Gabriel Obiols

Characterization of neural cell adhesion molecule (NCAM) expression in thyroid follicular cells : induction by cytokines and over-expression in autoimmune glands

NCAM (CD56) is a cell surface glycoprotein of the immunoglobulin superfamily expressed on neuroendocrine and natural killer (NK) cells which has considerable molecular heterogeneity due to differential splicing and post‐translational modifications. NCAM has been detected in the thyroid follicular cells (thyrocytes) immunohistological. We report here the molecular form, the modulation by cytokines and the levels of expression in thyroid pathology. By using a panel of MoAbs to NCAM on Western blots from thyrocyte extract we have determined that these cells express the 140‐ and 180‐kD forms of NCAM, Exposure of primary cultures of thyrocytes to interferon‐gamma (IFN‐Γ), and even more, to the combination of IFN‐Γ plus tumour necrosis factor‐alpha (TNF‐α) induced a clear increase in the expression of NCAM as assessed by FACS analysis. NCAM expression in thyrocytes was assessed by immunofluorescence in 59 surgical specimens of thyroid glands, and was found increased in 11/17 (64%) of Graves’, in 5/25 (20%) of multinodular goitre (MNG) and in occasional adenoma glands. No correlation was found with the expression of HLA class I. class II or the degree of lymphocytic infiltration scored in adjacent sections, but it was often seen in areas infiltrated by macrophages. In conclusion, NCAM is an adhesion molecule whose expression is clearly increased in thyrocytes in autoimmune glands, probably as a consequence of exposure to cytokines locally released. Since one of the forms of NCAM expressed by thyrocytes has the capability to generate intracellular signal it may play a role in normal thyroid function. In addition, NCAM may facilitate the recognition of thyrocytes by lymphocytes, particularly by NK CD56+ lymphocytes.

Preoperative Thyrotropin Serum Concentrations Gradually Increase from Benign Thyroid Nodules to Papillary Thyroid Microcarcinomas Then to Papillary Thyroid Cancers of Larger Size

We evaluated the preoperative serum thyrotropin (TSH) levels in 386 patients operated on for nodular thyroid disease (NTD). TSH levels for cases with final benign disease and differentiated thyroid carcinoma (DTC) were compared. No evidence of cancer was detected in 310 patients (80.3%), whereas malignancy was present in 76 cases (19.7%). Mean TSH concentration was 1.36 ± 1.62 mU/L in benign patients and 2.08 ± 2.1 in cases with malignant lesions (P = 0.0013). The group of malignancy was subdivided in papillary thyroid carcinoma (PTMC) versus thyroid cancer of larger size (TCLS). Mean TSH was 1.71 ± 1.52 in PTMC and 2.42 ± 2.5 in TCLS. Significant differences were found when all groups (benign, PTMC and TCLS) were compared (P < 0.001). However, pairwise comparisons between them showed that differences were only significant between benign and TCLS groups (P < 0.01). In conclusion, TSH levels were higher in patients with a final diagnosis of DTC. Moreover, it appears that there exists an increment in tumor size as a function of increment in the TSH level.

Vía de señalización dependiente de la proteincinasa de activación mitogénica en el carcinoma papilar de tiroides. De las bases moleculares a la práctica clínica

