Graves’ Ophthalmopathy: A Comprehensive Role For Platelet-Derived Growth Factors

Autoimmune thyroid disorders include Graves’ disease (GD), which leads to hyperthyroidism, and Hashimoto’s thyroiditis, which leads to hypothyroidism. The incidence of autoimmune hypothyroidism is around five times higher than the incidence of autoimmune hyperthyroidism, but together these autoimmune thyroid disorders are the most common autoimmune conditions in the Western population and affect around 2-5% of the population. Estimates for the incidence of autoimmune hyperthyroidism vary considerably, but a recent review estimates an incidence of 80/100.000/year for women and 8/100.000/year for men in Caucasian populations. Graves’ disease (GD) is the most common cause of hyperthyroidism in Western and other iodine-sufficient populations, and accounts for ~80% of cases of hyperthyroidism

PDGF enhances orbital fibroblast responses to TSHR stimulating autoantibodies in Graves' ophthalmopathy patients

Purpose: Thyroid-stimulating hormone receptor (TSHR) stimulating autoantibodies are associated with Graves' ophthalmopathy (GO), the orbital manifestation of Graves' disease (GD). TSHR autoantibody levels and orbital TSHR expression levels correlate positively with GO disease activity. Platelet-derived growth factors (PDGF) are increased in GO and potently activate orbital fibroblast effector functions. We investigated the possible relationship between PDGF and TSHR expression on orbital fibroblasts and how that influences the immunopathological effects of TSHR autoantibodies on orbital fibroblast activity. Methods: Orbital fibroblasts were stimulated with PDGF-AA, PDGF-AB, and PDGF-BB, and TSHR expression was determined by flow cytometry. Stimulatory effects of bovine TSH and GD immunoglobulins on orbital fibroblasts (with or without PDGF-BB preincubation) were determined by IL-6, IL-8, chemokine (C-C motif) ligand (CCL)-2, CCL5, CCL7, and hyaluronan ELISA. The TSHR blocking antibody K1-70 and the cAMP inhibitor H89 were used to determine involvement of TSHR signaling. Results: PDGF-AB and PDGF-BB stimulation increased TSHR expression on orbital fibroblasts, whereas PDGF-AA did not. Furthermore, stimulation with bovine TSH and immunoglobulins from GD patients induced IL-6, IL-8, CCL2, and hyaluronan production by orbital fibroblasts, and PDGF-BB preincubation enhanced this response of orbital fibroblasts. Blocking studies with a TSHR blocking antibody and a cAMP inhibitor inhibited these effects, indicating the involvement of TSHR signaling and thus of TSHR stimulating autoantibodies herein. Conclusions: These findings indicate that PDGF-B containing PDGF isoforms amplify the immunopathological effects of TSHR-stimulating autoantibodies in GO patients by stimulating TSHR expression on orbital fibroblasts. Copyrigh

Orbit-infiltrating mast cells, monocytes, and macrophages produce PDGF isoforms that orchestrate orbital fibroblast activation in graves' ophthalmopathy

Purpose: Platelet-derived growth factors (PDGF) are regulators of fibroblast activity that may be involved in the pathophysiology of Graves' ophthalmopathy (GO). We unraveled the expression and origin of PDGF family members in GO orbital tissue and investigated the effect of PDGF isoforms on IL-6 and hyaluronan production and proliferation by orbital fibroblasts. Methods: PDGF-A, PDGF-B, PDGF-C, PDGF-D, PDGF-Rα, and PDGF-Rβ expression was determined by real-time quantitative PCR and PDGF-A and PDGF-B protein expression was determined by Western blot in orbital tissues. Orbital tissues were immunohistochemically stained for PDGF-A and PDGF-B expression, together with stainings for T cells, monocytes, B cells, macrophages, and mast cells. Effects of PDGF-AA, PDGF-AB, and PDGF-BB on orbital fibroblast proliferation and IL-6 and hyaluronan production were examined. Finally, effects of PDGF-BB- and PDGF-AA-neutralizing antibodies on IL-6 and hyaluronan production in GO whole orbital tissue cultures were tested. Results: GO orbital tissue showed increased PDGF-A and PDGF-B mRNA and protein levels. Increased numbers of PDGF-A- and PDGF-B-positive monocytes, macrophages, and mast cells were present in GO orbital tissue. PDGF-BB stimulated proliferation and hyaluronan and IL-6 production by orbital fibroblasts the most, followed by PDGF-AB and PDGF-AA. Finally, in particular imatinib mesylate and PDGF-BB-neutralizing antibodies reduced IL-6 and hyaluronan production by whole orbital tis

