Thyroid Diseases: Marinò M

Bartalena L, Baldeschi L, Dickinson AJ, Eckstein A, Kendall-Taylor P, Marcocci C, Mourits MP, Perros P, Boboridis K, Boschi A, Currò N, Daumerie C, Kahaly GJ, Krassas G, Lane CM, Lazarus JH, Marinò M, Nardi M, Neoh C, Orgiazzi J, Pearce S, Pinchera A, Pitz S, Salvi M, Sivelli P, Stahl M, von Arx G, Wiersinga WM. · Department of Clinical Medicine, University of Insubria, Varese, Italy. · Thyroid. · Pubmed #18341379 No free full text.

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Bartalena L, Marcocci C, Tanda ML, Piantanida E, Lai A, Marinò M, Pinchera A. · Department of Clinical Medicine, University of Insubria, Varese, Italy. · J Endocrinol Invest. · Pubmed #16075933 No free full text.

Abstract: Graves' ophthalmopathy (GO), the most frequent extrathyroidal manifestation of Graves' disease, is a disorder of autoimmune origin, the pathogenic mechanisms of which are still incompletely understood. Although GO is severe in only 3-5% of affected individuals, quality of life is severely impaired even in patients with mild GO. Management of severe GO can be either medical or surgical (orbital decompression, eye muscle or lid surgery). Medical management relies on the use of high-dose systemic glucocorticoids or orbital radiotherapy, either alone or in combination. Studies carried out in the last 5 yr have shown that glucocorticoids are more effective through the i.v. route than through the oral route. However, particular attention should be paid to possible liver toxicity of i.v. glucocorticoids. Recent randomized clinical trials have, with one exception, confirmed that orbital radiotherapy is an effective and safe therapeutic procedure for GO. At variance with previous encouraging data, recent randomized clinical trials have shown that currently available SS analogs are not very effective in the management of GO. Antioxidants might have a role, at least in mild forms of GO. Particular attention should be paid to correction of risk factors (cigarette smoking, thyroid dysfunction, radioiodine therapy) involved in GO progression.

Marcocci C, Marinò M, Rocchi R, Menconi F, Morabito E, Pinchera A. · Department of Endocrinology and Metabolism, University of Pisa, Pisa, Italy. · J Endocrinol Invest. · Pubmed #15165004 No free full text.

Abstract: Treatment of severe Graves' ophthalmopathy (GO) is a complex therapeutic challenge and, in spite of any efforts, about one third of patients are disappointed with the outcome of treatment. Glucocorticoids (GC), orbital radiotherapy (RT), or a combination of both, are most frequently used for their immunosuppressive effects. Novel immunosuppressive treatment procedures (or novel modalities of established treatments) are reviewed in the present article. GC has recently been used by the i.v. route and this treatment modality has been shown to be more effective and better tolerated than the oral route. Promising preliminary results have been reported by some authors with somatostatin analogs, octreotide and lanreotide. The number of patients treated so far is limited, most of the results have been obtained in nonrandomized or uncontrolled studies, and comparison with other validated methods of treatment is also needed. Because of the pathogenic role of cytokines, cytokine antagonists, currently evaluated in other autoimmune diseases, have been tested with positive results also in a small series of GO patients. The use of antioxidants might also be envisioned in the future, since in vitro studies have shown that oxygen free radicals might be involved in GO. Based on the shared antigen(s) theory, total thyroid ablation, by removing the bulk of shared antigens(s), might be beneficial for the course of GO. New data on recently performed placebo-controlled studies on orbital radiotherapy are discussed, together with studies on long-term safety of orbital radiotherapy.

Marinò M, Chiovato L, Lisi S, Altea MA, Marcocci C, Pinchera A. · Department of Endocrinology, University of Pisa, Italy. · J Endocrinol Invest. · Pubmed #15164998 No free full text.

Abstract: One of the hypothesis to explain the pathogenesis of Graves' ophthalmopathy (GO) was formulated by Joseph P. Kriss in the early 1970s. He postulated that the initiating event in the pathogenesis of GO is the deposition and accumulation of thyroglobulin (Tg) in orbital tissues, followed by an autoimmune reaction against Tg. In the last 30 yrs several studies have addressed this hypothesis, through various, different experimental approaches, raising results that are both in favor and against the possibility that Tg plays a role in the pathogenesis of GO. The finding that intact Tg is present in orbital tissues of GO patients supports Kriss' hypothesis, although the role of Tg as an autoantigen seems to be unlikely, as GO is not significantly associated with serum TgAb and mice immunized with Tg do not develop GO. Whether Tg is indeed involved in the pathogenesis of GO remains to be established. Our current view is that, provided that Tg plays a role, it is unlikely the only factor involved and Tg in orbital tissues may rather reinforce or worsen a damage initiated by other mechanisms.

