Romana T. Netea-Maier

Discovery of common variants associated with low TSH levels and thyroid cancer risk

To search for sequence variants conferring risk of nonmedullary thyroid cancer, we focused our analysis on 22 SNPs with a P < 5 × 10−8 in a genome-wide association study on levels of thyroid stimulating hormone (TSH) in 27,758 Icelanders. Of those, rs965513 has previously been shown to associate with thyroid cancer. The remaining 21 SNPs were genotyped in 561 Icelandic individuals with thyroid cancer (cases) and up to 40,013 controls. Variants suggestively associated with thyroid cancer (P < 0.05) were genotyped in an additional 595 non-Icelandic cases and 2,604 controls. After combining the results, three variants were shown to associate with thyroid cancer: rs966423 on 2q35 (OR = 1.34; Pcombined = 1.3 × 10−9), rs2439302 on 8p12 (OR = 1.36; Pcombined = 2.0 × 10−9) and rs116909374 on 14q13.3 (OR = 2.09; Pcombined = 4.6 × 10−11), a region previously reported to contain an uncorrelated variant conferring risk of thyroid cancer. A strong association (P = 9.1 × 10−91) was observed between rs2439302 on 8p12 and expression of NRG1, which encodes the signaling protein neuregulin 1, in blood.

Quantitative and qualitative differences in protein expression between papillary thyroid carcinoma and normal thyroid tissue

In order to better understand basic mechanisms of tumor development and identify potential new biomarkers, we have performed difference gel electrophoresis (DIGE) and peptide mass fingerprinting on pooled protein extracts from patients with papillary thyroid carcinoma (PTC) compared with matched normal thyroid tissue. Image analysis of DIGE gels comparing PTC and matched normal thyroid tissue protein indicated that 25% of the protein spots were differentially expressed at a 2.5‐fold cutoff and 35% at two‐fold. Comparison between two different pools of protein from normal thyroid tissues revealed differential protein expression of only 4% at 2.5‐fold and 6% at two‐fold cutoff. One hundred ninety‐two protein spots were identified by MALDI‐TOFMS, representing 90 distinct proteins. Excluding albumin, globins and thyroglobulin, imaging software determined 31 proteins to be differentially expressed at the two‐fold (or greater) level. Individual gel comparisons (PTC vs. matched normal) from five patients established that 15/31 (48%) of these proteins exhibited statistically significant differential expression. Previously identified molecular markers in this group of proteins include cathepsin B, cytokeratin 19, and galectin‐3. Novel differentially expressed proteins include S100A6, moesin, HSP70 (BiP), peroxiredoxin 2, protein phosphatase 2, selenium binding protein 1, vitamin D binding protein, and proteins involved in mitochondrial function. The use of two‐dimensional gel electrophoresis (2DGE) revealed a significantly altered protein mass and/or pI in 10%–15% of proteins, suggesting alternatively spliced forms and other posttranslational modification of proteins revealed by this approach. We confirmed S100A6 as a potentially useful biomarker using immunohistochemical analysis (85% sensitivity and 69% specificity for distinguishing benign from malignant thyroid neoplasms). In summary, proteomic analysis of PTC using DIGE and mass spectrometry has confirmed several known biomarkers, uncovered novel potential biomarkers, and provided insights into global pathophysiologic changes in PTC. Many of the differences observed would not have been detected by genomic or other proteomic approaches.

A Meta-Analysis of Thyroid-Related Traits Reveals Novel Loci and Gender-Specific Differences in the Regulation of Thyroid Function

