SUN-560 Thyrotoxic Hashimoto's Disease: Is It Graves' Thyrotoxicosis or "Hashitoxicosis"?

We have postulated that a defect in specific antigenic induction of suppressor T lymphocytes may account for the immunoregulatory disorder in autoimmune thyroid disease. In this context, we have measured the proliferative responses of peripheral blood mononuclear cells (PBMC) to the synthetic peptides corresponding to the extracellular domain of the TSH receptor (TSHR) and recombinant glutamate decarboxylase (GAD65) by means of 3H thymidine incorporation. We have also studied the antigenic activation of CD4+ and CD8+ T lymphocytes by measuring human leukocyte antigen-DR (HLA-DR) expression on the cell surface by flow cytometric analysis. PBMC obtained from 47 patients with Graves' disease (GD) [including 19 hyperthyroid GD (hyper GD)], 18 with Hashimoto's thyroiditis (HT), 7 with nontoxic nodular goiter (NG), 18 with insulin-dependent diabetes (IDDM), and 20 normal controls (N), were cultured for 7 days in the presence or absence of the pool peptides representing 3 different segments of TSHR or GAD65 at final concentration of 30 micrograms/mL or 10 micrograms/mL. The proportion of subjects whose PBMC gave a positive proliferative response with a stimulation index (SI) of over 2.3 (i.e. above the mean +2 SD for N) to TSHR peptides was significantly higher in the hyper GD group than among euthyroid GD (eu GD), HT, IDDM, and N group. The corresponding differences in mean SI provided analogous results, showing significant responses above normal in only hyper GD. The CD4+ T lymphocytes from hyper GD group were significantly more activated by TSHR peptides compared to eu GD, HT, IDDM, and N, and this induction correlated to their thyroid hormone levels. Quite differently, the activation of CD8+ T lymphocytes from both hyper GD and eu GD group in response to TSHR peptides was impaired compared to HT, IDDM, and the N group; in contrast to the findings with CD4+ T lymphocytes, this was independent of thyroid hormone levels. On the other hand, while the CD8+ T lymphocytes from GD and N groups were activated equally by GAD65, the activation of CD8+ T lymphocytes from the IDDM group by GAD65 was impaired compared to the GD and N groups. In conclusion, the activation of CD8+ T lymphocytes from GD and IDDM by relevant antigens (i.e. TSHR peptides for GD and GAD65 for IDDM) was impaired, but not by irrelevant antigens (i.e. GAD65 for GD and TSHR peptides for IDDM). There was also a modest stimulation of CD8+ T cells from all groups by tetanus toxoid and cardiac myosin light chain peptide, both irrelevant antigens.(ABSTRACT TRUNCATED AT 400 WORDS)

Thyroid function in autoimmune thyroid disease (AITD) among patients with Down syndrome (DS) sometimes exhibits fluctuations over time, but the underlying cause of this variability remains unknown. Thyrotoxicosis is often associated with Graves disease (GD) and Hashitoxicosis. Thyroid-stimulating hormone (TSH) receptor antibody (TRAb) includes thyroid-stimulating antibody (TSAb) and TSH-stimulation blocking antibody (TSBAb), causing GD or hypothyroidism, respectively. We report two patients of DS showing discordance between thyroid function and TRAbs. Case 1: A 16-year-old boy with DS was diagnosed with Hashimoto thyroiditis at age 6, with positive TSAb and TSBAb. At age 7, GD was diagnosed based on thyrotoxicosis and elevated I123 uptake. His TRAb became negative after 18 months of methimazole treatment. During follow-up, recurrent thyrotoxicosis with positive TRAb/TSAb improved transiently without intensified treatment. Methimazole was discontinued at age 14, but hypothyroidism with positive TSAb and negative TSBAb became evident. Case 2: An 18-year-old woman with DS was diagnosed with Hashimoto thyroiditis at age 13. She experienced recurrent transient thyroiditis with positive TRAb/TSAb. Although TSAb remained positive and TSBAb negative, hypothyroidism continued without GD. In conclusion, our findings indicate that awareness of the potential discordance between TSAb/TSBAb status and thyroid function is important for deciding appropriate treatment in DS patients with unstable thyroid function and AITD.

