Malignant Thyroid Cells Research Articles

Comparison of mechanical properties of normal and malignant thyroid cells

Cancer is a disease of uncontrolled cell proliferation causing approximately 13% of deaths worldwide. Cancer cell mechanics is currently an important topic of investigation in cancer diagnostics as a possible tool to distinguish malignant cells from normal cells in addition to increasing our understanding of pathophysiology of the disease. Our study, based on Atomic Force Microscopy (AFM) measurements on cells, shows that malignant thyroid cells are 3- to 5-fold softer in comparison to primary normal thyroid cells depending on duration between cell seeding and AFM experiments. These results reveal cultivation period as an important factor that influences cell mechanics and which must be considered when comparing cells. Investigation of actin cytoskeleton by fluorescent labelling revealed differences in organization of actin between malignant and normal thyroid cells, which may be directly contributing to alteration of cell mechanics in cancer cells.

TSH-suppressive treatment in differentiated thyroid cancer. A dogma under review

TSH-suppressive therapy (ST) is part of the treatment protocol for differentiated thyroid carcinoma (DTC). There is however little evidence of its clinical effectiveness. On the other side, ST is not free from side effects related to the subclinical hyperthyroidism status induced in patients for a long time period. Because of this, widespread use of ST in all patients has recently been questioned, and individualized treatment based on the characteristics of each particular case has been proposed.Action of thyroid hormones (THs) depends on their binding to specific nuclear receptors. However, a second pathway mediated by a membrane receptor located in integrin αVβ3 has recently been established. It has been postulated that the proliferative and angiogenic effects attributed to THs would depend on this second mechanism. It is not known whether the tumorigenic action shown in other neoplasms may have an impact on DTC.The objective of this study was to review the relationship between ST and cancer, particularly as regards tumor evolution and occurrence of second primary neoplasms. One of the future challenges in the field of DTC will be to establish the specific role of TSH and THs in malignant thyroid cells, in order to be able to define more optimized therapeutic schemes.

Tratamiento supresor de la TSH en el cáncer diferenciado de tiroides. Un dogma en revisión

TSH-suppressive therapy (ST) is part of the treatment protocol for differentiated thyroid carcinoma (DTC). There is however little evidence of its clinical effectiveness. On the other side, ST is not free from side effects related to the subclinical hyperthyroidism status induced in patients for a long time period. Because of this, widespread use of ST in all patients has recently been questioned, and individualized treatment based on the characteristics of each particular case has been proposed.Action of thyroid hormones (THs) depends on their binding to specific nuclear receptors. However, a second pathway mediated by a membrane receptor located in integrin αVβ3 has recently been established. It has been postulated that the proliferative and angiogenic effects attributed to THs would depend on this second mechanism. It is not known whether the tumorigenic action shown in other neoplasms may have an impact on DTC.The objective of this study was to review the relationship between ST and cancer, particularly as regards tumor evolution and occurrence of second primary neoplasms. One of the future challenges in the field of DTC will be to establish the specific role of TSH and THs in malignant thyroid cells, in order to be able to define more optimized therapeutic schemes.

Osteoprotegerin (OPG) and Related Proteins (RANK, RANKL and TRAIL) in Thyroid Disease

Osteoprotegerin (OPG), receptor activator of nuclear factor kappaB (RANK), RANK ligand (RANKL), and TNF related apoptosis inducing ligand (TRAIL) are the key proteins in the development of bone metastases. Osteoprotegerin improves tumor cell survival and inhibits TRAIL induced apoptosis of cancer cell lines. It also binds to RANKL and inhibits its interaction with RANK (cell surface receptor), which influences osteoclast formation and function. The aim of the present study was to characterize the expression of OPG, RANK, RANKL, and TRAIL in benign and malignant thyroid tissue and thyroid cell lines. Archived thyroid tissue from 79 patients (60 differentiated thyroid cancers and 19 benign thyroids) was stained for OPG, RANK, RANKL, and TRAIL by immunohistochemistry. Staining was assessed semiquantitatively and correlated with pathology. Western blots were performed on three human thyroid cell lines: Nthy-ori 3-1 (thyroid follicular epithelium), FTC-133 (follicular thyroid carcinoma), and K1E7 (subclone of papillary thyroid carcinoma). Osteoprotegerin and RANKL staining was cytoplasmic; RANK staining was nuclear; and TRAIL staining was predominantly cytoplasmic. All four proteins were expressed in benign and malignant tissue. There was significant difference in RANK expression between malignant and benign tissue (8 vs. 84%, respectively; Fisher's exact test; P < 0.001). RANKL expression was significantly increased in malignant tissue (Fisher's exact test; P = 0.04). Western blotting showed OPG expression in all cell lines and TRAIL in none. RANK and RANKL were not detected in papillary and follicular cell lines, respectively. Nuclear expression of RANK protein in thyroid tissue is paradoxical; it could be due to nuclear migration after ligand binding. Suppression of RANK and increased expression of RANKL in malignant tissue warrant further investigation.

