Unraveling the Genetic Susceptibility to Autoimmune Thyroid Diseases: CTLA-4 Takes the Stage

167 OVER THE PAST DECADE there has been an explosion of studies on the genetics of complex diseases. Many complex diseases show familial aggregation and are believed to develop by an interaction of susceptibility genes and environmental triggers. Therefore, identifying the susceptibility genes causing complex diseases will likely lead to unraveling the primary abnormalities triggering them. Among the complex diseases the autoimmune thyroid diseases (AITDs) are particularly attractive for genetic studies for several reasons: (1) The phenotype of the major AITDs (Graves’ disease [GD], Hashimoto’s thyroiditis [HT]) is clearly defined, unlike in some other complex diseases (e.g., in systemic lupus erythematosus [SLE]); (2) The AITDs are common (1), and commonly segregate in families (up to 50% of first-degree relatives of GD and HT patients have thyroid antibodies [TAbs] [2,3]), thus enabling collection of large numbers of families for linkage studies; (3) The AITDs comprise several interrelated conditions including GD, HT, and TAb production. These AITD phenotypes often cosegregate within the same family, thus enabling geneticists to vary these parameters in the analysis in order to study the relationships between them. For example, one can study whether the genes for GD, HT, and TAb are common or unique for each disorder, or whether TAbs are a precursor of clinical disease or represent a different disease entity (4); (4) Once a suspected gene is identified, its function in thyroid tissue can be easily studied as thyroid tissues from AITD patients are readily available. Cytotoxic T-lymhocyte–associated molecule-4 (CTLA-4) is a critical molecule in the activation of T cells by antigens. Antigen presenting cells (APCs) activate T cells by presenting to the T-cell receptor an antigenic peptide bound to an HLA class II protein on the cell surface (5). However, for the T cells to be activated by the antigen a second signal is required and these costimulatory signals can be provided by the APCs themselves or other local cells (5). The costimulatory signals are provided by a variety of proteins that are expressed on APCs (e.g., B7-1, B7-2, CD-40) and interact with receptors (CD28, CTLA-4, ICOS, and CD-40L) on the surface of CD41 T lymphocytes during antigen presentation (5). CD28 and CTLA-4 are related glycoproteins of the immunoglobulin superfamily with opposing functions. Whereas the binding of B7 to CD28 on T cells augments T-cell activation, the binding of B7 to CTLA-4 is thought to downregulate Tcell activation. Indeed, activation of CTLA-4 by cross-linking inhibited T-cell responses, and blockade of CTLA-4-B7 interactions enhanced T-cell proliferation (6). Moreover, knockout mice lacking the CTLA-4 gene develop massive Tcell lymphoproliferation resulting in splenomegaly, lymphadenopathy, and lymphoid infiltration into many organs (7). While initially it was thought that CTLA-4 suppressed T-cell activation by competing with CD28 for binding to the B7 molecules, it is now apparent that this suppression is an active process and is not simply due to competition with CD28 for the B7 ligands. Recent data show that CTLA-4 activation during antigen presentation leads to inhibition of interleukin-2 (IL-2) production, a necessary cytokine for T-cell activation. CTLA-4 engagement also prevents T cell progression through the cell cycle thus directly inhibiting T-cell proliferation (8). The suppressive effects of CTLA-4 on T-cell activation have raised the possibility that sequence changes altering CTLA-4 function could lead to augmentation of immune responses and development of autoimmunity. Indeed, treatment of female NOD mice at the onset of insulitis with antiCTLA-4 or a monoclonal antibody specific for B7-2 (a CD28 ligand) suppressed the development of diabetes (31). Recently, there have been several reports demonstrating a weak association between the CTLA-4 gene and the AITDs (9,10–12). Earlier studies found an association between a microsatellite marker located near the CTLA-4 gene and GD, giving a relative risk of 2.1 to 2.8 (9,12). With the identification of an alanine/threonine (G/A) polymorphism inside the CTLA-4 leader peptide, this polymorphism was also tested for association with the AITDs. The ala (G) polymorphism was found to be associated with GD with a relative risk of 2.0 (11). These reports have been consistent in several populations (9,12). Two groups have also recently claimed that the G allele of the CTLA-4 single nucleotide polymorphism (SNP) was specifically associated with Graves’ ophthal-