Imaging-guided bioreactor for de-epithelialization and long-term cultivation of <i>ex vivo</i> rat trachea.

Abstract

Recent synergistic advances in organ-on-chip and tissue engineering technologies offer opportunities to create <i>in vitro</i>-grown tissue or organ constructs that can faithfully recapitulate their <i>in vivo</i> counterparts. Such <i>in vitro</i> tissue or organ constructs can be utilized in multiple applications, including rapid drug screening, high-fidelity disease modeling, and precision medicine. Here, we report an imaging-guided bioreactor that allows <i>in situ</i> monitoring of the lumen of <i>ex vivo</i> airway tissues during controlled <i>in vitro</i> tissue manipulation and cultivation of isolated rat trachea. Using this platform, we demonstrated partial removal of the rat tracheal epithelium (<i>i.e.</i>, de-epithelialization) without disrupting the underlying subepithelial cells and extracellular matrix. Through different tissue evaluation assays, such as immunofluorescent staining, DNA/protein quantification, and electron beam microscopy, we showed that the epithelium of the tracheal lumen can be effectively removed with negligible disruption in the underlying tissue layers, such as cartilage and blood vessel. Notably, using a custom-built micro-optical imaging device integrated with the bioreactor, the trachea lumen was visualized at the cellular level, and removal of the endogenous epithelium and distribution of locally delivered exogenous cells were demonstrated <i>in situ</i>. Moreover, the de-epithelialized trachea supported on the bioreactor allowed attachment and growth of exogenous cells seeded topically on its denuded tissue surface. Collectively, the results suggest that our imaging-enabled rat trachea bioreactor and localized cell replacement method can facilitate creation of bioengineered <i>in vitro</i> airway tissue that can be used in different biomedical applications.