Marcocci C, Bartalena L, Marinò M, Rocchi R, Mazzi B, Menconi F, Morabito E, Pinchera A. · Department of Endocrinology and Metabolism, University of Pisa, Pisa, Italy. · Ophthal Plast Reconstr Surg. · Pubmed #12439051 No free full text.

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Marinò M, Pinchera A, McCluskey RT, Chiovato L. · Department of Endocrinology, University of Pisa, Italy. · Thyroid. · Pubmed #11272097 No free full text.

Abstract: Megalin, a member of the low density lipoprotein endocytic receptor family, is expressed on the apical surface of thyroid epithelial cells, directly facing the follicle lumen, where colloid is stored in high concentrations. Studies in vivo and with cultured thyroid cells have provided evidence that megalin expression on thyroid cells is TSH-dependent. Thyroglobulin (Tg), the major protein component of the colloid and the precursor of thyroid hormones, binds to megalin with high affinity and megalin mediates in part its uptake by thyrocytes. Tg internalized by megalin avoids the lysosomal pathway and is delivered by transepithelial transport (transcytosis) to the basolateral membrane of thyrocytes, from which it is released into the bloodstream. This process competes with pathways leading to thyroid hormone release from Tg molecules, which occurs following internalization of Tg molecules from the colloid by other means of uptake (fluid phase endocytosis or endocytosis mediated by low affinity receptors) that result in proteolytic cleavage in the lyosomes. During transcytosis of Tg, a portion of megalin (secretory component) remains complexed with Tg and enters the circulation, where its detection may serve as a tool to identify the origin of serum Tg in patients with thyroid diseases. Tg endocytosis via megalin is facilitated by the interaction of Tg with cell surface heparan sulfate proteoglycans, which occurs via a carboxyl terminal heparin binding site of Tg functionally related with a major megalin binding site. Although autoantibodies against megalin can be found in the serum of approximately 50% of patients with autoimmune thyroiditis, a role of megalin in this and other thyroid diseases remains to be established.

Marinò M, McCluskey RT. · Pathology Research Laboratory, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts 02129, USA. · Am J Physiol Cell Physiol. · Pubmed #11029276 links to  free full text

Abstract: Thyroglobulin (Tg), the thyroid hormone precursor, is synthesized by thyrocytes and secreted into the colloid. Hormone release requires uptake of Tg by thyrocytes and degradation in lysosomes. This process must be precisely regulated. Tg uptake occurs mainly by micropinocytosis, which can result from both fluid-phase pinocytosis and receptor-mediated endocytosis. Because Tg is highly concentrated in the colloid, fluid-phase pinocytosis or low-affinity receptors should provide sufficient Tg uptake for hormone release; high-affinity receptors may serve to target Tg away from lysosomes, through recycling into the colloid or by transcytosis into the bloodstream. Several apical receptors have been suggested to play roles in Tg uptake and intracellular trafficking. A thyroid asialoglycoprotein receptor may internalize and recycle immature forms of Tg back to the colloid, a function also attributed to an as yet unidentified N-acetylglucosamine receptor. Megalin mediates Tg uptake by thyrocytes, especially under intense thyroid-stimulating hormone stimulation, resulting in transcytosis of Tg from the colloid to the bloodstream, a function that prevents excessive hormone release.

Lisi S, Menconi F, Altea MA, Agate L, Molinaro E, Castagna MG, Taddei D, Grasso L, Pinchera A, Elisei R, Marinò M. · Department of Endocrinology, University of Pisa, Pisa, Italy. · J Endocrinol Invest. · Pubmed #15505986 No free full text.

Abstract: When thyroid follicles are intact, some colloidal thyroglobulin (Tg) reaches the circulation by megalin-mediated transcytosis and is to various extents complexed with megalin secretory components. In contrast, in papillary thyroid cancer (PTC), serum Tg is not complexed with megalin because it is directly secreted by tumor cells. Here we attempted to use measurement of megalin secretory components to distinguish PTC patients with thyroid remnant plus metastases from those with thyroid remnant only, after thyroidectomy and before 131I ablation. Tg values in anti-Tg antibodies (TgAb)-free sera from 5 PTC patients with thyroid remnant plus metastases and 12 PTC patients with thyroid remnant only were measured following pre-adsorption with uncoupled protein A beads or with protein A beads coupled with antimegalin antibodies. The degree of Tg pre-adsorption with antimegalin antibodies was minimal, with no substantial differences between the two groups. Thus, we concluded that measurement of megalin secretory components is unlikely to be useful to identify the origin of serum Tg in PTC patients after thyroidectomy.

