Liraglutide improves cardiac function in patients with type 2 diabetes and chronic heart failure.

PURPOSE: To compare the effect of liraglutide, sitagliptin and insulin glargine added to standard therapy on left ventricular function in post-ischemic type-2 diabetes mellitus patients. METHODS: We evaluated 32 type-2 diabetes mellitus Caucasians with history of post-ischemic chronic heart failure NYHA class II/III and/or left ventricular ejection fraction ≤45 %. Participants underwent laboratory determinations, electrocardiogram, echocardiogram, Minnesota Living with Heart Failure questionnaire and 6 min walking test at baseline and following 52 weeks treatment. Patients were treated with standard therapy for chronic heart failure and were randomized to receive liraglutide, sitagliptin and glargine in addition to metformin and/or sulfonylurea. RESULTS: Liraglutide treatment induced an improvement in left ventricular ejection fraction from 41.5 ± 2.2 to 46.3 ± 3 %; P = 0.001). On the contrary, treatment with sitagliptin and glargine induced no changes in left ventricular ejection fraction (41.8 ± 2.6 vs. 42.5 ± 2.5 % and 42 ± 1.5 vs. 42 ± 1.6 %, respectively; P = NS). Indexed end-systolic LV volume was reduced only in liraglutide-treated patients (51 ± 9 vs. 43 ± 8 ml/m2; P < 0.05). Liraglutide treatment induced also a significant increase in the anterograde stroke volume (39 ± 9 vs. 49 ± 11 ml; P < 0.05), whereas no differences were observed in the other two groups. Cardiac output and cardiac index showed a significant increase only in liraglutide-treated patients (4.4 ± 0.5 vs. 5.0 ± 0.6 L/min; P < 0.05 and 1.23 ± 0.26 vs. 1.62 ± 0.29 L/m2; P = 0.005, respectively). Liraglutide treatment was also associated with an improvement of functional capacity and an improvement of quality of life. CONCLUSIONS: These data provide evidence that treatment with liraglutide is associated with improvement of cardiac function and functional capacity in failing post-ischemic type-2 diabetes mellitus patients.

Stage of change and motivation to healthy diet and habitual physical activity in type 2 diabetes.

Lifestyle changes to healthy diet (HD) and habitual physical activity (HPA) are recommended in type 2 diabetes mellitus (T2DM). Yet, for most people with diabetes, it may be difficult to start changing. We investigated the stage of change toward healthier lifestyles according to Prochaska's model, and the associated psychological factors in T2DM patients, as a prerequisite to improve strategies to implement behavior changes in the population. A total of 1,353 consecutive outpatients with T2DM attending 14 tertiary centers for diabetes treatment completed the validated EMME-3 questionnaire, consisting of two parallel sets of instruments to define the stage of change for HD and HPA, respectively. Logistic regression was used to determine the factors associated with stages that may hinder behavioral changes. A stage of change favoring progress to healthier behaviors was more common in the area of HD than in HPA, with higher scores in action and maintenance. Differences were observed in relation to gender, age and duration of disease. After adjustment for confounders, resistance to change toward HD was associated with higher body mass index (BMI) (odds ratio (OR) 1.05; 95 % confidence interval (CI) 1.02-1.08). Resistance to improve HPA also increased with BMI (OR 1.06; 95 % CI 1.03-1.10) and decreased with education level (OR 0.74; 95 % CI 0.64-0.92). Changing lifestyle, particularly in the area of HPA, is not perceived as an essential part of treatment by many subjects with T2DM. This evidence must be considered when planning behavioral programs, and specific interventions are needed to promote adherence to HPA.

Insulin sensitivity, and ß-cell function in relation to hemoglobin A1C.

