The enigma of persistent symptoms in hypothyroid patients treated with levothyroxine: A narrative review.

A significant minority of patients with hypothyroidism report persistent symptoms despite achieving normal thyroid biochemistry after levothyroxine (L-T4) replacement. Four principal lines of thinking, which are not mutually exclusive, may explain this enigma. The 'low tissue liothyronine hypothesis' emphasizes the potential imperfections of L-T4 replacement therapy that may lead to hypothyroidism in some tissues such as the brain, while others (eg hypothalamus) are euthyroid. The 'Somatic Symptom and Related Disorders hypothesis' draws attention to an incidental coexistence of a diagnosis of Somatic Symptom and Related Disorders in patients with treated hypothyroidism. The 'autoimmune neuroinflammation hypothesis' highlights the potential consequences of inflammatory mediators due to thyroid autoimmunity (the commonest cause of hypothyroidism) on the brain. The 'comorbidities and psychosocial hypothesis' implicates a variety of physical and psychosocial factors that have been noted to be associated with a diagnosis of hypothyroidism, which may be primarily the cause of persistent complaints. Over the past twenty years, a great deal of time and effort has been expended pursuing the 'low tissue liothyronine hypothesis', which has failed to yield results that translate to patient benefits. This has skewed the balance in clinical practice, in favour of pursuing answers relating to L-T4 and liothyronine combination treatment, while the alternative explanations have been downplayed and potentially useful interventions have been given insufficient attention.

Primary hypothyroidism and quality of life.

In the 1970s, treatment with thyroid extract was superseded by levothyroxine, a synthetic L form of tetraiodothyronine. Since then, no major innovation has emerged for the treatment of hypothyroidism. The biochemical definition of subclinical hypothyroidism is a matter of debate. Indiscriminate screening for hypothyroidism has led to overdiagnosis and treatment initiation at lower serum levels of thyroid-stimulating hormone (TSH) than previously. Adverse health effects have been documented in individuals with hypothyroidism or hyperthyroidism, and these adverse effects can affect health-related quality of life (QOL). Levothyroxine substitution improves, but does not always normalize, QOL, especially for individuals with mild hypothyroidism. However, neither studies combining levothyroxine and liothyronine (the synthetic form of tri-iodothyronine) nor the use of desiccated thyroid extract have shown robust improvements in patient satisfaction. Future studies should focus not only on a better understanding of an individual's TSH set point (the innate narrow physiological range of serum concentration of TSH in an individual, before the onset of hypothyroidism) and alternative thyroid hormone combinations and formulations, but also on autoimmunity and comorbidities unrelated to hypothyroidism as drivers of patient dissatisfaction. Attention to the long-term health consequences of hypothyroidism, beyond QOL, and the risks of overtreatment is imperative.

Autoimmune Hypercalcemia Due to Autoantibodies Against the Calcium-sensing Receptor.

CONTEXT: Autoimmune hypocalciuric hypercalcemia (AHH) is an acquired disorder caused by the presence of blocking autoantibodies against the calcium-sensing receptor (CaSR). Few cases of this condition have been described to date in the literature. OBJECTIVE: The objectives of this study were to describe 2 patients in whom the presence of AHH was suspected and to assess the patients for the presence of CaSR antibodies. METHODS: CaSR antibodies were detected and characterised by immunoprecipitation assays, CaSR peptide ELISAs, and functional assays based on the calcium-stimulated accumulation of inositol-1-phosphate in a mammalian cell line expressing the CaSR. RESULTS: Both patients presented with an acquired form of hypocalciuric hypercalcemia. Mutational analyses of CASR, GNA11, and AP2S1 for familial hypocalciuric hypercalcemia were negative. According to the presence of Hashimoto's disease in 1 patient and latent autoimmune diabetes of adulthood and thyroid autoimmunity in the other, AHH was suspected. Immunoprecipitation assays detected CaSR antibodies in both patients. Analysis of the antibody binding sites revealed 2 main epitopes at amino acids 41-69 and 114-126. Preincubation with purified CaSR antibodies against epitope 114-126 resulted in a significant decrease in inositol-1-phophate accumulation upon calcium-stimulation of mammalian cells expressing the CaSR, suggesting that the antibodies had receptor-blocking activity. CONCLUSIONS: AHH is to be suspected in patients with an acquired biochemical pattern of PTH-dependant hypocalciuric hypercalcemia, especially in those with other concomitant autoimmune diseases. Diagnosis by means of detecting CaSR antibodies may help to better characterise this probably under-reported condition.

