Thyroid-stimulating immunoglobulins as a reliable predictive blood test for thyroid eye disease

Abstract

Background:

Thyroid stimulating hormone, T3, T4, and thyroid auto-antibodies are essential in diagnosis of immune thyroiditis, Graves’ and Hashimoto’s. These, however, may not correlate with thyroid eye disease or TED. In recent years, a new blood test for thyroid stimulating immunoglobulins (TSI) is in use. However, there is no sufficient clinical evidence of their role in TED. The purpose of the current study is to evaluate the correlation between TSI and TED activity and to examine if TSI is a predictor for the severity of the disease.

Methods:

A retrospective analysis of electronic medical records of all patients who attended TED clinic at Sheba Medical Center, Israel between 2017 and 2022 and had at least one TSI sample was performed. Data included demographics, comprehensive ophthalmic examination, clinical activity scores (CASs), laboratory tests, and quality of life assessment by the Graves’ Orbitopathy QOL questionnaire.

Results:

60 patients with 43 females (72%) were included. TSI at presentation correlated with eyelid retraction, IOP (Pearson correlation, 0.369, P< =0.01), and CAS (Pearson correlation, 0.392, P = 0.029). TSI values >500% were found to be significantly correlated with the use of medical treatment including high-dose steroids (P = 0.05), steroid treatment according to the European Group on Graves’ Orbitopathy protocol (P = 0.02), and radioactive iodine (P = 0.03). Moreover, it was also significantly correlated with surgical procedures including decompression (P = 0.016) and strabismus surgeries (P = 0.024).

Conclusions:

TSI is correlated with more severe clinical presentation and higher likelihood of medical or surgical treatment. Therefore, TSI can be considered as a predictor factor for the disease. Routing testing for TSI may be warranted for all TED patients.

Thyroid eye disease (TED), also known as Graves orbitopathy, is an autoimmune multisystem disorder.[1] It is usually seen in patients with hyperthyroidism, but it can also be diagnosed in patients with euthyroid, hypothyroid, or chronic autoimmune thyroiditis.[1,2]

The specific pathogenesis of the disease remains unknown.[1] However, the common hypothesis is that the same antibodies that attack the thyroid gland and cause systemic problems also recognize orbital fat and extraocular muscles as antigenic, causing orbital inflammation, eye muscle expansion, and fat expansion.[1] These antibodies are specific for fibroblast surface thyroid-stimulating hormone receptor (TSH-R) and insulin-like growth factor-1 receptor (IGF-1R).

In the past, the most common blood tests for evaluating TED patients were thyroid stimulating hormone (TSH), free T3 (fT3), and free T4 (fT4). However, with the understanding that these blood tests are not correlated with the severity of the disease, new blood tests were searched.

The TSH Receptor Antibody (TRAb) was discovered over a half century ago and is thought to play a major key role in the pathogenesis of TED due to the fact that fibroblast and adipose orbital tissue of TED patients were thought to express higher levels of TSHR than normal.[3] TSI is a stimulating type of TRAb which induces thyroid growth and increases thyroid hormone production and therefore thought to have a clinical relevance in the disease.[4]

A few studies investigating the correlation between TSI and the clinical activity score (CAS) found that TSI exhibits greater sensitivity than TRAb.[5] However, there is not enough evidence to support the role of TSI, disease activity, severity, and prognosis.

Therefore, the purpose of this current study was to evaluate the correlation between TSI and TED and to examine whether TSI can predict the severity of the disease and treatment outcome.

Methods

Study participants

This is a retrospective cohort study. Patients diagnosed with TED and treated at the multidisciplinary TED clinic at Sheba Medical Center between 2017 and 2021 and who have at least TSI result at presentation were included.

In this multidisciplinary clinic, on each visit, all patients were examined by consultant endocrinologists and ophthalmologists. The preferred management for each patient is a joint decision of all consultants after a comprehensive discussion.

Data regarding comprehensive eye examination, disease activity, and ancillary blood tests were collected and analyzed.

The study was approved by the local institutional review board (IRB) of Sheba Medical Center.

Assessing disease activity

TED activity was evaluated at each visit using CAS.[6] The CAS is based on the classical signs of inflammation, and it consists of seven items: spontaneous pain behind the globe, pain on attempted upward gaze, redness of the conjunctiva, redness of the eyelid, chemosis, swelling of the caruncle, and eyelid swelling. One point is given for each item that is present. CAS scores <3 indicate inactive GO, while scores ≥3 indicate active GO. There is an expanded 10-point CAS scoring system for patients who have subsequent assessments, with additional points allocated for decreases in visual acuity (VA), worsening diplopia, and increasing proptosis, as well as for scores ≥4 indicative of clinically active disease.

Patients with active disease were treated according to the EUGOGO protocol,[7] while patients with acute optic neuropathy were treated with high-dose intravenous methylprednisolone 1000 mg daily for 3 days and then 1 mg/kg of methylprednisolone tablets tapering down for a long time according to patients’ reaction to steroids, and they were referred to medial wall decompression in case of no response to methylprednisolone.