A Case of Mixed Papillary Thyroid Tumor and Squamous-Cell Carcinoma

The occurrence of a mixed tumor containing papillary thyroid carcinoma (PTC) and primary squamous-cell carcinoma (SCC) is rare because there is no squamous epithelium in the thyroid gland. Reported herein is a 30-year-old female with mixed PTC and primary sec of the thyroid presented as thyroid incidentaloma. Fine-needle aspiration biopsy of the thyroid nodule revealed the presence of malignant thyroid cells. The histopathological examination following total thyroidectomy yielded two mixed, morphologically distinct histotypes that included PTC and sec. After total thyroidectomy, the patient underwent radioactive iodine therapy. No recurrence or metastasis occurred during the 20-month follow-up period after the operation.

Thyrocyte morphomethric analysis significance in differential diagnosis of thyroid carcinoma

BACKGRAUND/AIM: Conventional cytomorphology of thyroid cell aspirates is limited in preoperative differential diagnosis of follicular adenomas (FA) and hyperplastic adenomatoid nodular goiters from well differentiated thyroid follicular carcinoma (FTC) and follicular variant of thyroid papillary carcinoma (PTC). This is the reason of inaccurate presurgical differential diagnosis and in the same cases of inadequate operative managament. The aim of the study was to evaluate the role of quantitative estimation of thyreocite nuclear features (cariomorphometry) in thyroid aspirated smears in preoperative differential diagnosis of benign from malignant thyroid lesions. A total of 48 patients with thyroid nodular disease underwent fine needle aspiration biopsy for cytomorphology, cariomorphometric analysis of the aspirates, and histopathologic explorations conducted fully postoperatively. On the basis of cytomorphology classification the patients were divided into three groups: benign (B), n = 8; malignant (M), n = 15, and suspicious for malignancy (S), n = 25. Using a microscope connected to a computerized video system, mean nuclear area, the nuclear area coefficient of variation (NACV) and anisocariosis ratio were measured and calculated. In all the 15 patients with cytologically malignant results the diagnosis of PTC was confirmed histopathologically. All cytologically benign lesions were confirmed histopathologically. Thyroid carcinoma was found in 15 out of 25 patients with suspicious lesions. The highest mean values of nuclear area were in the PCT (90.74 +/- 26.71 microm2), and were significantly different from all other groups (p < 0.001). The mean nuclear area in FTC was 69.20 +/- 27.31 microm2 and was significantly higher than in the benign adenomatous group (p < 0.01). There was no significant difference in mean nuclear area between FTC and FA, but there was a significant difference in NACV between these two groups (FTC: 39.46% vs FA: 23.42%,p < 0.001). In 27 out of 30 patients with thyroid carcinoma higher values of NACV than 18% were found. Preoperatively cariomorphometry is a useful method in differential diagnosis of thyroid carcinoma from benign lesions, as a complementary method to convencional cytodiagnostics. The NACV showed highest sensitivity as a parameter of malignant thyroid cell transformation.

Does Fine-Needle Aspiration Cytology Optimize the Surgical Management of Thyroid Disorders in Endemic Goiter Region?