Immunosuppressive therapy of autoimmune hypoparathyroidism in a patient with activating autoantibodies against the calcium-sensing receptor.

CONTEXT: Activating antibodies directed at the extracellular calcium-sensing receptor (CaSR) have been described in autoimmune hypoparathyroidism in the setting of isolated hypoparathyroidism or autoimmune polyglandular syndrome type 1. MATERIALS AND METHODS: A 34-year-old female presented with hypocalcaemia (6.0 mg/dL) and hypomagnesaemia (1.1 mg/dL) accompanied by low serum PTH (2.4 pg/mL) as well as urinary calcium and magnesium wasting. She was diagnosed with hypoparathyroidism, which was refractory to standard therapy. She was started on 60 mg prednisone and 150 mg azathioprine treatment daily on suspicion of an autoimmune aetiology. The patient was tested for CaSR antibodies. RESULTS: The patient was positive for CaSR antibodies of the IgG1 subtype, which stimulated phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and inositol phosphate (IP) accumulation. Post-treatment with prednisone and azathioprine, her serum calcium and magnesium normalized, as did her CaSR antibody titre and antibody-mediated stimulation of ERK1/2 phosphorylation and IP accumulation. CONCLUSION: This is the first demonstration of CaSR antibody-mediated hypoparathyroidism responsive to immunosuppressive therapy, adding to the evidence that autoimmune hypoparathyroidism can be, in some cases, reversible and not the result of autoimmune parathyroid destruction.

Calcium-Sensing Receptor Autoantibodies in Patients with Autoimmune Polyendocrine Syndrome Type 1: Epitopes, Specificity, Functional Affinity, IgG Subclass, and Effects on Receptor Activity.

A major manifestation of autoimmune polyendocrine syndrome type 1 (APS1) is hypoparathyroidism, which is suggested to result from aberrant immune responses against the parathyroid glands. The calcium-sensing receptor (CaSR), which plays a pivotal role in maintaining calcium homeostasis by sensing blood calcium levels and regulating release of parathyroid hormone (PTH), is an autoantibody target in APS1. In this study, the aim was to characterize the binding sites, specificity, functional affinity, IgG subclass, and functional effects of CaSR autoantibodies using phage-display technology, ELISA, and bioassays. The results indicated that CaSR autoantibody binding sites were at aa 41-69, 114-126, 171-195, and 260-340 in the extracellular domain of the receptor. Autoantibodies against CaSR epitopes 41-69, 171-195, and 260-340 were exclusively of the IgG1 subclass. Autoantibody responses against CaSR epitope 114-126 were predominantly of the IgG1 with a minority of the IgG3 subclass. Only autoantibodies recognizing CaSR epitopes 114-126 and 171-195 affected receptor activity; inositol-phosphate accumulation was increased significantly in HEK293-CaSR cells, and PTH secretion from PTH-C1 cells was reduced significantly when either were incubated with purified Ab and Ca2+ compared with Ca2+ alone. In conclusion, although the majority of APS1 patients do not have CaSR-stimulating autoantibodies, the hypoparathyroid state in a small minority of patients is the result of functional suppression of the parathyroid glands.

Autoantibodies against the calcium-sensing receptor and cytokines in autoimmune polyglandular syndromes types 2, 3 and 4.

