To investigate the effect of serine/threonine-protein kinase B-Raf (BRAF)-activated long-chain non-coding RNA (lncRNA-BANCR) on apoptosis and autophagy in thyroid carcinoma cells and the underlying mechanisms.
 Methods: RT-PCR was used to detect the expression of lncRNA-BANCR in thyroid carcinoma and normal thyroid tissues. The association between lncRNA-BANCR and clinicopathological data was analyzed in patients with thyroid cancer. Cell counting kit-8 (CCK-8) was used to detect the effect of lncRNA-BANCR on the proliferation of thyroid cancer cells. The effect of lncRNA-BANCR on the apoptosis of thyroid carcinoma cells was detected by flow cytometry. Transwell invasion assay was used to detect the effect of lncRNA-BANCR on the invasive ability of thyroid cancer cells. Western blot was used to detect the changes of autophagy proteins LC3-I and LC3-II after the lncRNA-BANCR expression was suppressed.
 Results: Compared with normal thyroid tissues, the expression level of lncRNA-BANCR in thyroid carcinoma tissues was elevated (P<0.05). The expression of lncRNA-BANCR was positively related to the pathological stage of thyroid carcinoma and the lymph node metastasis. Inhibition of lncRNA-BANCR expression attenuated the proliferation and invasion ability of thyroid cancer cells (both P<0.05); but the apoptosis was enhanced (P<0.05); the expression levels of autophagy protein LC3-I and LC3-II were also increased (P<0.05).
 Conclusion: The expression level of lncRNA-BANCR affects the proliferation, invasion and apoptosis of thyroid cancer cells through modulation of autophagy behavior.
Apoptosis ; Autophagy ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Humans ; Neoplasm Invasiveness ; Proto-Oncogene Proteins B-raf ; metabolism ; RNA, Long Noncoding ; metabolism ; Serine ; metabolism ; Threonine ; metabolism ; Thyroid Gland ; cytology ; metabolism ; Thyroid Neoplasms ; metabolism ; pathology

Diagnostic and therapeutic use of radioiodine in the management of thyroid disorders depends on the ability of thyroid cells to concentrate radioiodine, a process that is regulated by the intracellular increase in cAMP. We hypothesized that theophylline, a drug known to increase intracellular cAMP via inhibition of phosphodiesterase, could increase thyroidal radioiodine uptake. We tested this effect in vivo, using C57BL/6j mice, and in vitro, using Fisher rat thyroid (FRTL-5) cells. One mouse received 2.5mg theophylline i.p., whereas a control mouse received only saline. Twenty-hours after theophylline, mice were injected with 10mu Ci Na(125)I in 0.1 mL saline through the tail vein. Mean thyroidal (125)I activity was 3.3-fold higher in theophylline-treated mice than in their respective controls. Radioiodine uptake and intracellular cAMP production of FRTL-5 cells were increased by a relatively low concentration of theophylline (1mu M). Intracellular cAMP increased up to 30 min and then declined in response to 1mu M theophylline. Sera from theophylline-treated mice stimulated (125)I uptake and intracellular cAMP production by FRTL-5 cells. These findings show that theophylline can enhance radioiodine uptake by thyrocytes in vivo and in vitro. The in vitro effects of theophylline on both radioiodine uptake and cAMP production in a dose-dependent manner are consistent with an action mediated by phosphodiesterase inhibition.
Animals ; Blood Proteins/pharmacology ; Cells, Cultured ; Cyclic AMP/metabolism ; Female ; Iodine Radioisotopes/*diagnostic use/pharmacokinetics ; Mice ; Mice, Inbred C57BL ; Theophylline/*pharmacology ; Thyroid Gland/cytology/*metabolism ; Vasodilator Agents/*pharmacology

OBJECTIVETo study the effect of different selenium intake on the expression of apoptosis protein Fas/FasL in experimental autoimmune thyroiditis (EAT) rats' thyroid with adequate iodine.

METHODSThirty-two female lewis rats were divided stochastically into 4 groups as C group, M group, Se(+) + M group, Se(-) + M group, respectively, and pretreated with feedstuffs containing different concentrations of selenium (Se(+) + M group 2 mg/kg, C and M group 0.20 mg/kg, Se(-) + M group 0.02 mg/kg, respectively) for two weeks, and immunized the rats with porcine thyroglobulin (pTg) to establish an EAT model. The thyroid gland was sampled, embedded in mineral wax and sliced, and the expression of Fas/FasL was measured with immunohistochemistry.

RESULTSBoth the expressions of Fas and FasL of EAT rats were significantly increased as compared with control group. The expression of Fas in rats' thyroid follicular cells with EAT was down-regulated as the increased selenium intake (optical density: 0.059 +/- 0.006), the expression of Fas of Se(+) + M group (0.036 +/- 0.004) was significantly inhibited (q = 11.591, P = 0.000), and expression of Fas was lower in the Se(+) + M group than Se(-) + M group (0.050 +/- 0.005) (q = 7.055 , P = 0.000). Effect of selenium on FasL was not identified.