The aim of this study was to determine the diagnostic value of fine-needle aspiration cytology (FNAC) before thyroidectomy in an endemic goiter region. One hundred patients with preoperative FNAC of thyroid nodules who underwent thyroidectomy were recruited. FNAC were classified into five groups. 0, no thyroid cells; 1, normal thyroid cells; 2, degenerative thyroid cells without evidence of malignacy; 3, follicular or oncocytary neoplasia; and 4, malignant thyroid cells. FNAC was compared with postoperative histopathological diagnoses. Only 76% of the FNAC allowed an adequate cytological examination. In 15 patients (15%), carcinomas were found in the postoperative histopathological diagnosis (including four follicular carcinomas). In the 48 patients of FNAC groups 3 and 4, nine carcinomas (18.7%) were found (including four follicular carcinomas). In the 28 patients of groups 1 and 2, there was only one papillary carcinoma (3.5%). In the 24 patients of group 0, there were two papillary, two follicular, and one anaplastic carcinomas (total of 20.8%). The sensitivity, specificity, and likelihood ratio (LR) of the FNAC for benign nodules were 90%, 40.9%, 0.24, respectively. The LR for malignant nodules was 13.2, and that for follicular neoplasia was 0.55. Despite the high prevalence of carcinoma in an endemic goiter region, FNAC disappointed its diagnostic expectation. The lower specificity of FNAC may be caused by a higher prevalence of thyroid nodules in an endemic goiter region or by the absence of a specialized cytopathologist.

Crystallization and preliminary X-ray diffraction analysis of rat protein tyrosine phosphatase eta.

The rat protein tyrosine phosphatase eta (rPTPeta) is a cysteine-dependent phosphatase which hydrolyzes phosphoester bonds in proteins and other molecules. rPTPeta and its human homologue DEP-1 are involved in neoplastic transformations. Thus, expression of the protein is reduced in all oncogene-transformed thyroid cell lines and is absent in highly malignant thyroid cells. Moreover, consistent with the suggested tumour suppression role of PTPeta, inhibition of the tumorigenic process occurs after its exogenous reconstitution, suggesting that PTPeta might be important for gene therapy of cancers. In this study, the catalytic domain of rPTPeta was produced in Escherichia coli in soluble form and purified to homogeneity. Crystals were obtained by the hanging-drop vapour-diffusion method. Diffraction data were collected to 1.87 A resolution. The crystal belongs to space group P2(1)2(1)2(1), with unit-cell parameters a = 46.46, b = 63.07, c = 111.64 A, and contains one molecule per asymmetric unit.

Expression of Aurora kinases in human thyroid carcinoma cell lines and tissues

The Aurora kinases are involved in the regulation of cell cycle progression, and alterations in their expression have been shown to associate with cell malignant transformation. In the present study, we demonstrated that human thyrocytes express all 3 Aurora kinases (A, B and C) at both protein and mRNA level and this expression is cell cycle-regulated. An increase in the protein level of the 3 kinases was found, with respect to normal human thyrocytes (HTU5), in the human cell lines derived from follicular (FTC-133), papillary (B-CPAP) and anaplastic (8305C) thyroid carcinomas, but not in cells derived from a follicular adenoma (HTU42). These observations were mirrored in RT-PCR experiments for Aurora-A and B. In contrast, Aurora-C mRNA levels were not significantly different among the different cell types analyzed, suggesting that posttranscriptional mechanism(s) modulate its expression. The expression at the protein level of all 3 Aurora kinases was significantly higher in 3 thyroid papillary carcinomas with respect to normal matched tissues obtained from the same patients. Similar modifications, at the mRNA level, could be observed in 7 papillary carcinoma tissues for Aurora-A and B, but not for Aurora-C. In conclusion, we demonstrated that normal human thyrocytes express all 3 members of the Aurora kinase family, and their expression is amplified in malignant thyroid cell lines and tissues. These results suggest that the Aurora kinases may play a relevant role in malignant thyroid cancers, and may represent a putative therapeutic target for thyroid neoplasms.

