Doctoral Dissertations

Date of Award

5-1994

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Biomedical Sciences

Major Professor

Virginia Godfrey

Committee Members

J. Erby Wilkinson, Liane B. Russell, Eugene M. Rinchik

Abstract

The X-linked recessive mutation scurfy (sf) results in a phenotype characterized by a rapidly fatal immune disorder involving the skin and lymphoid systems. Previous work has shown that T lymphocytes generated in a genetically sf thymic microenvironment mediate disease in the scurfy mouse. In order to understand better the immunobiology of the scurfy mouse, we have undertaken a series of experiments that investigated the role of the thymus and its components in the etiology and expression of scurfy disease. Initially, we utilized nude, SCID and sf-SCID mice to create models in which the thymic components had been manipulated. Results from experiments involving the transplantation of scurfy fetal thymi into H-2-compatible nude and SCID mice indicated that scurfy disease acts upon the fetal thymic environment as early as day-14 in development. Further, experiments involving the selective transfer of wild-type (+/Y) or scurfy thymic components demonstrated that both sf-derived T cell precursors, and a genetically sf thymic microenvironment were necessary for disease expression. To investigate the function of the T cell mediators in the scurfy disorder, we evaluated the respective roles of CD4 or CD8 single-positive T cells by treating scurfy neonates with monoclonal antibodies directed against the CD4 or CD8 molecules and by breeding the scurfy mutation onto mice that lacked either CD4+ or CD8- T cells. Results implicated CD4+CD8- T cells as the critical effector cells in the pathogenesis of scurfy disease. Also, using monoclonal antibodies, we established that CD4+CD8- T cells in the scurfy mouse exhibit an activated (and thus reactive) phenotype in vivo. Further, scurfy mice and CD8-less scurfy mice overexpressed mRNA messages and proteins for the cytokines IL-2, IL-3, IL-4, IL-6, IL-10, GM-CSF and TNF-α in their skin and/or lymphoid organs. We speculate that the scurfy mutation results in an alteration in normal thymic ontogeny, leading to the release of autoaggressive T cells. To test this, we examined the ability of scurfy mice to delete T cell clones that express Vβ elements reactive with endogenous I-E and Mls antigens. Our results demonstrated an increased number of CD4+ T cells in the lymph nodes of scurfy mice that bore these potentially self-reactive Vbeta; elements. While this phenomenon is suggestive of a failure in the process of thymic negative selection, other results indicated that scurfy T cells are capable of apoptosis. Finally, we have attempted to consolidate the map location of sf on the proximal X chromosome by following the segregation pattern of the mutation in relation to that of cloned loci using an interspecific Mus musculus/Mus spretus backcross. Recombination frequencies generated from over 250 backcross progeny implied that sf is inseparable from Gf-1, and proximal of Otc (spf). From these studies, we propose that the scurfy mutation results in a failure in the process of thymic negative selection thereby allowing the release of autoreactive T cells that provoke a generalized wasting disease via the overproduction of cytokines in the skin and lymphoid organs.

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