Strabismus surgeries and eyelid surgeries were performed, mainly in the nonactive phase of the disease.

Blood tests

The serum levels of TSH, fT4, and fT3 were measured with chemiluminescent microparticle immunoassays that have analytic sensitivities of 0.02 mIU/L, 0.4 ng/dl, and 1 pg/ml, respectively (Architect Analytical System Third Generation kits, Abbott Diagnostics, Abbott Park, IL). TSH levels were measured with an FDA-validated bioassay (Thyretain, Quidel Corp., San Diego, CA).

Serum TSI activity was measured with a functional cell-based TSHR bioassay (Thyretain, Quidel, CA, USA) according to the manufacturer’s instructions. Briefly, the levels of TSI activity were measured in triplicate with the Infinite M200 microplate reader (Tecan, Crailsheim, Germany). All measured values were corrected for the plate’s internal autoluminescence by reduction of the mean value in blank wells. The results were reported as percentage of specimen-to-reference ratio (SRR %). SRR % values were calculated according to the following formula: SRR % = Average TSI specimen relative light units (RLU)/average reference standard RLU × 100 as previously described. The patient’s serum was considered positive for the presence of TSI activity if the resultant SRR % measured ≥140% over the reference control. TSI above 500% was considered a high level.

Statistical analysis

Quantitative variables were described as mean ± standard deviation (SD). Categorical variables were described as absolute and relative frequencies. Matched pair test was performed to compare TSI at first visit versus last visit. Changes in TSI and changes in various clinical characteristics were calculated by comparing the result at the first visit to the clinic to the result at the last clinic visit. The correlations between TSI and various selected parameters were tested with Pearson’s correlation analysis.

The overall significance level was set to an alpha of 0.05. The statistical analysis was carried out with Microsoft Excel 2017 (Microsoft Corporation, Redmond, WA) and IBM SPSS software version 24.0 (SPSS, Inc., Chicago, IL).

Results

Demographics

Sixty patients (44 females, 73.33%) were evaluated in the TED multidisciplinary clinic during the 2-year study period and had at least one TSI test result. The mean age ± SD of the entire cohort was 49.26 ± 18.53 years (range 16–87). Twenty-three patients (28.3%) were smokers.

Thyroid blood tests at presentation

The mean TSH, T3, and T4 levels at first TED clinic visit were 3.85 ± 11.70 (0-67) mIU/L, 6.45 ± 5.51 (2-35) mIU/L, and 15.58 ± 5.32 (2-29) mIU/L accordingly. The mean level of TSI at the first visit was 1210.89 ± 1188 (71-4292). Forty-seven patients (73%) had TSI more than 500% at presentation.

There was a significant positive correlation between TSI at presentation and clinical characteristics such as IOP (intraocular pressure), MRD₁ (margin to reflex distance), PF (palpebral fissure), and Hertel measurements (P < 0.01, 0.013, <0.01, 0.050, Pearson’s correlation). There was a positive correlation between the CAS at presentation and the TSI level (0.392, 0.029). These correlations were not significant for TSH, T3, and T4. Table 1 summarizes the correlation at presentation.

Table 1.

Correlations between TSI at presentation and clinical characteristics

Clinical characteristic		Correlation to TSI		P (Pearson’s correlation)
Log MAR (VA)		0.017		0.908
HRR		-0.240		0.116
MD		-0.004		0.980
IOP		0.369		<0.01*
Hertel		0.429		0.050*
MRD1		0.354		0.013*
PF		0.455		<0.01*
CAS		0.392		0.029*

*Log MAR=Log of Minimum Angle of Resolution; HRR=Hardy Rand and Rittler; MD=Mean deviation; IOP=Intraocular pressure; MRD1=Margin reflex distance1; PF=Palpebral fissure; CAS=Clinical activity score

Patients with a TSI level of >500% at presentation were more likely to undergo surgical intervention or radiation: Decompression (9 patients vs. 0 patients, P = 0.016, chi_squre), strabismus (5 patients vs. 0 patients, P = 0.024, chi_squre), and radioactive iodine (7 patients vs. 0 patients, P = 0.037, chi_squre). Similarly, they received more IVGC per EUGOGO protocol (10 patients vs. 0 patients, P = 0.042, Chi-square) and more high-dose steroids of 1 gram methylprednisolone (9 patients vs. 1 patient, P = 0.050, chi_squre).

TSI at last visit

19 patients had TSI levels at their last clinical visit. The mean TSH, T3, and T4 levels at last TED clinic visit were 2.64 ± 3.77 (0-21) mIU/L, 4.62 ± 0.640 (3-6) mIU/L, and 14.36 ± 2.55 (9-19) mIU/L accordingly. The mean level of TSI activity at last visit was 497 ± 1442 (94-1706). Twenty percent (20%) out of total study patients had TSI more than 500% at the last visit.