Lisi S, Marinò M, Pinchera A, Mazzi B, Di Cosmo C, Sellari-Franceschini S, Chiovato L. · Department of Endocrinology, Pisa, Italy. · Thyroid. · Pubmed #12097194 No free full text.

Abstract: One of the hypotheses that explains the pathogenesis of thyroid-associated ophthalmopathy (TAO) is that thyroglobulin (Tg) is transported through a retrograde lymphatic route to orbital tissues (OT), where it elicits autoimmune damage. In a previous study we demonstrated the presence of intact Tg of thyroid origin in OT from three patients with TAO. The present study was undertaken to investigate this issue further, by increasing the number of patients, by analyzing the distribution of Tg in OT, and by investigating possible relations between the presence of Tg in OT and the clinical features of patients. OT was obtained from seven patients with TAO who underwent decompressive orbitotomy via a transpalpebral approach. Patients were designated P10 to P16. Inflamed palpebral skin, retrobulbar fibroadipose tissue and extraocular muscle surgical samples were collected separately. Tissue extracts were prepared by homogenization and analyzed for the presence of Tg using two different techniques. We first performed immunoprecipitation experiments, in which a rabbit polyclonal anti-Tg antibody was used to capture Tg on protein A and a mouse monoclonal anti-Tg antibody was used to re-veal captured Tg by Western blotting. Intact 330-kd Tg was detected in retrobulbar fibroadipose tissue extracts from three patients (P10, P11, and P16), whereas no Tg was detected in retrobulbar fibroadipose tissue extracts from the remaining four patients. Tg was not detected in the extraocular muscle extracts from all patients studied. In addition, intact 330-kd Tg was found in the inflamed palpebral skin extract from one patient (P10). No Tg was detected in OT extracts from two patients without thyroid or eye disease and in abdominal adipose tissue extracts from two obese patients without thyroid or eye disease. We then searched for Tg by enzyme-linked immunosorbent assay (ELISA), using the same antibodies used for immunoprecipitation. Tg was detected in retrobulbar fibroadipose tissue extracts from four patients (P10, P11, P12, and P16) and in the inflamed palpebral skin extract from patient P10, in amounts ranging from approximately 125 to approximately 400 pg/microg of tissue protein. Again, Tg was not detected in extraocular muscle extracts. A positive gradient between Tg in OT and Tg in the serum was observed in patient P12. Using an ELISA approach, we found that Tg in OT from three TAO patients (P10, P11, and P12) contained thyroxine (T4) residues (mean T(4) CONTENT: 2.42 molecules per molecule of Tg), indicating that Tg had originated in the thyroid. Combining the results obtained in our previous and present study, we found a possible relation between the presence of Tg in OT and the previous treatment with radioiodine. Thus, of the seven patients (3 in the previous and 4 in the present study) in whose OT Tg was found, six had been treated with radioiodine, whereas of the three patients with no Tg in their OT none had been treated with radioiodine. In conclusion, Tg was found in OT extracts from patients with TAO by immunoprecipitation in three of seven cases and by ELISA in four of seven cases. Tg was found in retrobulbar fibroadipose tissue, but not in extraocular muscles. There was a relation between the presence of Tg in OT and the previous treatment with radioiodine. Our results support the hypothesis that Tg may play a role as a coantigen in the pathogenesis of TAO. Further studies are needed to investigate this possibility.

Marinò M, Chiovato L, Mitsiades N, Latrofa F, Andrews D, Tseleni-Balafouta S, Collins AB, Pinchera A, McCluskey RT. · Pathology Research Laboratory, Massachusetts General Hospital, Harvard Medical School, Charlestown 02129, USA. · J Clin Endocrinol Metab. · Pubmed #10999849 links to  free full text

