration of the paracellular permeability induced by sodium caprate (C₁₀) and cytochalasin-D, permeation enhancing factors, can be elucidated. Furthermore, by tilting the microchamber device 90 degrees, the vertical cell section and microchambers were imaged in the same focal plane, allowing for live visualization of the passage of fluorescent substances across the cell monolayer. This technique is expected to be useful for investigating the relationship between paracellular permeability and cell morphology, which is unattainable through conventional methods.