There was a significant positive correlation between TSI at last visit and clinical characteristics at last visit, like MRD₁ and PF (P = 0.02, <0.01, Pearson’s correlation), and significant negative correlation with HRR (P < 0.01, Pearson’s correlation). There was a positive correlation between the CAS at last visit and the level of TSI. These correlations were not statistically significant for TSH, T3, and T4. Table 2 summarizes the correlation at last visit.

Table 2.

Correlations between TSI at last visit and clinical characteristics

Clinical characteristic		Correlation to TSI		P (Pearson’s correlation)
Log MAR		0.079		0.672
HRR		-0.676		<0.01*
MD		-0.050		0.822
IOP		0.114		0.521
Hertel		0.119		0.699
MRD1		0.401		0.023*
PF		0.556		<0.01*
CAS		0.661		0.010 *

*Log MAR=Log of Minimum Angle of Resolution; HRR=Hardy Rand and Rittler; MD=Mean deviation; IOP=Intraocular pressure; MRD1=Margin reflex distance1; PF=Palpebral fissure; CAS=Clinical activity score

TSI trend

There were significant differences in TSI at presentation versus at last follow-up (P < 0.01, paired samples t-test matched pairs). Fig. 1 presents the change in TSI per patient and for all groups.

Figure 1.

Change in TSI per patient and for the whole group

The change in TSI was positively significantly correlated with the change in clinical characteristics like change in Hertel measurements, change in MRD1, and change in PF (P = 0.024, <0.01, <0.01, Pearson correlation). Moreover, there was significant positive correlation between the change in TSI and the change in CAS (P < 0.01, Pearson correlation).

Discussion

The purpose of the current study was to evaluate TSI levels with TED clinical activity, severity, and different treatment modalities. Higher levels of TSI were associated with disease severity, steroids treatment, and radiation.

In the last decade, it is known that the TRAB plays an important role in the pathogenesis of GD by causing thyroid stimulation and inducing hyperthyroidism.[3,4] There are three types of TRAB: natural, stimulating, or blocking.[8] TSI stimulates the TSH receptor, thus inducing thyroid growth and increasing thyroid hormone production. On the other hand, blocking TSH-R-Ab (TBII) acts as an antagonist to the TSH receptor.[9] TRAB immunoassay does not distinguish between stimulating and blocking autoantibodies; therefore, the diagnostic performance of TRAB as a biomarker has certain limitations.[9] Specific differentiation of TSI from other types of autoantibodies is thought to be important for the accurate diagnosis of GD.[10,11]

However, only a few studies examined the role of TSI in the management of TED patients.[5,12,13,14,15] Ponto KA et al. examined the correlation between TSI and TBII to both clinical activity score and clinical severity score.[16] In their study, TSI showed more significant association with clinical features of GO than TBII. Another study by Ponto KA et al., published at 2015, found that high levels of TSI identify patients with recent DON, which required urgent treatment.[13] Additional studies examined the correlation between TSI and CAS, finding a positive correlation between them.[17] This is in accordance with our findings.

In this current study, TSI levels were found to be correlated with clinical characteristics including increasing IOP, eyelids measurements (PF and MRD1), and proptosis (Hertel) at presentation. Moreover, the change in TSI during the follow-up period was found to be significantly correlated with the change in the CAS. However, these correlations were not found with thyroid function blood panel (TSH, T3, and T4). This may show that TSH, T3, and T4 may not correlate with the clinical characteristics, while TSI is.

Patients with high TSI levels at their first visit in the clinic underwent more surgical procedures compared to patients with low TSI levels at presentation. Moreover, they also needed more medical therapy including steroids treatment as well as radioactive iodine. This may implicate that patients with high TSI are prone to more medical and surgical treatments. Therefore, patients with high TSI at their presentation should be under very close follow-up and maybe earlier medical and surgical intervention should be considered in these cases.

TSI blood test is a relatively expensive test compared to the basic thyroid function blood panel. Moreover, when compared to TRAB examination, in the United States (USA), TSI blood test is more expensive.[12] However, according to this study, this blood test is more accurate to predict the clinical features and treatment of the disease. Therefore, it should be recommended to examine TSI levels at the presentation of a TED patient. The result of TSI level can help to find the patients who need close follow-up.

Limitations of the current study stem from its retrospective design. In addition, we included only patients with TSI examination with relatively short follow-up. This is due to the fact that the multidisciplinary TED clinic in Sheba Medical Center was initiated only in 2016. Further long-term studies should be done.

Conclusion

In this current study, we described correlation between TSI and clinical features of the TED as well as the CAS. Moreover, patients with high TSI are prone to undergo more medical and surgical therapies. Therefore, TSI may be considered as a good predictor factor for the activity and severity of the disease. It is recommended to add this blood test to the regular follow-up blood tests of patients with TED.

Compliance with ethical standards

Research involving human participants and/or animals

The study was approved by the local institutional review board (IRB) of Sheba Medical Center.

Informed consent

All patients signed an informed consent prior to study inclusion.

The manuscript has not been published elsewhere, has not been evaluated for publication previously by another journal, and was not submitted simultaneously for publication elsewhere.

Conflicts of interest:

There are no conflicts of interest.

Funding Statement

Nil.