Diagnose, Therapie und Nachsorge des differenzierten Schilddrüsenkarzinoms

For early diagnosis of thyroid cancer, ultrasonography (US) and US-guided fine-needle aspiration biopsy are the methods of choice. Thyroid scintigraphy using Tc-99m pertechnetate can underline the necessity of surgery in case of hypofunctioning nodules. Treatment of thyroid cancer includes total thyroidectomy and staging lymphadenectomy, in the case of lymph node metastases, radical neck dissection of the ipsilateral side. Four weeks after surgery, if TSH exceeds a value of 50 mU/l, with the exception of papillary thyroid cancer pT1a (TNM 1997), radioiodine remnant ablation using activities between 2960 and 3700 MBq I-131 is performed in all other cases. As growth of benign and malignant thyroid cells depends on TSH stimulation, thyroid hormone therapy using TSH suppressive doses (TSH, <0.03 mU/l) follows radioiodine remnant ablation. Additional fractionated external radiation therapy (50 Gy) may be administered in advanced cases (e.g., pT4 N1M0; TNM 1997). Standard follow-up of differentiated thyroid cancer includes measurement of serum thyroglobulin, US of the neck and I-131 whole-body scintigraphy (I-131 WBS). With about 98% the sensitivity of thyroglobulin is very high under TSH stimulation. In case of elevated thyroglobulin, US is the method of choice to detect local recurrences and lymph node metastases of the neck. At defined intervals of follow-up or in case of increasing thyroglobulin, I-131 WBS will be performed under TSH stimulation. With the availability of recombinant TSH (exogenous TSH stimulation) the need to withdraw thyroid hormone over a period of 3-4 weeks (endogenous TSH stimulation) is no longer necessary to perform I-131 WBS. However, in about 20-40% of cases or in the course of disease after several radioiodine therapies, recurrences or metastases may be or become iodine negative. In this case, cationic complexes such as Tc-99m Sestamibi or Tc-99m Tetrofosmin are available to detect less differentiated metastases. In the course of dedifferentiation of malignant thyroid cells, the ability of iodine uptake decreases and uptake of glucose increases. This elevated glucose metabolism can be imaged using FDG PET. Today the combination of PET (metabolic imaging) and CT (morphologic imaging) using PET/CT fusion imaging is the method of choice to image iodine-negative metastases.

Evaluation of Peroxisome Proliferator-Activated Receptor-γ Expression in Benign and Malignant Thyroid Pathologies

Impairment of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) function through a dominant negative PAX-8/PPAR-gamma fusion gene or other events resulting in wild-type PPAR-gamma downregulation has been implicated in malignant thyroid cell transformation. The aim of our study was to perform a systematic evaluation of PPAR-gamma mRNA and protein expression in normal thyroid tissue as opposed to benign thyroid pathologies of different functional status and thyroid malignancy, to gain further insights into a putative physiological role of PPAR-gamma in the thyroid and to define whether PPAR-gamma could serve as a marker of thyroid cell differentiation. Ten cold benign (CTN) and 10 toxic (TTN) thyroid nodules and corresponding normal thyroid tissues, 10 follicular thyroid cancers (FTC), 10 papillary thyroid cancers (PTC) and 8 Graves' disease (GD) thyroids were studied by real-time polymerase chain reaction (PCR), immunohistochemistry and reverse transcriptase (RT)-PCR (PAX-8/PPAR-gamma fusion gene). PPAR-gamma mRNA expression was demonstrated in all samples. When comparing benign nodular and normal thyroid tissue of the same patient no significant difference in PPAR-gamma mRNA expression was observed. PPAR-gamma mRNA levels were similar in CTN and FTC. In contrast, PPAR-gamma mRNA expression was downregulated in 9 of 10 PTC and all GD samples, whereby at least 4 fold downregulation (compared with normal and benign nodular thyroid tissues) was observed in the latter. Immunohistochemistry showed an increased, patchy PPAR-gamma nuclear staining in CTNs and TTNs and only faint staining in the corresponding normal thyroid tissues. A diffuse and weak PPAR-gamma staining pattern was observed in all GD samples. No PAX-8/PPAR-gamma rearrangements were detected in any of the 68 thyroid tissue samples. In conclusion PPAR-gamma mRNA and protein expression levels are not concordant in benign thyroid nodular disease. Furthermore there is no clear-cut association of PPAR-gamma mRNA expression with follicular thyroid tumorigenesis. Absence of a PAX-8/PPAR-gamma fusion gene in the series of 68 thyroid samples is in agreement with the suggestion of PAX-8/PPAR-gamma rearrangement being restricted to a subset of follicular thyroid cancers. The marked downregulation of PPAR-gamma in GD warrants further investigation and could be linked, for example, with changes in apoptosis.

