Differentiated thyroid cancers and their metastases frequently exhibit reduced iodide uptake, impacting on the efficacy of radioiodine ablation therapy. PTTG binding factor (PBF) is a proto-oncogene implicated in the pathogenesis of thyroid cancer. We recently reported that PBF inhibits iodide uptake, and have now elucidated a mechanism by which PBF directly modulates sodium iodide symporter (NIS) activity in vitro. In subcellular localisation studies, PBF overexpression resulted in the redistribution of NIS from the plasma membrane into intracellular vesicles, where it colocalised with the tetraspanin CD63. Cell-surface biotinylation assays confirmed a reduction in plasma membrane NIS expression following PBF transfection compared with vector-only treatment. Coimmunoprecipitation and GST-pull-down experiments demonstrated a direct interaction between NIS and PBF, the functional consequence of which was assessed using iodide-uptake studies in rat thyroid FRTL-5 cells. PBF repressed iodide uptake, whereas three deletion mutants, which did not localise within intracellular vesicles, lost the ability to inhibit NIS activity. In summary, we present an entirely novel mechanism by which the proto-oncogene PBF binds NIS and alters its subcellular localisation, thereby regulating its ability to uptake iodide. Given that PBF is overexpressed in thyroid cancer, these findings have profound implications for thyroid cancer ablation using radioiodine.

International audienceSphingosine 1-phosphate (S1P) receptor expression and the effects of S1P on migration were studied in one papillary (NPA), two follicular (ML-1, WRO) and two anaplastic (FRO, ARO) thyroid cancer cell lines, as well as in human thyroid cells in primary culture. Additionally, the effects of S1P on proliferation, adhesion and calcium signalling were addressed in ML-1 and FRO cells. All cell types expressed multiple S1P receptors. S1P evoked intracellular calcium signalling in primary cultures, ML-1 cells and FRO cells. Neither proliferation nor migration was affected in primary cultures, whereas S1P partly inhibited proliferation in ML-1 and FRO cells. Low nanomolar concentrations of S1P inhibited migration in FRO, WRO and ARO cells, but stimulated ML-1 cell migration. Consistently, S1P 1} and S1P 3}, which mediate migratory responses, were strongly expressed in ML-1 cells and S1P 2}, which inhibits migration, was the dominating receptor in the other cell lines. The migratory effect in ML-1 cells was mediated by G i} and phosphatidylinositol 3-kinase. Both S1P and the S1P 1}-specific agonist SEW-2871 induced Akt phosphorylation at Ser-473. However, SEW-2871 failed to stimulate migration, whereas the S1P 1}/S1P 3} antagonist VPC 23019 inhibited S1P-induced migration. The results implicate that aberrant S1P receptor expression may enhance thyroid cancer cell migration and thus contribute to the metastatic behaviour of some thyroid tumours

PBF is an additional prognostic indicator in differentiated thyroid cancer that is transforming in vitro and tumorigenic in vivo.

Ramasamy, S., S. Singh, P. Taniere, M. J. S. Langman, and M. C. Eggo. Sulfide-detoxifying enzymes in the human colon are decreased in cancer and upregulated in differentiation. Am J Physiol Gastrointest Liver Physiol 291: G288 -G296, 2006. First published February 23, 2006 doi:10.1152/ajpgi.00324.2005.-H 2S is highly toxic and selectively inhibits butyrate oxidation in colonocytes. Ineffective detoxification may result in mucosal insult, inflammation, and ultimately in colorectal cancer (CRC). Rhodanese can detoxify H 2S and is comprised of two isoenzymes: thiosulfate sulfurtransferase (TST) and mercaptopyruvate sulfurtransferase (MST). Using specific antisera to discriminate TST from MST, we found that only TST could detoxify H2S. In sections of normal colon, both enzymes were located on the luminal mucosal surface, and they were expressed in the colonocytes but not in the mucin-secreting goblet cells. Expression of both enzymes was focally lost in ulcerative colitis and markedly reduced in advanced colon cancer, the disease progression correlating with the decreased expression of MST and TST. In HT-29 cells, a human colon cancer cell line, TST activity and expression were significantly increased by butyrate and by histone deacetylase inhibition, agents that promote HT-29 cell differentiation. Sulfide (0.1 mM) also increased TST activity, but higher sulfide concentrations (0.3-3 mM) were toxic. Preincubation in butyrate to increase TST expression, decreased sensitivity of the cells to sulfide toxicity. We conclude that decreased expression of TST (or MST) is a tumor marker for CRC. TST expression is increased in colonocyte differentiation. Dysregulation of TST expression and activity resulting in inability to effectively detoxify could be a factor in the cell loss and inflammation that accompany ulcerative colitis and ultimately then in CRC. colorectal cancer; thiosulfate sulfurtransferase; butyrate; HT-29 IN THE HUMAN COLON, anaerobic bacteria produce hydrogen sulfide (H 2 S) (25), which inhibits the oxidation of short-chain fatty acids (SCFA) (18,19). SCFAs are the main energy source (Ͼ70% ) for colonocytes (18) and also contribute 5-10% of the total energy supplies in humans. Inhibition of their oxidation in the colon will result in colonocyte starvation and death, resulting in inflammation. This, the "energy deficiency" hypothesis of Roediger (18), is postulated to lead to ulcerative colitis (UC). Many studies have shown that patients with UC have increased colorectal cancer (CRC) risk, with Prior et al. (16) estimating this as an 11-fold excess. On the basis of these studies, we suggest that the expression of enzymes important in sulfide detoxification is essential to preserve a healthy mucosa and that derangements in expression may accompany UC and CRC.We recently showed that colonic mucosal rhodanese can detoxify hydrogen sulfide (15) and thus might protect against mucosal injury. Rhodanese is a mitochondrial enzyme, present in all living organisms from bacteria to humans, and is thought to play a cen...

Adipocytes contain large lipid droplets in their cytoplasm. When cultured, they float on top of the medium, clump together, and do not gain equal and sufficient access to the medium. Morphological changes cannot be observed and the majority of adipocytes undergo cell lysis within 72 h of isolation. We have used a ceiling culture method for human mature adipocytes which uses their buoyant property to allow them to adhere to a floating glass surface, where they remain viable for several weeks. Using confocal immunofluorescence microscopy we showed the cellular expression and subcellular localization of leptin in ceiling-cultured adipocytes. The secretion of leptin was increased from ceiling cultures following tumour necrosis factor-treatment. Proliferation of mature human adipocytes in serum-containing medium was demonstrated by incorporation of bromodeoxyuridine, 2% of adipocytes showing positive incorporation after 4 h labelling. Proliferation was also evident from the budding of daughter cells. Apoptosis in the ceiling cultures was increased by 48 h serum deprivation (30-35 vs 10-15% in the control) and was assayed by propidium iodide staining and terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein nick-end labelling. Lipolysis, analysed by liquid scintillation counting, was increased by forskolin (10 µM for 90 min) and lipogenesis, shown by autoradiography, was stimulated by insulin (10 and 100 nM for 4 h). These findings indicate that ceiling-cultured adipocytes maintain adipocyte-specific functions and that ceiling culture, which overcomes the shortcomings of adipocyte suspension culture, can be used to study adipocyte cell biology.