Complementary DNA Expression Array Analysis Suggests a Lower Expression of Signal Transduction Proteins and Receptors in Cold and Hot Thyroid Nodules

Abstract

Autonomously functioning thyroid nodules are characterized by an increased proliferation and function, which is predominantly caused by constitutively activating TSH receptor mutations leading to an activation of cAMP. In contrast to autonomously functioning thyroid nodules, cold thyroid nodules are functionally inactive and less differentiated. Their molecular cause is still unknown. To further investigate the pathophysiological aspects of autonomously functioning thyroid nodules and to elucidate the molecular etiology of cold thyroid nodules, it is essential to identify genes with differential expression in autonomously functioning thyroid nodules and cold thyroid nodules and to compare this expression to that in normal surrounding tissue. The list of possible candidates for differential regulation ranges from growth factors and their receptors to transcription factors or oncogenes. Therefore, we evaluated the potential of cDNA arrays and studied the expression of 588 known genes from 6 different classes of proteins in thyroid nodules characterized for their function. Forty-seven genes showed a differential expression between nodular and surrounding tissue identified by the expression arrays. The differential expression of 15 transcripts was verified by real-time PCR. About 25% of the transcripts determined by LightCycler PCR are considered false positives because data from PCR and array analysis did not agree. This indicates the reliability of cDNA expression arrays to identify differentially expressed genes in thyroid nodules compared with their surrounding tissue. The 15 selected genes were additionally quantified by real-time PCR in 7 additional cold thyroid nodules, autonomously functioning thyroid nodules, and their surrounding tissues. The highest number of differentially expressed genes was in the group of signal transduction proteins (4 of 38 detectable genes) and extracellular cell signaling and communication proteins (2 of 62 detectable genes). In contrast, transcripts of other classes of proteins were unchanged (e.g. DNA-binding molecules and stress responses). Most of the transcripts were down-regulated in autonomously functioning thyroid nodule and cold thyroid nodule compared with the respective surrounding tissue. This finding could be the result of a dominant activation of a signal transduction pathway, with the cAMP pathway being the likely candidate for autonomously functioning thyroid nodules. The qualitatively similar pattern of changes in this limited number of genes in autonomously functioning thyroid nodules and cold thyroid nodules could suggest a similar dominant activation of a specific signaling cascade in cold thyroid nodules as the constitutively activating mutations in autonomously functioning thyroid nodules.

Gene expression in thyroid autonomous adenomas provides insight into their physiopathology

“…First, where the two microarray analyses overlap, their results are similar, even though different samples and different arrays and protocols for samples processing were used. Second, we confirmed several regulations already identified previously (Deleu et al, 2000;Eszlinger et al, 2001Eszlinger et al, , 2004O'Donovan et al, 2002;Finley et al, 2004;Mazzanti et al, 2004;Detours et al, 2005). Moreover, several findings are in line with data obtained in different but related systems such as thyroid cells or other cells chronically stimulated by the cAMP pathway (Yamazaki et al, 2003).…”

“…Signal transduction proteins were regulated in both directions with more genes downregulated than upregulated. The greater number of downregulated genes has been reported before (Eszlinger et al, 2001). It can be related to the decrease in lymphocytic population and/or to a simplification of signal transduction in the tumor cell.…”

“…The low frequency of altered regulations already observed in autonomous adenomas (Eszlinger et al, 2001) and in Tg-A2aR mice (Goffard et al, 2004) reflects the physiology of such tumors. The metabolism of the adenomas is normal if stimulated.…”

A multi-gene approach to differentiate papillary thyroid carcinoma from benign lesions: gene selection using support vector machines with bootstrapping

“…Therefore, we decided to use a balanced mixture of samples from malignant and benign tumors and normal thyroid tissue to mimic the clinical situation, where the material from any of these may be obtained and shall be properly classified. This large 180-array dataset is derived respectively from de novo studies ( n =40), previously published own microarray data ( n =124; Eszlinger et al 2001 , 2004 , Jarzab et al 2005 ), and accessible datasets published by other authors ( n =16; Huang et al 2001 ).…”

Increased expression of pro-angiogenic factors and vascularization in thyroid hyperfunctioning adenomas with and without TSH receptor activating mutations

“…An increased blood supply provided by enhanced vascularization is a known feature of these tumors. Indeed, studies in small series of toxic adenomas have revealed changes in the secretion or expression of the vascular endothelial growth factor (VEGF) [7,8] and plateletderived growth factors (PDGF) [9], but the global gene expression profile of these factors and their receptors in these tumors has never been investigated. Moreover, there are no data regarding the influence of the TSH receptor mutations on the tumor vascularization and the expression of the angiogenic factors as well as their relationship with other markers of thyrocyte differentiation such as sodium iodide symporter (NIS), TSHR, Thyroglobulin (Tg), and Thyroperoxidase (TPO).…”