The rat tyrosine phosphatase η increases cell adhesion by activating c-Src through dephosphorylation of its inhibitory phosphotyrosine residue

The expression of the receptor protein tyrosine phosphatase r-PTPeta is drastically reduced in rat and human malignant thyroid cells, whereas its restoration reverts the neoplastic phenotype of retrovirally transformed rat thyroid cells. Moreover, reduced levels and loss of heterozygosity of DEP-1, the human homolog of r-PTPeta, have been found in many human neoplasias. Here, we report that the r-PTPeta protein binds to c-Src in living cells and dephosphorylates the c-Src inhibitory tyrosine phosphorylation site (Tyr 529), thereby increasing c-Src tyrosine kinase activity in malignant rat thyroid cells stably transfected with r-PTPeta. Tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin was enhanced in r-PTPeta-expressing cells. This was associated with increased adhesion of malignant r-PTPeta-transfected thyroid cells vs both untransfected cells and cells stably transfected with an inactive r-PTPeta mutant. Treatment of rat thyroid cells with the c-Src inhibitor PP2 decreased cell adhesion to a higher extent in r-PTPeta-transfected cells than in mock-transfected or stably transfected cells with the inactive r-PTPeta mutant, indicating that r-PTPeta regulates cell-substratum adhesion by activating c-Src. Interestingly, the extent of both c-Src dephosphorylation at Tyr 529, FAK and paxillin phosphorylation, and the increased cell adhesion were associated with the degree of r-PTPeta expression.

Activation of the hepatocyte growth factor (HGF)-Met system in papillary thyroid cancer: biological effects of HGF in thyroid cancer cells depend on Met expression levels.

Met, the receptor for hepatocyte growth factor (HGF), is overexpressed in approximately 90% papillary thyroid carcinomas. To investigate the role of the HGF-Met system in these tumors, we examined HGF and Met expression in a variety of primary cultures, normal or malignant thyroid cells, and tissue specimens and analyzed the different HGF effects (promotion of mitogenesis, branching morphogenesis, and cell motility and invasion). In cancer specimens, HGF was produced at high levels by tumor stromal cells, and Met was constitutively phosphorylated in malignant cells. No HGF production was found in a panel of malignant thyroid cancer cells. Biological effects of HGF were examined in papillary cancer cell cultures with either high or low Met expression. High-Met cells were more sensitive to the growth effect of HGF (ED50 = 3-5 ng/ml and 10-15 ng/ml in high- or low-Met cells, respectively). Moreover, only high-Met cells underwent branching morphogenesis in response to HGF. In contrast, high-Met cells showed a reduced migration. Met down-regulation by small interfering RNAs resulted in enhanced cell migration and abrogation of branching morphogenesis in response to HGF. Conversely, Met overexpression impaired cell migration while favoring branching morphogenesis and cell adherence to substrate. These data suggest that both Met and HGF are overexpressed in papillary thyroid carcinomas, that Met is frequently activated in these carcinomas and may favor tumor growth, and that the abundance of Met expression may differently regulate cell growth, morphogenesis, and migration in response to HGF.

Application of the Cre/loxP system to enhance thyroid-targeted expression of sodium/iodide symporter.

Radioiodide uptake activity mediated by the human Na(+)/I(-) symporter (hNIS) in thyroid follicular cells is the basis for effective (131)I therapy in thyroid cancer. However, radioiodide therapy is not effective in patients with thyroid cancer displaying low or absent hNIS expression. This study assessed the Cre/loxP system for enhancing thyroid-targeted hNIS expression driven by the thyroglobulin (Tg) promoter. The following three recombinant adenoviruses (rAd) were constructed: rAd-Tg-hNIS drives hNIS expression by the Tg promoter; rAd-Tg-Cre drives Cre expression by the Tg promoter; and rAd-CMV-loxP-hNIS drives hNIS expression by the cytomegalovirus (CMV) promoter after Cre-mediated excision of an intervening loxP-GFP-Zeo-loxP. Immortalized normal and malignant rat thyroid cell lines and primary cultures of normal human thyroid and human follicular adenoma cells were investigated. We found that the relative promoter activity of Tg vs. CMV is critical for the efficacy of the Cre/loxP system. In cells with weak Tg promoter activity, coinfection of rAd-Tg-Cre and rAd-CMV-loxP-hNIS induced higher hNIS expression than single infection of rAd-Tg-hNIS. Finally, Tg promoter activity was partially restored in malignant thyroid cells by forced expression of the paired domain-containing transcription factor (Pax-8), allowing the Cre/loxP system to mildly enhance radioiodide uptake.