BACKGROUND AND AIMS: The A1C diagnostic criterion for identifying individuals at increased risk for diabetes, introduced by the American Diabetes Association in 2010, was not defined on the basis of the principal pathophysiological abnormalities responsible for the development and progression of type 2 diabetes; we therefore wished to gain a deeper insight into the metabolic abnormalities characterizing the group of at risk individuals with an A1C value of 5.7-6.4%. METHODS AND RESULTS: As many as 338 non-diabetic offspring of type 2 diabetic patients were consecutively recruited. Insulin secretion was assessed using both indexes derived from oral glucose tolerance test (OGTT), and intravenous glucose tolerance test (IVGTT). Insulin sensitivity was measured by hyperinsulinemic euglycemic clamp. As compared with subjects with A1C <5.7%, individuals with A1C of 5.7-6.4% exhibited lower insulin sensitivity after adjusting for age, gender and body mass index (BMI). Insulin secretion estimated from the OGTT, did not differ between the two groups. By contrast, as compared with subjects with A1C <5.7%, the acute insulin response (AIR) during an IVGTT and both IVGTT-derived and OGTT-derived disposition indexes were reduced in individuals with A1C of 5.7-6.4% after adjusting for age, gender and BMI. As A1C increased to ≥ 5.7%, a sharp decrease in insulin sensitivity and ß-cell function, measured as disposition index, was observed. CONCLUSIONS: Caucasian individuals with A1C ≥ 5.7% exhibit both core pathophysiological defects of type 2 diabetes i.e. insulin resistance and ß-cell dysfunction.

Increased carotid intima-media thickness in the physiologic range is associated with impaired postprandial glucose metabolism, insulin resistance and beta cell dysfunction.

BACKGROUND: Carotid Intima-Media Thickness (C-IMT) is a reliable predictor of cardiovascular events. We examined if increased C-IMT was associated with defects in glucose metabolism in non-diabetic subjects independently of age. METHODS: In 366 Caucasian non-diabetic subjects of the CARAMERIS study, we measured glucose response during a 75 g-Oral Glucose Tolerance Test (OGTT), insulin sensitivity index (ISI, by Matsuda Index), Liver Insulin Resistance Index (Liver-IR), insulin secretion by ΔAUC Ins0-120/Glu0-120 (ΔI/ΔG) and beta cell function (Disposition Index, DI). RESULTS: Subjects were divided in two groups according to the median age (AGE1 ≤ 45 y; AGE2 > 45 y). Only 5 subjects in AGE1 and 32 in AGE2 had C-IMT > 0.9 mm. Compared to AGE1, AGE2 had a worse cardio-metabolic profile, increased cholesterol, glucose and insulin concentrations, blood pressure and C-IMT. Both ΔI/ΔG ratio and DI were significantly reduced in AGE2. By considering tertiles of C-IMT in each AGE group (G1-G3, where G3 comprised the highest C-IMT), we found that G3 showed increased OGTT glucose profiles and Liver IR, decreased ISI and DI, compared to G1 in each AGE group. CONCLUSIONS: Increased C-IMT, but within normal ranges, is associated independently of age with altered postprandial glucose profile, increased peripheral and hepatic insulin resistance, decreased b-cell function. C-IMT measurement should become a routine analysis even in younger subjects to predict the risk of cardio-metabolic disease.

[Hyperglycaemia and Acute Coronary Syndrome]. / L'iperglicemia nelle Sindromi Coronariche Acute.

Hyperglycaemia in patients with Acute Coronary Syndrome (ACS)is common, and is an independent predictor of mortality and morbidity in patients both with and without diabetes mellitus. Hyperglycaemia may be a marker of pre-existing diabetes or glucose intolerance or may also represent a transient stress response mediated through the autonomic nervous system with release of adrenal corticosteroids and catecholamines. Several evidences suggest that an intensive control of hyperglycaemia results in a significant improvement of the adverse outcomes in the short and long term. In fact, an intensive metabolic treatment can counteract the negative effects of hyperglycaemia. However, the main difficulty to intensive glucose control in patients with ACS remains hypoglycaemia that is associated with an increased risk of mortality and myocardial re-infarction. No definitive data are available about the beneficial effects of insulin intensive treatment. Therefore, randomized multicenter clinical trials will be needed to definitively establish whether intensive glucose control will reduce the associated increased mortality rate and higher rates of complications in hospitalized ACS patients with hyperglycaemia.

Comparison of A1C, fasting and 2-h post-load plasma glucose criteria to diagnose diabetes in Italian Caucasians.

