TSH Receptor Antibodies as Measured in the Thyroid Stimulating Immunoglobulin (TSI) Reporter Bioassay Thyretain are not Detected in Patients with Euthyroid Graves’ Disease

Research Article

Austin J Clin Ophthalmol. 2014;1(5): 1024.

TSH Receptor Antibodies as Measured in the Thyroid Stimulating Immunoglobulin (TSI) Reporter Bioassay Thyretain are not Detected in Patients with Euthyroid Graves’ Disease

Hooshang Lahooti1, Jeffrey A Houtz2, Huy A Tran3, Ilhem Al-Honchari1, Simon D Lytton2, Bernard Champion1 and Jack R Wall1*

1Department of Medicine, University of Sydney, Australia

2Department of Medicine, SeraDiaLogistics, Germany

3Department of Clinical Chemistry, Newcastle University, Australia

*Corresponding author: Jack R Wall, Department of Medicine, University of Sydney, Nepean Hospital, PO Box 63, Penrith NSW 2751, Australia

Received: April 30, 2014; Accepted: June 16, 2014; Published: June 18, 2014


Background: The ophthalmopathy associated with Graves’ hyperthyroidism is most likely a T-cell mediated disorder, although most investigators believe that auto antibodies directed against the TSH Receptor (TSH-R) expressed on the surface of the orbital fibroblasts and pre-adipocytes are the main driver of the orbital reactions. Cases of ophthalmopathy without TSH-R binding antibodies and not closely associated with Thyroid Stimulating Immunoglobulin’s (TSI), challenge the notion that ophthalmopathy is perpetrated by TSI circulating in peripheral blood and TSI producing plasma cells resident in the orbital tissues. One way to address a possible role of TSH-R antibodies in the development of ophthalmopathy is to study patients with so-called “Euthyroid Graves’ eye Disease (EGD)”, who have the same eye disorder but with normal thyroid function and no features of thyroid autoimmunity during long term follow up.

Methods: The cell-based reporter bioassay Thyretain ™ was used to assess TSI in serum of the patients (n=50) and control subjects (n=20); The patient groups; EGD (n=13), of whom 6 were first tested early in the course of their eye disorder (< 6 months) and 7 tested ≥ 6 months after the onset of eye symptoms, Graves’ disease with (n=13), and without (n=12), ophthalmopathy and euthyroid relatives from a previously studied family having a high prevalence of thyroid autoimmunity and ophthalmopathy (WH-Fam n=12). Thyretain ™-TSI results were expressed as per cent of the sample to reference ratio (SRR %), a positive test being taken as an SRR% of > 140%.

Results: Only 3 of 13 (23 %) patients initially diagnosed with EGD tested TSI positive in one or more serum samples, and all 3 became hyperthyroid at the time of the positive test or soon after. Among the remaining 10 EGD patients who tested TSI negative, none developed thyroid autoimmunity during follow up visits 6months to 10 years after initial diagnosis. Normal subjects and euthyroid relatives from WH-Fam all tested TSI negative whereas 9 out of 12 (75%) Graves’ Hyperthyroidism (GH) and 11 out of 13 (86%) Graves’ Ophthalmopathy (GO) patients tested TSI positive.

Conclusion: These findings of positive TSI among patients with GH and GO, but not among patients with EGD with the exception of the 3 patients who developed Graves’ hyperthyroidism, strongly suggests that TSH-R antibodies cannot play a major role in the pathogenesis of what is best called endocrine or auto immuneophthalmopathy. Keywords: Ophthalmopathy; Thyroid autoimmunity; TSH receptor antibodies; Euthyroid Graves’ disease; Thyretain TM TSI reporter bioassay


Euthyroid Graves’ Disease (EGD) is an autoimmune condition of the eye and orbital tissues in the absence of thyroid dysfunction and is presumed to be the same eye disorder that is associated with Graves’ hyperthyroidism, where it is known as “Graves’ ophthalmopathy”. Some of these patients do develop Graves’ hyperthyroidism or Hashimoto’s thyroiditis on long term follow up [1] but, in a more recent study [2], none of 7 patients developed any evidence of thyroid dysfunction or autoimmunity during up to 11 years of follow up. In these patients, Thyroid Peroxidase (TPO), Thyroglobulin (TG) and TSH receptor (TSH-R) antibodies measured as TSH-R binding antibodies (TRAb), and as Thyroid-Stimulating Immunoglobulin (TSI) measured in a cAMP reporter bioassay, were always negative [2]. The pathogenesis of Graves’ ophthalmopathy, which is presumed to be the same eye disorder but associated with hyperthyroidism, is best explained by the action of T-lymphocytes and antibodies that target auto antigens shared between the thyroid and orbit muscle tissue, such as the TSH-R expressed on the orbital pre-adipocytes and fibroblasts [3-6], the main candidate antigen. However, the TSH-R is expressed in non-thyroid tissues as well and anti-TSH-R antibodies are not detected in all patients with Graves’ ophthalmopathy at the time of diagnosis [7], nor in patients with Hashimoto’s thyroid it is with mainly upper eyelid signs and symptoms [8,9]. For this reason, the conspicuous absence of TSH-R antibodies upon multiple visits of patients with EGD implies that anti-TSH-R antibodies are neither necessary nor sufficient for the pathogenesis of ophthalmopathy associated with thyroid autoimmunity.

TSI by Thyretain ™ -reporter bioassay (Thyretain-TSI) were previously shown to be functional indicators of the activity and severity of Graves’ ophthalmopathy; Furthermore, the combination of negative TRAb and positive Thyretain-TSI is closely associated with ophthalmopathy (Lytton et al [10-12] while patients testing positive for TRAb and negative for Thyretain-TSI had Graves’ disease without orbitopathy [12]. However, Thyretain-TSI is also found in newly diagnosed Graves’ disease patients showing no clinical signs of eye disease. In the present study we have used the Thyretain ™ bioassay to assess the TSI levels in sera from 13 patients with EGD, including several seen on repeated visits within a few months of the onset of symptoms. The TSI levels of these EGD patients were compared with normal controls and with three other patient groups; hyperthyroid Graves’ disease patients with and without ophthalmopathy, and 12 of their euthyroid relatives, belonging to single family with a high prevalence of thyroid autoimmunity.

Clinical Subjects and Methods

Clinical subjects

We studied sera from; 1) 13 patients with euthyroid Graves’ disease, 9 females and 4 males aged 43-70 (mean age 56years). Multiple serum samples from 6 of the patients and single samples from 7 patients were tested in single Thyretain TSI reporter bioassays. All 13 patients were diagnosed as “euthyroid Graves’ disease” at the time of the initial blood sample. Although 3 patients (nos. 1,2 and 3 Table 1), subsequently converted to Graves’ hyperthyroidism (2 patients) or had developed Graves’ disease many years earlier (1 patient).