A study on the antagonistic targeting of thymic stromal lymphopoietin (TSLP) as a potential therapeutic strategy for Asthma bronchiale

摘要

In recent years, Thymic stromal lymphopoietin (TSLP) has emerged as a master switch for actuating cellular inflammation and local remodeling in asthmatic airways. Originally discovered as a growth and proliferation factor of the B cells, thymic stromal lymphopoietin (TSLP) is now implicated in immune system related effects on several cells including B cells, basophils, CD4+, CD8+ and natural killer T cells, dendritic cells, eosinophils, epithelial cells and mast cells. The structural and immune cells express TSLP at the site of allergen entry in the airways. TSLP regulates the T helper 2 (Th2) cells humoral immunity by upregulation of a ligand, OX40L, on the surface of the dendritic cells (DCs), which activates the naive Th0 cells to active Th2 lymphocytes. The role of TSLP in the promotion of Th2 responses has been panoptically studied in the context of several allergic disorders. The experimental work in this thesis is focused on the creation of tools to study/manipulate the signaling of murine TSLP/TSLPR system and thus help us in a better understanding of targeting the TSLP induced signal transduction during allergic inflammation. The studies involved development of a TSLPR expressing luciferase gene based murine cellular model, as well as an inducible TSLP secreting epithelial-dendritic cell based model as readouts. The process involved production, purification and characterization of murine TSLP (mTSLP) as a fusion protein with maltose binding protein (MBP) i.e. mTSLP-MBP and mutant I37E mTSLP-MBP. Exo-mTSLPR domain soluble protein was also produced as a competitive inhibitor and it was then used to generate an anti murine TSLPR antibody for the detection of mTSLPR receptors on the cells. These tools have been successfully tested on the readout systems as mentioned above. The I37E mTSLP-MBP mutant showed partially competitive effects in the cellular readouts. The mTSLPR showed an effective blocking of mTSLP induced signaling. The anti mouse TSLPR antibody could bind to the TSLPR receptors on the cells but it was found to be non inhibitory in nature. By generating the above mentioned inhibitory proteins, novel strategies have been developed to analyze and specifically inhibit TSLP induced immunological activation. The results from these cellular model based studies are in agreement with those obtained from studies involving similar molecules designed for studying human TSLP-TSLPR system in our group in the past. These results cumulatively prove that a successful blockade of the human/murine TSLP-TSLPR system is possible. These results offer an insight into multifaceted roles of TSLP and could potentially pave the way for therapeutic manipulations of allergic disorders.