CONCLUSIONIncreasing the intake of selenium might decrease the expression of Fas on thyroid follicular cells and restrain the development of EAT.

Animals ; Apoptosis ; Fas Ligand Protein ; biosynthesis ; Female ; Rats ; Rats, Inbred Lew ; Selenium ; pharmacology ; therapeutic use ; Thyroid Gland ; cytology ; metabolism ; Thyroiditis, Autoimmune ; drug therapy ; metabolism

This study is to investigate the activation effect of butyl-p-hydroxybenzoate (Bpb) on cAMP-dependent cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel gating. A stably transfected Fischer rat thyroid (FRT) epithelial cell lines co-expressing human CFTR and a green fluorescent protein mutant with ultra-high halide sensitivity (EYFP) were used to measure CFTR-mediated iodide influx rates. Bpb was identified as an effective activator of wild-type CFTR chloride channel, it can correct delta F508-CFTR gating defects but not processing defect. Bpb can't potentiate G551D-CFTR channel gating. The activity was reversible and dose-dependent. The study also provided clues that Bpb activates CFTR chloride channel through a direct binding mechanism. Our study identified Bpb as a novel structure CFTR activator. Bpb may be useful for probing CFTR channel gating mechanisms and as a lead compound to develop pharmacological therapy for CFTR-related disease.
Animals ; Cell Line ; Cystic Fibrosis Transmembrane Conductance Regulator ; genetics ; metabolism ; Dose-Response Relationship, Drug ; Epithelial Cells ; metabolism ; Green Fluorescent Proteins ; genetics ; metabolism ; Ion Channel Gating ; drug effects ; Mutation ; Parabens ; administration & dosage ; pharmacology ; Rats ; Rats, Inbred F344 ; Thyroid Gland ; cytology

The aim of the present study was to investigate whether the physiological features of Ano1 were affected by enhanced green fluorescent protein (EGFP) fusing at Ano1 C-terminal. The eukaryotic expression vectors of Ano1 and EGFP-Ano1 were constructed, and these plasmids were transfected into Fischer rat thyroid follicular epithelial (FRT) cells using liposome. The expression and location of Ano1 were examined by using inverted fluorescence microscope. The ability of Ano1 to transport iodide was detected by kinetics experiment of fluorescence quenching. The results showed that both Ano1 and EGFP-Ano1 were expressed on FRT cell membrane and could be activated by Ca(2+). There was no significant difference of the ability to transport iodide between Ano1 and EGFP-Ano1. These results suggest Ano1 and EGFP-Ano1 have similar physiological feature.
Animals ; Anoctamin-1 ; Cell Membrane ; physiology ; Chloride Channels ; metabolism ; Epithelial Cells ; physiology ; Genetic Vectors ; Green Fluorescent Proteins ; metabolism ; Microscopy, Fluorescence ; Plasmids ; Rats ; Recombinant Fusion Proteins ; metabolism ; Thyroid Gland ; cytology ; Transfection

B-RafV600E mutant is found in 40-70% of papillary thyroid carcinoma (PTC) and has an important role in the pathogenesis of PTC. The sodium iodide symporter (NIS) is an integral plasma membrane glycoprotein that mediates active iodide transport into the thyroid follicular cells, and B-RafV600E has been known to be associated with the loss of NIS expression. In this study, we found that B-RafV600E inhibited NIS expression by the upregulation of its promoter methylation, and that specific regions of CpG islands of NIS promoter in B-RafV600E harboring PTC were highly methylated compared with surrounding normal tissue. Although DNA methyltransferase 3a and 3b (DNMT3a,3b) were not increased by B-RafV600E, DNMT1 expression was markedly upregulated in PTC and B-RafV600E expressing thyrocytes. Furthermore, DNMT1 expression was upregulated by B-RafV600E induced NF-kappaB activation. These results led us to conclude that NIS promoter methylation, which was induced by B-RafV600E, is one of the possible mechanisms involved in NIS downregulation in PTC.
Base Sequence ; Carcinoma/*genetics/metabolism/pathology ; Cells, Cultured ; DNA (Cytosine-5-)-Methyltransferase/analysis/*genetics/metabolism ; DNA Methylation ; Down-Regulation ; *Gene Expression Regulation, Neoplastic ; Humans ; Molecular Sequence Data ; *Point Mutation ; Promoter Regions, Genetic ; Proto-Oncogene Proteins B-raf/*genetics/metabolism ; Symporters/analysis/*genetics/metabolism ; Thyroid Gland/cytology/metabolism/pathology ; Thyroid Neoplasms/*genetics/metabolism/pathology ; Up-Regulation

OBJECTIVETo observe the effects of amitrole on the transcription of thyroglobulin (tg), thyroid peroxidase (tpo), Na(+)/I- symporter (nis), Na(+)/I- symporter (nis), thyroid-stimulating hormone receptor (tshr), thyroid transcription factor 1 (ttf-1) and paired-domain protein-8 (pax-8) genes in FRTL-5 cells and investigate the mechanism of amitrole for intervening in thyroid hormone activity.