The association of low vitamin D status with autoimmune thyroid disease (AITD), which includes Hashimoto's thyroiditis (HT) and Graves' disease (GD), is controversial. The aim of this study was to investigate the correlation between vitamin D insufficiency and AITD, HT, and GD. Our cross-sectional study included patients who visited a tertiary hospital for thyroid evaluation or follow-up. A total of 776 patients in whom serum 25-hydroxyvitamin D [25(OH)D], thyroid function, and anti-thyroid antibodies had been measured were analyzed retrospectively. Vitamin D insufficiency was defined as serum 25(OH)D level <75 nmol/L. The prevalence of vitamin D insufficiency was significantly higher in the 369 patients with AITD than in the 407 without AITD (46.1% vs. 37.1%, p=0.011), and higher in the 221 patients with HT than in those with GD or non-AITD (48.9% vs. 41.9%, 37.1%, p=0.017). Among HT cases, patients with overt hypothyroidism had a significantly higher prevalence of vitamin D insufficiency (60.4% vs. 44.1%, 21.7%, 37.1%, respectively, p <0.001) and lower 25(OH)D levels (80.1 ± 47.7 vs. 99.34 ± 61.2, 110.3 ± 69.9, 99.6 ± 53.7 nmol/L, respectively, p=0.009) compared with those with euthyroidism and subclinical hypothyroidism or those without AITD. Serum 25(OH)D levels were significantly negatively correlated with serum thyroid stimulating hormone (TSH) levels after adjustment for age, sex, body mass index, and sampling season (r= -0.127, p=0.013). Vitamin D insufficiency was associated with AITD and HT, especially overt hypothyroidism. Low serum vitamin D levels were independently associated with high serum TSH levels.

The Pathogenesis of Graves’ Disease

P.1.137 Autoimmune thyroiditis (AIT) and affective disorders — a prospective pilot study

Association of PD-1 (rs36084323) gene polymorphism with autoimmune thyroid diseases.

Osteopontin (OPN) is recognized as a potent immunoregulator of autoimmune disease. In the study, we tried to explore the association of serum OPN levels with autoimmune thyroid disease, including Graves' disease (GD) and Hashimoto's thyroiditis (HT), in an ethnic Chinese population. We enrolled 131 patients with GD, 33 patients with HT and 123 healthy controls. Serum OPN, B cell-activating factor (BAFF) and interferon (IFN)-α levels were quantified. Graves' disease patients with high thyroid function at the time of sample collection were defined as having active GD, while the other patients were defined as having inactive GD. Serum OPN levels were higher in active GD than in inactive GD and the control groups (P=0.001 and P=0.018, respectively). In GD, significant associations of OPN levels with thyroid-stimulating hormone receptor antibody (TSHRAb) levels were observed in women (r=-0.344, P=0.002, and r=0.440, P=0.004, respectively) but not in men. Osteopontin levels were associated with BAFF levels only in women with GD or HT (r=0.506, P<0.001 and r=0.430, P=0.025, respectively), but not in men with GD or HT. Serum OPN levels were upregulated in active GD, and serum OPN levels were associated with thyroid function and TSHRAb levels in GD. Additionally, OPN levels were correlated with BAFF levels in GD and HT. The associations of OPN levels with clinical phenotypes of GD and BAFF levels showed a dimorphic pattern.

