Multi-organ immunity on a 3D-printed multi-tissue chip with tubing-free impeller pump

Multi-organ-on-chip systems (MOOCs) have the potential to mimic communication between organ systems and reveal mechanisms of health and disease. However, many existing MOOCs are challenging for non-experts to implement, due to complex tubing, electronics, or pump mechanisms. In addition, few MOOCs have incorporated immune organs such as the lymph node (LN), limiting their applicability to critical events such as vaccination. Here we developed a 3D-printed, user-friendly device and companion tubing-free impeller pump to co-culture two or more tissue samples, including a LN, under a recirculating common media. Native tissue structure and immune function were incorporated by maintaining slices of murine LN tissue ex vivo in 3Dprinted mesh supports for at least 24 hr. In a two-compartment model of a LN and an upstream injection site, vaccination of the multi-tissue chip was similar to in vivo vaccination in terms of locations of antigen accumulation and acute changes in activation markers and gene expression in the LN. We anticipate that in the future, this flexible platform will enable models of multi-organ immune responses throughout the body. Overall design: We compared the response to vaccination in murine lymph nodes, where the vaccination was performed in a multi-tisse chip, in vivo, or in standard well plate (wells). Our model vaccine was comprised of R848 as the adjuvant mixed with rhodamine-labeled ovalbumin (Rho-OVA) as the antigen. PBS was used as a negative control for vaccination. Following 6 hrs, RNA was isolated from whole lymph nodes after in vivo vaccination or sliced lymph nodes after on-chip and well plate conditions. Each sample from in vivo vaccination was comprised of two whole lymph nodes from two mice pooled together. Each sample from both on-chip and well plate vaccination was comprised of 6 total lymph node slices from two mice pooled together. This experiment was replicated three times. RNA was analyzed by bulk RNA sequencing.