CD34(−) Orbital Fibroblasts From Patients With Thyroid-Associated Ophthalmopathy Modulate TNF-α Expression in CD34(+) Fibroblasts and Fibrocytes

PURPOSE: Orbital fibroblasts from patients with Graves' disease (GD-OF) express many different cytokines when treated with bovine thyrotropin (bTSH). The present study aimed to determine why TNF-α cannot be induced by bTSH in GD-OF. METHODS: Fibrocytes and GD-OFs were cultivated from donors who were patients in a busy academic medical center practice. Real-time PCR, Western blot analysis, reporter gene assays, cell transfections, mRNA stability assays, ELISA, and flow cytometry were performed. RESULTS: We found that bTSH induces TNF-α dramatically in fibrocytes but is undetectable in GD-OF. The induction in fibrocytes is a consequence of increased TNF-α gene promoter activity and is independent of ongoing protein synthesis. It could be attenuated by dexamethasone and the IGF-1 receptor inhibiting antibody, teprotumumab. When separated into pure CD34(+) OF and CD34(−) OF subsets, TNF-α mRNA became highly inducible by bTSH in CD34(+) OF but remained undetectable in CD34(−) OF. Conditioned medium from CD34(−) OF inhibited induction of TNF-α in fibrocytes. CONCLUSIONS: Our data indicate that CD34(−) OF appear to release a soluble(s) factor that downregulates expression and induction by bTSH of TNF-α in fibrocytes and their derivative CD34(+) OF. We proffer that CD34(−) OF produce an unidentified modulatory factor that attenuates TNF-α expression in GD-OF and may do so in the TAO orbit.