METHODSFRTL-5 cells were treated with amitrole at 0.001, 0.01 and 0.1 mg/ml for 24 h, respectively, after which the cells were collected for extraction of the total RNA. RT-PCR was used to examine the effects of amitrole on the transcription of tg, tpo, nis, tshr, pax-8 and ttf-1 genes in FRTL-5 cells.

RESULTSAmitrole significantly induced tg gene transcription at all the doses, but produced no obvious effects on tpo and nis gene transcription. At the concentration of 0.1 mg/ml, amitrole significantly reduced pax-8 and tshr gene transcription but increased ttf-1 gene transcription.

CONCLUSIONThe effects of amitrole on thyroid hormone activity may be related with its actions on tg, ttf-1, tshr and pax-8 gene transcription.

Amitrole ; toxicity ; Animals ; Cells, Cultured ; Enzyme Inhibitors ; toxicity ; Epithelial Cells ; cytology ; drug effects ; metabolism ; Nuclear Proteins ; genetics ; Rats ; Rats, Inbred F344 ; Receptors, Thyrotropin ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Thyroglobulin ; genetics ; Thyroid Gland ; cytology ; Thyroid Nuclear Factor 1 ; Transcription Factors ; genetics ; Transcription, Genetic ; drug effects

Members of the inhibitors of differentiation (Id) family of helix-loop-helix (HLH) proteins are known to play important roles in the proliferation and differentiation of many cell types. Thyroid-stimulating hormone (TSH) regulates proliferation and differentiation by activating TSH receptor (TSHR) in thyrocytes. In this study, we found that Id2, one of the Id family proteins, is a major target for regulation by TSH in FRTL-5 thyroid cells. TSH rapidly increases the Id2 mRNA level in FRTL-5 thyroid cells but the Id2 protein showed biphasic regulatory patterns, being transiently reduced and subsequently induced by TSH treatment. Transient reduction of Id2 protein was noted within 2 hr of TSH treatment and was mediated by proteasomal degradation. Moreover, reduced Id2 expression correlated with the activity of the phosphatidylinositol 3 kinase pathway, which is activated by TSH. Although TSH increases the activity of the Id2 promoter, TSH-induced activation of this promoter was independent of c-Myc. Id2 did not alter TTF-1- and Pax-8-mediated effects on the regulation of the Tg promoter. Thus, in summary, we found that TSH regulates Id2 expression, but that Id2 does not alter the expression of thyroid-specific genes, such as Tg, in FRTL-5 thyroid cells.
1-Phosphatidylinositol 3-Kinase/metabolism ; Animals ; Cattle ; Cell Differentiation ; Cell Proliferation ; *Gene Expression Regulation ; Inhibitor of Differentiation Protein 2/metabolism ; Insulin/metabolism ; Paired Box Transcription Factors/metabolism ; Promoter Regions, Genetic ; Proto-Oncogene Proteins c-myc/metabolism ; Rats ; Thyroglobulin/metabolism ; Thyroid Gland/*cytology ; Thyrotropin/*metabolism

Aim of the present study is to investigate activation effect of nobiletin on cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel activity. CFTR-mediated iodide influx assay and patch-clamp tests were done on FRT cells stably co-transfected with human CFTR and EYFP/H148Q. Nobiletin potently activated CFTR chloride channel activity in a dose- and time-dependent manner. The CFTR blocker CFTR(inh)-172 could completely reverse the effect. Preliminary mechanism study indicated that nobiletin activated CFTR chloride channel through a direct binding way. In addition, ex vivo tests done on mice trachea showed that nobiletin time-dependently stimulated submucosal gland fluid secretion. Nobiletin may be a therapeutic lead compound in treating CFTR-related diseases including disseminated bronchiectasis.
Animals ; Benzoates ; pharmacology ; Cystic Fibrosis Transmembrane Conductance Regulator ; antagonists & inhibitors ; drug effects ; metabolism ; Dose-Response Relationship, Drug ; Epithelial Cells ; metabolism ; Exocrine Glands ; secretion ; Flavones ; administration & dosage ; pharmacology ; Humans ; Mice ; Patch-Clamp Techniques ; Rats ; Rats, Inbred F344 ; Thiazolidines ; pharmacology ; Thyroid Gland ; cytology ; Time Factors ; Trachea ; secretion