OBJECTIVE: The frequency of autoimmunity against the parathyroid glands in patients with polyglandular autoimmunity that is not due to autoimmune polyendocrine syndrome type 1 (APS1) is unclear. To investigate this, this study aimed to determine the prevalence of autoantibodies against parathyroid autoantigens, calcium-sensing receptor (CaSR) and NACHT leucine-rich-repeat protein 5 (NALP5), in a large group of patients with non-APS1 polyendocrine autoimmunity. Possible occult APS1 was investigated by cytokine autoantibody measurement and AIRE gene analysis. DESIGN, SUBJECTS AND MEASUREMENTS: Subjects were 178 patients with APS2, 3 or 4, and 80 healthy blood donors. Autoantibodies against the CaSR, NALP5 and cytokines were measured by immunoprecipitation, radioligand binding assays or ELISA, respectively. RESULTS: Four patient samples (2.2%), but none of the controls, were positive for CaSR autoantibodies. NALP5 autoantibodies were not detected in any participant. Eleven patients (6.2%) had cytokine autoantibodies, but none of the control samples was positive. None of the patients with cytokine autoantibodies had any known or novel mutations in the AIRE gene. CONCLUSIONS: The low prevalence of CaSR autoantibodies indicate a very low level of subclinical parathyroid autoimmunity in APS types 2, 3 and 4. In addition, autoantibodies against cytokines constitute an uncommon feature of non-APS1 polyglandular autoimmunity.

Genome-wide association studies of autoimmune vitiligo identify 23 new risk loci and highlight key pathways and regulatory variants.

Vitiligo is an autoimmune disease in which depigmented skin results from the destruction of melanocytes, with epidemiological association with other autoimmune diseases. In previous linkage and genome-wide association studies (GWAS1 and GWAS2), we identified 27 vitiligo susceptibility loci in patients of European ancestry. We carried out a third GWAS (GWAS3) in European-ancestry subjects, with augmented GWAS1 and GWAS2 controls, genome-wide imputation, and meta-analysis of all three GWAS, followed by an independent replication. The combined analyses, with 4,680 cases and 39,586 controls, identified 23 new significantly associated loci and 7 suggestive loci. Most encode immune and apoptotic regulators, with some also associated with other autoimmune diseases, as well as several melanocyte regulators. Bioinformatic analyses indicate a predominance of causal regulatory variation, some of which corresponds to expression quantitative trait loci (eQTLs) at these loci. Together, the identified genes provide a framework for the genetic architecture and pathobiology of vitiligo, highlight relationships with other autoimmune diseases and melanoma, and offer potential targets for treatment.

Early-onset hypoparathyroidism and chronic keratitis revealing APECED.

Early diagnosis of potentially life-threatening autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is crucial, but is often delayed due to the clinical heterogeneity of the disorder. Even in the absence of the classic disease triad of chronic mucocutaneous candidiasis, hypoparathyroidism, and adrenocortical insufficiency, a diagnosis of APECED should be considered in children who have hypoparathyroidism and chronic keratitis, with a past medical history showing a mild and transient Candida infection.

Dominant Mutations in the Autoimmune Regulator AIRE Are Associated with Common Organ-Specific Autoimmune Diseases.

The autoimmune regulator (AIRE) gene is crucial for establishing central immunological tolerance and preventing autoimmunity. Mutations in AIRE cause a rare autosomal-recessive disease, autoimmune polyendocrine syndrome type 1 (APS-1), distinguished by multi-organ autoimmunity. We have identified multiple cases and families with mono-allelic mutations in the first plant homeodomain (PHD1) zinc finger of AIRE that followed dominant inheritance, typically characterized by later onset, milder phenotypes, and reduced penetrance compared to classical APS-1. These missense PHD1 mutations suppressed gene expression driven by wild-type AIRE in a dominant-negative manner, unlike CARD or truncated AIRE mutants that lacked such dominant capacity. Exome array analysis revealed that the PHD1 dominant mutants were found with relatively high frequency (>0.0008) in mixed populations. Our results provide insight into the molecular action of AIRE and demonstrate that disease-causing mutations in the AIRE locus are more common than previously appreciated and cause more variable autoimmune phenotypes.

IL-14 and IL-16 are expressed in the thyroid of patients with either Graves' disease or Hashimoto's thyroiditis.

