Using ex vivo lymph node slices to monitor changes in molecular transport after vaccination

Abstract

Abstract Understanding dynamics in the lymph node has the potential to reveal disease mechanisms and inform the design of immunotherapies. To this end, our laboratory combines ex vivo slices of murine lymph node (LN) tissue with novel microfluidic culture systems to study dynamic responses to events such as vaccination or the growth of a tumor. We have developed systems for co-culture of tissue slices from multiple organs, e.g. tumor and LN, and to deliver inflammatory stimuli to local regions of ex vivo cultures. Here, we used the latter system to assess changes in molecular diffusion through the LN after vaccination with adjuvants of varied mechanism. Mice were vaccinated with ovalbumin adjuvanted with Alum, Poly I:C, Complete Freud’s Adjuvant (CFA), or a vehicle control (saline). Lymph node slices were collected on day 4, and the diffusion coefficients of a set of non-binding dextrans were measured by using microfluidic integrated optical imaging (microIOI), a method developed previously by our laboratory (Ross et. al, Anal Chim Acta, 2019). Smaller dextrans (3 and 10 kDa), but not larger ones (70 kDa), diffused significantly faster after vaccination with Poly I:C than with Alum or saline. These data suggested an expansion of the extracellular space with an upper limit of 6.0 nm, the Stokes radius of 70 kDa dextran. Consistent with this prediction, the diffusion coefficients of an antibody immunotherapy and its isotype control (IgG, 150 kDa) remained unaltered after vaccination. Next, we will test the extent to which changes in the stromal network correlate with the changes in diffusivity after vaccination, and continue to explore the use of ex vivo LN slices to inform models of intercellular communication in this organ and immunotherapy design.