BACKGROUND AND AIMS: The American Diabetes Association (ADA) has revised criteria for diagnosis of type 2 diabetes recommending an A1C cut point of ≥6.5% in addition to criteria based on glucose levels. We compared A1C, fasting plasma glucose (FPG) or 2-h post-challenge glucose (2-hPG) criteria for the diagnosis of diabetes in a cohort of Italian Caucasians. METHODS AND RESULTS: A total of 1019 individuals without known diabetes completed an oral glucose tolerance test (OGTT) and had A1C measured. Moderate agreement existed for A1C and FPG criteria for diagnosis of type 2 diabetes (κ coefficient = 0.522), with 85.5% of individuals classified as not having diabetes by both A1C and FPG criteria, and 5.8% classified as having diabetes by both A1C and FPG criteria. Discordant classifications occurred for 5.5% of individuals who had an A1C ≥ 6.5% and FPG <126 mg dl(-1), and for 3.2% who had an A1C <6.5% and FPG ≥126 mg dl(-1). Modest agreement existed for A1C and 2-hPG criteria for diagnosis of type 2 diabetes (κ coefficient = 0.427), with 81.8% of individuals classified as not having diabetes by both A1C and 2-hPG criteria, and 6.0% classified as having diabetes by both A1C and 2-hPG criteria. The area under the receiver operating characteristic curve of A1C for identifying subjects with diabetes according to FPG or 2-hPG criteria was 0.856 and 0.794, respectively. Modest agreement existed for A1C and FPG and/or 2-hPG criteria for diagnosis of type 2 diabetes (κ coefficient = 0.446). CONCLUSIONS: A1C ≥ 6.5% demonstrates a moderate agreement with fasting glucose and 2-hPG for diagnosing diabetes among adult Italian Caucasians subjects.

One-hour post-load plasma glucose levels are associated with elevated liver enzymes.

BACKGROUND AND AIMS: Glucose-tolerant subjects who have 1-h post-load glucose levels ≥155 mg dl(-1) (normal glucose tolerance (NGT)-1h-high) are at an increased risk of developing type 2 diabetes. Prospectively conducted studies indicated that high levels of liver enzymes are predictors of a tendency to develop type 2 diabetes; however, it is unknown whether the NGT-1h-high subjects are at increased risk for secreting higher levels of liver biomarkers. METHODS AND RESULTS: In this study, oral glucose tolerance tests (OGTTs) were performed in a cohort of 1000 non-diabetic Caucasians and levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and gamma-glutamyltransferase (GGT) were measured in these subjects. The NGT-1h-high subjects had increased levels of ALT and GGT, but not AST, as compared with the NGT-1h-low. Following adjustment for age and gender, the ALT, AST and GGT levels were all found to be significantly correlated with body mass index (BMI), waist circumference, blood pressure, triglycerides as well as fasting and post-challenge glucose and insulin levels. In a logistic regression analysis adjusted for age and gender, NGT-1h-high subjects were found to be at increased risk of having ALT levels in the highest quartile as compared with NGT-1h-low subjects (odds ratio (OR) = 1.71; 95% confidence interval (CI): 1.16-2.52). In addition, NGT-1 h-high subjects exhibited an increased risk for having GGT levels in the highest quartile (OR = 1.50; 95%CI: 1.02-2.17). These associations remained significant after adjustment for BMI, blood pressure and lipids, but were not significant following further adjustment for an insulin sensitivity index. NGT-1h-high subjects were at increased risk of having AST levels in the highest quartile as compared with NGT-1h-low subjects (OR = 1.51; 95%CI: 1.04-2.22). This association ceased to be significant following adjustment for BMI, blood pressure and lipids. CONCLUSIONS: These data suggest that a 1hPG ≥ 155 mg dl(-1) cut-off may facilitate the identification of NGT individuals at risk of developing liver abnormalities.

IGF-1 levels link estimated glomerular filtration rate to insulin resistance in obesity: a study in obese, but metabolically healthy, subjects and obese, insulin-resistant subjects.