Abstract: After its endocytosis from the colloid, some thyroglobulin (Tg) is transcytosed intact across thyrocytes, accounting in part for its presence in the circulation. We previously showed that megalin (gp330), an endocytic Tg receptor, mediates apical to basolateral Tg transcytosis. Here we investigated whether a portion of megalin remains combined with Tg after its transcytosis, using studies with cultured thyroid cells and in vivo observations. FRTL-5 cells, a rat thyroid cell line, cultured on filters in dual chambers form tight junctions and exhibit features of polarity, with expression of megalin exclusively on the upper (apical) surface. After the addition of unlabeled Tg to the upper chamber and incubation at 37 C, some Tg was transcytosed intact across FRTL-5 cells into the lower chamber. Two antimegalin ectodomain antibodies precipitated transcytosed Tg in fluids collected from the lower chamber. After the addition of Tg to surface-biotinylated FRTL-5 cells, an anti-Tg antibody and the two antimegalin ectodomain antibodies precipitated high molecular mass biotinylated material in fluids collected from the lower chamber, corresponding to much of the megalin ectodomain, as well as smaller amounts of lower molecular mass material. The results indicate that Tg transcytosed across FRTL-5 cells remains complexed with megalin ectodomain components, which we refer to as megalin secretory components. In aminotriazole-treated rats, which develop increased megalin-mediated Tg transcytosis, antimegalin antibodies precipitated some of the Tg in the serum. Tg was also precipitated by antimegalin antibodies in sera from patients with Graves' disease, in which we found increased megalin expression on the apical surface of thyrocytes. In contrast, in thyroidectomized patients with metastatic papillary thyroid carcinoma, in whom Tg is directly secreted by neoplastic thyroid cells into the circulation rather than transcytosed, serum Tg was not precipitated by antimegalin antibodies. The detection of Tg-megalin complexes may help identify the source of serum Tg in patients with thyroid diseases.

Bartalena L, Baldeschi L, Dickinson A, Eckstein A, Kendall-Taylor P, Marcocci C, Mourits M, Perros P, Boboridis K, Boschi A, Currò N, Daumerie C, Kahaly GJ, Krassas GE, Lane CM, Lazarus JH, Marinò M, Nardi M, Neoh C, Orgiazzi J, Pearce S, Pinchera A, Pitz S, Salvi M, Sivelli P, Stahl M, von Arx G, Wiersinga WM, Anonymous00022. · Department of Clinical Medicine, University of Insubria, 21100 Varese, Italy. · Eur J Endocrinol. · Pubmed #18299459 links to  free full text

This publication has no abstract.

Lisi S, Botta R, Pinchera A, Di Cosmo C, Perri A, De Marco G, Menconi F, Marinò M. · Department of Endocrinology and Metabolism, University of Pisa, 56124 Pisa, Italy. · J Endocrinol Invest. · Pubmed #18075286 No free full text.

Abstract: The LDL receptor-associated protein (RAP) is involved in secretion of thyroglobulin (Tg) from the thyrocyte to the colloid. Disruption of the RAP gene in mice results in a reduced Tg content within the colloid, leading to subclinical hypothyroidism and histological alterations resembling early goiter. Here we studied the entire coding sequence of RAP in genomic DNA samples from 18 patients with primary hypothyroidism not due to thyroid autoimmunity or dysgenesis. The control group included 21 subjects with no evidence of thyroid alterations. Eleven different polymorphisms with amino-acid substitution and 4 different missense polymorphisms without amino-acid substitution were found in various regions of the RAP gene. Only one polymorphism in exone 7 (V311M) was observed exclusively in patients, but it had been previously reported in normal subjects as well. The remaining polymorphisms were found either both in patients and controls or only in controls and had not been previously reported. The frequency of the various polymorphisms did not differ significantly between patients and controls. Based on these findings, we conclude that alterations of the RAP gene are not a common cause of hypothyroidism, although it cannot be excluded that other, rarer alterations with a pathogenic effect exist, and that they should be investigated in a larger number of patients.

Marcocci C, Pinchera A, Marinò M. · Department of Endocrinology and Metabolism, University of Pisa, Pisa, Italy. · Nat Clin Pract Endocrinol Metab. · Pubmed #17452970 No free full text.

Abstract: BACKGROUND: A 59-year-old woman with a history of nodular goiter developed thyrotoxic symptoms while on levothyroxine therapy. Her thyrotoxicosis persisted after levothyroxine withdrawal, so she was given methimazole and, once euthyroid, underwent near-total thyroidectomy. Histological examination revealed a nodular variant of Graves' disease. Proptosis, eyelid swelling and diplopia appeared 2 months after surgery. These symptoms worsened, and the patient was initially given four intravenous pulses of glucocorticoids, which resulted in a transient amelioration of her eye symptoms. After glucocorticoid withdrawal, however, the patient's eye motility worsened and there was a reduction of visual acuity in the left eye. She was then referred to our hospital for further advice and treatment. INVESTIGATIONS: Complete thyroid and ophthalmological evaluation, computerized visual field analysis, CT scan of the orbits, routine blood tests, search for occult fecal blood, blood tests for hepatitis B and C virus markers, measurements of serum non-organ-specific autoantibodies and serum anti-TSH-receptor antibodies, and liver ultrasonography. DIAGNOSIS: Nodular Graves' disease with severe, active Graves' orbitopathy complicated by optic neuropathy. MANAGEMENT: Intravenous glucocorticoid therapy for 3 consecutive days, followed by once-weekly pulses of intravenous glucocorticoids over a 10-week period, and then by oral prednisone treatment on alternate days for 2 months. During the first 2 weeks of intravenous glucocorticoid therapy the patient received orbital irradiation. Therapy resulted in optimized visual acuity and a moderate improvement of soft-tissue inflammatory signs and symptoms, whereas proptosis and eye motility improved only slightly. The patient is now scheduled for orbital decompression and rehabilitative surgery.

