BACKGROUND:We compared the analytical and clinical performance of 3 porcine thyroid receptor antibody (TRAb) methods (1 second-and 2 new third-generation systems) with the conventional TRAb assay based on the human recombinant TSH receptor (hTRAK).
Doxorubicin remains the most extensively used drug in the chemotherapy of thyroid cancer. However, drug resistance often limits the efficacy of chemotherapy in clinical practice. Several anticancer drugs exert their cytotoxic effect by triggering Fas-mediated apoptosis in some cell types. However, no investigations have been conducted to determine whether doxorubicin causes apoptosis in thyroid carcinomas. In the present study, we assessed the cytotoxic and apoptotic effects of doxorubicin on two thyroid cancer cell lines (FTC 238 and FTC 133). Cytotoxic effects of doxorubicin were evaluated by a 3-(4,5 dimethylthiazol-2yl) 2-5 diphenyltetrazolium bromide (MTT) assay. Apoptosis was quantified by fluorescein isothiocyanate-conjugated annexin V/flow cytometric analysis and by DNA fragmentation. Fas expression was measured by flow cytometric analysis. After a 24-hour incubation, doxorubicin induces a dose-dependent cytotoxicity in the two cell lines. Treatment with doxorubicin (0.5 and 1 microM) for 24 hours induced cell apoptosis and upregulated Fas expression. A significant correlation was found between the fluorescence intensity values obtained with annexin V staining and those observed for Fas expression (r = 0.996; p < 0.001 or r = 0.957; 0.02 < p < 0.05 for FTC 238 or FTC 133 cells, respectively). In conclusion, doxorubicin exerts its cytotoxic effects, at least partly, through Fas-mediated apoptosis in thyroid cancer cells. These results may have clinical implications for thyroid cancer therapy.
ATP-binding cassette (ABC) transporters [P-glycoprotein and multidrug resistance (MDR)-associated proteins (MRPs)] confer MDR to tumor cells. In this work, we investigated doxorubicin resistance in three thyroid carcinoma cell lines. The effects of sodium butyrate (NaB) on doxorubicin-induced cytotoxicity and on transcription of three MDR genes were also studied. Thyroid cell lines established from anaplastic (8505C) and two poorly differentiated follicular (FTC 238 and FTC 133) cancers were cultured for 24 or 48 h in the presence of NaB (0, 0.25, 0.5 and 1 mM) alone or combined with increased doses of doxorubicin. Cytotoxicity was assessed using the MTT test. MDR1, MRP1 and MRP2 mRNA expression was studied by RT-PCR. After a 24- or 48-h incubation, doxorubicin alone induced cytotoxicity in the three cell lines. NaB significantly (p<0.0001) increased the doxorubicin-induced cytotoxicity. MRP1 transcripts were expressed in the three non-treated cell lines. MDR1 and MRP2 mRNAs were both present in 8505C, but absent in FTC 133 or FTC 238 cell lines, respectively. Treatment with NaB for 24 or 48 h induced no change in MRP1 and MRP2 levels, but increased MDR1 expression in 8505C and FTC 238 cell lines comparably to alkaline phosphatase activity. In conclusion, MRP1 and sometimes MDR1 and MRP2 are expressed in the tested cell lines. NaB potentiates doxorubicin-induced cytotoxicity independently of the ABC transporters. The combination of doxorubicin and NaB might have clinical implications for thyroid cancer therapy.