BACKGROUND AND AIMS: Metabolically healthy but obese (MHO) subjects have a favourable cardio-metabolic risk profile, but whether they are also at lower risk for kidney dysfunction is still questionable. METHODS AND RESULTS: A total of 106 MHO, 122 normal-weight and 212 insulin-resistant obese (IRO) subjects were stratified on the basis of their insulin sensitivity and body mass index (BMI). The CKD-EPI equation was used to estimate glomerular filtration rate (eGFR) and ISI index was used to estimate insulin sensitivity. eGFR was significantly lower in IRO as compared to MHO subjects after adjusting for age, gender and BMI (P = 0.008). In a logistic regression model adjusted for age, gender and BMI, IRO subjects showed an increased risk of having eGFR in the lowest quartile (odds ratio (OR) 1.91, 95% confidence interval (CI) 1.01-3.58; P = 0.04) as compared with MHO subjects. This association was maintained when waist, lean body mass, blood pressure, HDL cholesterol, triglyceride, fasting glucose and insulin levels were additionally included into the model (OR 2.49, 95%CI 1.17-5.27; P = 0.01), but its independence was not retained with further inclusion of insulin-like growth factor-1 (IGF-1) levels (OR 2.16, 95%CI 0.93-5.04; P = 0.07) No differences in eGFR were observed between non-obese and MHO individuals. CONCLUSIONS: These results indicate that heterogeneity in obese phenotypes may account for conflicting evidence regarding the significance of obesity as a risk factor for chronic kidney disease. Our findings suggest that obesity is associated with lower kidney function only when insulin sensitivity is reduced, and that plasma IGF-1 is likely to be an important mechanism linking the IRO phenotype with reduced eGFR.

[The HPA axis in the pathogenesis of chronic fatigue syndrome]. / L'asse ipotalamo-ipofi si-surrene nella patogenesi della Sindrome da Stanchezza Cronica.

Chronic fatigue syndrome (CFS) is a clinical syndrome characterized by profound disabling chronic fatigue associated with a wide array of other physical symptoms. Its etiology is currently unknown. Among the various hypotheses, considerable interest has been placed in the hypothalamus-pituitary-adrenal axis as a possible target of the pathogenesis of CFS. This article reviews the available scientific evidence about a role of hypothalamic-pituitary-adrenal axis in the pathogenesis of chronic fatigue syndrome.

Sudden progression from impaired glucose tolerance to type 2 diabetes after discontinuation of administration of anti-tumor necrosis factor-alpha antibody infliximab.

We present the case of a 45-year-old man with psoriasis and psoriatic arthritis and concomitant impaired fasting glucose (IFG) and impaired glucose tolerance (IGT). In this patient, refractory to DMARD's, infliximab was started to control the arthritis. After achieving clinical remission of the disease, infliximab was discontinued and a 75 g- oral glucose tolerance test (OGTT) was performed. After the test, we observed a conversion from IFG/IGT glucose tolerance status to type 2 diabetes. No diet, lifestyle or therapy modifications were made during the observation period. Autoimmune diabetes was ruled out by serum antibodies determination and body weight remained constant, sustaining a protective role in infliximab in the worsening of glucose tolerance.

Elevated one-hour post-load plasma glucose levels identifies subjects with normal glucose tolerance but early carotid atherosclerosis.

OBJECTIVE: To examine whether individuals with normal glucose tolerance (NGT), whose 1-h post-load plasma glucose is >or=155 mg/dl, or with impaired glucose tolerance (IGT) have an increased carotid intima-media thickness (IMT), as compared with NGT individuals with 1-h post-load plasma <155 mg/dl. METHODS: Atherosclerosis risk factors, oral glucose tolerance test (OGTT), and ultrasound manual measurement of IMT were analyzed in 400 non-diabetic Caucasians. RESULTS: As compared with individuals with a 1-h post-load plasma glucose <155 mg/dl, NGT individuals with a 1-h post-load plasma glucose >or=155 mg/dl exhibited higher hsCRP (2.0+/-1.5 vs. 1.5+/-1.0, P=0.008), and IMT (0.82+/-0.20 vs. 0.71+/-0.16; P=0.006), and lower insulin sensitivity (71+/-39 vs. 105+/-57; P<0.0001), and IGF-1 levels (214+/-88 vs. 176+/-49; P<0.03). No significant differences were observed in metabolic and cardiovascular risk factors between IGT and NGT subjects with a 1-h post-load glucose >or=155 mg/dl. Of the three glycemic parameters, 1-h and 2-h post-load glucose, but not fasting glucose, were significantly correlated with IMT. In a stepwise multivariate regression analysis in a model including age, gender, and a variety of atherosclerosis risk factors, the three variables that remained significantly associated with IMT were age (P<0.0001), BMI (P<0.0001), and 1-h post-load glucose (P=0.02) accounting for 20.2% of its variation. CONCLUSIONS: NGT subjects with a 1-h post-load glucose >or=155 mg/dl have an atherogenic profile similar to IGT individuals. These data suggest that a cutoff point of 155 mg/dl for the 1-h post-load glucose during OGTT may be helpful in the identification of NGT subjects at increased risk for cardiovascular disease.