Menconi F, Marinò M, Pinchera A, Rocchi R, Mazzi B, Nardi M, Bartalena L, Marcocci C. · Department of Endocrinology, University of Pisa, Via Paradisa 2, 56100 Pisa, Italy. · J Clin Endocrinol Metab. · Pubmed #17299076 links to  free full text

Abstract: CONTEXT: Graves' orbitopathy (GO) is probably caused by autoimmune reactions against autoantigen(s) shared by thyroid and orbital tissues sustained by intrathyroidal autoreactive T-lymphocytes infiltrating the orbit. Total thyroid ablation (TTA) may be beneficial for GO through removal of shared antigen(s) and autoreactive T-lymphocytes, but randomized studies are lacking. OBJECTIVE: Our objective was to evaluate the effects of TTA in patients with GO treated with iv glucocorticoids (GC). DESIGN/SETTING: A prospective, single-blind, randomized study was conducted at a referral center. PATIENTS/INTERVENTIONS: Sixty patients with mild to moderate GO were randomized into: 1) near-total thyroidectomy (TX); or 2) TX plus (131)I (TTA) groups, and then treated with iv GC. Patients were evaluated 3 and 9 months after iv GC. MAIN OUTCOME MEASURE: Overall improvement of GO at 9 months was the main outcome measure. RESULTS: The distribution of GO outcome at 9 months was significantly more favorable in TTA than in TX patients (P = 0.0014 by chi(2) test). A cumulative significant (P = 0.0054) difference between the two groups at 3 and 9 months was found using a generalized linear model. Radioiodine uptake test and thyroglobulin assay in a patient sample showed complete ablation in the majority of TTA, but not of TX patients. CONCLUSIONS: Compared with thyroidectomy alone, TTA is followed by a better outcome of GO in patients given iv GC. Whether TTA maintains this advantage in the long-term remains to be established.

Botta R, Lisi S, Pinchera A, Segnani C, Cianferotti L, Altea MA, Menconi F, Mattii L, Corsini GU, Marcocci C, Dolfi A, Bernardini N, Marinò M. · Department of Endocrinology and Metabolism, University of Pisa, Via Paradisa 2, 56124, Pisa, Italy. · Thyroid. · Pubmed #17123336 No free full text.

Abstract: The low density lipoprotein (LDL) receptor-associated protein (RAP) is an endoplasmic reticulum (ER)-resident molecular chaperone for several LDL receptor family members and it also binds to thyroglobulin (Tg), the thyroid hormone precursor. Disruption of the RAP gene in thyrocytes results in impaired Tg secretion. To gain further insights into the function of RAP in the thyroid, we investigated whether its expression in thyrocytes is regulated by thyroid-stimulating hormone (TSH), a feature common to all proteins involved in thyroid hormone secretion. We found by immunofluorescence that in FRTL-5 cells cultured in the presence of TSH, RAP is expressed intracellularly. The levels of expression increased after exposure to TSH, beginning at 48 hours, in a concentration-dependent manner as observed by immunofluorescence and Western blotting. Expression of RAP was also increased by TSH in primary cultures of human thyrocytes as observed by Western blotting. In hypothyroid mice with high serum TSH, RAP was markedly increased compared with euthyroid mice as observed by immunohistochemistry and Western blotting. Based on these findings, we concluded that RAP is expressed by thyrocytes in a TSH-dependent manner, both in cultured thyroid cells and in vivo.

McKeag D, Lane C, Lazarus JH, Baldeschi L, Boboridis K, Dickinson AJ, Hullo AI, Kahaly G, Krassas G, Marcocci C, Marinò M, Mourits MP, Nardi M, Neoh C, Orgiazzi J, Perros P, Pinchera A, Pitz S, Prummel MF, Sartini MS, Wiersinga WM, Anonymous00103. · Centre for Endocrine and Diabetes Sciences, University Hospital of Wales, Cardiff CF 4 4XN, UK. · Br J Ophthalmol. · Pubmed #17035276 No free full text.