Galectin-3: A promising marker of thyroid malignancy

Background: Galectin-3 is an endogenous beta-galactoside binding lectin implicated in neoplastic transformation and tumor progression. High levels of this lectin have recently been found in malignant thyroid tumors, but not in normal or benign thyroid tissue, suggesting galectin-3 as a promising presurgical marker of thyroid malignancy. Methods: We analyzed immunohistochemically galectin-3 expression in thyroid tissue using a monoclonal antibody. The total of 108 tissue specimens included 55 cases of thyroid carcinoma (30 papillary, 15 follicular, and 10 anaplastic type), 15 samples of follicular adenoma, 15 samples of normal thyroid tissue, and 23 thyroid tissue specimens from human fetuses (16 to 37 weeks of intrauterine life). Results: The results showed galectin-3 expression in 20/30 papillary carcinomas, 11/15 follicular carcinomas, 10/10 anaplastic carcinomas, and 4/15 follicular adenomas. Thyroid follicular cells in normal adult and fetal tissue were negative. Conclusions: These results further confirm that galectin-3 expression is a feature of malignant thyroid cells, and that immunohistochemical detection of galectin-3 could be useful in thyroid carcinoma diagnostics. The absence of galectin-3 in thyroid cells during fetal development suggests that galectin-3 is expressed de novo during malignant transformation of thyroid epithelium, thus it should not be considered an oncofetal antigen.

The role of Fas-mediated apoptosis in thyroid disease.

Recent evidence has emphasized the importance of apoptosis in the maintenance of tissue homeostasis and the pathogenesis of malignant and immune diseases. Autoimmune thyroid diseases such as Hashimoto's thyroiditis and Graves' disease, as well as other autoimmune endocrine diseases, have been associated with dysregulation of apoptotic signaling pathways. In particular, dysfunction of the Fas apoptotic pathway or production of soluble factors including soluble Fas and soluble Fas ligand may be involved in the pathogenesis of these disorders. On the other hand, malignant thyroid cells may avoid Fas-mediated suicide possibly by expression of inhibitors of apoptosis and evade the immune system by inducing apoptosis on infiltrating lymphocytes. The delicate balance between cell proliferation and cell death through the Fas pathway may also play an important role in the control of thyroid cell mass and goitrogenesis. This review analyzes the current evidence on the role of Fas-mediated apoptosis in the pathogenesis of thyroid diseases including Hashimoto's thyroiditis, Graves' disease, thyroid cancer and goiter. However, the exact mechanisms involved in the regulation of apoptosis in thyroid disease remain unclear. Further investigation is needed.

Estrogen promotes growth of human thyroid tumor cells by different molecular mechanisms.

Thyroid tumors are about 3 times more frequent in females than in males. Epidemiological studies suggest that the use of estrogens may contribute to the pathogenesis of thyroid tumors. In a very recent study a direct growth stimulatory effect of 17beta-estradiol was demonstrated in FRTL-5 rat thyroid cells. In this work the presence of estrogen receptors alpha and beta in thyroid cells derived from human goiter nodules and in human thyroid carcinoma cell line HTC-TSHr was demonstrated. There was no difference between the expression levels of estrogen receptor alpha in males and females, but there was a significant increase in expression levels in response to 17beta-estradiol. Stimulation of benign and malignant thyroid cells with 17beta-estradiol resulted in an increased proliferation rate and an enhanced expression of cyclin D1 protein, which plays a key role in the regulation of G(1)/S transition in the cell cycle. In malignant tumor cells maximal cyclin D1 expression was observed after 3 h, whereas in benign cells the effect of 17beta-estradiol was delayed. ICI 182780, a pure estrogen antagonist, prevented the effects of 17beta-estradiol. In addition, 17beta-estradiol was found to modulate activation of mitogen-activated protein (MAP) kinase, whose activity is mainly regulated by growth factors in thyroid carcinoma cells. In response to 17beta-estradiol, both MAP kinase isozymes, extracellular signal-regulated protein kinases 1 and 2, were strongly phosphorylated in benign and malignant thyroid cells. Treatment of the cells with 17beta-estradiol and MAP kinase kinase 1 inhibitor, PD 098059, prevented the accumulation of cyclin D1 and estrogen-mediated mitogenesis. Our data indicate that 17beta-estradiol is a potent mitogen for benign and malignant thyroid tumor cells and that it exerts a growth-promoting effect not only by binding to nuclear estrogen receptors, but also by activation of the MAP kinase pathway.

Rat protein tyrosine phosphatase eta suppresses the neoplastic phenotype of retrovirally transformed thyroid cells through the stabilization of p27(Kip1).

