Peripheral Tissue Module

The goal of the Peripheral Tissue module is the creation of a three-dimensional model that reliably mimics the immune process in peripheral tissues. An important component of the immune response is the capture of antigens by antigen-presenting cells (APCs). The APCs engulf and process antigen and may then traffic to the closest lymph node(s), where they interact with T and/or B cells to initiate antigen-specific immune responses. An essential aspect of the PT module is the reproduction of this process, by allowing autonomous generation of resident macrophages and APCs such as migratory dendritic cells (DC). The DCs have the capacity to migrate out of the site of antigen presentation and reach localized T and B cells to initiate antigen-specific immune responses. Tissue constructs based on endothelial cells and a 3D matrix have shown the in vitro potential for autonomous generation of monocyte-derived DCs and macrophages. In the PT module, based on one monolayer of endothelial cells grown to confluency over a 3D collagen matrix, monocytes from total PBMCs selectively extravasate in and differentiate into either resident macrophages or migratory DCs with potent antigen-presenting capacity.

The Peripheral Tissue module provides a good representation of what occurs in vivo:

  • Autonomous DC generation from monocytes without exogenous factors being added;
  • Similar DC development kinetics;
  • A wide array of DC differentiation and maturation states;
  • A natural selection of hematopoietic DC precursors from whole peripheral blood mononuclear cells (PBMCs) due to the presence of an endothelium;
  • Presence of a 3D extracellular matrix environment;
  • Extensive phenotypic analyses showing that the APCs in the PT module are essentially identical to that found in human dermal skin explants.

The HUVEC endothelium in the PT module is quiescent for most applications. However, this endothelium can be activated with various inflammatory signals such as interluekin-1β or TNFα. This leads to neutrophil migration into the endothelium. We show that the confluent endothelium on the collagen-membrane is quiescent via a neutrophil migration assay; thus it does not have a high inflammatory background. When endothelial cells are grown in the presence of an inflammatory mediator, they upregulate adhesion molecules like E-selectin, VCAM-1, and ICAM-1. Chemokines such as IL-8 are also produced, an attractant for the body’s main inflammatory cell, the neutrophil. However, when endothelial cells are cultured in a non-inflammatory environment or with constructs that have a low endotoxin level such as LPS, they do not express these molecules that promote neutrophil transendothelial trafficking.

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In vivo, neutrophils are only recruited into inflamed tissues. If neutrophil migration across the endothelium is evident in vitro, the endothelium must have upregulated neutrophil migration-promoting molecules in response to any one of a number of possible inflammatory mediators in the environment. The collagen-membrane PT structures are not activated in their native form, precisely what is desired in most cases to assess the innate responses of various adjuvants, vaccines, biologics, biologicals, chemicals and cosmetic formulations.

However, the Peripheral Tissue module can be used as a model of inflammation as well. We have tested this hypothesis for inflammation induced by age-related macular degeneration. We have found that drug delivery micro-particles accumulate at inflammatory sites and release complement inhibitors using both our in vitro peripheral tissue module and in vivo animal studies.