Lack of TSH inhibition by exogenous L-T4 in a patient with follicular thyroid carcinoma.

We report a case of a male patient thyroidectomized for follicular thyroid carcinoma and presenting extremely elevated serum thyrotropin levels under L-T4 suppressive therapy. Administration of L-T3 in increasing amounts resulted in a significant decrease of serum TSH levels. The nature of the possible molecular defects underlying this unusual condition and pitfalls arising from the failure of L-T4 therapy to inhibit TSH secretion in a patient in post-surgical follow-up for follicular carcinoma are discussed.

Proteomic analysis of human thyroid cell lines reveals reduced nuclear localization of Mn-SOD in poorly differentiated thyroid cancer cells.

Differential protein arrays between nuclear extracts of human thyroid cell lines obtained from tumors with different degree of differentiation were exploited to define molecular alterations occurring during thyroid tumor progression. Nuclear extracts from the well differentiated TPC-1 (from papillary carcinoma) and the poorly differentiated ARO (from anaplastic carcinoma) cells showed an overall similar pattern of protein expression as revealed by two-dimensional gel electrophoresis analysis. However, manganese-superoxide dismutase (Mn-SOD) was clearly identified by mass spectrometry procedures as significantly less expressed in ARO compared to TPC-1 cells. A reduced expression of Mn-SOD in the nuclear compartment was confirmed by Western blot and immunofluorescence analysis. A similar expression pattern of nuclear Mn-SOD was detected by immunohistochemistry in human thyroid tumors, with the lowest or absent detection in anaplastic carcinomas. Moreover, the levels of nuclear Mn-SOD in tumor cells were lower than in the normal thyrocytes. These data indicate that an altered nuclear expression of Mn-SOD parallels, together with changes in other elements of the antioxidant protective system, the loss of differentiation occurring during the progression of thyroid tumors.

Evaluation of a DHPLC-based assay for rapid detection of RET germline mutations in Italian patients with medullary thyroid carcinoma.

Causative gain-of-function mutations of the RET tyrosine-kinase receptor gene have been reported in more than 95% of inherited cases of medullary thyroid carcinoma (MTC; OMIM# 155240). Most RET activating mutations are clustered in mutational "hot spots" in exons 10, 11, 13, 14, 15 and 16 and are usually detected by single-strand conformation polymorphism (SSCP) followed by direct sequencing. To improve sensitivity, time and costs of mutational screening we have developed a denaturing high performance chromatography (DHPLC) protocol, based on the detection of heteroduplex molecules by ion-pair reverse-phase liquid chromatography under partially denaturing conditions. The mutational screening of RET exons 10, 11, 13-16 was performed in a total of 111 subjects, including 45 MTC patients and 49 relatives with known RET mutations and 17 individuals, being at risk of hereditary MTC and carrying unknown RET alleles. Heteroduplex peaks with a distinct and reproducible DHPLC elution profile allowed the detection of both rare and common RET mutations. Overall, the DHPLC-based methodology showed a high level of sensitivity and accuracy, nearing 100%. Furthermore, our protocol showed the ability to identify: 1) all the mutated codons of RET located in the "hot spots" domain; 2) the different point mutations occurring in the same codon of RET gene; 3) less frequent or rare mutations; 4) polymorphisms. As such, it can be proposed as a relatively simple and highly accurate method for a rapid genetic testing for members of MTC families.

Slow growth of benign thyroid nodules after menopause: no need for long-term thyroxine suppressive therapy in post-menopausal women.

