Another interesting aspect of the CD34 fibrocyte is its expression of multiple “thyroid-specific” proteins. Fernando and colleagues [46] reported that those cultivated from circulating peripheral blood mononuclear cells express not only TSHR but also low levels of Tg mRNA. Levels of the transcript are considerably lower than those found in thyroid tissue. Tg resolved on polyacrylamide gel electrophoresis as a 305 kDa protein. Both ¹²⁵Iodine and ³⁵S methionine can be incorporated into Tg expressed by fibrocytes. siRNAs targeting Tg attenuated the synthesis of the protein. Tg gene promoter is active in fibrocytes. Compared with Tg levels found in fibrocytes, those in orbital fibroblasts from donors with GD were substantially lower. When GD-orbital fibroblasts were sorted into pure CD34⁺ and CD34⁻ populations, Tg mRNA expression segregated to CD34⁺ fibroblasts. Tg expression is considerably greater in CD34⁺ fibroblasts than in the mixed cell population found in parental cultures. Two additional thyroid-specific proteins have been detected in the protein expression repertoire of fibrocytes, namely sodium iodide symporter (NIS) and thyroid peroxidase (TPO) [47]. Levels of NIS and TPO mRNAs are substantially lower than those found in thyroid tissue and FRTL-5 cells in culture. However they are adequately abundant to result in translation into detectable protein. They are considerably higher than those detected in dermal fibroblasts or orbital fibroblasts from healthy individuals and those with TED. The expression of multiple autoantigens suggested that a specific transcription factor might be involved, namely the autoimmune regulator protein (AIRE) [44]. AIRE is a major determinant of intra-thymic education that allows the negative selection of autoreactive thymocytes. In fibrocytes, AIRE mRNA and protein are detectable but at extremely low levels compared to thymus. Interruption of AIRE expression using small interfering RNAs resulted in substantial reductions in the steady state mRNA levels of TSHR, Tg, TPO, and NIS (Fig. 3.3). Further, knocking down AIRE expression also reduced levels of PAX8 and TTF1, the two transcription factors closely associated with expression of these proteins in thyroid epithelium. To confirm the importance of AIRE in this specific context, fibrocytes cultivated from an individual with APS 1 syndrome were also interrogated for levels of thyroid proteins and found to be significantly below those found in an unaffected first degree relative. AIRE expression in fibrocytes is the consequence of an active AIRE gene promoter and remarkably stable AIRE mRNA. The implications of these findings to the biology of fibrocytes and the tissues which they infiltrate include the suggestion that AIRE is an important component of extra thyroidal autoimmunity related to peripheral loss of tolerance to thyroid-associated proteins such as thyroglobulin, TPO, and TSHR. Further, they suggest that fibrocytes may be capable of synthesizing thyroid hormones or their derivatives. The array of self-antigens expressed by fibrocytes does not appear to be limited to those associated with autoimmune thyroid disease. Fernando et al. reported recently that IA-2 and ICA69 can also be detected in these cells, findings with implications for type 1 diabetes mellitus [48]. In addition to their expression of these proteins, fibrocytes express high levels of MHC Class II and can present antigens to T cells [49].

Despite substantial efforts to understand better the mechanisms responsible for the initiation of GD and the factors that overarch involvement of the orbit in TED, current knowledge remains mired in uncertainty. Much of this can be attributed to an absence of preclinical models that faithfully recapitulate the human disease. These deficits have resulted in substantial unmet need in how we treat this disease. Especially during the active phase when surgery is frequently discouraged, medical options are limited. Nonspecific anti-inflammatory agents, including corticosteroids, nonsteroidal agents (PGHS-2 inhibitors), and antimetabolites may lessen the symptoms directly caused by inflammation. But none of these agents is thought to alter the clinical course of TED. More recently, targeted strategies, many repurposed from other diseases, have been used on an ad hoc basis. Anti-CD20 monoclonal antibody (rituximab) is the currently best-studied among these and the results of two separate prospective trials are eagerly awaited. Other biologics, such as those that interrupt cytokine networks, have yet to be studied in a manner that could lead to conclusions about their efficacy. Their roles in the therapy of active TED remain to be established. Further, all of these candidate agents are associated with potentially serious side effects. Thus, we continue to rely on surgical intervention for ocular rehabilitation.

Recognition of the potential involvement of fibrocytes in GD and TED widens the horizons for future inquiry into disease mechanisms. It may be possible to directly target specific aspects of their behavior as potential mediators of the immune reactivity systemically as well as within the orbit space. To be certain, TED is a complex autoimmune condition with numerous molecular and cellular underpinnings. We are only now beginning to understand the underlying processes. Further insights into the potential roles fibrocytes play in the disease, especially the initial phases, should bring us closer to developing more effective treatments.