In recent years, significant progress has been made in elucidating the genetic bases promoting tumorigenesis in various human neoplasms. Constitutive activation of the mitogen-activated protein kinase (MAPK) signaling pathway is a major event in the carcinogenesis of papillary thyroid carcinoma (PTC), the most prevalent endocrine malignancy. Affected elements include RET/PTC rearrangements and point mutations of the Ras and BRAF genes. Mutations in these genes are found in over 70% of PTC. Chromosomal RET rearrangements, called RET/PTC, result in constitutive ligand-independent activation of RET kinase, which was the first genetic anomaly detected in PTC and is found in 5-70% of tumoral samples. Although less frequent, the activation of other tyrosine kinase receptors, such as NTRK1, c-Met or EGFR, has also been reported in PTC. The BRAF mutation represents the most common genetic alteration found in PTC. More than 90% of BRAF mutations lead to a change of a valine to a glutamic acid at position 600 (V600E). Finally, Ras is the least affected molecule in the pathway. A relationship between clinical behavior and these genetic alterations has been proposed. Thus, the BRAF mutation is associated with a more aggressive PTC phenotype and is correlated with poorer outcomes. However, no clear association has been found between RET/PTC and clinical features. The discovery of these alterations opens the way to new therapeutic strategies, especially to treat those patients in whom conventional therapy is not effective. Several new drugs are being tested, such as small molecule tyrosine kinase inhibitors. Some of these recently developed agents have begun to be used with promising results.En los ultimos anos se ha empezado a descubrir las bases geneticas de la carcinogenesis en diferentes neoplasias humanas. La desregulacion de la via de senalizacion dependiente de la proteincinasa de activacion mitogenica (MAPK) es un mecanismo fundamental en la oncogenesis del carcinoma papilar de tiroides (CPT), el tumor endocrino mas frecuente. Concretamente, tres integrantes de la via –el receptor tirosincinasa RET, Ras y BRAF– se encuentran implicados en mas del 70% de los casos. El reordenamiento cromosomico de RET, denominado RET/PTC, ocasiona la activacion enzimatica de esta tirosincinasa. Fue la primera alteracion genetica especifica del CPT y se encuentra en un 5-70% de las muestras tumorales. En menor proporcion se han encontrado activaciones constitutivas de otros receptores tirosincinasa, como NTRK1, c-met o EGFR. La mutacion de BRAF es la alteracion genetica mas frecuente en el CPT. En la mayoria de los casos, la mutacion ocasiona un cambio de valina por acido glutamico en la posicion 600 (V600E). Finalmente, Ras es la molecula menos afectada de la via. La presencia de estas alteraciones se correlaciona, en algunos casos, con el comportamiento clinico de la enfermedad. Asi, la mutacion de BRAF condiciona una mayor agresividad y un peor pronostico. No hay tanto consenso en el papel de RET/PTC. Por otro lado, el descubrimiento de estas alteraciones abre la puerta a nuevas estrategias terapeuticas, en especial para los pacientes en quienes el tratamiento convencional resulta inefectivo. Entre las nuevas opciones se encuentran los inhibidores de las tirosincinasas. Algunos farmacos de esta familia han empezado a ser utilizados con resultados esperanzadores.

γσ Lymphocytes in endocrine autoimmunity: evidence of expansion in Graves' disease but not in type 1 diabetes

Endocrine autoimmune disorders are mediated by T cell‐dependent responses to organ‐specific antigens, but the mechanisms initiating the process remain unknown. Lymphocytes whieh use the γδ heterodimer as T ceii receptor (TCR) for antigen constitute a distinct subset of T cells whose function remains elusive. In order to investigate their possible involvement in endocrine auloimmunity we have determined the proportion of γδ T cells in the peripheral biood of 23 patients with type 1 (insulin‐dependent) diabetes mellitus (type‐1 DM) and 30 patients with autoimmune thyrotoxicosis (Graves’ disease). T lymphocyte TCR expression was assessed by fluorescence‐activated flow eytometry on peripheral blood mononuclear cells using MoAbs UCHTI (CD3), TCR 51 (γδ TCR), WT31 and βF1 (αβ TCR) and both the percentage of T cells expressing γδ and the ratio γδ/αβ were calculated. In the diabetie patients γδ cells were not significantly different from (he control group (7.7 ± 54%versus 8.0 ± 5.5%) of T eells, P NS). There was no relation between the proportion of γδ lymphoeytes and the presence ol’ islet cell antibodies (ICA) in the sera. The Graves’ patients showed a tendency towards a higher proportion of γδ T lymphocytes than the controls (γδ/αβ ratios: 0.095 ± 0.047 versus 0.063 ± 0.022, P= 0 03). In 14 Graves’ patients the number of γδ were measured in paired samples of peripheral and inlrathyroidal lymphocytes, demonstrating an expansion of γδ within the thyroid glands (0.21 ± 0.3 versus 0.095 ± 0.047, P= 0.032). Immunohistochemical studies showed that γδ celts were scattered among the predominant αβ lymphoeytes infiltrating the thyroid gland and that they aeeount for 10% of intraepitheliai lymphocytes. No relation was found between the increase of γδ lymphocytes and any clinieai features.