Benign thyroid nodules represent a very common disorder, the management of which is still controversial. The aim of the present work was to evaluate by ultrasound examination the volume changes of thyroid nodules in post-menopausal women presenting single palpable nodular goiter of recent onset (less than 6 months from diagnosis). Forty-three patients received L-T4-treatment, 38 represented the no-treatment group. Long-term follow up (3 and 5 yr) did not show any significant change in the mean volume nodule in these patients. In the no-treatment group, the mean nodule volumes were stable over time from baseline to 5 yr. No significant difference was observed at any follow-up evaluation between thyroid hormone treated and untreated patients. After 1 yr of treatment, a significant decrease (p = 0.0275) in mean nodule volume occurred only for nodules with a baseline volume lower than 1.5 ml. The frequency of clinically relevant nodule size variation showed a more frequent decrease (13.9%) at 1 yr in the L-T4 group, as compared to the no-treatment group (2.6%), while the proportion of increased volume at 1 yr was higher in the untreated than in the L-T4 group (5.3% vs 2.3%). This inverse relationship between the 2 groups was not statistically significant (p = 0.076). In conclusion, an arrest in the growth of benign thyroid nodules occurs in the majority of women after menopause. Only a very limited number of these patients may benefit from thyroid hormone suppressive treatment.

Novel somatic MEN1 gene alterations in sporadic primary hyperparathyroidism and correlation with clinical characteristics.

Primary hyperparathyroidism (pHPT) is a common endocrine disease that in more than 95% of cases is sporadic and only in some cases is caused by inherited disorders, isolated or as part of multiple endocrine neoplasia (MEN1 and 2). Somatic mutations of MEN1 gene have also been described in sporadic parathyroid tumors. In our study, we examined the presence of alterations in MEN1 gene in a series of 39 patients who had undergone surgery for sporadic pHPT (35 with parathyroid adenoma or hyperplasia, 4 with a carcinoma). A genotype-phenotype correlation was also analysed. After DNA extraction from paraffin-embedded tissues, we amplified by PCR and sequenced the exons 2-10 of the MEN1 gene. Somatic MEN1 mutations were detected in 6 of the 35 patients with a benign parathyroid lesion examined (17.1%), whereas no alterations were found in the carcinomas. Four novel MEN1 gene mutations were identified as follows: one frameshift mutation (222insT, exon 2), one frameshift deletion (912delTA, exon 5), one in-frame deletion (835del18, exon 4) and one missense mutation (P291A, exon 6). In addition, one missense mutation (L89R, exon 2) and one nonsense mutation (Q536X, exon 10) were previously reported. Moreover, two polymorphisms were also found: one allele carried a R171Q polymorphism (1/39 tumors), while a D418D polymorphism (GAC/GAT) was found in 15 and 8 tumors in hetero (CT) and homozygosity (TT), respectively. In no case (mutations and/or polymorphisms) did we find a genotype-phenotype correlation. In conclusion, our data demonstrate the presence of somatic alterations of the MEN1 tumor suppressor gene in about one fifth of benign sporadic parathyroid tumors. The absence of a genotype-phenotype correlation, however, suggests the involvement of other genetic/epigenetic factors for the full expression of the disease.

Expression of adenylyl cyclase types III and VI in human hyperfunctioning thyroid nodules.

Hyperfunctioning thyroid nodules are characterized by the presence of spontaneous somatic mutations responsible for constitutive activation of the cAMP pathway. However, alterations affecting other elements of the cAMP signaling system may counteract the effects of the mutations. In this study, the expression of the adenylyl cyclase (AC) types III and VI was investigated by Western blot in 18 hyperfunctioning thyroid nodules; in 12 samples, we also assessed the presence of TSH receptor (TSHR) or gsp mutations and levels of AC VI and III mRNA. We found that the expression of nodular AC VI (but not AC III) was significantly lower (85.1% of normal, P=0.014) than the expression of both adenylyl cycles types of perinodular tissue from the same patients. Slightly, but not significant differences were detected in nodules with or without mutations and AC protein levels generally showed correlation with the levels of the transcripts detected by RT-PCR. In addition, AC III and AC VI expression levels within a given nodule were characterized by a significant positive correlation. These findings indicate that a diminished expression of AC type VI may be part of the mechanisms occurring in the hyperfunctioning nodules, independently of the presence of TSHR or gsp mutations, which influence the resulting phenotype.