The r-PTPeta gene encodes a rat receptor-type protein tyrosine phosphatase whose expression is negatively regulated by neoplastic cell transformation. Here we first demonstrate a dramatic reduction in DEP-1/HPTPeta (the human homolog of r-PTPeta) expression in a panel of human thyroid carcinomas. Subsequently, we show that the reexpression of the r-PTPeta gene in highly malignant rat thyroid cells transformed by retroviruses carrying the v-mos and v-ras-Ki oncogenes suppresses their malignant phenotype. Cell cycle analysis demonstrated that r-PTPeta caused G(1) growth arrest and increased the cyclin-dependent kinase inhibitor p27(Kip1) protein level by reducing the proteasome-dependent degradation rate. We propose that the r-PTPeta tumor suppressor activity is mediated by p27(Kip1) protein stabilization, because suppression of p27(Kip1) protein synthesis using p27-specific antisense oligonucleotides blocked the growth-inhibitory effect induced by r-PTPeta. Furthermore, we provide evidence that in v-mos- or v-ras-Ki-transformed thyroid cells, the p27(Kip1) protein level was regulated by the mitogen-activated protein (MAP) kinase pathway and that r-PTPeta regulated p27(Kip1) stability by preventing v-mos- or v-ras-Ki-induced MAP kinase activation.

Similar and divergent patterns in the regulation of matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of MMP-1 gene expression in benign and malignant human thyroid cells.

An imbalance between the activity of matrix metalloproteinases (MMPs) (proteolytic enzymes that degrade protein components of the extracellular matrix) and their inhibitors, the tissue inhibitors of metalloproteinases (TIMPs), may be one of the mechanisms responsible for tumor cell invasion. We have investigated the regulation of MMP-1 and TIMP-1 gene expression in benign and malignant (follicular, anaplastic, and papillary) human thyroid cells. As expected of cells with invasive potential, detectable MMP-1 messenger RNA (mRNA) levels were observed in malignant cells under basal conditions, in contrast to undetectable levels in benign cells. Exposure of these cells, for 1 h, to the active phorbol ester, phorbol 12-myristate 13-acetate (TPA, 100 nmol/L), acting via protein kinase C (PKC), elicited an increase in MMP-1 mRNA, with a peak stimulation after a 3- to 4-h culture period. Epidermal growth factor (EGF, 25 ng/mL), however, acting via protein tyrosine kinase (PTK), stimulated such gene expression in malignant cells but failed to do so in benign cells. TIMP-1 mRNA was not significantly altered by the TPA-PKC, EGF-PTK, or TSH-protein kinase A (PKA) pathways in malignant cells. In benign cells, however, TPA induced a small, though significant, increase in TIMP-1. The MMP-1 stimulation by EGF and lack of TPA-induced rise in TIMP-1 in malignant cells, in sharp contrast to the effects obtained in benign thyrocytes, seems to indicate that the MMP: TIMP balance favors a more extensive extracellular matrix protein breakdown by malignant thyrocytes, as expected of cells exhibiting invasive capacity. TSH (10-500 microU/mL) failed to significantly influence basal MMP-1 or TIMP-1 mRNA levels, but it caused a dose-dependent inhibition in TPA- and EGF-induced MMP-1 mRNA in malignant cells, and TPA-stimulated MMP-1 and TIMP-1 in benign cells. The repressive action of TSH on MMP-1 mRNA was mimicked by forskolin and 8-bromo-cAMP and was abrogated by the PKA inhibitor, H-89, suggesting that the TSH inhibitory action is PKA-mediated. In conclusion, the present study provides novel data on MMP-1 and TIMP-1 gene expression and their modulation by the major signal transduction pathways operating in human thyroid cells. Similar and divergent patterns have emerged in the regulation of such gene expression in benign and malignant human thyrocytes, in many instances in accord with the concept of MMP playing the role of stimulating, and TIMP inhibiting, cell invasion. Although MMP-1 may be just one of the many factors responsible for tumor cell invasion, the present findings demonstrating the possibility, at least in vitro, of repressing MMP gene expression may have important clinical ramifications.

Similar and Divergent Patterns in the Regulation of Matrix Metalloproteinase-1 (MMP-1) and Tissue Inhibitor of MMP-1 Gene Expression in Benign and Malignant Human Thyroid Cells

Similar and Divergent Patterns in the Regulation of Matrix Metalloproteinase-1 (MMP-1) and Tissue Inhibitor of MMP-1 Gene Expression in Benign and Malignant Human Thyroid Cells