Engineered human lymph node stroma model for examining interstitial fluid flow and T cell egress

Abstract

The lymph node (LN) performs essential roles in immunosurveillance throughout the body. Developing in vitro models of this key tissue is of great importance to enhancing physiological relevance in immunoengineering. The LN consists of stromal populations and immune cells, which are highly organized and bathed in constant interstitial flow. The stroma, notably the fibroblastic reticular cells (FRCs) and the lymphatic endothelial cells (LECs), play crucial roles in guiding T cell migration and are known to be sensitive to fluid flow. During inflammation, interstitial fluid flow rates drastically increase in the LN. It is unknown how these altered flow rates impact crosstalk and cell behavior in the LN, and most existing in vitro models focus on the interactions between T cells, B cells, and dendritic cells rather than with the stroma. To address this gap, we developed a human engineered model of the LN stroma consisting of FRC-laden hydrogel above a monolayer of LECs in a tissue culture insert with gravity-driven interstitial flow. We found that FRCs had enhanced coverage and proliferation in response to high flow rates, while LECs experienced decreased barrier integrity. We added CD4+ and CD8+ T cells and found that their egress was significantly decreased in the presence of interstitial flow, regardless of magnitude. Interestingly, 3.0 {micro}m/s flow, but not 0.8 {micro}m/s flow, correlated with enhanced inflammatory cytokine secretion in the LN stroma. Overall, we demonstrate that interstitial flow is an essential consideration in the lymph node for modulating LN stroma morphology, T cell migration, and inflammation.