Lymphoid Tissue Equivalent Module

Jump To: Mimicking Lymph Node Functionality | Results of Testing

In a natural human immune system, the lymph nodes are activated when the body is threatened by a pathogen, producing lymphocytes that help the body fight off infection. In the MIMIC™ System, the role of the lymph nodes is played by the Lymphoid Tissue Equivalent (LTE) module, an immunological tissue construct that functionally reproduces the environment of a human lymph node shown below with the right cell types in the right places at the right times. The core module for MIMIC™ systems is the Lymphoid Tissue Equivalent module.

In its current form, the LTE module comprises T cells, B cells, APCs and/or FDCs. When the DCs present their antigen to the right receptor-matched T cells, a chain reaction occurs: the antigen activates the T cells, which then activate the B cells to form antibodies. Immune stimulation results in the production and release of cytokines by lymphocytes. These cytokines and antibodies are testable predictive immune responses. After these cytokines are produced in the LTE module, they are then removed for testing and classification based on the criteria needed.

Mimicking Lymph Node Functionality

In the human body, two- and three-way interactions of key cells (dendritic cells, B cell, T cells and FDCs) occur in spatially segregated regions of the lymph nodes in a sequential order as shown below. These are important considerations if one is to develop a model to test human immunity in a model system. The LTE module is an engineered tissue construct that is essentially an artificial lymph node. The LTE module merges the right cell growth media with the appropriate cells in a suitable microenvironment.

After the antigen-presenting cells become activated in the Peripheral Tissue (PT) module, the media containing the activated dendritic cells is removed from the PT module and placed into the LTE module. In the LTE module , the dendritic cells induce antigen-specific cell expansion and cytokine production of T cells, and expansion, differentiation, and production of antigen specific antibodies for B cells.

The activated lymphocytes, cytokine profiles, and antibodies are fully characterized using well-accepted, commercially available methods (FACS, ELISpot and ELISA). Results are typically available within 7–21 days.

Sometimes individuals fail to show a response when tested with in vitro models of human immunity. The MIMIC™ system can help resolve why this occurs. With the help of our research partners, VaxDesign has been able to determine which cells are important in the immune response (shown above between FDCs, T cells, and B cells). Not only are cell types and cell environment important to a functional MIMIC™ system, but the timing of the interactions between the cell-cell interactions plays an important role as well.

Step 1: Ag uptake by DCs

Step 2: In Vitro DC activation of T cells

Step 3: In Vitro Ag-specific Ab responses

Results of Testing

VaxDesign is developing assay systems for the production of fully human antibody-secreting cells against naive and recall antigens. We routinely demonstrate that APCs trigger the differentiation of naive antigen-specific CD4+ and CD8+ T cells into helper and cytotoxic T lymphocytes (CTL), respectively, by measuring lymphoproliferation, effector function and intracellular cytokine production by these T cells. An example of gp120-specific CD4 T cell responses is shown below using HBV as the irrelevant Ag. Furthermore, we find that the magnitude of antigen-specific CD4+ T cell responses detected in our in vitro cultures increased following vaccination (individuals donate blood pre- and post-vaccination) suggests that specific responses generated in MIMIC™ assays are reflective of the immune status of the donor.

Considering the possibility that the MIMIC™ system could be used to generate T cell populations for cloning or clinical use, we have shown various mechanisms, such as the removal of regulatory T cells or inclusion of pro-T cell cytokines, to rapidly increase the number of antigen-specific helper T cells in MIMIC assays. The LTE module enables researchers to dissect mechanisms of action via inclusion or exclusion of various cell types.

We have also demonstrated the presence of multifunctional in vitro T cells in response to vaccine stimulus. Multifunctional CD8 T cell responses were observed by multi-parameter flow in response to vaccine stimulus for both a recall challenge (Flumist) and a naïve challenge (YF-VAX) in the in vitro MIMIC™ system. The magnitudes of both total and multifunctional naïve CD8 T cell responses were significantly increased via a second re-stimulation with DCs with the cultured cells, a process that resembles prime/boost vaccination strategies. These in vitro datasets are consistent with previous multi-parameter flow cytometry studies that demonstrate the delineation of T cells into distinct functional populations, helping assess the quality of the T cell response to vaccination.

We have extensive data that the MIMIC™ technology can be used to generate humoral responses specific for recall and primary antigens. The MIMIC™ assay system can routinely be used for the generation of rapid, sensitive, and robust culture method for the induction of either recall or naive antigen-specific B cell response against a variety of protein antigens.

To obtain physiologic immune responses, we have optimized the following experimental conditions:

• cell types
• kinetics
• culture media
• Number of cells for recall and naive responses
• DC:T cell ratio
• CD4+ T cell help
• Microenvironment
• FDCs