Thyroid hormone is essential for normal metabolism and development, and overt abnormalities in thyroid function lead to common endocrine disorders affecting approximately 10% of individuals over their life span. In addition, even mild alterations in thyroid function are associated with weight changes, atrial fibrillation, osteoporosis, and psychiatric disorders. To identify novel variants underlying thyroid function, we performed a large meta-analysis of genome-wide association studies for serum levels of the highly heritable thyroid function markers TSH and FT4, in up to 26,420 and 17,520 euthyroid subjects, respectively. Here we report 26 independent associations, including several novel loci for TSH (PDE10A, VEGFA, IGFBP5, NFIA, SOX9, PRDM11, FGF7, INSR, ABO, MIR1179, NRG1, MBIP, ITPK1, SASH1, GLIS3) and FT4 (LHX3, FOXE1, AADAT, NETO1/FBXO15, LPCAT2/CAPNS2). Notably, only limited overlap was detected between TSH and FT4 associated signals, in spite of the feedback regulation of their circulating levels by the hypothalamic-pituitary-thyroid axis. Five of the reported loci (PDE8B, PDE10A, MAF/LOC440389, NETO1/FBXO15, and LPCAT2/CAPNS2) show strong gender-specific differences, which offer clues for the known sexual dimorphism in thyroid function and related pathologies. Importantly, the TSH-associated loci contribute not only to variation within the normal range, but also to TSH values outside the reference range, suggesting that they may be involved in thyroid dysfunction. Overall, our findings explain, respectively, 5.64% and 2.30% of total TSH and FT4 trait variance, and they improve the current knowledge of the regulation of hypothalamic-pituitary-thyroid axis function and the consequences of genetic variation for hypo- or hyperthyroidism.

Modulation of inflammation by autophagy: Consequences for human disease

ABSTRACT Autophagy and inflammation are 2 fundamental biological processes involved in both physiological and pathological conditions. Through its crucial role in maintaining cellular homeostasis, autophagy is involved in modulation of cell metabolism, cell survival, and host defense. Defective autophagy is associated with pathological conditions such as cancer, autoimmune disease, neurodegenerative disease, and senescence. Inflammation represents a crucial line of defense against microorganisms and other pathogens, and there is increasing evidence that autophagy has important effects on the induction and modulation of the inflammatory reaction; understanding the balance between these 2 processes may point to important possibilities for therapeutic targeting. This review focuses on the crosstalk between autophagy and inflammation as an emerging field with major implications for understanding the host defense on the one hand, and for the pathogenesis and treatment of immune-mediated diseases on the other hand.

The hypercoagulable state in Cushing's disease is associated with increased levels of procoagulant factors and impaired fibrinolysis, but is not reversible after short-term biochemical remission induced by medical therapy.

CONTEXT Cushings disease (CD) is accompanied by an increased risk of venous thromboembolism. Surgery is the primary treatment of CD. OBJECTIVE The aim of the study was to compare hemostatic parameters between patients with CD and controls and to evaluate the effect of medical treatment of CD on hemostasis. DESIGN AND SETTING During 80 d, stepwise medical treatment was applied with the somatostatin analog pasireotide, the dopamine agonist cabergoline, and ketoconazole, which suppresses adrenocortical steroidogenesis, at four university medical centers in The Netherlands. PATIENTS Seventeen patients with de novo, residual, or recurrent CD were included. MAIN OUTCOME MEASURES We measured urinary free cortisol and parameters of coagulation and fibrinolysis. RESULTS Patients with CD had significantly higher body mass index (P < 0.001), shortened activated partial thromboplastin time (P < 0.01), and higher levels of fibrinogen, Factor VIII, and protein S activity (P < 0.05) compared to healthy control subjects. In addition, fibrinolytic capacity was impaired in patients with CD as reflected by prolonged clot lysis time (P < 0.001) and higher levels of plasminogen activator inhibitor type 1, thrombin-activatable fibrinolysis inhibitor, and α2-antiplasmin (P < 0.01). There were no statistically significant differences in von Willebrand factor:antigen, antithrombin, and protein C activity. After 80 d, 15 of 17 patients had normalized urinary free cortisol excretion. Despite biochemical remission, only slight decreases in antithrombin (P < 0.01) and thrombin-activatable fibrinolysis inhibitor (P < 0.05) levels were observed. Other parameters of coagulation and fibrinolysis did not change significantly. CONCLUSIONS The hypercoagulable state in patients with CD, which is explained by both increased production of procoagulant factors and impaired fibrinolysis, is not reversible upon short-term biochemical remission after successful medical therapy. This may have implications for the duration of anticoagulant prophylaxis in patients with (cured) CD.

Impaired quality of life in patients in long-term remission of Cushing's syndrome of both adrenal and pituitary origin: a remaining effect of long-standing hypercortisolism?

