Open-source, three-dimensionally printed manifolds for exposure studies using human airway epithelial cells.

RATIONALE: In vitro studies using air-liquid interface (ALI) cultures enable controlled investigation of human airway epithelial cell (HAEC) responses to clinically relevant exposures. Commercial in vitro exposure systems provide precise and reproducible dosage but require significant investment. Exposure science may benefit from a more accessible customisable open-source exposure system. We present three-dimensionally (3D) printed manifolds for applying a range of exposures uniformly across standard, commercially available 6- and 24-well plates with ALI culture inserts. METHODS: A chamber-style exposure system and the designed manifolds were evaluated for exposure uniformity via simulations and deposition of nebulised fluorescein isothiocyanate (FITC)-labelled dextran. Chamber and manifolds were manufactured using 3D stereolithography printing. Cannabis concentrate vapour was generated from three different vaporisers and applied to well plates using the manifold system. Calu-3 cells and primary HAECs were cultured on Transwell inserts for exposure studies. RESULTS: The manifolds produced less variation in simulations and physical deposition of FITC-dextran aerosol across well plates compared to the chamber system. Distinct doses of cannabis concentrate vapour were delivered to well plates with minimal variation among wells. Whole tobacco smoke exposure using the manifold system induced functional changes in Calu-3 barrier function, cytokine production (interleukin (IL)-6 and IL-8) and cell membrane potential. Cannabis smoke led to reduced primary HAEC barrier function in a dose- and strain-dependent manner. CONCLUSIONS: Our data demonstrate the feasibility and the validity of our open-source 3D printed manifolds for use in studying multiple exposures and position our designs as an accessible option in parallel with commercially available systems.