OBJECTIVES: Cytokines have an important role in orchestrating the pathophysiology in autoimmune thyroid disease. The aim of the current study was to analyse the expression of interleukin (IL)-14 and IL-16 in the thyroid tissue of patients with Graves' disease (GD), Hashimoto's thyroiditis (HT) or multinodular goitre (MNG) and in that of normal individuals, in patients' intrathyroidal CD4(+) and CD8(+) T cells, and in patient and normal cultured thyroid follicular cells. METHODS: The expression of IL-14 and IL-16 mRNA and protein was investigated using reverse transcription-polymerase chain reaction (RT-PCR) amplification, and Western blotting and ELISAs, respectively. RESULTS: IL-14 mRNA expression was detected in thyroid tissue from 8/9 GD, 3/4 HT and 3/13 MNG patients and 1/6 normal individuals, and IL-16 mRNA expression in thyroid tissue from 9/9 GD, 4/4 HT and 9/13 MNG patients and 4/6 normal individuals. IL-14 mRNA expression was detected in intrathyroidal CD4(+) and CD8(+) T cells from 2/2 GD and 2/2 HT patients, while IL-16 mRNA was detected in samples from 1/2 HT patients but not in those from either patient with GD. IL-14 and IL-16 mRNA expression was found in thyroid follicular cells derived from 2/2 patient with GD and 1/1 normal individual. IL-14 protein was detected in thyroid tissue from 6/6 GD, 1/1 HT and 0/6 MNG patients and 0/6 normal individuals, and IL-16 protein in thyroid tissue from 6/6 GD, 1/1 HT and 1/6 MNG patients and 0/6 normal individuals. Expression of IL-14 protein was stimulated in thyroid follicular cells derived from two patients with GD and one normal individual by peripheral blood mononuclear cell (PBMC)-conditioned medium. Treatment of thyrocytes from two patients with GD and one normal individual with PBMC-conditioned medium and tumour necrosis factor (TNF)-α stimulated IL-16 protein expression. In normal thyrocytes, IL-16 protein synthesis was induced also by IL-1ß, IL-17A, IL-4 and transforming growth factor (TGF)-ß. CONCLUSIONS: The data provide evidence that the intrathyroidal production of IL-14 and IL-16 is associated with the pathogenesis of autoimmune thyroid disease. Thyroid follicular cells display the ability to express IL-14 and IL-16 mRNA and can be stimulated to express IL-16 protein, by a panel of cytokines, and IL-14 protein, by as yet unidentified factors.

Regulatory T cells in vitiligo: Implications for pathogenesis and therapeutics.

Vitiligo is a hypomelanotic autoimmune skin disease arising from a breakdown in immunological self-tolerance, which leads to aberrant immune responses against melanocytes. Regulatory T cells (Tregs) are crucial to the development of self-tolerance and so are major foci in the study of autoimmune pathogenesis of vitiligo. This review will summarise recent findings concerning the role of Tregs in the pathogenesis of vitiligo. In addition, as antigen-specific Tregs are a potential route for the reinstatement of immune tolerance, new strategies that expand or induce de novo generation of Tregs and which are currently being investigated as therapies for other autoimmune diseases, will be discussed. These approaches will highlight the opportunities for Treg cell-based therapeutics in vitiligo.

Thyroid abnormalities.

Thyroid abnormalities and nonthyroidal illness complicate human immunodeficiency virus (HIV) infection. Among the effects that result from HIV and other opportunistic infections, distinctive features of HIV infection include early lowering of reverse tri-iodothyromine (T3) levels, with normal free T3 levels. Later, some patients develop an isolated low free thyroxine level. After highly active antiretroviral therapy, the immune system reconstitutes in a way that leads to dysregulation of the autoimmune response and the appearance of Graves disease in 1% to 2% of patients. Opportunistic thyroid infections with unusual organisms are most commonly asymptomatic, but can lead to acute or subacute thyroiditis.

Impact of month of birth on the development of autoimmune thyroid disease in the United Kingdom and Europe.