Thyrotropin receptor mutations and thyroid hyperfunctioning adenomas ten years after their first discovery: unresolved questions.

Ten years after the first description of activating mutations in the thyroid stimulating hormone receptor (TSHR) gene in sporadic autonomous hyperfunctioning thyroid adenomas, there is general agreement in assigning a major pathogenic role of this genetic abnormality, acting via the constitutive activation of the cAMP pathway, in both the growth and functional characteristic of these tumours. From the beginning, however, the pathophysiological and clinical relevance of somatic TSHR mutations has been debated and some arguments still exist against a fully causative role of these mutations and the practical value of detecting these mutations for the diagnosis, treatment and prognosis of thyroid hot nodules. Some major issues will be examined herein, including (a) the frequency of TSHR alterations in various reports showing that the genetic abnormality underlying the pathogenesis of a substantial subset of thyroid tumours has yet to be identified; (b) the limitations of the present experimental models, which suggest greater caution in the interpretation of in vitro results; (c) the still unresolved question of absence of genotype-phenotype correlation. Clarification of these issues may hopefully provide new and useful tools for improving the clinical management of this disease.

APE/Ref-1 is increased in nuclear fractions of human thyroid hyperfunctioning nodules.

Apurinic/apyrimidinic endonuclease APE/Ref-1 is a multifunctional protein provided with DNA repair, transcription-factor regulation and anti-apoptotic activities. We have previously reported that, in thyroid cells, TSH regulates both the synthesis and nuclear translocation of APE/Ref-1. We have also shown that nuclear levels of this protein are reduced both in thyroid carcinoma tissues and cell lines. In the present study, APE/Ref-1 expression and cellular localization were analysed by Western blot in hyperfunctioning thyroid nodules from patients with toxic adenoma and/or toxic multinodular goiter. The total content of APE/Ref-1 protein was increased in the majority of the hyperfunctioning tissues with respect to normal adjacent tissue. There was also an increase in the nuclear levels of APE/Ref-1, suggesting enhanced cytoplasm-to-nucleus translocation of the protein in addition to its increased rate of synthesis. These results demonstrate that the phenomenon of nuclear translocation of APE/Ref-1 hypothesized on the basis of cell culture experiments does actually occur in vivo. Together with previous observations in thyroid carcinomas and tumoral cell lines, our findings suggest a two-stage model of APE/Ref-1 behaviour during malignant thyrocyte transformation: an early stage characterized by simple hyperplasia and upregulation of APE/Ref-1 in the nuclear compartment of the cell and a later stage in which nuclear levels of the protein drop to below-normal levels as the cell becomes progressively undifferentiated.

Similarities and differences in the phenotype of members of an Italian family with hereditary non-autoimmune hyperthyroidism associated with an activating TSH receptor germline mutation.

Constitutively activating germline mutations of the TSH receptor (TSH-R) are considered the cause of hereditary non-autoimmune hyperthyroidism. In this study, 10 members (8 affected and 2 unaffected) of an Italian family with hereditary non-autoimmune hyperthyroidism were investigated for the presence of mutations in the TSH-R gene. The clinical features of the disease were also analyzed. PCR-amplified fragments of the TSH-R gene were obtained from genomic DNA extracted from peripheral blood leukocytes of each family member and analyzed by direct nucleotide sequencing and restriction analysis. An identical germline TSH-R mutation was detected in all the patients with hyperthyroidism but in none of the unaffected family members. The mutation was heterozygotic and determined the substitution of valine for methionine (codon 463; ATG-->GTG) in the second transmembrane domain of the TSH-R. When expressed in chinese hamster ovary (CHO) cells, the Val463 mutant TSH-R induced constitutive activation of the TSH receptor. Analysis of the clinical features of our family and those of other families with hereditary non-autoimmune hyperthyroidism, including one with the same Val463 mutation, revealed wide variability in the phenotypical expression of the disease. Our findings indicate that an activating germline mutation in the TSH-R gene plays a key role in hereditary non-autoimmune hyperthyroidism although the onset of clinical manifestations and the evolution of the disease seem to depend heavily on other factors, thus far unidentified. The absence of a clear correlation between mutant genotypes and phenotypic expression of the disease currently limits the prognostic value of genetic testing in families with hereditary non-autoimmune hyperthyroidism.