Platelet-derived growth factor-BB: A stimulus for cytokine production by orbital fibroblasts in graves' ophthalmopathy

Purpose. Graves' ophthalmopathy (GO) is characterized by the infiltration of immune cells into the orbit, a process in which cytokines play a central role. Orbital fibroblasts are potent producers of cytokines on different stimuli. Recently, the authors showed increased expression of the PDGF-B chain in GO orbital tissue. The dimeric PDGF-BB molecule has been described to activate the NF-κB pathway, which is well recognized for its role in regulating cytokine production. This study was conducted to determine the role of PDGF-BB in the production of proinflammatory cytokines by orbital fibroblasts in GO. Methods. Orbital, lung, and skin fibroblasts were stimulated with PDGF-BB, and cytokine (IL-1β, IL-6, IL-8, IL-16, CCL2, CCL5, CCL7, TNF-α) production was measured by ELISA. Involvement of NF-κB activation through PDGF signaling was investigated by electrophoretic mobility shift assay, specific NF-κB inhibitors, and the PDGF-receptor kinase inhibitor imatinib mesylate. Results. IL-6, IL-8, CCL2, CCL5, and CCL7 production by orbital fibroblasts was increased by PDGF-BB stimulation, whereas IL-16, IL-1β, and TNF-α production was not affected. PDGF-BB induced NF-κB activity in orbital fibroblasts, and both NF-κB inhibitors and imatinib mesylate reduced PDGF-BB-induced cytokine production. Similar, but less vigorous, effects of PDGF-BB on cytokine production were observed in lung and skin fibroblasts. Conclusions. PDGF-BB is a potent inducer of proinflammatory cytokines via the NF-κB pathway in orbital fibroblasts, whereas cytokine production by fibroblasts from other anatomic locations showed a moderate response. These data suggest a possible role for PDGF-BB in regulating orbital inflammation in GO and identify the PDGF signaling cascade as a therapeutic target in GO

Histamine induces NF-κB controlled cytokine secretion by orbital fibroblasts via histamine receptor type-1

Mast cells and their products are likely to be involved in regulating orbital fibroblast activity in Graves' Ophthalmopathy (GO). Histamine is abundantly present in granules of mast cells and is released upon mast cell activation. However, the effect of histamine on orbital fibroblasts has not been examined so far. Orbital tissues from GO patients and controls were analyzed for the presence of mast cells using toluidine blue staining and immunohistochemical detection of CD117 (stem cell factor receptor). Orbital fibroblasts were cultured from GO patients and healthy controls, stimulated with histamine and cytokines (IL-6, IL-8, CCL2, CCL5, CCL7, CXCL10 and CXCL11) were measured in culture supernatants. Also hyaluronan levels were measured in culture supernatants and hyaluronan synthase (HAS) and hyaluronidase (HYAL) gene expression levels were determined. In addition, histamine receptor subtype gene expression levels were examined as well as the effect of the histamine receptor-1 (HRH1) antagonist loratadine and NF-κB inhibitor SC-514 on histamine-induced cytokine production. Mast cell numbers were increased in GO orbital tissues. Histamine stimulated the production of IL-6, IL-8 and CCL2 by orbital fibroblasts, while it had no effect on the production of CCL5, CCL7, CXCL10, CXCL11 and hyaluronan. Orbital fibroblasts expressed HRH1 and loratadine and SC-514 both blocked histamine-induced IL-6, IL-8 and CCL2 production by orbital fibroblasts. In conclusion, this study demonstrates that histamine can induce the production of NF-κB controlled-cytokines by orbital fibroblasts, which supports a role for mast cells in GO