Abstract
The thymus plays a crucial role in preventing autoimmune reactions in a variety of species. Through the expression of the autoimmune regulator (Aire), a protein found in medullary thymic epithelial cells (mTECs), the thymus is able to express tissue-restricted peripheral self-antigens (TRAs). Through their interaction with these TRAs, developing thymocytes that target host antigens can be tagged for clonal deletion. Graft versus host disease (GvHD), an autoimmune reaction occurring in approximately 40% of allogeneic stem cell transplantation patients, is an immune mediated reaction in which donor T cells recognize the host as antigenically foreign, causing donor T cells to expand and attack host tissues. GvHD is often associated with significant morbidity and mortality, and because the thymus is a target tissue of T-cell alloimmunity and can be severely damaged in many autoimmune diseases, we focused on significant morphological and cellular changes within the thymus as possible contributors to the continuing proliferation and survival of alloimmune T-cells in GvHD.
Previous data collected from a mouse model of GvHD showed a reduction in expression of Aire and a reduced diversity of TRAs in the thymus. We also found that the thymus is significantly smaller in size when compared to wild-type mice, which correlates to overall thymic involution as both the cortical and medullary TEC levels were reduced. This reduction of Aire expression also correlated with a reduction in thymically-derived FoxP3+NrpI+regulatory T-cells, which were significantly reduced in both spleen and blood of GvHD mice when compared to bone marrow transplant controls (blood p < 0.0005, spleen p < 0.0001). SEMA4A, a transmembrane protein known to interact with Nrp1 to promote regulatory T-cell function and survival, was also significantly changed in GvHD compared to healthy controls (p = 0.014). In addition, cytokine data collected from a GvHD mouse model also showed increases in several chemokines associated with thymic atrophy and aging, indicating that thymii found in GvHD mice are subject to significant damage due to autoimmune reactions.
Interestingly, there is evidence of thymic damage in mouse models of colitis as well. Ulcerative colitis is a form of inflammatory bowel disease caused by T-cell infiltration into the colon, resembling some of the pathophysiology of G.I. GvHD. Thymii from DSS colitis mice showed a similarly significant reduction of Aire and FoxP3 expression, suggesting a new, more prominent role for thymic damage in colitis severity (p < 0.005). This damage was observed to occur in a generalized way, as FACS analysis of thymic epithelial cell types showed that both medullary and cortical epithelial cells in diseased thymus were reduced, not just mTECs. The reduction in expression of Aire protein is due directly to RNA reduction, with a mean 8 fold reduction of Aire RNA expression in colitis mice compared to healthy mice (p = 0.0154).
While SEMA4A was seen to be increased in GvHD, there was a significant reduction in expression in colitis thymii (p=0.0138). Reductions in SEMA4A have previously been shown to cause impaired immune cell differentiation and function, followed by impaired homeostasis and function of FoxP3+T-cells, indicating that this reduction in SEMA4A may be the cause of reduced FoxP3 expression in the colitis thymus. It was also observed that NrpI expressing FoxP3 cells in the spleen and blood were unchanged in colitis mice, indicating that reduction in FoxP3 expressing cells in colitis are not entirely due to thymic damage. This is supported by cytokine data, in which no inflammatory or aging related cytokines were changed in colitis thymii.
The significant thymic involution and reduced expression of the Aire protein in the thymus is a likely contributor to overall increases of autoimmune T-cells in both colitis and GvHD. Though colitis and GvHD clearly use different pathways to affect the thymus, thymic damage plays a significant role in both diseases. The reduced expression of the FoxP3 protein also indicates a role for loss of Aire in lowered immune tolerance, contributing to the overall autoimmune phenotype in both diseases.
Disclosures
No relevant conflicts of interest to declare.
Characterization of the expression of cytokeratins 5, 8, and 14 in mouse thymic epithelial cells during thymus regeneration following acute thymic involution