This project is a continuation of studies into the immunobiology of murine intestinal intraepithelial lymphocytes (IELs) as a model for understanding human intestinal T cells in health and disease. The studies described here are derived from work done during the previous funding period, and are designed to address specific unresolved aspects of intestinal T cell development, and to obtain detailed information into intestinal immune-endocrine interactions in the regulation of local intestinal immunity and gastrointestinal disease. The goals of this project are:
Specific Aim 1 - to characterize regional sites of local intestinal IEL maturation and development; to determine the lineage relationships between IEL precursors and mature intestinal T cells using intestine- and bone marrow-derived precursors; to determine the potential contribution of mature peripheral T cells and immature thymocytes to the overall intestinal T cell pool. These studies will use transgenic mice which express high levels of a green fluorescent protein in adoptive transfer experiments to trace IEL developmental pathways through phenotypic studies, and by molecular analyses of IEL Vgamma,delta, and beta TCR repertoires.
Specific Aim 2 - to systematically characterize the intestinal thyrotropin (thyroid- stimulating hormone [TSH])-mediated immune-endocrine network in terms of local hormone production and utilization in phenotypically-defined cell populations; to study their immunoregulatory effects on intestinal IELs; to delineate biochemical mechanisms of TSH-mediated intestinal T cell activation. RT-PCR analyses of hormone receptor gene expression and hormone binding assays will be used to accurately delineate pathways of intercellular communication; RNase protection assay, ELISA, and ELISPOT assays will be used to identify cytokine and inflammatory mediators activated by TSH; and immunoprecipitation of phosphorylated Jak/STAT regulatory elements will be done to identify TSH receptor- mediated IEL activation.
Specific Aim 3 -to further explore the capacity of hypothalamus-pituitary-thyroid hormones to influence or alter gastrointestinal autoimmunity and inflammation using established animal models of those diseases. Phenotypic analyses will be done using lymphocytes from gastrointestinal inflammatory sites, and characterization of hormone-activated regulatory and effector cells will be identified through adoptive transfer experiments.
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