Clinical significance of RET/PTC and p53 protein expression in sporadic papillary thyroid carcinoma

Aims:  Rearranged during Transfection (RET)/papillary thyroid carcinoma (PTC) and p53 are two genes involved in the pathogenesis of PTC. It has been suggested that RET/PTC expression is associated with higher rates of local extension and lymph node involvement, whereas p53 mutations are more frequent in poorly differentiated and anaplastic carcinomas. In addition, experimental studies have shown that p53 activity can modify the behaviour of PTC carrying RET/PTC. The aim of this study was to investigate the expression of both RET/PTC and p53 in order to evaluate their usefulness as prognostic factors.

Differences in the Form of Presentation between Papillary Microcarcinomas and Papillary Carcinomas of Larger Size

Papillary thyroid carcinomas (PTCs) with a diameter ≤1 cm are referred to as papillary microcarcinomas (PTMCs). The prognostic factors for PTMCs have not been defined. Different clinical and histopathologic variables were studied in 152 PTCs, including 74 PTMCs and 78 PTCs of larger size. We found that PTMCs are associated with less multifocality (P = .046) and bilaterality (P = .003), fewer lymphadenectomies (P < .001), and a higher rate of incidental tumours (P < .001). Moreover, patients with a low aggressive profile were significantly older than the remaining patients (54 ± 13.7 years versus 45.8 ± 13.1 years; P = .001). In conclusion PTMCs show significant differences compared to PTCs of larger size in the form of presentation. Furthermore, it is possible that the classic risk factors, which are well validated in PTCs, such as age, must be cautiously interpreted in the current increasing subgroup of PTMCs.

Expression of p21cip1, p27kip1, and p16INk4a Cyclin-Dependent Kinase Inhibitors in Papillary Thyroid Carcinoma: Correlation with Clinicopathological Factors

In a variety of human malignancies, aberrant expression of proteins involved in the control of cell-cycle progression has been reported. In this study, p21cip1, p27kip1, and p16INk4a cyclin-dependent kinase inhibitors were analyzed to evaluate their usefulness in clinical management of papillary thyroid carcinoma (PTC). Archived material derived from 46 cases of PTC was analyzed immunohistochemically. Protein expression was ascertained on tissue microarrays, and results were correlated with clinicopathological features of the patients. Positive immunostaining was observed in 14 (30,4%) p21cip1, 26 (56,5%) p27kip1, and 14 (30,4%) p16INk4a cases. No significant correlation between p21cip1 or p27kip1 and clinical factors was found. In contrast, p16INk4a expression showed a significant correlation with initial extension of the disease. Therefore, 45.8% of patients with loco-regional extension were p16INk4a positive, whereas overexpression was only seen in 15.7% of cases with intrathyroid disease (p < 0.05). Moreover, all patients with simultaneous p16INk4a positivity and lack of p27kip1 staining (four patients) presented lymph node metastases. In contrast, only 12 (28.5%) of the remaining patients showed lymph node tumor involvement. In conclusion, p16INk4a expression suggests extrathyroid neck extension of PTC. This effect is enhanced when p27kip1 is negative. We think that their analysis by immunohistochemistry could be useful in the management of patients with PTC.

Preoperative TSH level and risk of thyroid cancer in patients with nodular thyroid disease: nodule size contribution ☆

BACKGROUND Many reports have supported the relationship between high preoperative TSH levels and risk of thyroid cancer in nodular thyroid disease (NTD). OBJECTIVES We investigated whether TSH levels are related to the risk of differentiated thyroid carcinoma (DTC) in patients who have undergone total thyroidectomy for NTD. The relationship between TSH and size of malignant nodule was investigated. Finally, we assessed whether TSH levels are related to DTC and presence of additional benign nodules. PATIENTS AND METHODS A retrospective study of 980 patients was conducted. Variables included age at diagnosis, TSH level, nodule size, gender, final histology (benign versus DTC), and type of malignancy. RESULTS Malignancy was present in 261 (26.6%) patients. These patients had higher median TSH levels as compared to those with no malignancy (1.61 mU/L (0.9-2.5) versus 0.9 mU/L (0.3-1.6); p-value<0.001). TSH was higher in patients with DTC in whom the largest nodule was malignant than in patients in whom the largest nodule was benign (1.80 mU/L (1.1-2.6) versus 1.38 mU/L (0.7-2.1) respectively; p-value=0.025). A significant correlation was seen between malignant nodule size and TSH level, but not between TSH levels and size of the largest benign nodule. CONCLUSIONS Our study supported an association between preoperative TSH levels and risk of DTC in patients with NTD. There was also a direct relationship between malignant nodule size and TSH levels. By contrast, no relationship was found between the size of benign nodules and TSH levels.