Abstract: BACKGROUND: This study was performed to determine clinical features of dysthyroid optic neuropathy (DON) across Europe. METHODS: Forty seven patients with DON presented to seven European centres during one year. Local protocols for thyroid status, ophthalmic examination and further investigation were used. Each eye was classified as having definite, equivocal, or no DON. RESULTS: Graves' hyperthyroidism occurred in the majority; 20% had received radioiodine. Of 94 eyes, 55 had definite and 17 equivocal DON. Median Clinical Activity Score was 4/7 but 25% scored 3 or less, indicating severe inflammation was not essential. Best corrected visual acuity was 6/9 (Snellen) or worse in 75% of DON eyes. Colour vision was reduced in 33 eyes, of which all but one had DON. Half of the DON eyes had normal optic disc appearance. In DON eyes proptosis was > 21 mm (significant) in 66% and visual fields abnormal in 71%. Orbital imaging showed apical muscle crowding in 88% of DON patients. Optic nerve stretch and fat prolapse were infrequently reported. CONCLUSION: Patients with DON may not have severe proptosis and orbital inflammation. Optic disc swelling, impaired colour vision and radiological evidence of apical optic nerve compression are the most useful clinical features in this series.

Lisi S, Botta R, Agretti P, Sellari-Franceschini S, Marcocci C, Pinchera A, Marinò M. · Department of Endocrinology, University of Pisa, Pisa, Italy. · J Endocrinol Invest. · Pubmed #16075925 No free full text.

Abstract: It has been proposed that thyroglobulin (Tg) may be involved in the pathogenesis or the progression of Graves' ophthalmopathy (GO). According to this hypothesis, following its release from the thyroid, Tg would reach orbital tissues, thereby eliciting an autoimmune aggression. In support of this, we recently found that intact Tg is present in orbital tissues of patients with GO, where it is complexed with glycosaminoglycans. In this study, we searched for additional Tg binding sites in orbital tissues, using primary cultures of orbital and skin fibroblasts from 7 GO patients who had undergone orbital decompression. Biotin-labeled Tg bound to both skin and orbital fibroblasts in a saturable manner, with constants of dissociation of approximately 75 nmol/l for skin fibroblasts and approximately 40 nmol/I for orbital fibroblasts. In an attempt to identify Tg binding sites, fibroblast extracts were blotted onto membranes that were incubated with biotin-labeled Tg, which bound especially to a protein migrating at approximately 300 kDa, present in both orbital and skin fibroblast extracts. Because no appreciable inhibition of binding of biotin-labeled Tg was produced by unlabeled Tg, we concluded that binding was poorly specific and it is unlikely to be involved in the pathogenesis of GO.

Marinò M, Morabito E, Altea MA, Ambrogini E, Oliveri F, Brunetto MR, Pollina LE, Campani D, Vitti P, Bartalena L, Pincheral A, Marcocci C. · Department of Endocrinology, University Hospital of Pisa, Pisa, Italy. · J Endocrinol Invest. · Pubmed #15952415 No free full text.

Abstract: We report a case of acute hepatitis of autoimmune origin which occurred in a 43-yr-old woman during iv glucocorticoid (GC) pulse therapy for Graves' ophthalmopathy (GO). Prior to therapy, liver function tests were normal with no previous history of liver disorders or conditions predisposing to GC-associated liver damage. After the administration of a 4.7-g cumulative dose of methylprednisolone acetate, there was a marked increase of liver enzymes, prompting immediate discontinuation of iv GC. Nevertheless, liver enzymes increased further, reaching a peak 45 days later, with values 30- to 50-fold greater than those prior to therapy, associated with evidence of impaired liver function. Liver biopsy showed a marked lymphocytic infiltration, likely indicating an autoimmune hepatitis. Based on the assumption that following GC-induced immune suppression, autoimmune hepatitis might have been precipitated by sudden re-activation of the immune system during interpulse periods, we treated the patient with im and then oral GC, in order to re-induce immune suppression. Within three days from re-institution of GC therapy, there was a marked reduction of liver enzymes and amelioration of liver function. Complete normalization was achieved two months later, while the patient was still receiving a low maintenance dose of oral prednisone.

Marinò M, Lisi S, Pinchera A, Marcocci C, Menconi F, Morabito E, Macchia M, Sellari-Franceschini S, McCluskey RT, Chiovato L. · Department of Endocrinology, University of Pisa, Pisa, Italy. · Thyroid. · Pubmed #14588099 No free full text.

