Proangiogenic effect of thyrotropin receptor stimulating antibody in human umbilical vein endothelial cells.

Objective: This study aims to investigate the role of TRAb in the angiogenesis associated with Graves' disease (GD) and to elucidate its underlying mechanisms.

Methods: Human thyroid follicular epithelial cells (Nthy-ori 3-1) and human umbilical vein endothelial cells (HUVECs) were treated with the monoclonal thyroid-stimulating antibody M22 and thyroid-stimulating hormone (TSH) at various concentrations. Cell viability, migration, and tube formation were evaluated using CCK-8, wound healing, and tube formation assays, respectively. Protein expressions of TSHR receptor (TSHR) and phosphorylated AKT (p-AKT) in M22-induced HUVECs were quantified via Western blotting. Proteomic analysis was employed to identify changes in protein expression profiles and relevant signaling pathways in GD specimens. Immunofluorescence assays were conducted to detect and localize the expressions of CD34 and PROX1 in GD specimens and normal thyroid tissues.

Results: M22 stimulated the proliferation of Nthy-ori 3-1 cells and HUVECs in a dose-dependent manner, while TSH exhibited an inverted U-shaped dose-response effect. M22 also dose-dependently promoted angiogenesis, and more effectively induced tube formation in HUVECs compared to TSH, although the difference was not statistically significant. A total of 16 proteins were significantly upregulated and 24 were downregulated in M22-induced HUVECs. Notably, PROX1, the most significantly upregulated protein, is closely associated with angiogenesis. Immunofluorescence confirmed that PROX1 was significantly more expressed in thyroid tissues from GD patients compared to normal tissues adjacent to papillary thyroid cancer (PTC), and it co-localized with CD34.

Conclusion: TRAb enhances angiogenesis and upregulates PROX1 expression in HUVECs, suggesting a novel possible mechanism for goiter formation in GD.