PROJECT SUMMARY/ABSTRACT
Local interactions in the immune system determine whether an immune response is protective or
destructive, fighting infection or initiating autoimmunity. Adaptive immunity begins in the lymph node (LN), a
highly organized and dynamic tissue. Currently, it is difficult to parse the role of inflammatory mediators or
rationally design therapies for chronic inflammatory disease such as artherosclerosis, rheumatoid arthritis and
multiple sclerosis, which together affect 5 – 7% of the Western population. We hypothesize that analyzing local
responses ex vivo in intact tissue will provide information not easily obtained from current methods (in vitro/in
vivo). Such experiments require new tools to analyze dynamics in the immune system, which we develop by
combining expertise in bioanalytical chemistry, microfluidics, and immunology.
In this project, we will develop a novel ex vivo model of immunity, using a hybrid of microfluidic
culture and LN slices. In Aim 1, we will establish long-term culture coupled with analysis methods for live
murine and human LN slices. Slice culture offers the advantage of preservation of the extracellular
microenvironment and any matrix-bound signals. We will optimize long-term culture (7-21 days) for murine LN
slices and human tonsil slices to maintain high viability, low cellular activation markers at rest, and ability to
respond to inflammatory and antigen-specific stimuli. In Aim 2, we will develop a novel microfluidic system for
on-demand local stimulation of LN slices. We will improve the spatial resolution of our previously developed
device, to target clusters 2 – 10 cells in diameter (20 – 100 µm lateral resolution) using short- and long-term
stimulation. We will also enable on-demand selection of delivery zone by using a mobile port, making the
whole tissue accessible with minimal handling. In Aim 3, we will validate the hybrid microfluidic-tissue slice
system for analysis of inflammatory responses and anti-inflammatory therapies. We will compare the
inflammatory response to a pro-inflammatory cytokine, TNF-a, in slices versus cell cultures and in vivo
systems. Finally, we will test the extent to which the model provides new information to guide immunotherapy,
by using the hybrid tissue-chip system with mouse and human tissue to compare the effects of competing
TNF-a inhibitors (anti-TNF-a monoclonals or soluble TNF-a receptor).
Combining local microfluidic stimulation with tissue slice technology produces the first experimental
platform for analysis of spatially organized signaling and cell-cell interactions in live LN tissue. This innovative
platform will advance both basic and translational biomedical research: locally delivered cytokines will serve as
a much-needed model of acute or chronic inflammation, and locally delivered immunotherapies will guide the
design of targeted drug-loaded nanoparticles. This technology is broadly applicable for a host of inflammatory
diseases, including rheumatoid arthritis, Chron’s disease, multiple sclerosis, Alzheimer’s disease, and cancer.