Abstract: The presence of thyroglobulin (Tg) in orbital tissues of patients with thyroid-associated ophthalmopathy (TAO) supports a role of Tg in TAO pathogenesis. To search for Tg-binding sites in orbital tissues, because Tg is a heparin-binding protein, we investigated its binding to glycosaminoglycans (GAGs) that are abundant in orbital tissues: chondroitin sulfate B (CSB) and C (CSC) and hyaluronic acid (HA). Both in solid phase and solution phase assays purified human Tg bound to GAGs. In solid-phase assays, binding was increased by coincubation with heparin or GAGs in solution, or with an antibody against a Tg heparin-binding sequence (Arg2489-Glu2503), possibly suggesting crosslinking of Tg molecules induced by GAGs or by the presumably bivalent antibody. Orbital tissue extracts from TAO patients that contained Tg were subjected to high-salt treatment, which resulted in separation of Tg from GAGs, as observed by column chromatography. After separation from GAGs, the Tg in orbital tissue extracts acquired the ability to bind to immobilized CSB, and heparin enhanced binding, resembling the findings with purified human Tg. Therefore, we conclude that GAGs provide binding sites for Tg in orbital tissues, which may explain the presence of Tg in orbital tissues of patients with TAO.

Marcocci C, Bartalena L, Rocchi R, Marinò M, Menconi F, Morabito E, Mazzi B, Mazzeo S, Sartini MS, Nardi M, Cartei F, Cionini L, Pinchera A. · Department of Endocrinology, University of Pisa, 56124 Pisa, Italy. · J Clin Endocrinol Metab. · Pubmed #12915636 links to  free full text

Abstract: We investigated the long-term side-effects of orbital radiotherapy (OR) in 204 patients with Graves' ophthalmopathy (GO), irradiated from 1972-1996 [44 by cobalt unit (CU) and 160 by linear accelerator (LA), mostly combined with glucocorticoids], with a 5- to 25-yr follow-up (median, 11 yr). Cataract was observed in 21 patients (10%) 3-21 yr after OR, with a higher (not significant) prevalence in CU-treated patients (18% vs. 8% in LA-treated patients). The prevalence of cataract was higher, although not significantly, in CU-treated patients aged less than 60 yr, but not in LA-treated patients, compared with the general population. Mild, asymptomatic retinopathy was observed in 1 of 7 patients (14%) with diabetes and hypertension, in 1 of 31 patients (3%) with hypertension alone, and in 0 of 11 patients with diabetes alone. No tumors were observed in 157 patients submitted to computed tomography scan of orbital and adjacent regions. In conclusion, OR is a safe treatment, not associated with an increased frequency of cataract, provided a high voltage apparatus is used. Hypertension, especially if associated with diabetes, may represent a relative contraindication, as it may cause retinopathy. Although no secondary tumors were detected, due to the long latency of radiation-induced tumors, OR should be restricted to patients older than 35 yr.

Marinò M, Lisi S, Pinchera A, Mazzi B, Latrofa F, Sellari-Franceschini S, McCluskey RT, Chiovato L. · Department of Endocrinology, University of Pisa, Italy. · Thyroid. · Pubmed #11288989 No free full text.

Abstract: Thyroid-associated ophthalmopathy (TAO) is thought to have an autoimmune pathogenesis because of its association with autoimmune thyroid disease, in particular with Graves' disease. Nevertheless, the nature of the autoimmune reaction is unclear, and a target orbital autoantigen has not been conclusively identified. A widely discussed hypothesis is that antigens constitutively shared by the thyroid and orbital tissues are targets of an autoimmune reaction. It has been also postulated that a thyroid-soluble antigen, namely thyroglobulin (Tg), is transported to orbital tissues through the lymphatics, where it accumulates and elicits autoimmune damages in susceptible individuals. Here we have investigated whether Tg is present in orbital tissues from patients with TAO. Retrobulbar tissue specimens were obtained from three patients with Graves' disease and TAO who underwent decompressive orbitotomy, and at autopsy from two patients with no thyroid or eye disease. All patients with TAO had been previously treated with radioiodine to control Graves' hyperthyroidism. Western blot analysis with a monoclonal anti-Tg antibody showed the presence of intact Tg, both in soluble and membrane-associated fractions of orbital tissue extracts from the patients with TAO, in amounts estimated to range from approximately 320 to approximately 900 pg/microg of tissue protein. In contrast, Tg was not detected in orbital tissue extracts from patients with no thyroid or eye disease. Tg was also demonstrated in orbital tissue extracts from two of three patients with TAO by enzyme-linked immunosorbent assay (ELISA), in amounts estimated to range from approximately 450 to approximately 1000 pg/microg of protein. In addition, Tg in orbital tissue extracts from patients with TAO was immunoprecipitated by a rabbit anti-Tg antibody, suggesting that it retained its native conformation. An anti-thyroxine (T4) antibody captured in solid-phase Tg from orbital tissue extracts, showing that it contained thyroid hormone residues and had therefore originated in the thyroid. Tg-anti-Tg immune complexes were not found in orbital tissues, suggesting that if an autoimmune reaction to Tg occurs in TAO, it is likely to be cell mediated.