A coexistence of mitral valve prolapse (MVP) with autoimmune thyroid disease (AITD) has been described, but there are not sufficient data to explain this association. The aim of the present study was to investigate the prevalence of MVP in patients with AITD and to evaluate whether any correlation between MVP and certain immunological parameters exists. M-mode, two-dimensional Doppler echocardiography was performed in 29 patients with Graves' disease (GD), 35 with Hashimoto's thyroiditis (HT), 20 with nonautoimmune goiter, and 30 normal controls. Serum samples were examined for antinuclear antibodies (ANA), antibodies against extractable nuclear antigen (ENA), antiphospholipid antibodies (aCL), rheumatoid factor (RF), thyroid autoantibodies (TAAb), immunoglobulins and C3, C4. Eight of 29 GD patients and 8 of 35 HT patients had MVP, while none of the control group and 2 of 20 of the simple goiter group had MVP (p < 0.05). ANA were detected at low titers in 5 of 8 in MVP(+) GD versus 3 of 21 in MVP(-) GD (p < 0.05). In the HT group the MVP(+) patients had a significantly higher incidence of ANA and ENA, 5 of 8 and 2 of 8 versus 5 of 27 and 0 of 27 of MVP(-) patients, respectively, p < 0.05. A statistically significant higher incidence of aCL was found in HT MVP(+) patients. (3/8) versus HT MVP(-) 1/27, p < 0.05. RF levels (immunoglobulin A [IgA]) were significantly higher in MVP(+) patients. The association of MVP with nonorgan-specific autoantibodies indicates that MVP may also be an autoimmune disease. It is possible that patients with AITD who also have MVP may be at an increased risk to develop systemic autoimmunity.

TSH Receptor Antibodies: Relevance & Utility

IntroductionInterferon (IFN)-β is known as an environmental trigger for the occurrence of autoimmune thyroid disease (AITD). However, the association of another type-1 IFN, IFN-β, with AITD is unknown.Material and methodsIn the study, we explored the association of serum IFN-β levels with AITD in an ethnic Chinese (i.e., Taiwanese) population. We enrolled 160 patients with Graves’ disease (GD), 47 patients with Hashimoto’s thyroiditis (HT), and 119 healthy controls. Serum IFN-β and B-cell activating factor (BAFF) levels were quantified in healthy controls at the baseline and in patients with AITD either prior to receiving medication or while under medication. Thyroid function and thyroid-stimulating hormone receptor antibody (TSHRAb) levels were measured at the time of serum collection.ResultsSerum IFN-β levels were lower in the HT group than in the control group (p = 0.031). A significant inverse correlation was observed between IFN-β and TSHRAb levels in men with GD (r = –0.433, p = 0.044). Serum IFN-β levels were also negatively associated with BAFF levels in men with GD, HT, and AITD (r = –0.320, p = 0.032; r = –0.817, p = 0.047; and r = –0.354, p = 0.011, respectively), but not in women with GD, HT, or AITD.ConclusionsSerum IFN-β levels were lower in HT patients. Correlations of serum IFN-β with TSHRAb and BAFF levels were found to be gender-specific. Further well-designed studies with larger sample sizes are required to confirm our findings.

The N-glycome of immunoglobulin G (IgG), the most abundant glycoprotein in human blood serum, reflects pathological conditions of autoimmunity and is sensitive to medicines applied in disease therapy. Due to the high sensitivity of N-glycosylation, the IgG N-glycan profile may serve as an indicator of an ongoing inflammatory process. The IgG structure and its effector functions are strongly dependent on the composition of N-glycans attached to the Fc fragment, and the binding of antigens is regulated by Fab sugar moieties. Because of the crucial role of N-glycans in IgG function, remodeling of its N-oligosaccharides can induce pathological changes that ultimately contribute to the development of autoimmunity; restoration of their physiological structure is critical to the reduction of disease symptoms. Our recently published data have shown that the pathology of autoimmune thyroid diseases (AITDs), including Hashimoto’s thyroiditis (HT) and Graves’ disease (GD), is accompanied by alterations of the composition of IgG N-glycans. The present study is a more in-depth investigation of IgG glycosylation in both AITDs, designed to determine the relationship between the severity of thyroid inflammation and IgG N-glycan structures in HT, and to assess the impact of immunosuppressive therapy on the N-glycan profile in GD patients. The study material consisted of human serum samples collected from donors with elevated anti-thyroglobulin (Tg) and/or anti-thyroperoxidase (TPO) IgGs without symptoms of hypothyroidism (n=68), HT patients characterized by high autoantibody titers and advanced destruction of the thyroid gland (n=113), GD patients with up-regulated IgG against thyroid-stimulating hormone receptor (TSHR) before (n=62) and after (n=47) stabilization of TSH level as a result of methimazole therapy (study groups), and healthy donors (control group, n=90). IgG was isolated from blood serum using protein G affinity chromatography. N-glycans were released from IgG by PNGase F digestion and analyzed by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) after 2-aminobenzamide (2-AB) labeling. UPLC-MS chromatograms were integrated into 25 peaks (GP) in the Waters UNIFI Scientific Information System, and N-glycans were assigned based on the glucose unit values and mass-to-charge ratios (m/z) of the detected ions. The Kruskal-Wallis non-parametric test was used to determine the statistical significance of the results (p<0.05). The obtained results suggest that modifications of IgG sialylation, galactosylation and core-fucosylation are associated with the severity of HT symptoms. Methimazole therapy implemented in GD patients affected the IgG N-glycan profile; as a result, the content of the sialylated and galactosylated oligosaccharides with core fucose differed after treatment. Our results suggest that N-glycosylation of IgG undergoes dynamic changes during the intensification of thyroiditis in HT, and that in GD autoimmunity it is affected significantly by immunosuppressive therapy.