OBJECTIVE The determinants that cause impaired quality of life (QOL) in patients in long-term remission of Cushings syndrome (CS) are unknown. The aim of this study was to get more insight into the patient and disease characteristics related to impaired QOL in these patients. DESIGN Cross-sectional study. METHODS The QOL of 123 patients in remission of CS (age 52.2 ± 12.0 years, 106 women, duration of remission 13.3 ± 10.4 years, 80% pituitary CS), assessed with seven validated questionnaires, was compared with the QOL of an age- and sex-matched control group (n=105). To investigate the influence of the aetiology of CS on QOL, patients in remission of pituitary and adrenal CS were compared. Furthermore, the influence of hormonal deficiencies, treatment strategy, duration of remission, gender and age on QOL was investigated. RESULTS QOL in the total patient group and each patient subgroup was significantly worse on practically all dimensions of questionnaires compared with the control group (P<0.05), except for patients in remission of pituitary CS without hormonal deficiencies who had an impaired QOL on 50% of the QOL dimensions. Subgroup analysis revealed no difference in QOL between different patient groups, especially no difference between patients in remission of adrenal and pituitary CS. Female gender and a shorter duration of remission had a negative influence on QOL in the patient group. CONCLUSIONS QOL remains impaired in patients in long-term remission of CS regardless of aetiology, presence of hormonal deficiencies and treatment strategies. More research is needed to establish the causes.

Standardized Multidisciplinary Evaluation Yields Significant Previously Undiagnosed Morbidity in Adult Women with Turner Syndrome

CONTEXT Besides short stature and gonadal dysgenesis, Turner syndrome (TS) is associated with various abnormalities. Adults with TS have a reduced life expectancy, mainly related to structural abnormalities of the heart and aorta, and an increased risk of atherosclerosis. OBJECTIVE Our objective was to investigate the yield of an initial standardized multidisciplinary screening in adult TS patients. DESIGN AND SETTING This was an observational study at a multidisciplinary care unit for adult women with TS. PARTICIPANTS Participants were adult women with TS (n = 150). Mean age was 31.0 ± 10.4 yr, with 47% karyotype 45,X. INTERVENTIONS All women were consulted by an endocrinologist, a gynecologist, a cardiologist, an otorhinolaryngologist, and when indicated, a psychologist. The screening included magnetic resonance imaging of the heart and aorta, echocardiography, electrocardiogram, dual-energy x-ray absorptiometry, renal ultrasound, audiogram, and laboratory investigations according to international expert recommendations. MAIN OUTCOME MEASURES New diagnoses and prevalence of TS-associated morbidity were evaluated. RESULTS Thirty percent of patients currently lacked medical follow-up, and 15% lacked estrogen replacement therapy in the recent last years. The following disorders were newly diagnosed: bicuspid aortic valve (n = 13), coarctation of the aorta (n = 9), elongation of the transverse aortic arch (n = 27), dilation of the aorta (n = 34), osteoporosis (n = 8), osteopenia (n = 56), renal abnormalities (n = 7), subclinical hypothyroidism (n = 33), celiac disease (n = 3), glucose intolerance (n = 12), dyslipidemia (n = 52), hypertension (n = 39), and hearing loss warranting a hearing aid (n = 8). Psychological consultation was needed in 23 cases. CONCLUSIONS Standardized multidisciplinary evaluation of adult women with TS as advocated by expert opinion is effective and identifies significant morbidity. Girls with TS benefit from a careful transition to ongoing adult medical care.

The role of [(18)F]-2-fluoro-2deoxy-d-dglucose-positron emission tomography in thyroid nodules iwht indeterminate fine-needle aspiration biopsy: sysematic review and meta-analysis of the literature