CONTEXT: Viral/bacterial infection is proposed as a trigger for the autoimmune thyroid diseases (AITD): Graves' disease (GD) and Hashimoto's thyroiditis (HT). Previous studies in European Caucasian AITD subjects found higher birth rates in the autumn/winter, suggesting those born in the autumn/winter experience increased viral/bacterial exposure after birth, impacting upon immune system development and predisposing to AITD later in life. OBJECTIVE: Month of birth effects were investigated in three independent European Caucasian AITD datasets. DESIGN: Variation in GD and HT onset was compared across months and seasons, with fluctuations across all 12 months analyzed using a Walter-Elwood test. SETTING: The study was conducted at a research laboratory. PATIENTS: National UK Caucasian AITD Case Control Collection (2746 GD and 502 HT compared with 1 423 716 UK births), National UK Caucasian GD Family Collection (239 GD and 227 unaffected siblings), and OXAGEN AITD Caucasian Family Collection (885 GD, 717 HT, and 794 unaffected siblings of European Caucasian decent). MAIN OUTCOME MEASURES: Case-control and family-based association studies were measured. RESULTS: No consistent month of birth effects were detected in GD females or males across all three collections. In HT females from the OXAGEN AITD Caucasian Family Collection, slightly higher birth rates were detected in autumn (Walter's test statistic = 7.47, P = .024) however, this was not seen in the HT females from the case-control cohort. CONCLUSION: Our results suggest in UK/Northern European Caucasian GD subjects, month of birth does not impact on AITD development. Although some month of birth effects for HT females in one collection cannot be excluded, only further work in larger European Caucasian AITD collections can confirm these effects.

Pendrin and NIS antibodies are absent in healthy individuals and are rare in autoimmune thyroid disease: evidence from a Danish twin study.

OBJECTIVE: Antibodies against thyroglobulin, thyroid peroxidase and the TSH receptor are accepted as pathophysiological and diagnostic biomarkers in autoimmune thyroid disease (AITD). In contrast, the prevalence, aetiology and clinical relevance of autoantibodies against the human sodium-iodine symporter (NISAb) and pendrin (PenAb) remain unclear. The objectives of the study were to investigate the presence of NISAb and PenAb in Danish twins, with and without AITD, to study whether the published variations in NISAb and PenAb frequencies were related to differences in methodology or study populations, and to evaluate whether the presence of NISAb or PenAb most likely results from genetic or nongenetic factors. METHODS: Sera from 93 patients with AITD and 230 healthy controls were evaluated for NISAb and PenAb using radioligand binding assays (RBA). RESULTS: Patients with AITD had a higher prevalence than the controls: NISAb: 17% vs 0% (P < 0·001) and PenAb: 11% vs 0% (P < 0·001). Subdividing according to cause of AITD yielded similar results: 20% (11/56) of patients with Graves' disease (GD) and 14% (5/37) of patients with Hashimoto's thyroiditis (HT) had NISAb, (P < 0·05, vs control population). Seven of 56 (13%) patients with GD and three of 37 (8%) patients with HT had PenAb (P < 0·05 vs control population). No twin pairs were concordant for NISAb or PenAb, not even among twin pairs concordant for AITD. CONCLUSIONS: In accord with studies using the same RBAs, the frequency of NISAb and PenAb was low in Danish patients with AITD and absent in healthy individuals, suggesting that differences between studies rely on assay differences. The skewed distribution of NISAb and PenAb within AITD concordant twin pairs suggests that NISAb and PenAb are likely attributable to the effects of environmental factors acting in genetic susceptible individuals.

Graves' disease following immune reconstitution or immunomodulatory treatment: should we manage it any differently?

Graves' disease and other disorders of thyroid function may occur following treatment with novel anticancer agents or during periods of lymphocyte recovery after lymphopenia. There are three main settings for such lymphocyte reconstitution: recovery after a bone marrow or haematopoietic stem cell transplant, alemtuzumab treatment and the use of highly active antiretroviral therapy (HAART) for human immunodeficiency virus infection. The available evidence suggests that Graves' disease behaves as normal in most of these cases and should be treated conventionally, but it may follow a more favourable course in those receiving alemtuzumab or HAART. As spontaneous or drug-induced remission may be more likely in these two settings, first-line treatment should usually consist of an antithyroid drug.

Prevalence and clinical associations of calcium-sensing receptor and NALP5 autoantibodies in Finnish APECED patients.