Polymorphic variants of cytotoxic T-lymphocyte antigen-4 (CTLA-4) and forkhead box protein P3 (FOXP3) genes are implicated in dysregulated immune homeostasis and autoimmune disorders. We analyzed the association between CTLA-4 rs231775 and FOXP3 rs3761548, rs3761549 polymorphisms and predisposition to autoimmune thyroid disease (AITD), inclusive of Hashimoto's thyroiditis (HT) and Graves' disease (GD) in South-Indian population. A total of 355 AITD subjects (comprising 275 HT and 80 GD) and 285 randomly selected age- and sex-matched control subjects were genotyped for the aforementioned polymorphisms by PCR-RFLP method. The rs231775 "G" allele was preponderant in HT and GD subjects when compared with controls and exerted a dominant influence on the susceptibility to HT (p = 0.009) and GD (p = 0.02), respectively. There was no allelic association of rs3761548 and rs3761549 polymorphisms with AITD susceptibility, albeit a significant difference in genotype distribution with respect to rs3761549. Haplotype analysis revealed an increased frequency of rs3761548 "C"-rs3761549 "T" in HT and GD subjects, thereby associating it with disease predisposition (p = 0.03). Epistatic interaction analysis by multifactor dimensionality reduction approach revealed redundancy between CTLA-4 and FOXP3 genes in influencing the susceptibility to AITD. The genetic variation in CTLA-4 gene with reference to rs231775 polymorphism contributes to an increased predisposition to HT and GD. Also, in conjunction with FOXP3 gene variants it seems to influence the susceptibility to HT and GD respectively. The significance of these findings in combination with antithyroid antibody screening could plausibly contribute towards meticulous case-finding for effective treatment of HT and GD.