Indeterminate results at fine‐needle aspiration biopsy (FNAB) of thyroid nodules pose a clinical dilemma, because only 20% to 30% of patients suffer from malignancy. Previous studies suggested that the false‐negative ratio of [18F]‐2‐fluoro‐2‐deoxy‐D‐glucose–positron emission tomography (FDG‐PET) is very low; therefore, it may help identify patients who would benefit from (hemi)thyroidectomy. A systematic literature search was performed in 5 databases. After assessment, the identified studies were analyzed for heterogeneity, and the extracted data of test characteristics were pooled using a random‐effects model. Threshold effects were examined, and publication bias was assessed. The query resulted in 239 records, of which 6 studies met predefined inclusion criteria. Data from 225 of the 241 described patients could be extracted. There was mild to moderate heterogeneity in study results (inconsistency index [I2] = 0.390‐0.867). The pooled prevalence of malignancy was 26%. Pooled sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were 95% (95% confidence interval [95% CI], 86%‐99%), 48% (95% CI, 40%‐56%), 39% (95% CI, 31%‐47%), 96% (95% CI, 90%‐99%), and 60% (95% CI, 53%‐67%), respectively. Sensitivity increased to 100% for the 164 lesions that measured >15 mm in greatest dimension. There was no evidence of threshold effects or publication bias. A negative FDG‐PET scan in patients who had thyroid nodules >15 mm with indeterminate FNAB results excluded thyroid cancer in a pooled population of 225 patients. Conversely, a positive FDG‐PET result did not identify cancer, because approximately 50% of these patients had benign nodules. The authors concluded that the incorporation of FDG‐PET into the initial workup of such patients before surgery deserves further investigation. Cancer 2011.

Discovery and validation of protein abundance differences between follicular thyroid neoplasms.

Distinguishing between benign follicular thyroid adenoma (FTA) and malignant follicular thyroid carcinoma (FTC) by cytologic features alone is not possible. Molecular markers may aid distinguishing FTA from FTC in patients with indeterminate cytology. The aim of this study is to define protein abundance differences between FTC from FTA through a discovery (proteomics) and validation (immunohistochemistry) approach. Difference gel electrophoresis (DIGE) and peptide mass fingerprinting were performed on protein extracts from five patients with FTC and compared with six patients with FTA. Individual gel comparisons (i.e., each FTC extract versus FTA pool) were also performed for the five FTC patients. Immunohistochemical validation studies were performed on three of the identified proteins. Based on DIGE images, 680 protein spots were matched on individual gels. Of these, 102 spots showed statistically significant differences in abundance between FTC and FTA in the individual gel analyses and were therefore studied further. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to identify 54 of these protein spots. Three candidates involved in protein folding (heat shock protein gp96, protein disulfide isomerase A3, and calreticulin) were studied by immunohistochemistry. Moderate calreticulin immunohistochemical staining was the best single marker with a high negative predictive value (88%); combining all three markers (any marker less than moderate staining) had the best positive predictive value (75%) while still retaining a good negative predictive value (68%). With DIGE, we identified 54 proteins differentially abundant between FTC and FTA. Three of these were validated by immunohistochemistry. These findings provide further insights into the diagnosis, prognosis, and pathophysiology of follicular-derived thyroid neoplasms.

Trends in incidence and mortality of thyroid carcinoma in The Netherlands between 1989 and 2003: Correlation with thyroid fine-needle aspiration cytology and thyroid surgery

A persistent increase in incidence of thyroid carcinoma (TC) has been reported worldwide. The aim of our study was to assess trends in incidence and mortality of TC in The Netherlands between 1989 and 2003 and to examine whether these trends correlate with changes in diagnostic practices such as changes in the number of fine needle aspiration biopsies (FNAB) and/or thyroid surgeries. Population‐based incidence and mortality data were retrieved from the Netherlands Cancer Registry. Data concerning FNAB and thyroid surgeries were obtained through the nationwide network and registry of histo‐ and cytopathology in The Netherlands (PALGA). Overall, the incidence of TC remained unchanged. However, there was a slight increase in incidence of papillary TC of 2.1% per year (p < 0.001) particularly in stage I tumors, possibly, in part, because of a marked increase in use of FNAB. Appropriate iodine intake, reduced radiation exposure and a more conservative diagnostic approach toward asymptomatic thyroid nodules may explain why this increase is less pronounced compared to other countries. Incidence of other subtypes of TC decreased (follicular TC, 1.3% per year, p = 0.02 and anaplastic TC, 7.1% per year, p = 0.006) or remained unchanged (medullary TC). The number of FNABs per year progressively increased from 1,093 in 1989 to 4,123 in 2003, whereas the number of thyroid surgeries decreased from 3,419 in 1989 to 2,825 in 2003. The mortality rates decreased by 2.3% per year (p = 0.01). The decrease in incidence of both follicular and anaplastic TC is assumed to be largely responsible for the decrease in TC mortality rates.