CONTEXT: Previous studies have identified the calcium-sensing receptor (CaSR) and NALP5 as parathyroid autoantibody targets in patients with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED). However, although NALP5 antibodies have been associated with the occurrence of hypoparathyroidism (HP) in APECED, it is unclear whether CaSR antibodies are a specific or sensitive marker for APECED-associated HP. OBJECTIVE: The objective of the study was to identify associations between the presence of CaSR and NALP5 antibodies and the disease manifestations and demographic characteristics of Finnish APECED patients. DESIGN, SUBJECTS, AND METHODS: This was a case-control study including 44 APECED patients and 38 age- and sex-matched healthy controls. Antibodies against the CaSR and NALP5 were detected using immunoprecipitation assays and radioligand binding assays, respectively. RESULTS: CaSR and NALP5 antibodies were detected in 16 of 44 (36%) and 13 of 44 (30%) patients, respectively. No statistically significant associations were found between the presence of CaSR or NALP5 antibodies and the disease manifestations of APECED including HP (P > .05). For the diagnosis of HP, CaSR and NALP5 antibodies had specificities of 83% and 50%, respectively, and sensitivities of 39% and 26%, respectively. A significant association between both a shorter APECED and HP duration (<10 y) and positivity for CaSR antibodies was noted (P = .019 and P = .0061, respectively). CONCLUSION: Neither CaSR nor NALP5 antibodies were found to be specific or sensitive markers for HP in APECED. Further investigations are required to determine the exact role of the autoimmune response against the CaSR and NALP5 in the pathogenesis of this autoimmune syndrome.

Acquired hypocalciuric hypercalcemia in a patient with CKD.

We present a case of an 82-year-old woman with elevated parathyroid hormone (PTH) levels, hypocalciuria, hypercalcemia, and stage 3 chronic kidney disease. Hypocalciuria initially was attributed to chronic kidney disease, and hypercalcemia was attributed to primary hyperparathyroidism. Subsequent laboratory studies showed autoantibodies in the patient's serum directed against the calcium-sensing receptor (CaSR). Functional testing in a CaSR-transfected human embryonic kidney-293 cell line showed that the patient's antibodies inhibited CaSR-mediated intracellular signaling that ordinarily would have been stimulated by extracellular calcium ions. Her serum calcium and PTH levels were normalized by treatment with the calcimimetic cinacalcet. We advise consideration of the presence of inhibitory autoantibodies directed at the CaSR in patients with hypercalcemic hyperparathyroidism and unexplained hypocalciuria or with confounding conditions affecting interpretation of urinary calcium measurement. A calcimimetic is an effective treatment for the hypercalcemia and elevated PTH levels in acquired hypocalciuric hypercalcemia caused by inhibitory anti-CaSR autoantibodies.

Low frequency of pendrin autoantibodies detected using a radioligand binding assay in patients with autoimmune thyroid disease.

CONTEXT: Pendrin is a transmembrane protein located at the apical end of the thyrocyte in which it mediates the efflux of iodide through the thyroid follicular cell. Recently pendrin was described as a significant antibody target in Japanese patients with Graves' disease (GD) or autoimmune hypothyroidism (AH) using an immunoblotting assay. However, a subsequent study failed to verify this in autoimmune thyroid disease (ATD) patients of Tunisian origin. OBJECTIVE: The aim of the current study was to evaluate a UK population of patients with ATD for the presence of pendrin autoantibodies using a novel radioligand binding assay (RBA). RESULTS: Sera from 71 GD and 66 AH patients and 28 healthy controls were evaluated for pendrin autoantibody reactivity in RBAs. The results indicated that 8.8% of patients with ATD (9.9% GD and 7.6% AH) were positive for pendrin autoantibodies. Overall, the frequency of pendrin autoantibodies did not differ significantly between the ATD patient cohorts and the healthy control group: P = .186 and P = .317 for GD and AH patients, respectively. CONCLUSION: Pendrin autoantibodies, detected using a novel RBA, are not widely prevalent in UK patients with ATD, nor do they differ in frequency between GD and AH. These autoantibodies are therefore unlikely to be a useful marker for disease diagnosis, although the role that pendrin may play as an autoantigen in the initiation or maintenance of thyroid autoimmunity remains to be established.