Tissue engineering is a dichotomy of scaffold-based and scaffold-free cultures. Scaffold-based cultures form highly organotypic structures but with low uniformity and throughput. Conversely, scaffold-free cultures create consistently sized and shaped cell aggregates with limited spheroid-like structure and function, thus restricting their use for accurate disease modeling. We hypothesized that combining aspects of each culturing format, we would produce highly organotypic structures of consistent size and shape for use in pulmonary modeling. To this end, we created a novel easily scalable, minimalistic design for a micropatterned hydrogel dish that increases reliability and efficiency in 3D cell culture. Our suspension cultures are a unique improvement on current methods to produce organotypic structures. By using soluble non-gelling concentration of basement membrane (BM), we created an organotypic lung model from three stable cells representing epithelial, vascular, and stromal cell populations in substantially less time a currently reported methods with similar properties. We observed that soluble BM promotes emergence of lumina comparable to mammalian lung airspaces. Using hypoxia induction techniques, we provide evidence for formation of branching, perfusable vasculature in pulmonary aggregates supplemented with soluble BM. Aside from these structural traits, we observed increased proliferation, survival, and 3D growth of aggregates. These results were established using IF, biotinylated-Matrigel and proteomic techniques. As proof of concept, we applied this method in modeling of lung fibrosis using TGF-Beta induction followed by testing two investigational antifibrotic drugs. Our results demonstrate a novel 3D culture method that creates organotypic models from stable cell lines. We anticipate this technology to pioneer creation of novel suspension-based organoids fostering consistent, expedited 3D culture. Recently we used our cells in a customizable miniaturized high-resolution 3D printed microfluidic device capable of 2-fold serial dilutions which we validated using a permeabilization assay. Proving these technologies have the potential to expedite patient-derived organoids for personalized drug screening using lung-on-a-chip assays.

Novel organotypic lung triculture technologies paving the way to personalized medicine

ACS Fall 2021

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.