Construction of an in vitro lymphoid organoid: studying innate-adaptive immune cell interaction in a 3D culture system

Lead Research Organisation: University of Nottingham
Department Name: School of Medicine

Abstract

The immune system is made up of two components, innate and adaptive. Innate immune cells, mainly dendritic cells, fight pathogens using defences that are quickly mobilised and triggered by cell surface molecules (receptors) that recognise a broad spectrum of pathogens. Adaptive immune cells, such as T cells, on the other hand provide a more specific and long-term protection against pathogens. Although the classification of immunity into innate and adaptive has served to simplify and facilitate the description of many immunological events, it is becoming increasingly evident that the interaction between innate and adaptive immune cells is crucial for mounting appropriate immune responses. This, therefore, necessitated the establishment of suitable culture systems and animal models for studying such interactions, and the use of these systems has so far been useful in understanding the interrelationships between dendritic cells and T cells. However, in trying to better recreate what happens in the human lymph nodes (the small organs where dendritic cells and T cells meet), we are proposing to construct a three-dimensional culture system that resembles a lymphoid organoid. This will hopefully be a more physiological system and one that will allow the development of further applications for studying cell-cell interactions. If successful, this system will eventually reduce the number of some animal experimentations. The development of such a system is timely given that the 7th amendment to the EU Cosmetics Directive will impose a marketing ban on new ingredients tested on animals from 2009 onwards. Our initiative is supported by Unilever, a company which has considerable experience and vested interest in the development of model systems for predicting the allergenic potential of new healthcare products.

Technical Summary

Dendritic cells (DCs) are key regulators of the immune system. For instance, they are capable of stimulating lymphocytes to generate antigen-specific cell-mediated and humoral immune responses against pathogens and tumour cells and instruct T cells to tolerate self-antigens. Interaction between DCs and T cells takes place in lymph nodes (LN) which are aggregates of structured lympho-reticular tissue. The particular structure of LN and the presence of extra-cellular matrix (ECM) facilitate/guide cell migration, cell proliferation, cell differentiation, intracellular signalling and cell-cell interaction, which collectively determine the outcome of immune responses. However, in the vast majority of current in vitro models of DC-T cell interaction, these cells are studied in 2D (i.e. in tissue culture plates) and more importantly in the absence of ECM, conditions which do not restore the 3D architecture and many other properties of LN. Therefore, there is a real need to develop novel in vitro cell culture systems to better simulate the microenvironment in which these cells co-exist. Thus, the aim of this proposal is to engineer a lymphoid organoid comprising DCs co-cultured with naïve T cells in the presence of ECM, and within a perfusable 3D multi-cell environment, that is amenable to interactive control and monitoring of emergence, maturation and interaction of cells.
 
Description 1- We have shown that the presence of relevant extracellular matrix proteins (ECM) change the phenotype/functional properties of human dendritic cells (DCs) enabling them to maintain a more immature phenotype at resting state with significantly better endocytic ability. The latter is partlydue to up regulation of key endocytic receptors such as MR and DD-SIGN.

2- We have also shown that ECM treated DCs (in 2D and 3D) are superior in antigen uptake andT cell activation and establish a more efficient 'immunological synapse' with T cells. This knowledge allowed us to development a more sensitive in vitro T cell proliferation assay for testing sensitizers and model antigens.

3- We have developed a range of 3D scaffolds with different architecture, mechanical properties and surface chemistry and have used them for sequential seeding of different cells types. Cell seeded scaffolds have been incorporated inside a bioreactor which was designed 'in house'.
Exploitation Route Use of scaffolds for 3D culture of immune cells
Sectors Pharmaceuticals and Medical Biotechnology

 
Description Our findings during this project led to better understanding of how important human immune cells (also called antigen presenting cells or dendritic cells) interact with 3D synthetic scaffolds and a number of our own body's proteins. This knowledge underpinned new collaborations and research projects in developing novel immune competent tissue models that can be use d for disease modelling and testing new drug leads.
First Year Of Impact 2002
Sector Pharmaceuticals and Medical Biotechnology
Impact Types Societal

 
Description Construction of a miniaturized human lymph node model as an alternative to the Local Lymph Node Assay
Amount £120,000 (GBP)
Funding ID NC/K500318/1 
Organisation National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs) 
Sector Private
Country United Kingdom
Start 09/2012 
End 08/2016
 
Description Construction of a miniaturized human lymph node model as an alternative to the Local Lymph Node Assay
Amount £120,000 (GBP)
Funding ID NC/K500318/1 
Organisation National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs) 
Sector Private
Country United Kingdom
Start 09/2012 
End 08/2016
 
Description Local Immunomodulation around implants by innovative auxiliary hydrogel-based systems encapsulating autologous and phenotype controlled macrophages.
Amount £5,600,000 (GBP)
Funding ID 602694 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 10/2013 
End 09/2017
 
Description Personalized And/Or Generalized Integrated Biomaterial Risk Assessment
Amount € 7,992,471 (EUR)
Funding ID 760921 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 12/2017 
End 01/2022
 
Description A microfluidic device for studying DC-T cell interaction under flow condition in 2D and 3D 
Organisation Harvard University
Country United States 
Sector Academic/University 
PI Contribution We are working on optimisation of a multi-channel microfuidc device that enables co-culture of different immune cells (e.g. T cells and dendritic cells) within a hydrogel under flow conditions. The device is also amenable to real-time imaging which provides an opportunity for monitoring the kinetics of DC-T cell cross--talk
Start Year 2013