Marinò M, Barbesino G, Pinchera A, Manetti L, Ricciardi R, Rossi B, Muratorio A, Braverman LE, Mariotti S, Chiovato L. · Department of Endocrinology, University of Pisa, Italy. · Thyroid. · Pubmed #11041457 No free full text.

Abstract: We previously showed that myasthenia gravis (MG) has a mild clinical expression when associated with autoimmune thyroid diseases (AITD). In the present study we have investigated the frequency of thyroid-associated ophthalmopathy (TAO) in patients with Graves' disease (GD) associated with MG as compared with GD patients without MG. A total of 418 patients with GD were studied, 31 with MG and 387 without MG. TAO was evaluated by physical examination, exophthalmometry, computerized tomography, and computerized visual fields assessment. The overall prevalence of TAO was similar in GD patients with MG (61.2%) and in those without MG (56.4%). When the analysis was restricted to GD patients with ocular MG, a greater frequency of TAO was found (84.6%), compared with GD patients without MG or with GD patients with generalized MG, although the differences did not reach the statistical significance. GD patients with MG had a significantly greater prevalence (12.9%) of euthyroid ophthalmopathy (clinically overt ophthalmopathy without previous and/or current hyperthyroidism) than those without MG (3.1%; p = 0.003). The results suggest a preferential association between the ocular manifestations of GD and MG, which may be due to immunological cross-reactivity against common autoimmune targets in the eye muscle as well as to a common genetic background.

Marinò M, Chiovato L, Friedlander JA, Latrofa F, Pinchera A, McCluskey RT. · Pathology Research Laboratory, Massachusetts General Hospital, Harvard Medical School, Charlestown 02129, USA. · J Clin Endocrinol Metab. · Pubmed #10404822 links to  free full text

Abstract: Megalin (gp330) is a multiligand receptor found on the apical surface of selected epithelial cells, including thyroid cells. We recently showed that megalin is a high-affinity receptor for thyroglobulin. Megalin is capable of inducing autoantibodies, as shown in the rat model, Heymann nephritis. Based on this consideration and on the knowledge that autoantibodies against several thyroid antigens develop in patients with autoimmune thyroid diseases, we searched for antimegalin antibodies in 78 patients with autoimmune and nonautoimmune thyroid diseases. We developed an assay, based on flow cytometry, to measure binding of serum IgGs to L2 cells, a rat carcinoma cell line that expresses abundant megalin. After incubation of L2 cells with serum samples and then with fluorescein isothiocynate-conjugated antihuman IgG Fc-specific antibody, the mean fluorescence intensity (MFI) was determined. Using results obtained in sera from 32 normal subjects, we established a cutoff value for MFI (50.62), above which, tests were considered positive. Significantly elevated values were found in 18 patients, including 13 of 26 patients with autoimmune thyroiditis (50.0%) and in 2 of 19 patients with Graves' disease (10.5%). Furthermore, 2 of 19 patients with nontoxic goiter (10.5%) and 1 of 14 patients with differentiated thyroid cancer (7.14%) had MFI values greater than 50.62, associated with the presence of circulating antithyroid autoantibodies. As a control cell line, we used Chinese hamster ovary cells, which do not express megalin. We found that, among the 18 patients with positive tests for binding to L2 cells, only 1 patient with nontoxic goiter had significant binding of serum IgGs to Chinese hamster ovary cells. Binding of serum IgGs to L2 cells was significantly reduced by coincubation with purified megalin in 15 of 18 positive patients (83.33%) and by a rabbit antimegalin antibody in 11 patients (61.11%). Further and more conclusive evidence that positive tests (MFI >50.62) for binding to L2 cells were attributable to serum antimegalin antibodies was demonstrated by immunoprecipitation experiments. After incubation of serum samples with L2 cell extracts, incubation with antihuman IgG Fc-specific agarose beads resulted in immunoprecipitation of megalin in all the 18 positive patients, but not in normal subjects, as assessed by Western blotting using a monoclonal antibody against megalin. Furthermore, the intensity of the band corresponding to megalin precipitated by serum IgGs in the above 18 patients was significantly correlated with the L2 binding MFI. This is the first clear-cut demonstration of antibodies against megalin in humans. Further studies are needed to determine whether antimegalin antibodies have pathogenic significance or diagnostic value in autoimmune thyroid diseases.