Utilidad de la determinación intraoperatoria de parathormona en el tratamiento quirúrgico del hiperparatiroidismo primario por adenoma de paratiroides

Fundamento y objetivo La exploracion quirurgica de las 4 paratiroides es un procedimiento demasiadoagresivo para la mayoria de los casos de hiperparatiroidismo primario (HPTP) cuya causa es un adenoma preoperatoriamente localizado. Recientemente la determinacion intraoperatoria de paratormona (PTH) ha demostrado ser una herramienta util en el tratamiento de estos pacientes y permitiria el uso de tecnicas quirurgicas minimamente invasivas, con una menor morbilidad. El objetivo de nuestro trabajo es la valoracion de la utilidad de la determinacion intraoperatoria de la PTH en el abordaje quirurgico del HPTP. Pacientes y metodo Se incluyo a 27 pacientes consecutivos, diagnosticados de HPTP causado por un adenoma de paratiroides. El estudio de localizacion consto de ecografia cervical y gammagrafia con Tc-MIBI. Durante la intervencion, se determino la PTH en el momento de la induccion anestesica y 5 y 10 min despues de la exeresis del adenoma. Un descenso de la PTH mayor del 50% a los 10 min se considero criterio de curacion. La PTH se determino por un metodo quimioluminimetrico (Advantage, Nichols). El tiempo necesario para la obtencion del resultado fue de 20 min. Resultados En los 27 casos no existio hipercalcemia 24 h despues de la intervencion, por loque se consideraron curados. La PTH disminuyo mas de un 50% en todos ellos. En un caso, la PTH se mantuvo elevada despues de extirpar una lesion que se habia localizado preoperatoriamente. El dictamen patologico fue que se trataba de un tejido paratiroideo normal. La continuacion de la exploracion quirurgica permitio encontrar un adenoma en el lado contralateral. La PTH posterior fue menor del 50%. Por tanto, de las 28 determinaciones, la PTH fue predictiva del resultado quirurgico en la totalidad de los casos. Conclusiones La determinacion intraoperatoria de PTH es util en el abordaje quirurgico del HPTP y permite el uso de tecnicas quirurgicas minimamente invasivas.

Graves’ Disease TSHR-Stimulating Antibodies (TSAbs) Induce the Activation of Immature Thymocytes: A Clue to the Riddle of TSAbs Generation?

Graves’ disease (GD) is an autoimmune thyroid disease defined by the production of stimulating autoantibodies to the thyroid-stimulating hormone receptor (TSHR) (TSAbs) that induce a sustained state of hyperthyroidism in patients. We previously demonstrated that TSHR, the target of this autoimmune response, is also a key susceptibility gene for GD, probably acting through thymic-dependent central tolerance. We also showed that TSHR is, unexpectedly, expressed in thymocytes. In this report, we confirm the expression of TSHR in thymocytes by protein immunoblotting and quantitative PCR, and show that expression is confined to maturing thymocytes. Using functional assays, we show that thymic TSHR is functional and that TSAbs can stimulate thymocytes through this receptor. This new activity of TSAbs on thymocytes may: 1) explain GD-associated thymic enlargement (hyperplasia), and 2) suggest the provocative hypothesis that the continuous stimulation of thymocytes by TSAbs could lead to a vicious cycle of iterative improvement of the affinity and stimulating capability of initially low-affinity antibacterial (e.g., Yersinia) Abs cross-reactive with TSHR, eventually leading to TSAbs. This may help to fill one of the gaps in our present understanding of unusual characteristics of TSAbs.