We studied the lymphocyte-induced alterations in hormonal metabolism and the production of tumour necrosis factor alpha (TNF-alpha) during coculture of thyrocytes and autologous lymphocytes from 20 patients with Graves' disease and from five normal subjects. Thyroglobulin (Tg) mRNA was assessed by slot-blot analysis under TSH stimulation. Tg, tri-iodothyronine (T3) and cAMP secretion in the presence of TSH were measured by RIA after 3 or 5 days of coculture. TNF-alpha levels produced after 5 days incubation were also assayed in lymphocyte culture and coculture media. Lymphocytes isolated from peripheral blood (PBLs) altered the production of Tg, T3 and cAMP in autologous thyrocytes. Intrathyroidal lymphocytes (ITLs) decreased Tg and cAMP secretion but had no effect on T3 secretion. The reductions in Tg and cAMP levels obtained with mechanically isolated ITLs (M-ITLs) were generally higher than those obtained with ITLs isolated by dispase (D-ITLs). No difference was seen between Graves' disease and normal cocultures. PBLs secreted large concentrations of TNF-alpha, larger than those obtained with M-ITLs whereas D-ITLs produced low amounts of this cytokine. In coculture, TNF-alpha levels were lower than those observed in lymphocyte culture. Significant correlations were obtained between TNF-alpha levels and the decrease in Tg, T3 and cAMP concentrations. The percentage of T lymphocytes was higher in PBLs and D-ITLs than in M-ITLs. B lymphocytes levels were higher in ITLs, especially M-ITLs, than in PBLs. TNF-alpha production by B lymphocytes was maximal in M-ITLs. In conclusion, lymphocytes induced a decrease in hormonal thyroid metabolism when cocultured with autologous thyrocytes. These perturbations may be attributed, at least partly, to TNF-alpha secreted by lymphocytes. TNF-alpha interacts via the adenylate cyclase pathway of TSH signal transduction.
Cis-diamminedichloroplatinum (II) (cisplatin) is a widely used anticancer drug which induces many side-effects, but its action on the thyroid gland is still unknown. We have investigated the effects of this drug on human thyrocytes cultured in monolayers or in follicles and stimulated with 200 microU TSH/ml. After 72 h in culture, different concentrations of cisplatin (15, 30 and 75 microM) caused partial or total inhibition of cyclic AMP (cAMP), thyroglobulin (Tg) and tri-iodothyronine (T3) production, whereas thyroxine levels increased in the medium of thyrocytes cultured as follicles. Small doses of the drug did not affect thyrocyte production. Decreases in neutral-red uptake by thyroid cells and in intracellular lactate dehydrogenase, alpha-hydroxybutyryldehydrogenase and creatine phosphokinase activities were induced by 30 and 75 microM cisplatin. These data show that high concentrations of cisplatin had a cytotoxic effect on thyrocytes. Cisplatin also induced inhibition of the production of cAMP, Tg and T3.
Adhesion molecules, such as Intercellular Adhesion Molecule-1 (ICAM-1), play an important role during the autoimmune process of Graves' disease (GD). So the objective of the study was to evaluate the time-course of the soluble ICAM-1 (sICAM-1) in GD. Concentrations of sICAM-1, thyroid hormones and TSAb (thyroid-stimulating antibodies) were determined in sera from 30 healthy controls, 41 untreated GD patients and after 3, 6, 12, 18 months of carbimazole therapy (no.=30), at relapse (no.=11) or 2 years after the end of therapy when remission (no.=13). Mean sICAM-1 concentration was significantly higher in untreated GD patients than in controls (mean+/-SD: 371+/-108 ng/ml vs 243+/-47 ng/ml, p<0.0001) until 6 months of therapy (289+/-102 ng/ml; NS). The number of positive patients (sICAM-1 levels>mean of the controls+2 SD) declined from 56% (23/41) at the time of the diagnosis to 10% (3/29) at 18 months. At relapse, mean sICAM-1 level significantly increased compared to that at 18 months of therapy (288+/-65 vs 236+/-59 ng/ml, p=0.005). At remission mean sICAM-1 level was significantly lower than in relapse patients (240+/-48 ng/ml, p=0.04); no patient displayed sICAM-1 positive values. In conclusion, sICAM-1 concentrations were increased in sera of newly diagnosed GD patients, declined significantly during carbimazole therapy and could again be increased at relapse. sICAM-1 could reflect an ongoing immune process and help to affirm the presence of an autoimmunity notably in some cases of TSAb negative patients. However its precise interest in clinical practice remains to be determined in further studies.
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