To estimate abnormalities in humoral or cellular immunity that relate to the etiologies of Graves' disease (GD) and Hashimoto's thyroiditis (HT), peripheral mononuclear cell subpopulations were enumerated by an immunofluorescence technique using monoclonal antibodies. Also antithyrotropin receptor antibodies (thyrotropin-binding inhibitor immunoglobulin, TBII) were measured by the radioreceptor assay according to Smith's method; and antithyroglobulin antibodies (TGHA) and antithyroid microsomal antibodies (MCHA), by the tanned red cell hemagglutination technique. The data obtained were analyzed for the count of peripheral total white cells, lymphocytes and granulocytes, and to the level of serum free thyroxine (T4), free T4 index (FT4I) and free triiodothyronine index (FT3I). Peripheral white cells tended to be decreased in some cases of GD and HT. The absolute count of lymphocytes was slightly increased in GD but did not change in HT. On granulocytes (neutrophils mainly), the absolute count was considerably decreased in hyperthyroid GD. In this disease, FT3I showed significantly positive correlations to the percentage and absolute count of peripheral lymphocytes, whereas FT4I revealed significantly negative correlations to the percentage and count of granulocytes. These facts indicate that lymphocytosis and granulocytopenia in GD might be ascribed partially to the direct effects of thyroid hormones. The percentage of peripheral OKT3-positive cells (common T lymphocytes) was significantly decreased in untreated cases of GD, showing negative correlations to FT4, FT4I and FT3I. The absolute count of OKT4-positive cells (inducer/helper T lymphocytes) was increased in untreated cases of GD and hyperthyroid and euthyroid cases receiving antithyroid drugs (ATD) for GD. But peripheral OKT8-positive cells (suppressor/killer T lymphocytes) was significantly decreased in HT. The percentage of peripheral OKT8-positive cells in GD was also decreased in untreated and remitted cases, but slightly more increased in ATD-medicated cases than in nonmedicated cases. The ratios of OKT4-positive cells to OKT8-positive cells (OKT4+/OKT8+ ratios) were significantly increased in untreated, ATD-medicated euthyroid and remitted cases of GD, and in HT. It seems that ATD may exert direct effects on OKT8-positive cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Several mechanisms are probably involved in determining the evolution of autoimmune thyroid disease (AITD) towards either hypothyroidism and the clinical syndrome known as Hashimoto's thyroiditis (HT) or toward hyperthyroidism and the symptoms of Graves' disease (GD). To gain further insight into such mechanisms we performed an exhaustive comparative analysis of the expression of key molecules regulating cell death (Fas, Fas ligand [FasL], Bcl-2) and apoptosis in both thyrocytes and thyroid infiltrating lymphocytes (TILs) from patients with either GD or HT. GD thyrocytes expressed less Fas/FasL than HT thyrocytes, whereas GD TILs had higher levels of Fas/FasL than HT TILs. GD thyrocytes expressed increased levels of the antiapoptotic molecule Bcl-2 compared to the low levels detected in HT thyrocytes. The opposite pattern was observed in GD (low Bcl-2) and HT (high Bcl-2) TILs. The patterns of apoptosis observed were consistent with the regulation of Fas, FasL, and Bcl-2 described above. Our findings suggest that in GD thyroid the regulation of Fas/FasL/Bcl2 favors apoptosis of infiltrating lymphocytes, possibly limiting their autoreactive potential and impairing their ability to mediate tissue damage. Moreover, the reduced levels of Fas/FasL and increased levels of Bcl-2 should favor thyrocyte survival and favor the thyrocyte hypertrophy associated with immunoglobulins stimulating the thyrotropin (TSH) receptor. In contrast, the regulation of Fas/FasL/Bcl2 expression in HT promotes thyrocyte apoptosis, tissue damage, and a gradual reduction in thyrocyte numbers leading to hypothyroidism. These findings help define key molecular mechanisms contributing to the clinical outcome of thyroid autoimmunity.

To characterize and compare autoimmune comorbidity patterns and temporal dynamics in patients with Hashimoto’s thyroiditis (HT) and Graves’ disease (GD) within an autoimmune polyglandular syndrome (APS) framework. We retrospectively analysed patients with an autoimmune thyroid disease (AITD) and at least one additional autoimmune disease fulfilling APS criteria, followed at a tertiary APS referral centre between 2000 and 2025. The prevalence of associated autoimmune diseases was compared between HT and GD. Time to development of subsequent autoimmune diseases was assessed using Kaplan–Meier analysis and Cox proportional hazards models, with adjustment for sex and age. A total of 1,057 patients (76% women; mean age 53.7 ± 16.4 years) were included, of whom 964 had HT and 93 GD. Type 1 diabetes was the most frequent autoimmune comorbidity, but was significantly less prevalent in GD than in HT (OR 0.48, 95% CI 0.31–0.75; FDR-adjusted p = 0.008). GD was selectively enriched for systemic sclerosis (OR 51.54, 95% CI 10.96–242.41; FDR-adjusted p < 0.001) and vitiligo (OR 3.46, 95% CI 1.82–6.55; FDR-adjusted p < 0.001). In patients in whom AITD was the first autoimmune manifestation (n = 333), GD was associated with a longer latency to additional autoimmune diseases and a lower hazard of subsequent autoimmunity compared with HT (HR 0.52, 95% CI 0.37–0.73; p < 0.001). Within APS-related AITD, HT and GD anchor distinct autoimmune phenotypes with different clustering patterns and temporal trajectories. These differences have relevant clinical implications for risk stratification and personalized surveillance of patients with autoimmune thyroid disease.