The role of IL-7/IL7R interactions in the development, maintenance and organization of the lymph node microenvironment.

The lymph node or ?gland? is a dynamic microenvironment of lymphocytes, dendritic cells and lymph node stromal cells where immune responses are initiated. Intercellular interactions between these cells are essential for the survival, homeostasis and function of all of these cell types, with disruption of these interactions leading to cell death. In order to study the involvement of locally produced cytokines (immune hormones) in the interdependency of these different cells we propose to generate mice in which lymph node stromal cells expressing cytokines are tagged by fluorescent proteins. The homeostasis of peripheral T cells is dependent on IL-7 which is produced by lymph node stromal cells. To visualize stromal cells we will generate IL-7 fluorescent protein mice. Utilizing multiphoton microscopy, tissue culture, gene expression analysis and flow cytometry we will analyze the molecular and cell biology of lymphocyte - stromal cell interactions during lymph node development, homeostasis and organization. Our experiments will test the hypothesis that IL-7/IL7R interactions between stromal cells and lymphocytes are required for the development of the lymph node microenvironment.

The main objectives of our proposal are:

i) To characterize lymph node stromal cell biology.

ii) To examine the role of IL-7/IL7R interactions in the development, organization and maintenance of the lymph node microenvironment.

iii) To characterize the molecular mechanism governing intercellular communication initiated by IL-7/IL7R signalling between lymphocytes and lymph node stromal cells.

A three dimensional network of lymph node stromal cells provides the microenvironment for the survival, homeostasis, activation and differentiation of lymphocytes. Conversely, communication between lymphocytes and lymph node stromal cells is also required for the survival of the stromal cell microenvironment. Despite the importance of lymph node stromal cells in the survival and function of peripheral lymphocytes little is understood about the development, maintenance and organization of stromal cells. Recently we have shown that IL-7/IL7R interactions have an important role in these processes. Here we will utilize IL-7 reporter mice and inducible IL-7 knock-out mice to test our hypothesis that IL-7/IL7R signalling is required for stromal cell development, organization and maintenance.

Our specific aims are:

i) To characterize IL-7 expressing lymph node stromal cells.

ii) To analyze the functional role of IL-7/IL7R signalling in lymph node stromal cell development, organization and maintenance.

iii) To utilize gene expression analysis, gene knock-out mice to analyze the molecular mechanisms governing the bi-directional interactions initiated by IL-7/IL7R signalling during neonatal development.

To achieve our aims we will utilize 4D multiphoton, 3D tissue culture, flow cytometry and gene expression analysis of material from appropriately genetically altered animals. The results of these studies will provide important new information on the development and function of the lymph node.